TRENDS IN BIOSCIENCES JOURNAL 5-1 2012 EDITION

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Trends in Biosciences
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Trends in Biosciences
Volume 5 No. 1 April, 2012
CONTENTS
MINI REVIEW
1. Indian Patent Act 2005 in Perspective of Biotechnological Researches, Health and Industry 1
Praveen Gupta and Anil Kumar Dhiman
2. An Emerging Scenario of Long-term Changes in Balanced Dietary Pattern and Food Requirement in Uttar Pradesh 4
Anil Kumar, B.S. Sachan, Jitendra Kumar and M.M. Rajput
RESEARCH PAPERS
3. Generation of Expressed Sequence Tags (ESTs) from Banana Fruit Tissue 8
Madhusudhan, R, Vivek B.C., Manjunath, B and Lalitha Anand
4. Genetic Variability and Character Association in Turmeric (Curcuma longa L.) 11
Abhishek Pratap Singh, V. P. Pandey, S. M. A. Rahman and Rashid Pervez
5. Induced Desynapsis in Capsicum annuum L. 14
Mohd Gulfishan and Ainul Haq Khan
6. Impact of Abiotic Factors on Population of Acridoid Fauna (Orthoptera) in Aligarh Fort, Uttar Pradesh, India 17
Md. Humayoon Akhtar, Mohd. Kamil Usmani and Md. Rashid Nayeem
7. In Vitro Evaluation of Anti Bacterial Properties of Anacardium occidentale L. 20
T.G. Nagaraja and S.S. Shaikh
8. Effect of Sowing Methods, Input and Varieties Levels on Growth, Yield Components, Yield and 22
Nutrient Uptake of Durum Wheat (Triticum durum Desf.)
Piyush Jain
9. Effect of Season on Helminth Parasitic Prevalence, Dominance, Means Intensity and 25
Abundance in Some Fresh Water Scaly Fish
Krishna Singh and Abha Mishra
10. Life cycle, Population Index and Feeding Activities of the Lime Butterfly, Papilio demoleus 31
(Lepidoptera: Rhopalocera: Papilionidae)
Durgesh Nandni, Arun Raghuwanshi and Vinoy Kumar Shrivastava
11. In Vitro Quantification of B-Cell Proliferation in Kidney Culture of Channa striatus Exposed to Different 35
Doses of KOV Antigen of Aeromonas hydrophila, using Lymphocyte Transformation Test (LTT)
S.A. Mastan
12. Pathogenicity and Mass Production of Entomopathogenic Nematode, Heterohabditis indica on Major Insects 38
of Agricultural Importance
Rishi Pal, G.N. Tiwari and C.S. Prasad

13. Bioefficacy of Cow Urine Decoction (CUDs) of Different Plants on Population Growth of Lipaphis 41
erysimi (Kalt.) on B. juncea under Field Conditions
Wajid Hasan and M.S. Ansari
14. Effect of Different Formulations of Bacillus thuringiensis on Larval Mortality of Diacrisia obliqua 45
Zeenat Warsi and Ajay Capoor
15. An Empirical Study of Land Degradation its Magnitudes and Casualities 47
Anil Kumar, B.S. Sachan and Keshav Prasad
16. Comparative Efficacy of Three Chemical Products on the Root-Knot Development and Plant Growth 51
of Green Gram, (Vigna radiata L.)
Mohd. Yaqub Bhat, A.H. Wani and Naseer Hussain Shah
17. Effect of Some Aqueous Plant Extracts on Egg Hatching of Meloidogyne incognita (Kofoid and White) 54
Chitwood, the Root-Knot Nematode
Harpreet Kaur and Anu Katoch
18. Effect of Bacterial Preparations on Development of Diacrisia obliqua 57
Zeenat Warsi and Ajay Capoor
19. Determination of Antimycobacterial Activity of Polyenzyme Preparation Immunoseb-S using In Vitro Methods 61
Anita Joshi, Shilpa Risbud, Bhavana Pradhan, Smita Deshpande and Renu Bharadwaj
20. Genetic Divergence Analysis in Faba Bean (Vicia faba L.) 64
B.K. Chaubey, C.B. Yadav, V.K. Mishra and K. Kumar
21. Evaluation of Different Substrate for Mass Multiplication of Trichoderma spp. 68
Anuradha Singh, Mohd Shahid, Vipul Kumar and Mukesh Srivastava
22. Compatibility of Entomopathogenic Nematodes (Nematoda: Rhabditida) with Pesticides and their 71
Infectivity against Lepidopteran Insect Pest
Rashid Pervez and S.S. Ali
23. Prevalence of Grain Moulds in Sorghum growing areas of Northern Karnataka 74
Y.S. Mahesh, S. Lingaraju, B. Ravi Kumar, B.Manjunath and M.G. Palakshappa
SHORT COMMUNICATIONS
24. Genetic Diversity in Bread Wheat [Triticum aestivum (L.) Em. Thell] 77
Hasan Tanveer, Arvind Kumar and Hamveer Singh
25. Relation of Temperature and Humidity to Postharvest Rot of Chickpea Caused by Fungi 79
Sarita Joshi and Kamlesh Gupta
Erratum 80
Author Index (Vol. 4, No. 1&2, 2011) 81
Subject Index (Vol. 4, No. 1&2, 2011) 84
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Trends in Biosciences 5 (1): 1-3, 2012
MINI REVIEW
Indian Patent Act 2005 in Perspective of Biotechnological Researches, Health and
Industry
PRAVEEN GUPTA* AND ANIL KUMAR DHIMAN**
*Mewar University, Chittorgarh (Rajasthan), e-mail: praveenshrihari@rediffmail.com
Gurukul Kangri University, Haridwar (U.K.), e-mail: akvishvakarma@rediffmail.com
ABSTRACT
It has been a challenge to understand the patentability matters
in health care area in particular Biopharmaceuticals in India.
Major biopharmaceutical companies have increased their
investment in research and development from several million
US dollars to several billion dollars in past decades. Now, these
days big biopharma companies are focusing on extensive
analysis of patentability and placing of strategies on patent
portfolio management as it showed a pay off strategy in long
run in health care area. Recently, Government’s focus to address
the needs of human health on emerging diseases, formulation
of legislations, availability of off-patented biosimilars, tax-free
incentives made this sector as a top priority sector for investment
for many of global investors. Thus, outcomes from the research
and development departments in way of patents are critical for
the health of biopharmaceuticals. Globally big Indian
Biopharmaceutical companies are also keeping eyes to patent
their biological inventions to increase their longevity and
monopoly in the market. Unfortunately, there are no clear
criteria’s for patentable matters in biotechnological inventions
under Indian patent act 2005.
Key words
Indian patent law, biotechnology, biopharma,
patenting
A patent is a monopoly right granted to a person who
has invented a new and useful article or an improvement of an
existing article or a new process of making an article. It consists
of an exclusive right to manufacture the new article invented
or manufacture an article according to the inventive process
for a limited period (Seth, 2012). It is an exclusive right granted
by the government to the patentee in respect of a new
invention, which may be product or process. A patent is
granted by the patent office in which you wish to protect
valuable invention. But according to Shivashankar, et al., 2011,
the assignee only enjoys the rights, if the owner can assign or
license the invention to the assignee. The inventor needs to
disclose the invention in written form with description in order
to obtain exclusive monopoly over the invention for a specific
duration.
Sharma and Garg, 2008 state that an invention should
possess novelty, must have followed inventive steps and be
of industrial applicability for patenting. But an invention that
is frivolous; or its primary or intended use or commercial
exploitation that could be contrary to public order or morality
Mr. Praveen Gupta has done M.Sc.
in Biotechnology from Maharaja
Sayagirao University of Baroda
Gujrat and persuing Ph.D in
biotechnology from Mewar
University, Chittorgarh, Rajsthan.
Presently he is managing project in
Rheumatoid Arthritis and Fertility
research at Apcegen Technologies
Private Limited, Indian Institute of
Technology (IIT) Kanpur. He has worked on the projects
for development and manufacturing of 10,000+
recombinant rabbit monoclonal antibodies (r-MAbs) at
2-5L stir tank bioreactor scale; Business area cellular
analysis & biomarkers as Bioprocess Lead, at Life
Technologies Inc (USA), Bangalore,he was awarded for
outstanding work in “Development of Low Cost
Diagnostic Products-r-Mabs Production Centre” at Life
Technologies Inc, USA.
that cause serious prejudice to human, animal, or plant life or
health or to the environment; and mere discovery of a scientific
principle; mere discovery of any new property or new use of
known substance; or mere arrangement or re-arrangement or
duplication of known devices each functioning independently
of one another in known way; along with a method of
agriculture and horticulture; invention relating to atomic
energy; topography of integrated circuits; a presentation of
information; method of playing game; a mathematical or
business method; a computer programme per se; and any
process for the medical, surgical, curative, prophylactic or
other treatment of human beings or any process for a similar
treatment of animal to render them free of disease are not
patentable.
Patenting in India
Historically saying according to Seth, 2012, the first act
relating to patent rights was passed in 1856 (Act VI of 1856)
which granted certain exclusive privileges to inventors of new
manufacture for a period of 14 years. This act was re-enacted
with modifications under Act No. XVI of 1859. The provisions
of this act were modeled basis of the English Patent Act, 1852

2 Trends in Biosciences 5 (1), 2012
wherein patent monopolies were termed as “exclusive
privileges”. In 1872, the Patterns and Design Protection Act
was passed followed by Invention and Designs Act 1888.
Subsequently, Indian Patents and Designs Act was passed
replacing all previous acts. During the period from 1911 to
1970, various amendments were made to this act. Later on,
based on the interim report submitted by a committee headed
by Dr. Bakshi Tekchand, amendments were made to this act
by the Act 32 of 1950. Subsequently, in 1959 Ayyangar’s report
was submitted containing recommendations for effecting
radical changes to the Patent law prevailing in India.
Eventually, The Patents Act, 1970 was passed and it came
into force on 20 th April 1972.
India is also a signatory to the Paris Convention for the
protection of industrial property, 1883, and the Patent
Cooperation Treaty, 1970. The patents act provides that any
invention that satisfies the criteria of newness, nonobviousness
and usefulness can be the subject matter of a
patent (Zacharias and Farias, 2002).
On March 26, 1999, Patents (Amendment) Act 1999 came
into force from 1 st January 1995. According to this amendment,
it is now possible to make an application for patent claiming
for a substance itself intended for use or capable of being
used as medicine or drug excepting the intermediate for a
preparation of drug. Exclusive marketing rights would be valid
for a period of five years or till the date of grant of patent or
date of rejection of the application for the grant of patent
whichever is earlier.
On 20 May 2003, the Patents (Amendment) Act 2002
came into force which was having following key amendments:
l
l
l
Extended patent life to 20 years from the date of filing
those were come under term of patent under Section 16.
“Process” defined under S. 3(1) in case of plants, were
then patentable while a “process” in case of diagnostic
and therapeutic were considered as non patentable.
After this amendment, a method or process of testing
during the process of manufacture was patentable.
There were two new grounds for revocation:
l
l
The complete specification does not disclose or wrongly
mentions the source or geographical origin of biological
material used for the invention.
The invention so far as claimed in any claim of the
complete specification was anticipated having regard
to the knowledge, oral or otherwise, available within
any local or indigenous community in India or
elsewhere.
It is seen that India has been able to attract the intention
of international investors especially in Biopharmaceutical area
in past few years. There are examples where big Indian pharma
were able to make business deal with International giants.
Hence, the most recent amendment Patent act 2005, in which
Indian legislature has tried to make it compliance fully with
TRIPS – the Trade related Intellectual Property Rights, came
in to force. According this act– one can go for patent of new
form of known substance which can show enhancement of
the known efficacy under section 3 which mean; salts, esters,
ethers, polymorphs, metabolites, pure form, particle size,
isomers, mixtures of isomers, complexes, combinations and
other derivatives of known substance shall be considered to
be the same substance, unless they differ significantly in
properties with regard to efficacy and will not be eligible for
patentable candidates. However, at the same time Indian Patent
Act 2005 has played a vital role to protect innovator & public
interest in health care area.
Patentable Subject Matters in B iote chnological
Researches
The Principle of Indian Patent Act 2005 is that a patent
should be useful for industry, novel, non-obvious and most
importantly properly disclosed according to patent law.
According to this law following subject matters are possible
to patent in biotechnological researches:
l
l
l
l
l
l
l
l
l
l
l
l
l
This act promotes “Process Innovations” rather than a
“Product Innovations”.
Any “life form” such as micro-organisms, animal cells,
is not patentable however, “Processes” based on microorganism
are fully patentable.
“Process” in or for manufacturing of products for
example; process for increasing glycosylation in
Bioreactor.
“Method” for example; specific purification step to purify
specific or all biomolecules.
“Technology” for example; production of recombinant
monoclonal antibodies.
“Compositions” for example; animal cell culture medium
compositions.
“Efficacy Enhancement” for example; formulation of
biotherapeutics.
“Tests” for example; specific diagnostic tests.
“Research tools” for example; screening system of
biomolecules.
“Gene Sequence”, Genetic “optimized sequences”,
Chimeric Promoters Sequences.
“Expression Vectors back bones”, Specific Epitope
Sequence, SNPs.
“Gene Therapy” & “Therapeutic Protein Sensors”’ and
“Efficacious Improvement”, examples – knockouts,
transgenic, anti-sense, target specific expression.

GUPTA and DHIMAN, Indian Patent Act 2005 in Perspective of Biotechnological Researches, Health and Industry 3
Indian Patenting Trends in Biotechnology
Patents granted to Indian organizations during 1990-
2002 in different sectors easily reveals the fact that chemicals
and pharmaceuticals were the major areas in which Indian
organizations had obtained patents. However, it is also
observed that Indian organizations also got patents in
biotechnology. But majority of these were overlapping patents
addressing other sectors - mainly the pharmaceuticals. The
main technological domains of patenting activity in
biotechnology were in micro-organism compositions;
macromolecular compounds; and biocide and plant
reproduction techniques.
There has a steady increase in activities in biotechnology
from the different data available. An analysis of Indian
patenting activity during 2003-04 shows again that in this
period also, pharmaceutical and chemical sectors were the
dominant areas of patenting activity. Pharmaceuticals had 213
patents (46% of total patents) during these two years while
chemical sector had 125 patents (27% of total patents).
Biotechnology sector was also well addressed with 48 patents
granted during this period. Other major sectors contributed
insignificant number of patents (Malviya, et al., 2010).
Impact on Health Sector
Most people in India prefer for the local medications
like ayurveda etc. because of low level income. Besides, the
prices of medicines were raised too high so the common people
can not afford to buy the modern medicines and antibiotics.
Though, ayurveda is getting back the attention from majority
of the people world wide (Dhiman, 2006), but modern medicines
do/can do better in emergency cases and deadly diseases.
However, many of the new medical researchers are now
targeting developed countries with promising profits for
medicines for lifestyle diseases whereas developing countries
are still in need of basic health care except three sectors i.e.,
food processing, pharmaceutical and agrochemicals. Indian
patent act allows product patent only. Only in these three
sectors process patent is allowed, as on today. India has only
process patent regime with relation to pharmaceuticals product
(Shivashankar, et al., 2011).
Biopharmaceutical organizations pour resources into
R&D of various molecules for the benefit of mankind. The
development of a biopharmaceutical goes through a series of
permutations and combinations resulting in uncertainties
which could be many and substantial. Maximizing the certainty
that a research-based manufacturer can obtain enforce, defend,
and make full, legitimate use of intellectual property rights
including patenting is essential to maintain the cycle of
innovation for the benefit of public health. Otherwise, in the
absence of strong intellectual property rights at each stage of
the innovation cycle, promise of biopharmaceutical innovation
could be lost (Andrade, et al., 2007). This needs proper
attentions.
India is slowly moving into global markets and
competing with international quality standards and prices.
Although R&D is an important factor to ensure a competitive
edge in the international arena, the future of the Indian
biopharmaceutical industry hinges on patent protection. There
are areas where company’s executives are working on
development of strategic patent portfolio. Recently, Nomura
Research Institute publish a methodology called “Technology
Heat Map” by which one can analyze the status of patent
application field with strategic step by step approach to select
specific technology fields (Masayuki, et al., 2004).
Moreover, recently forced Indian Patent Act has also
tried to compliance with TRIPS but still it fails to address the
problems of biotechnological researches where their
innovations are not encouraged with clearly defined guidelines
on biotechnological subject matter. Hence, more attention is
needed in this area so that biotechnological innovations could
prove themselves more useful to common people. But as
advocated by Dhiman, 2010, the ethics in biological researches
should not also be violated.
LITERATURE CITED
Andrade, C., Shah, N. and Chandra, S. 2007. The New Patent Regime
Implication for Patients in India. Indian journal of Psychiatry,
49(1): 56-59.
Dhiman, Anil Kumar. 2006. Ayurvedic Drug Plants, Daya Publishing
House, New Delhi.
Dhiman, Anil Kumar. 2010. Developing Trends and Emerging Needs of
Ethics in Biotechnology in Present Environment. In: ‘Advances
in Applied Biotechnology’ (eds Pradeep Parihar and Leena Parihar)
Agrobios India, Jodhpur. pp. 293-302.
Indian Patent History Information from Indian Patent Office Website.
Available at www.ipindia.nic.in/.
Malviya, Rishabha, Bhardwaj, Vineet, Srivastava, Pranati, Bansal,
Mayank and Sharma, Pramod Kumar. 2010. Biotechnological
Innovations Patent: A Review. International Journal of
Pharmaceutical Sciences Review and Research, 3(2): 131-33.
Masayuki, M., Yuji, M. and Keiichi, H. 2004. Strategic Intellectual
Property Portfolio management; Technology Appraisal by Using
the Technology Heat Map. NRI Papers No. 83. Nomura Research
Institute, Japan. Available at www.nri.co.jp/english/opinion/papers/
2004/pdf/np200483.pdf.
Seth, Karnika. 2012. History and Evolution of Patent Law–International
and National Perspectives. Available at www.sethassociates.com/
wp-content/.../history-and-evolution-of-patents.pdf
Sharma, Sudhir and Garg, Deepa. 2008. Patent Literacy A Techno-legal
Right for Scientific Community. Advanced Biotech, pp. 26-28.
Shivashankar, Murugesh, Mani, Dhandayuthapani and Mandal, Badal
Kumar. 2011. An overview on Intellectual Property Rights in
Pharmaceutical and Biotechnology Industries. Journal of Chemical
and Pharmaceutical Research, 3(2): 753-761.
The Patents (Amendment) Act. 2005. The Gazette of India, 15, 2005.
Available at ipindia.nic.in/ipr/patent/patent_2005.pdf.
Zacharias, Nilesh and Farias, Sandeep. 2002. Patents and the Indian
Pharmaceutical Industry. Business Briefing: Pharmatech, pp. 42-
47.
Recieved on 28-2-2012 Accepted on 3-4-2012

Trends in Biosciences 5 (1): 4-7, 2012
MINI REVIEW
An Emerging Scenario of Long-term Changes in Balanced Dietary Pattern and Food
Requirement in Uttar Pradesh
ANIL KUMAR 1 , B.S. SACHAN 2 , JITENDRA KUMAR 3 AND M.M. RAJPUT 4
1
Seed & Farm, C.S. Azad University of Agriculture And Technology, Kanpur
2
Deptt. of Agril. Economics, C.S. Azad University of Agriculture and Technology, Kanpur
3
Deptt. of Agril. Extension, C.S. Azad University of Agriculture and Technology, Kanpur
4
Deptt. of Agril. Economic B.N.V. College Rath Hamirpur (U.P.)
ABSTRACT
In this paper an attempt has been made to analyse the National
Sample Survey (NSS) data is respect of Uttar Pradesh and to
quantify the Nutritional food Problem in the state. The NSS
repo rt of the National Sample Survey Org anizations,
Government of India. The calorie and Protein content of food
items consumed per consumer unit is the main source of
information for this paper. It gives per capita food consumption
levels for 2009 and 2010, disaggregated by Rural and Urban
areas exhibited a more diversified food basket with significantly
higher levels of per capita consumption of edible oils, milk,
fruits and meat, fish and eggs increasing urbanization and
widening rural-urban disparity has reduced the consumption
per head for cereals and pulses. The consumption of cereals
per head declined while consumption for fruits and vegetables,
milk and eggs increased significantly. This shift has taken
place among both rural and urban consumers. The trend in
rising consumption of high value commodities has generated
higher growth in demand for edible oils, Horticulture and
livestock commodities. A structural shift balancing the food
production and demand. Structural changes in consumption
pattern, growth in the economy as well as sizeable additions to
population have increased to demand for livestock products in
Uttar Pradesh in the year 2010.
Key words
Dietary patterns, food, consumptions, demand,
production, growth
As per the 2010-2011 N.S.S. survey we have 187 million
person below the poverty line, defined mainly in tones of
adequate food. That mean we can say we have 187 million
persons are hungry. At the same time we have 60 million tones
of food grains in government stock this is absurd. More than
that it is obscene. This is indeed a failure of markets, of policies
and politics.
In empirical analysis of the economic aspects of the
problems of deficient nutrition, attention has to be focused
mainly on how nutritional levels are related to levels of income
or assets, total expenditure and expenditure on food in the
case of specific target groups and for different regions in the
country. Analysis of the behaviour of food expenditure in
relation to the total expenditure or Income of household, or a
per capita basis, has been a part of the conventional analysIs
in this area of research. However, an equally important aspect
Dr. Anil Kumar presently serving
as Assistant Director Seed and
Farms at C.S. Azad University of
Agriculture & Technology, Kanpur.
He has done M.Sc. (Ag.) and Ph.D
in Agril. Economics from Bundelkhand
University, Jhansi. He has
published more than ten research
papers in National and International
repute journals.
Dr. B.S. Sachan presently serving
as Assitt. Professor, Department of
Agricultural Economics and
Statistics at C.S. Azad University of
Agriculture and Technology,
Kanpur. He has completed M.Sc.
(Ag.) in Agril. Economics from C.S.
Azad University of Agril. & Tech.
Kanpur and Ph.D. from C.S.J.M.
University, Kanpur. He has
published seven research papers in National and
International journals.
for examination is the behaviour of such variables as energy
value of food, protein in take, calorie-protein balance, and
food composition in relation to the levels of food expenditures.
The data
The National Sample Survey Organisation (NSSO)
collects data on household consumption expenditure in India
by adopting sample survey techniques. Our study uses the
household level data of National Sample Survey (NSS) rounds
38, 43, 50 and 55 pertaining to the periods 1993, 1997-98, 2003-
04 and 2005 for the Uttar Pradesh.
Demand analysis
The Food Characteristics Demand System (FCDS) model
which is based on individual’s demand for energy, variety,
and tastes of foods is used for computing the demand
elasticities for food items for rural and urban households

KUMAR et al., An Emerging Scenario of Long-term Changes in Balanced Dietary Pattern and Food Requirement 5
(Appendix Table 1). These demand elasticities are used in
projecting the consumption per household for various food
items under the assumptions that (i) per capita total
expenditure (income) will grow at the rate of 5%, (ii) pace of
urbanization will be consistent with the recent historical trend,
(iii) the average annual per capita consumption for 2005 (round
55) us taken as the base consumption.
The annual consumption of the commodity is predicted as:
d = d x (1 + y x e) t
Where, d 1
is annual per capita consumption of a
commodity in year t; d 0
is annual per capita consumption of
the commodity in the base year, y is growth in per capita
expenditure/income; and e is the expenditure elasticity of
demand for the commodity. The per capita projected
consumption in the year ‘t’ is multiplied with the projected
population to arrive at human demand is arrived at by adding
the direct (human consumption) and the indirect (seed, feed,
industrial use and wastage etc.) demand for food items.
Dietary pattern changes
Table 1 gives per capita food consumption levels for
1983 and 1999 disaggregated by rural and urban. Urban areas
exhibited a more diversified food basket with significantly
higher levels of per capita consumption of edible oils, milk,
fruits, and meat, fish and eggs. Increasing urbanisation and
widening rural-urban disparity has reduced the consumption
per head for cereals and pulses. Between 2009 and 2010, the
consumption of cereals per head declined, while consumption
for fruits, vegetables, milk, edible oils, and meat, fish and eggs
increased significantly. This shift has taken place among both
rural and urban consumers; the trend in ‘rising consumption
of high value commodities has generated higher growth in
demand for edible oils, horticultural and livestock commodities,
Demand for foodgrains
The consumer demand for rice, wheat, coarse cereals,
and pulses for the year 2001 to 2021 corresponding to 5%
income growth at constant prices is presented in Table 2. In
Table 1.
Change in consumption of food in Uttar Pradesh
(Annual per capita food consumption in kg)
Rural
Urban
Item 2006 2010 %
Change
2006 2010 %
Change
Rice 45.6 53.9 +18.1 28.0 35.0 +25.1
Wheat 124.2 109.3 -12.0 110.3 97.8 -11.3
Coarse cereals 18.3 3.4 -81.6 3.2 0.6 -79.8
Total cereals 188.1 166.6 -11.4 141.5 133.4 -5.7
Pulses 15.1 13.8 -8.6 11.6 12.1 +4.2
Edible oils 3.8 6.2 +63.3 5.4 7.4 +37.3
Vegetables 55.8 81.4 +45.7 54.0 88.3 +63.3
Fruits 2.3 10.2 +345.2 3.6 14.9 +319.8
Milk 43.3 68.2 +57.4 55.5 84.1 +51.5
Meat, fish & 1.7 3.3 +94.8 3.5 4.4 +26.2
eggs
Sugar 15.0 12.4 -17.7 13.6 13.1 -3.6
the year 2006 the human demand for foodgrains is expected to
be 33.6 million tonnes (mt) which might increase to 37.7 mt in
the year 2011 and 47.4 mt in the year 2021. Domestic demand
is arrived by adding the direct demand (human consumption)
and the indirect demand (seed, feed, industrial use and
wastage). In the year 2006, the total domestic demand for rice
is expected to be 9.4 mt for wheat it would be 22.4 mt coarse
cereals 2, 2 mt for pulses 4, 6 mt By 2021, demand for rice is
expected to increase to 13. 7 wheat 30.7 mt, coarse cereals 3
and pulses 7.74 mt. The foodgrains demand in Uttar Pradesh
will increase from a level of 34mt in the 2001 to 38.5 mt in 2006,
43.4 mt in 2011.
48.8 mt in 2016 and 55 mt in the year 2021. If production
of rice, wheat and coarse cereals continues to grow at the
same pace as in 1990’s then the state will be able to meet its
demand of population for these commodities till 2016. But in
pulses production in 2006 is 2.6 mt, which is even short of
demand for population of the state in 2010.
To meet the growing domestic needs, the average yield
at the state level is required to be improved by 66% for rice.
48% and 77% for coarse cereals and 101 % pulses over the
next two decades.
Demand for livestock and fisheries products
Structural changes in consumption patterns, growth in
the economy as well as sizable additions to population have
increased the demand for livestock products (milk. meat and
eggs). Table 3 show the expected domestic demand for
livestock products in Uttar Pradesh.
Thousand tonnes while domestic demand in 2001 is
estimated to be 281 thousand tonnes which would rise to 349
thousand tonnes in 2006. Meat and milk are among the most
important commodities of the consumption basket as income
level increases due to high expenditure elasticities (Appendix
Table 2.
Demand for foodgrains in Uttar Pradesh
Item
Production
2006 2011 2016 2021
Human demand (million tonnes)
Rice 8.75 9.94 11.27 12.76
Wheat 19.72 21.79 23.95 26.37
Coarse cereals 1.11 1.23 1.47 1.61
Total cereals 29.59 32.95 36.56 40.62
Pulses 4.01 4.77 5.70 6.77
Food Grains 33.60 37.72 42.26 47.39
Indirect demand (million tonnes)
Rice 0.59 0.68 0.78 0.89
Wheat 2.64 2.97 3.33 3.74
Coarse cereals 1.12 1.35 1.64 1.98
Pulses 0.56 0.67 0.81 0.97
Domestic demand for (million tonnes)
Rice 9.35 10.62 12.05 13.66
Wheat 22.37 24.76 27.28 30.12
Coarse cereals 2.24 2.58 2.99 3.46
Total cerea 33.95 37.95 42.32 47.23
Pulses 4.57 5.44 6.51 7.74
Food Grains 38.52 43.39 48.83 54.97

6 Trends in Biosciences 5 (1), 2012
Table 3.
Demand for livestock and fisheries in Uttar
Pradesh
Item
Production
2006 2011 2016 2021
Milk (Million Tonnes) 16.20 19.57 23.82 28.76
Meat (Thousand Tonnes) 349 433 544 676
Fish (Thousand Tonnes) 63 78 98 122
Eggs (Million Number) 669 831 1043 1296
Feed (Million Tonnes 3.03 3.69 4.52 5.49
Note: Weight of egg is taken 60 gm.
Table 1). Thus, state should immediately lay emphasis on
increase in production of these two commodities In 2006
demand for fish is estimated to be 63 thousand tonnes and for
eggs 669 million numbers. In the year 2021, demand will grow
to about 122 thousand tonnes for fish, and 1296 millions eggs.
The demand of livestock in next two decades will grow by
more than two folds. In fish production, Uttar Pradesh is a
surplus state but for eggs, efforts to increase production
should continue in order to meet the challenges of future
increase in demand. The production of livestock and fisheries
products must be improved by 115-240% by 2021 over the
year 2001. Based on the demand projection for livestock
products, the requirement for feed is projected. The feed
demand is estimated about 2.Smt in 2001, which will grow to
about 3. 7mt in 2011 and 5. in the year 2021.
Demand for edible oils
The consumption of edible oils has increased from 3.8
kg/capita/year in 1983 to 6.2 kg/capita/year in 1999 in rural
areas and from 5.4 kg/capita/year in 1983 to 7.4 kg/capita/year
in the urban areas of Uttar Pradesh (Table 1). The shift is quite
significant even in rural areas and reflects high growth in
demand for edible oils and oilseeds. Demand for edible oils of
0.89 mt in 2001 is projected to grow to about 1.72mt in 2021
(Table 4) To match the demand of edible oils. Uttar Pradesh
farmers may have to increase production of oil seeds from
2.8mt in 2001 to a level of 5.34 mt by the year 2021. In the
current situation the production of oilseeds has been 1 .3Omt
in 1999, whereas the demand of oil seeds in 2001 is more than
double of the production. Special emphasis need to be laid on
oil seeds production in order to make Uttar Pradesh meet its
own demand for edible oil.
Demand for horticultural products
Table 4.
Having achieved significant increase in production of
Year
Demand for edible oils and seeds in Uttar Pradesh
Edible oils
(million tonnes)
Required edible
oilseeds
Production
(Million Tonnes)
Production
2006 1.04 3.25
2011 1.23 3.83
2016 1.46 4.53
2021 1.72 5.34
cereals, the state has started paying much greater attention to
the horticultural crops. This has increased the availability of
horticultural products and its consumption in both rural and
urban households. The annual per capita ëonsumption of
vegetables has increased from a level of 56 kg in 1983 to 81 kg
in 1999 in rural and from 54kg in 1983 to 88kg in 1999 in urban
area. More than three fold increases in consumption of fruits
were observed from a level of 2.3 kg in 1983 to 10.2 kg in 1999
among the rural consumer and from a level of 3.6 kg in 1983 to
14.9 kg in 1999 in urban (Table 1). The domestic demand for
vegetables and fruits are presented in Tables 5 and 6
respectively. The human demand for vegetables and fruits,
which was 12.8mt for vegetables and 1.8mt for fruits in the
year 2001, is projected to grow to about 25.5 mt for vegetables
and 3.5mt for fruits in the year 2021. The production of
vegetables and fruits in 1999 is of the time of 13 .8mt and 3.2
respectively. Although for fruits its performance is good, still
constant efforts are needed to maintain this production level
and also to increase the productivity. To meet human demand,
the state needs to plan the production target at about 28.4mt
for vegetables and 5.5mt for fruits on or before 2021. This
means, the production of vegetables and fruits must be
enhanced about 100% by 2021 over the year 2001.
Demand for sugarcane and its products
The domestic demand for sugar of 2.4mt in 2006 is
projected to grow to about 3.7mt in the year 2021. The
sugarcane production target needs to be fixed at 39mt in the
year 2021 from a level of 21.7mt in the year 2001 (Table 7). In
sugarcane production Uttar Pradesh is having huge surpluses
and after meeting its own demand the surplus can be used for
domestic or international exports.
Growth in food demand
The annual growth in domestic demand for various food
items during the period 2001-1 1 and 201 1-21 are computed
and presented in Table 8. The results revealed that the demand
for cereals is likely to increase with an annual growth of 2%
for wheat, 2 .5% for rice, and 2.8% for coarse cereals Looking
at the production and demand scenarios for cereals. The
demand for non-cereal commodities will increase much faster
than the growth in population. The demand for horticultural,
livestock products will increase with annual growth 3.5% to
4.5 %. To meet the growing demand for edible oils and sugar,
the production of oil seeds and sugarcane must grow at more
than 3% annually.
Table 5. Demand for vegetables in Uttar Pradesh
Year
Human demand
(Million Tonnes)
Required edible
oilseeds production
(Million Tonnes)
Production
2006 15.15 16.83
2011 18.00 20.00
2016 21.49 23.88
2021 25.51 28.35

KUMAR et al., An Emerging Scenario of Long-term Changes in Balanced Dietary Pattern and Food Requirement 7
Table 6.
Year
Demand for fruits in Uttar Pradesh
Human demand
(Million Tonnes)
Required edible
oilseeds production
(Million Tonnes)
Production
2006 2.16 3.09
2011 2.61 3.72
2016 3.17 4.52
2021 3.81 5.45
Table 7. Demand for fruits in Uttar Pradesh
Year
Sugar
(Million Tonnes)
Required edible
oilseeds production
(Million Tonnes)
Production
2006 2.38 25.12
2011 2.76 29.11
2016 3.21 33.79
2021 3.71 39.10
Production, Demand and Trade
Table 9% presents a consolidated picture of production
of commodities, domestic demand for commodities and net
surplus or deficit with Uttar Pradesh for trade. The state is
surplus in the production of rice (4.68mt), wheat (5.76mt),
coarse cereals (1.8Omt), sugar (8.44mt), fish (142 thousand
tones) and eggs (2l3miflion no.). In these commodities present
Level of efforts and growth in production, will ensure sufficient
surplus even for next 20 years. In fruits and milk production,
the state is marginally surplus of the tune of 0.65mt and 0.2Omt
respectively, while, net deficit is seen in the production of
pulses (-1.24mt), oilseeds (-1.46mt), vegetables (- 0.3 7mt) and
meat (119 thousand tonnes).
A structural shift in dietary pattern towards livestock,
fisheries, and horticultural products is already underway and
is being predicted to intensify further. Decline in per capita
consumption of cereals and rapid increase in consumption of
edible oils, fruits, vegetables, milk, meat, eggs and fish is
observed. Food demand challenges ahead are formidable
considering the non-availability of favourable environment,
which prove past growth, and shrinking resource base. The
state enjoys a surplus status in rice, wheat, sugarcane, fruits,
fish and eggs. It is marginally surplus in milk production, but
falls short in suppiy of vegetables, pulses, oil seeds and meat.
Table 8.
Annual growth in demand for agricultural
commodities (in per cent)
Commodities 2001-11 2011-21
Rice 2.56 2.53
Wheat 2.03 1.96
Coarse cereal 2.84 2.94
Pulses 3.53 3.58
Foodgrains 2.38 2.37
Sugarcane 3.32 3.35
Vegetable 2.97 2.97
Fruits 3.49 3.53
Milk 3.80 3.86
Meat, Fish and Eggs 3.84 3.91
Feed 4.41 4.52
Rice 3.95 4.04
Table 9.
Production, demand and net trade in agricultural
items in Uttar Pradesh (in million tonnes)
Commodities Production Demand Net trade
Rice 12.91 8.23 +4.68
Wheat 25.98 20.22 +5.76
Coarse cereal 3.75 1.95 +1.80
Cereal 42.64 30.40 +12.24
Pulses 2.60 3.84 -1.24
Foodgrains 45.24 34.24 +11.00
Oilseed 1.30 2.76 -1.46
Sugar 10.50 2.06 +8.44
Vegetable 13.8 14.17 -0.37
Fruits 3.21 2.56 +0.65
Milk 13.60 13.40 +0.20
Meat (th t) 162 281 -119
Fish (th t) 193 51 +142
Eggs (th t) 752 539 +213
Appendix Table 1. Expenditure elasticities of demand for Uttar
Pradesh
Item Rural Urban All
Rice 0.100 0.144 0.123
Wheat -0.084 -0.088 -0.079
Coarse cereal -0.211 -0.265 0.103
Pulses 0.447 0.469 0.471
Milk 0.612 0.649 0.587
Edible oil 0.396 0.422 0.402
Vegetables 0.439 0.492 0.462
Fruits 0.627 0.655 0.572
Meat, Fish and Eggs 0.805 0.752 0.793
Sugar 0.294 0.289 0.276
Source : Estimated from 55 th NSS round household data Characteristic
Demand System (FCDS). Kumar, 1998.
Appendix Table 2. Expenditure elasticities of demand for Uttar
Pradesh
Year Uttar Pradesh Uttrakhand (new) Uttar Pradesh (old)
1991 132.06 7.05 139.11
2001 165.80 8.71 174.51
2006 185.04 9.56 194.60
2011 206.58 10.52 217.10
2016 229.89 11.45 241.34
2021 255.98 12.49 268.46
Greater emphasis is needed on diversification towards pulses,
oilseeds, milk and vegetables to meet the growing demand for
these commodities. Emphasis has to be laid on diversification,
dissemination of productivity increasing technology, and
adequate development of general and marketing infrastructure
which will help in balancing the food production and demand.
LITERATURE CITED
FAQ. 2003. The State of Food Insecurity in the World 2003, Rome,
36pp. FSA, (2002), “Creative strategy and Development”, Food
Standards Agency, September, London.
HPA. 2001. “Eating for Health: A survey of eating habits and young
people in Northern Ireland”, Health Promotion Agency, March.
Shetty, Prakash S. 2002, “Nutrition transition in India”, Public Health
Nutrition, 5(1 A): 175-182.
World Food Programme. 2002. Food Insecurity Atlas of Urban India,
M.S. Swaminathan Research Foundation and World Food
Programme, Chennai.
Recieved on 2.1.2012 Accepted on 1.4.2012

Trends in Biosciences 5 (1): 8-10, 2012
Generation of Expressed Sequence Tags (ESTs) from Banana Fruit Tissue
MADHUSUDHAN, R 1 , VIVEK B.C. 2 , MANJUNATH, B 3 AND LALITHA ANAND
1
Department of Biotechnology, University of Agricultural Sciences, G.K.V.K., Bangalore
2
Department of Genetics and Plant Breeding, University of Agricultural Sciences, G.K.V.K., Bangalore
3
Department of Plant Pathology, University of Agricultural Sciences, G.K.V.K., Bangalore
email: rsmadhu@gmail.com
ABSTRACT
In the present investigation, a range of ESTs associated with
developmental events during fruit ripening in Musa acuminata
were generated. Out of the 26 recombinants obtained from
normalized cDNA library, 17 inserts were sequenced. Sequence
data were compared against the non-redundant combined
databases using the program BLAST and functions were
assigned to the ETS sequences generated. The generated
sequences pertained to both housekeeping genes as well as
genes involved in ripening metabolic pathways. Three
sequences coding for ACC synthase, pectate lyase and
polygalactouronase which are all known to be directly involved
in fruit ripening were obtained. Scores of the ESTs generated
were in the range of 42 to 519. From the analysis, it was found
that ten ESTs out of 17 had significant homology. These ten
ESTs can be used for further studies as a tool to isolate a
particular gene from genomic DNA of any organism. ESTs
generated in the present investigation will also help in
identifying genes involved in ripening. The genes thus
identified can be utilized in genetic engineering studies to
enhance shelf life of fruits.
Key words
Banana, expressed sequence tag, cDNA library,
sequencing and plasmid
Bananas and plantains are important sources of
carbohydrate for the millions of people of developing world.
It is the second most important fruit crop next to mango and
regarded as poor man’s apple or fruit of heaven.
Ripening represents an important phase of the banana
fruit development since it is a climacteric fruit. Delaying the
ripening processes in banana fruit is of interest to producers
because it allows more time for shipment of fruit from the
farmer’s fields to the grocers shelf and increases the shelf life
of the fruit for consumers. Molecular approaches now provide
the key not only for identifying ripening related genes but
also for their manipulation for banana improvement. With the
advent of recombinant DNA technology, it is now possible to
genetically modify banana and develop transgenic banana
with extended shelf-life.
Generation of Expressed Sequence Tags (ESTs) is a
powerful tool that has emerged from genomics research. EST
collections can reveal gene expression pattern, gene regulation
and sequence diversity, now that enriched libraries and high
throughput sequencing are available and are efficient means
of gene discovery in focused metabolic situations. It is
important to use this approach to unravel the genes expressed
during ripening as post harvest losses is one of the important
aspects in case of banana fruit and this information can be
used to design strategies for genetic manipulation for
enhanced shelf life.
MATERIALS AND METHODS
The Mysore AAB (Natibale) - fully matured fruit bunch
was bought from farmer’s field, Hessaraghatta and was used
for RNA isolation. The banana hands were separated from the
bunch and cleaned. The hands were then treated with Ethrel
(1.4 ml in 1lt of water) to initiate the ripening process. Samples
were drawn from four banana fruits from the treated day itself
which served as ‘0` day sample. Sampling was done by
removing the outer pericarp of the finger and retaining only
the fruit pulp. The pulp was cut into small pieces and packed
in a polythene bag and sealed. Then the packed polythene
bags containing the sample were dipped in liquid nitrogen for
10 mins to cease the physiological activity and stored at -
80 0 C.
RNA was extracted from the samples following the
method of Lopez-Gomez and Gomez-Lin, 1992. Banana pulp
was ground in a tissue homogenizer in presence of lysis buffer
containing 0.2% SDS, 50 mM Disodium EDTA, 150 mM Tris
borate (pH 7.5). To this homogenate 0.25 volume of absolute
alcohol, 0.11 volume of 5 M Potassium acetate, 1% b-
mercaptoethanol were added and vertexed until the tissue
was fully homogenized. Equal volume of (49:1) chloroform:
isoamyl alcohol was added and centrifuged at 14,000 rpm for
20 min. To the supernatant, equal volume of (1:1) phenol:
chloroform was added and centrifuged at 14,000 rpm for 20
min. equal volume of (49:1) chloroform: isoamyl alcohol was
added to supernatant and centrifuged at 14,000 rpm for 20
min. Aqueous phase was carefully recovered and the RNA
was precipitated with 10 M LiCl (3 M final concentration) and
kept at -20 0 C overnight or at -80 0 C for 1 hr. RNA was pelleted
by centrifugation at 14,000 rpm for 45 min at 4 0 C and washed
once with 3 ml of 3 M LiCl. RNA was resuspended in 1 ml
sterile DEPC water; 5 M Potassium acetate (0.3 M final
concentration) and 2.5 ml of absolute alcohol was added and
kept at -20 0 C overnight or at -80 0 C for 1 hr. Centrifuged at
14,000 rpm for 40 min and then washed once with 2 ml of 75%
alcohol. The pellet was dried at 37 0 C and resuspended in 200

MADHUSUDHAN et al., Generation of Expressed Sequence Tags (ESTs) from Banana Fruit Tissue 9
ml of sterile DEPC water. Isolated RNA was checked for integrity
after each isolation by denaturing gel electrophoresis
containing formaldehyde.
mRNA was isolated from the total RNA using the
Genelute mRNA miniprep kit in which oligo (dT) beads were
used to bind to poly (A) tail of mRNA. mRNA bound to oligo
(dT) beads was eluted using elution buffer and then utilized
for cDNA library preparation.
Double stranded cDNA was synthesized from mRNA
following the protocol proposed by Frohman, et al., 1998.
First strand cDNA was synthesized by using mRNA as
template in presence of Reverse Transcriptase and terminal
transferase. Second strand was synthesized using First strand
cDNA as template in presence of Taq polymerase and dNTPs
in a PCR reaction. The success of cDNA synthesis was
confirmed by using synthesized double stranded cDNA as
template in a PCR reaction along with specific primers for
ACC oxidase, b-galactosidase and expansin genes.
The ds cDNA synthesized was cloned into vector
PTZ57R. This was then used for transformation into E. coli
using MBI FERMENTAS cloning kit.
The recombinant colonies were picked from cDNA
library plate and were inoculated into LB broth. Overnight
grown culture was used for isolation of plasmids. Plasmids
were isolated from the white colonies and run on the agarose
gel along with control plasmids with insert (952 bp) insert and
without insert. Plasmid samples, which were on par with the
control having insert, were recombinants and those plasmid
samples, which were on par with control plasmid having no
insert, were non-recombinants. The plasmid samples
confirmed to be recombinant were given for single pass
sequencing to the Department of Biochemistry, IISc, Bangalore.
Sequence data were compared against the non-redundant
combined databases using the program BLAST (n) provided
by the NCBI site.
RESULTS AND DISCUSSION
Yield of RNA obtained was found to be in the range of
3.5 – 7.7ìg /g of fruit tissue. Quality of RNA was found to be
good with an A 260
/A 280
ratio of around 1.7 – 2.0. Based on OD
values obtained, it was concluded that RNA obtained was of
fair quality however slightly contaminated with protein
fraction. Similar approach for isolation of RNA from banana
has been reported by Nascimento et al., 1997 where the isolated
RNA was used in studying the expression of banana sucrose
phosphate gene during development and ripening.
The success of cDNA synthesis was confirmed by using
this as a template in a PCR reaction along with specific primers
for ACC oxidase, b-galactosidase and expansin genes. The
PCR amplification products obtained were electrophoresed.
We obtained b-galactosidase band at 850 to 900 bp range,
ACC oxidase at 900 to 950 bp range and expansin in the range
of 800 to 850 bp. Martin, et al., 1998 have reported the use of
gene specific primers for identification and verification of
differential display cDNAs. To minimize false positives, the
strategy avoids the use of impure Northern blot probes
obtained from PCR-amplified DD bands. To increase
throughput, the cloning of DD bands is replaced by a genespecific
primer approach and hybridization arrays are used in
place of Northern blots. Construction of representative cDNA
library is an important step in generation of ESTs. The source
of the mRNA for the cDNA library in ESTs generation is critical
and varies depending on the goal of the study. Since the
objective of the present investigation was to generate ESTs
for ripening genes, mRNAs from banana fruit pulp during
ripening stage were selected as the source material for cDNA
library preparation.
The cDNA library obtained by following the method
supplied in the library construction kit of MBI FERMENTAS
had both blue and white colonies. Screening procedure was
very efficient to identify the recombinant clones. Blue colonies
represented non-recombinant clones and white colonies
represented recombinant clones. This was possible since
disruption in b-galactosidase gene allows blue white selection
from library plate when plated with X-gal and IPTG.
Plasmids were isolated from the white clones and run on
the agarose gel along with control plasmids with insert (952bp
insert) and without insert. In Fig. 1, lane 14 and 15 are control
plasmid with insert and control plasmid without insert,
respectively. Remaining lanes contain plasmid samples
isolated from white clones. From the gel profile we can deduce
that all the plasmid samples in the gel are recombinants except
the plasmid samples in lanes 21 and 23.
Out of the 26 recombinants obtained, we sequenced 17
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28
Fig. 1.
Gel profile showing plasmid samples run along with
control plasmid with 952 bp insert and control plasmid
without insert (Lanes 14 and 15 respectively) Lanes 1
to 28 (except 14 and 15) –indicates plasmids isolated
from white clones Lanes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,
11, 12, 13, 16, 17, 18, 19, 20, 22, 24, 25, 26, 27, and
28 are reco mbinants Lanes 21 and 23 are no n-
recombinants

10 Trends in Biosciences 5 (1), 2012
inserts. On comparison with known sequences in the database
using BLAST (n), we found that these sequences
corresponded to 17 known plant sequences. We have therefore
been able to generate 17 ESTs from banana fruit.
ESTs have been generated in many plant species at
different developmental stages or from different plant parts.
5548 ESTs from Stevia rebaudiana (Brandle, et al., 2002), 1316
ESTs from castor (Lange, et al., 2000). 800 from Medicago
trunculata (Covitz, et al., 1998), 110 from Lotus japanicus
(Szczyglowski, et al., 1997), 5692 from poplar (Sterky, et al.,
1998), 130 from Zea mays (Keith, et al., 1993) and more than
176915 ESTs from Arabidposis thaliana.
Although there are no published reports of generation
of ESTs in banana 27 ESTs have been reported in the dbEST
of NCBI (Gupta, et al., personal communication). The sizes of
the ESTs generated were in the range of 104-291bp. Putative
functions have been assigned for only seven of these. This is
probably because of the small size of the ESTs generated.
The sizes of the ESTs generated are in the range of 100-
627bp. These studies, have been able to assign putative
functions to all the sequences generated. It can be seen that
these sequences encode for both house keeping genes as
well as genes involved in metabolic pathways. Three
sequences coding for ACC synthase, pectate lyase and
polygalacturonase were obtained which known to be directly
involved in fruit ripening.
Since in this study a normalized cDNA library used which
have not been able to identify gene sequences that are
predominantly expressed during ripening. Normalized cDNA
libraries are used to reduce the frequency of highly expressed
genes and to increase the rate of gene discovery (Soares, et
al., 1994). As a result we have been able to identify some
development specific genes and transcription factor like tif3
gene and MADs-box protein gene.
This has great significance since it is now becoming
increasingly evident that manipulation of genes coding for
transcription factor offers better regulation of events like fruit
ripening which are orchestrated by several genes (Giovannoni,
2001).
The score is the sum of the integer values of the aligned
aminoacids from the region of the sequences with highest
similarity. Each of the edited sequences was deposited in
dbEST and compared to the non-redundant nucleotide
sequences database by BLAST (n) searches. Deduced
aminoacid sequence homology between an EST and a known
sequence was deemed significant if the BLAST score was
greater than 80.
Empirical studies have generally indicated that BLAST
(x), BLAST (n) and TBLAST (n) scores of approximately 80 or
higher are worth for further investigation (Pearson, 1991).
In the present investigation, scores obtained pertaining
to ESTs generated is in the range of 42 to 519. Based on the
score analysis ten of the ESTs out of 17 had significant
homology. These ten ESTs can be used for further studies as
a tool to isolate a particular gene from genomic DNA of any
organism.
LITERATURE CITED
Brandle, J.E., Richman, A., Swanson, A.K. and Chapman, B.P. 2002.
Leaf ESTs from Stevia rebaudiana: a resource for gene discovery
in diterpene synthesis Plant Molecular Biology, 50 : 613-622.
Covitz, P.A., Smith, L.S. and Long, S.R.1998. Expressed Sequence Tags
from a root hair enriched Medicago truncatula cDNA library. Plant
Physiology, 117 : 1325-1332.
Giovannoni, J.J. 2001. Molecular biology of fruit maturation and
ripening. Annual Review of Plant Physiology and Plant Molecular
Biology, 52:725-749.
Keith, C.S., Hoang, D.O., Barret, B.M., Feigelman, B., Nelson, M.C.,
Thai, H. and Baysdorfer, C. 1993. Partial sequence analysis of 130
randomly selected maize cDNA clones. Plant physiology, 101: 329-
332.
Lange, M.B., Wildug, M.R., Stauber, E.J., Sanchez, C., Pouchnik, D.
and Croteau, R., 2000. Probing essential oil biosynthesis and
secretion by functional evaluation of expressed sequence tags from
mint glandular trichomes. Proceedings of National Academy of
Science. USA, 97(6) : 2934-2939.
Lopez-Gomez, R. and Gomez-Lim, M.A.1992. A method for extracting
intact RNA from fruits rich in polysaccharides using ripe mango
mesocarp. Hortscience, 27(5): 440-442.
Martin, K.J., Kwan, C.P., O’hare, M.J., Pardee, A.B. and Sage, R.1998.
Identification and verification of differential display cDNAs using
gene specific primers and hybridization arrays. Biotechniques, 24(6):
1018-1026.
Nascimento, J.R.O., Cordenunsi, B.R., Lajolo, F.M. and Alcocer,
M.J.C.1997. Banana sucrose-phosphate synthase gene expression
during fruit ripening. Planta, 203: 283-288.
Pearson, W.R. 1991. Identifying distantly related protein sequences.
Current Opinion in Structural Biology, 1: 321-326.
Soares, M.B., Bonaldo, M.F., Jelene, P., SU, L., Lawton, L. and
Efstratiadis, A.1994. Construction and characterization of a
normalized cDNA library. Proceedings of National Academy of
Science, USA, 91: 9228-9232.
Szczyglowski, K., Hamburger, D., Dapranov, P. and Bruijin, F. 1997.
Construction of a Lotus japonicus late nodulin expressed sequence
tag library and identification of novel nodule-specific genes. Plant
Physiology, 114 :1335-1346.
Recieved on 30-11-2011 Accepted on 25-01-2012

Trends in Biosciences 5 (1): 11-13, 2012
Genetic Variability and Character Association in Turmeric (Curcuma longa L.)
ABHISHEK PRATAP SINGH * , V. P. PANDEY , S. M. A. RAHMAN * AND RASHID PERVEZ ***
Department of Vegetable Science, N. D. University of Agriculture & Technology, Kumarganj Faizabad 224229
*
Krishi Vigyan Kendra, Gwaldam (Chamoli)-246 441, Uttarakhand
***
Indian Institute of Spices Research Calicut 673 012, Kerala
email : abhishek.veg@gmail.com
ABSTRACT
Studies on the genetic variability, heritability, genotypic
coefficient of variation, phenotypic coefficient of variation,
genetic advance of the associated character of the turmeric at
different site and estimate the curcumin and oleoresin content,
are evaluated in two seasons 2007-08 and 2008-09. Significant
differences were observed for sixteen characters among
seventeen genotypes in turmeric (curcuma longa L.). The
phenotypic coefficient variation (PCV) was higher than the
genotypic coefficient of variation (GCV) estimates for all the
traits studied. The high heritability accompanied with a high
genetic advance in per cent of mean was recorded for weight of
mother rhizome and weight of fresh rhizome per plant while
high heritability along with moderate genetic advance was
recorded for number of tillers per clump, plant girth, length
and width of mother rhizome, number and weight of fresh
rhizome per plant, number and weight of secondary rhizome
per plant, rhizome girth, dry matter (%). Rhizome yield had
positive and significant correlation with plant height, number
of leaves on main shoot, plant girth, length and width of mother
rhizome, number and weight of primary rhizome per plant,
number of secondary rhizome per plant and weight of fresh
rhizome per plant at phenotypic and genotypic levels, while
only one character viz number of tillers per clump showed
positively and significant association at genotypic level only.
The estimates of genetic parameters revealed scope for further
improvement of rhizome yield by selection.
Key words
Genetic variability, heritability, turmeric, Curcuma
longa
The genus Curcuma longa L. belongs to the family
Zingiberaceae, among the 40 species of curcuma, only two
species viz., Curcuma longa L. and C. aromatica Salisb. are
commercially produced. India is the largest producer, exporter
and consumer of turmeric in the world and accounts for more
than 46 per cent of the world trade.
Genetic variability is the backbone of any crop
improvement programme and effectiveness of selection
depends upon its nature and magnitude in the genetic material
at the disposal of plant breeder. Success in selection depends
primarily upon the magnitude of heritable portion of variability.
Heritability estimates provide information on transmission of
characters from parent to the progeny. Such estimate facilitates
evaluation of hereditary and environmental effect in
phenotypic variation and thus aid in selection. Heritability
estimates are used to predict expected advance under
selection so that breeders are able to anticipate improvement
from different selection intensity. Johnson, et al., 1955 have
suggested that heritability estimates in association with
genetic advance are much useful for selection than heritability
alone.
In view of this, the present study on the genetic
Table 1.
Analysis of variance for different characters of turmeric at different sites
S. Character
2007-08 2008-09
N.
Site 1 Site 2 Site 3 Site 4
R T E R T E R T E R T E
1. Plant height (cm) 29.3 500.7** 26.4 20.6 426.6** 36.4 28.5 453.6-7** 25.6 - 595.9** 17.7
2. Number of tillers per clump 0.2 2.7** 0.2 0.1 0.9** 0.08 0.1 0.9** 0.1 0.3 1.7** 0.1
3. Number of leaves on main shoot 0.2 2.2** 0.1 0.07 2.0** 0.2 0.1 0.9** 0.1 0.1 0.3** 0.02
4. Plant girth (cm) 0.1 3.0** 0.1 - 4.0** 0.2 0.7 7.4** 0.3 0.03 5.7** 0.3
5. Length of mother rhizome (cm) 0.03 5.1** 0.1 0.3 7.3** 0.1 0.09 6.9** 0.3 0.04 5.0** 0.04
6. Width of mother rhizome (cm) - 3.7** 0.2 0.02 2.1** 0.05 0.2 3.4** 0.1 0.3 1.3** 0.05
7. Weight of mother rhizome (g) 4.7 9174.7** 10.4 26.9 8044.5** 14.2 13.5 900.8** 5.6 25.6 8517.5** 9.2
8. Number of primary rhizomes/ plant 0.2 1.8** 0.08 0.07 1.2** 0.07 0.02 1.6** 0.1 0.05 1.5** 0.07
9. Weight of primary rhizomes/ plant (g) 148.8 4350.8** 82.3 0.6 2556.1** 2.1 23.9 2465.3** 11.7 3.2 2647.3** 10.8
10. Number of secondary rhizomes/ plant 0.7 20.0** 0.6 0.3 1.8** 0.1 0.1 3.0** 0.2 - 6.6** 0.2
11. Weight of secondary rhizomes/plant (g) 65.2 16582.9** 41.4 2.3 924.7** 4.3 19.3 1157.4** 32.3 22.6 1341.7** 17.5
12. Weight of fresh rhizome/plant (g) 24.7 131601.7** 439.8 28.6 91268.6** 51.4 1305.2 122465.1** 1218.1 4846.8 6203.0** 1667.1
13. Rhizome yield (q/ha) 23.8 6133.2** 26.7 1068.3 4670.8** 388.3 136.3 6059.2** 224.7 0.6 6117.6** 20.5
14. Dry matter (%) 0.05 41.7** 0.3 0.9 16.1** 0.4 - 34.2** 0.3 - 42.1** 0.3
15. Curcumin (%) 0.02 8.9** 0.2 0.02 8.5** 0.2 0.01 8.4** 0.1 0.3 8.1** 0.1
16. Oleoresin (%) - 32.4** 0.1 1.9 29.8** 1.9 0.2 32.3** 0.2 1.8 29.6** 2.1
*, ** Significant at 5% and 1% level, respectively, R- replication; T- treatment; E-error

12 Trends in Biosciences 5 (1), 2012
Table 2.
S.
No.
1. Plant height (cm)
Genetic parameters for yield and yield attributes in turmeric
Characters Site General
mean
2. Number of tillers per clump
3. Number of leaves on main shoot
4. Plant girth (cm)
5. Length of mother rhizome (cm)
6. Width of mother rhizome (cm)
7. Weight of mother rhizome (g)
8. Number of primary rhizomes
per plant
9. Weight of primary rhizome
per plant (g)
10. Number of secondary rhizome
per plant
11. Weight of secondary
Rhizomes per plant (g)
12. Weight of fresh rhizome
per plant (g)
13. Rhizome yield (q/ha)
14. Dry matter (%)
Range
Genotypic
coefficient of
variation
(GCV)
Phenotypic
coefficient
of variation
(PCV)
Heritability
(%)
Genetic
advance
Genetic
advance
in %
mean
1 97.22 72.50-118.89 11.20 12.38 81.8 20.28 20.86
2 97.18 72.28-110.48 10.16 11.90 72.8 17.36 17.87
3 96.95 73.12-112.70 10.67 11.87 80.7 19.14 19.74
4 88.78 67.83-113.79 13.54 14.34 89.1 23.37 26.32
1 3.52 2.41-5.51 22.90 25.41 81.2 1.49 42.51
2 3.24 2.35-4.11 14.44 16.91 72.9 0.82 25.39
3 3.70 2.50-4.78 12.83 14.41 79.2 0.87 23.52
4 3.29 1.65-4.76 18.95 22.08 73.6 1.10 33.47
1 9.45 7.92-10.93 7.67 8.509 81.2 1.34 14.23
2 8.91 7.22-10.05 7.62 9.12 70.1 1.17 13.17
3 7.91 6.97-8.99 5.99 6.90 75.3 0.84 10.71
4 8.09 7.55-8.73 3.18 3.74 72.3 0.45 5.57
1 5.16 3.83-7.51 16.43 17.98 83.4 1.59 30.89
2 4.62 3.24-7.77 21.16 23.17 83.4 1.83 39.797
3 5.70 4.20-10.51 23.27 25.44 83.7 2.50 43.84
4 5.49 3.76-9.31 21.16 23.21 83.1 2.18 39.73
1 6.17 4.46-10.72 18.21 18.96 92.2 2.22 36.01
2 5.71 4.62-11.52 23.46 24.29 93.3 2.66 46.68
3 6.22 4.35-10.80 20.76 22.35 86.3 2.47 39.72
4 5.77 3.98-10.19 19.40 19.78 96.2 2.26 39.20
1 3.76 2.12-7.10 25.01 27.49 82.8 1.76 46.88
2 3.13 2.35-5.62 22.81 24.014 90.2 1.39 44.64
3 3.91 2.42-6.98 23.25 25.17 85.3 1.73 44.25
4 3.51 2.77-5.23 15.88 17.25 84.7 1.05 30.11
1 54.93 21.19-251.18 87.13 87.33 99.5 98.38 179.09
2 49.78 16.11-243.26 90.01 90.32 99.3 91.97 184.77
3 39.36 20.60-290.00 38.00 38.47 97.6 30.44 77.33
4 48.48 21.24-247.14 95.13 95.34 99.6 94.80 195.56
1 5.96 4.69-7.22 11.10 12.10 84.1 1.25 20.98
2 3.50 2.73-4.84 14.98 16.95 78.1 0.95 27.28
3 3.57 2.60-5.26 17.11 19.19 79.6 1.12 31.45
4 3.56 2.58-4.90 17.03 18.71 82.9 1.13 31.95
1 95.35 38.31-175.92 34.25 35.55 92.8 64.83 67.99
2 68.69 41.75-172.22 36.78 36.84 99.7 51.96 75.65
3 75.25 51.99-179.52 32.91 33.22 98.1 50.539 67.16
4 71.44 41.91-175.24 35.93 36.23 98.4 52.45 73.42
1 7.68 4.91-16.50 28.72 30.33 89.7 4.30 56.03
2 6.38 4.68-7.54 10.01 11.80 71.9 1.11 17.49
3 7.00 5.12-8.63 12.05 13.54 79.2 1.54 22.11
4 6.08 2.45-8.01 20.70 22.39 85.5 2.39 39.44
1 100.01 28.48-290.16 64.30 64.62 99.0 131.81 131.80
2 59.78 28.53-86.44 25.37 25.61 98.2 30.96 51.78
3 65.49 37.60-92.92 25.60 27.03 89.7 32.718 49.95
4 63.50 35.90-100.15 28.65 29.40 95.0 36.530 57.52
1 263.48 91.39-1036.98 69.86 70.33 98.7 370.55 142.96
2 263.17 221.73-892.61 57.38 57.44 99.8 310.73 118.07
3 288.75 153.23-1050.17 60.29 61.49 96.1 351.65 121.78
4 266.90 119.71-772.00 46.02 48.50 90.1 240.11 89.97
1 322.94 261.33-432.72 12.09 12.20 98.3 79.79 24.70
2 315.60 235.38-393.92 10.36 12.10 73.4 57.74 18.29
3 321.43 230.68-431.57 11.88 12.76 86.7 73.23 22.78
4 321.68 231.67-428.97 12.13 12.21 98.7 79.89 24.83
1 21.21 17.80-30.81 15.16 15.38 97.2 6.53 30.79
2 19.91 17.01-23.96 9.95 10.44 90.8 3.89 19.54
3 21.38 18.16-29.65 13.62 13.88 96.2 5.88 27.52
4 21.35 17.74-26.39 15.15 15.38 97.1 6.56 30.75

SINGH et al., Genetic Variability and Character Association in Turmeric (Curcuma longa L.) 13
variability, heritability, genotypic coefficient of variation,
phenotypic coefficient of variation, genetic advance of the
associated character of the turmeric at different site and
estimate the curcumin and oleoresin content.
MATERIALS AND METHODS
Seventeen genotypes of turmeric was evaluated in a
complete randomize block design with three replications at
two different locations i.e. Experiment Station of Vegetable
Science, Narendra Deva University of Agriculture and
Technology, Kumarganj, Faizabad (U.P.) and Krishi Vigyan
Kendra Farm, Masodha, Faizabad for two consecutive seasons
i.e. 2007-08 and 2008-09.
Turmeric rhizome was planted in the month of June at a
spacing of 30 x 20 cm as per routine agronomic practice was
followed for each location. Observations were recorded on
five random plants for each replication on plant height (cm),
number of tillers per clump, number of leaves on main shoot,
plant girth (cm), length of mother rhizome (cm), width of mother
rhizome (cm), weight of mother rhizome (g), number of primary
rhizomes per plant, weight of primary rhizomes per plant (g),
number of secondary rhizomes per plant, weight of secondary
rhizomes per plant (g), weight of fresh rhizome per plant (g),
rhizome yield (q/ha) and dry matter (%). The various genetic
parameters, oleoresin (%) and Curcumin (%) content was also
analysis as per standard procedure.
RESULTS AND DISCUSSION
Significant differences were observed for sixteen
characters among seventeen genotypes in turmeric (Curcuma
longa L.). The phenotypic coefficient variation (PCV) was
higher than the genotypic coefficient of variation (GCV)
estimates for all the traits studied (Table 1).
The high heritability accompanied with a high genetic
advance in per cent of mean was recorded for weight of mother
rhizome and weight of fresh rhizome per plant while high
heritability along with moderate genetic advance was recorded
for number of tillers per clump, plant girth, length and width of
mother rhizome, number and weight of fresh rhizome per plant,
number and weight of secondary rhizome per plant, rhizome
girth, dry matter (%).
The genotypic coefficient of variation for plant height
was maximum in site 4 (13.52%), while, it was minimum in site
2 (10.16%) and phenotypic coefficient of variation was
maximum in site 4 (14.34) and minimum in site 3 (11.87%). The
highest genotypic (22.90%) and phenotypic (25.41%)
coefficient of variations (GCV and PCV) were observed in site
1. It was found that the estimates of heritability ranged from
72.90 to 81.20% (Table 2). Similar results were reported by
Burton and de Vane, 1953.
Each site showed high estimates of heritability and
genetic advance in per cent of mean except in site 2, which
exhibited medium heritability and low genetic advance in per
cent of mean for no. of secondary rhizomes per plant. This
study supported by Pandey, et al., 2002 and Sinkar, et al.,
2005. The heritability in broad sense for the weight of
secondary rhizomes (g) was high in all the environments while
genetic advance in per cent of mean was medium in site 2, 3
and 4 and high in site 1. High heritability estimates were
recorded in site 1 and site 2, while high genetic advance in per
cent of mean was reported in all the sites for oleoresin (%)
(Table 3). This study supported by Jaha, et al., 2001 and
Shanmugasundaran, et al., 2000.
Table 3.
Curcumin and oleoresin contents in the turmeric
in the form of : Mean (Range)
S. Content
2007-08 2008-09
N.
Site 1 Site 2 Site 3 Site 4
1. Curcumin (%) 5.61
(3.24-8.63)
5.49
(3.18-8.46)
5.45
(3.17-8.40)
5.43
(3.08-8.26)
2. Oleoresin (%) 8.65 8.83 8.63 8.41
(3.15-13.61) (3.14-13.59) (3.12-13.60) (2.80-13.58)
ACKNOWLEDGEMENT
The authors are thankful to Dr. J. Dixit, Professor,
Department of Vegetable Science, N.D.U.A & T. Kumargang,
Faizabad for providing facilities and Dr. Mithlesh Pandey,
Programme Coordinator, KVK Masodha, Faizabad for
encouragement.
LITERATURE CITED
Burton, G. W. and De Vane, E.W. 1953. Estimation heritability in tall
fescue (Fistuea arundicea) from replicated clonal materials. Agric.
J. 45: 178-181.
Jaha, J. C., Duttam, S. and Chatterjee, R. 2001. Genetic variability,
heritability and correlation studies in turmeric (Curcuma longa
L.). Research on Crops, 2(2): 220-225.
Johnson, H. W., Robinson, H. F. and Comstock, R. E. 1955. Estimates
of genetic and environmental variability in Soybean. Agron. J., 47:
314-318.
Pandey, V. S., Pandey, V. P., Singh, T. and Srivastava, A. K. 2002.
Genetic variability and correlation studies in turmeric (Curcuma
longa L.). In: Proc. Nat. seminar on strategies for increasing
production and export of spices. Calicut, Kerala. Oct., 24-26. pp.
195-200.
Shanmugasundaram, K. A., Thangaraj, T. and Chezhiyan, N. 2000.
Variability, heritability and genetic advance studies in turmeric.
South Indian Hort., 48(1/6): 88-92.
Sinkar, P. V., Haldankar, P. M., Khandekar, R. G., Ranpise, S. A.,
Joshi, G. D. and Mahale, B. B. 2005. Preliminary evaluation of
turmeric (Curcuma longa L.) varieties at Konkan region of
Maharashtra. Journal of Spices and Aromatic Crops, 14(1): 28-
33.
Recieved on 10.2.2012 Accepted on 12.4.2012

Trends in Biosciences 5 (1): 14-16, 2012
Induced Desynapsis in Capsicum annuum L.
MOHD GULFISHAN* AND AINUL HAQ KHAN
Cytogenetics and Mutation Breeding Laboratory, Department of Botany, A.M.U. Aligarh 202 002
*e-mail: fishan.amu@gmail.com
ABSTRACT
The seeds of Capsicum annuum L. were treated with 0.5%
aqueous solution of MMS for 3, 5 and 6 h. During the cytological
studies 2 desynaptic plant were isolated at 6 h treatment. In
plant-1, average number of univalents and bivalents per cell
were 13.00 and 6.50, at diakinesis and 15.80 and 4.52 at
metaphase-I, while in plan-2 it was 6.12 and 9.20 at diakinesis
and 10.80 and 7.80 at metaphase-I. At anaphase, equal
distribution of chromosome was observed only in 7.89 and
11.42% pollen mother cells (PMCs) in plant-1 and plant-2
respectively. Rest of PMCs shows irregular distribution of
chromosomes to the poles. Laggards at anaphase and
micronuclei at telophase were also recorded. The desynaptic
obtained during the present investigation, fit into the medium
strong type with reduced pollen fertility.
Key words
MMS, pollen fertility, desynapsis, laggards,
micronuclei.
Desynapsis is the cytogenetic phenomenon where the
homologous chromosomes pair at zygotene and pachytene
but fail to remain paired during subsequent stages resulting
in reduction in chiasma frequency or complete failure of
chiasma formation. In many cases, it is known to be governed
by a single pair of recessive gene or induced by either
environmental factors or by mutagens or brought about by
an interaction of the genotype and the environment or very
rarely by a dominant gene (Koduru and Rao, 1981). Desynapsis
during meiotic division has been reported in many plants such
as pearl millet (Singh, et al., 1977), chilli (Katiyar, 1977), rice
(Reddi, et al., 2000), Cicer arietinum L. (Kumar and Sharma,
2001), barley (Singh, 2002) and in Vigna mungo (Kumar and
Kesarwani, 2003). During the present investigation, the
desynaptic mutants has been isolated in Capsicum annuum
L. var. Pusa jwala, which is one of major spice crop of India
and of the world as well. The present paper reports the
cytogenetic characteristics of 2 induced desynaptic mutants.
MATERIALS AND METHODS
The seeds of Capsicum annuum L. var. Pusa jwala
obtained from IARI New Delhi, were treated with 0.5% freshly
prepared MMS solution for 3, 5 and 6 h. The seeds were
washed thoroughly in running tap water and excess of
moisture was blotted off and then sown in earthen pots. The
controls were maintained separately. At maturity, the young
flower buds of appropriate sizes were fixed in freshly prepared
Carnoy’s solution (Absolute Alcohol : Chloroform : Glacial
Acetic Acid in 6:3:1 ratio) for 24 hr and stored in 70% alcohol.
The anther were squashed in 2% propionocarmine
(Swaminathan, et al., 1954). The slides were made permanent
using n-Butyl Alcohol Schedule (Bhaduri and Ghose, 1954).
Studies on the pollen sterility were also conducted by staining
the pollen grain with 2% propionocarmine. Undersized and
empty pollen grains were considered to be sterile.
RESULTS AND DISCUSSION
During the present investigation, meiosis was perfectly
normal in control plant and12 bivalents were regularly formed
at diakinesis and metaphase (Fig.1a, b) followed by normal
segregation at anaphase. The advent of diakinesis in mutant
plant is marked by abrupt separation of paired chromosome
and many of them were observed as univalents (Fig.1c). In
plant-1 only univalent and bivalent were recorded while in
plant-2 associations of three and four chromosomes in
addition to bivalent and univalents in some pollen mother
cells (PMCs) and only univalents and bivalents in others were
observed. Metaphase-I was also irregular in both the
desynaptic plant. The univalents were not restricted to the
Table 1.
Mean and range of chromosomal associations at diakinesis and metaphase-I and pollen fertility in the desynaptic
mutants of Capsicum annuum L. var. pusa jwala
Plant 1 Diak. 130 13.00±0.36 6.50±0.45 _ _ 15.00
(2-17) (2-11)
Met. 150 15.80±0.64 4.52±0.28 _ _
(1-21) (1-10)
Plant 2 Diak. 130 6.12±o.39 9.20±0.24 0.55±0.06 0.13±0.03 18.00
(2-12) (3-15) (1-4) (0-3)
Met. 150 10.80±0.23 7.80±0.21 0.20±0.07 0.10±0.05
(1-18) (4-11) (1-4) (0-1)

Table 2.
MOHD GULFISHAN AND AINUL HAQ KHAN, Induced Desynapsis in Capsicum annuum L. 15
Chromosome distribution at anaphase-I in two desynaptic mutants of Capsicum-annuum L. var. Pusa jwala
Chromosome
Plant 1 Plant 2
Distribution No. of PMCs Per cent No. of PMCs Per cent
12:12 4 7.89 4 11.42
10:14 7 18.40 11 31.32
11:13 12 31.50 5 14.28
15:9 8 21.05 7 20.00
9:14+1L* 5 13.15 6 17.14
3:9+2 L* 4 10.50 3 8.57
L*= laggards.
metaphase plate instead scattered all over the cytoplasm
(Fig.1d, e) The univalents and bivalents varied considerably
from PMC to PMC in both plants and univalents occurred at
a greater frequency at metaphase-I than at diakinesis (Table
1). As a consequences of univalent formation at metaphase-I,
the following meiotic stages were also highly irregular. At
anaphase-I, normal 12:12 segregation was observed only in
7.89 and 11.42% in plant-1 and plant-2 respectively (Table 2),
while unequal and irregular distribution of chromosomes was
very common (Fig.1f). A variable number of laggards (Fig.1g)
and bridge were also noticed at anaphase-I. Telophase-1was
also not free from abnormalities and variable number of
micronuclei was observed (Fig.1h, i). Second phase of division
also exhibited abnormalities like bridge formation, stickiness,
Fig. 1.
Legends (a-i). a-b normal plant, c-i desynaptic plant.
1. Diakinesis showing 12 bivalents. 2. Metaphase-I
showing 12 bivalents. c-d. Univalents and bivalents. e.
Multivalent along with univalents and bivalents. f.
Unequal separation of chromosome at anaphase. g.
Laggards at anaphase. h. Micronuclei at telophase. i.
Multinuclei at telophase-II.
lagging chromosome and micronuclei etc. Pollen grains were
of variable sizes and pollen fertility was very much reduced
ranging from 15.0 to 18.0% in the 2 desynaptic plants while in
control it was recorded as 85%.
Since chromosome pairing was normal and complete at
pachytene followed by falling apart of some or all homologues
at the subsequent stages, the mutants described here can be
considered as desynaptic and the occurrence of univalents at
diakinesis and metaphase I can be attribute to failure of chiasma
formation. Depending upon the frequency of chromosome
pairing at diakinesis and metaphase I, Prakken, 1943
distinguished desynapsis as weak, medium strong and strong/
complete. The mutant plant of the present study fits into
medium strong type which has been previously described in
safflower (Prasad and Prasad, 1983), Chilli (Raja Rao and Aniel
Kumar 1983), chickpea (Kumar and Sharma 2001). The
distribution of univalents at metaphase I in the desynaptic
plants may be either polar or equatorial and may be influenced
by the number of bivalents per cell. When the bivalents are
few, the univalents tend to be polar but when many bivalents
occur, the univalents tend to be at equatorial plate (Ostergren
and Vigfusson 1953). In the present case, the univalents were
randomly distributed and were not seems to be effected by
number of bivalents per cell. At anaphase, unoriented and
disturbed plate formation leads to irregular chromosome
distribution to the poles (Kumar and Kesarwani 2003).
Presence of laggards was due to the abnormal spindle
formation and failure of chromosomal movement.
Thus, on the basis of the result it can be concluded that
desynapsis is a complicated phenomenon and, is due to a
variety of causes such as gene action, loss of chromosome
pairing, apomixes, structural and numerical changes of
chromosome in addition to environmental causes. In the
present study desynapsis was due to gene mutation because
meiosis of untreated and other treated plant did not show
such phenomenon despite grown in the same field, collected
at the same time, either same or another day. Environmental
effect is ruled out because both the normal and desynaptic
plant were grown under similar environmental conditions. Such
desynaptic plant might be useful for physiological studies on
chiasma formation and crossing over and in establishing
aneuploid lines for basic studies.

16 Trends in Biosciences 5 (1), 2012
LITERATURE CITED
Bhaduri, P.N. and Ghosh, P.N. 1954. Chromosomes squash in cereals.
Stain Technology, 29:269-276.
Katiyar, R.B. 1977. Radiocytogenetical studies in Capsicum: Induced
desynapsis. Caryologia, 30:347-350.
Koduru, P.R.K, and Rao, M.K. 1981. Cytogenetics of synaptic mutants
in higher plants. Theoretical and Applied Genetics, 59:197-214.
Kumar, G. and Kesarwani, S. 2003. Genetic analysis of induced desynaptic
mutant in Vigna mungo L. var. PDU.1. The Nucleus, 46:107-109
Kumar, G. and Sharma, V. 2001. Induced desynapsis in Cicer arietinum
L. Journal of Cytology and Genetics, 2:123-127.
Ostergren, G. and Vigfusson, E. 1953. On the position correlation of
univalents and quasi bivalents formed by sticky univalents. Hereditas,
39:33-50.
Prakken, R. 1943. Studies of asynaptics in rye. Hereditas, 29:475-495.
Prasad, G. and Prasad, J. 1983. Induced desynapsis in Safflower. Indian
Journal of Genetics and Plant Breeding, 43:276-278.
Raja Rao, K.G. and Aniel Kimar, O. 1983. Cytogenetics of spontaneous
desynaptic mutant in chillies (Capssicum annuum L.). Cytologia,
48:195-199.
Reddi, T.V.V, and Rao, D.R.M. 2000. Cytology of induced desynaptic
mutants in rice. Cytologia, 65:35-41.
Singh, R.B., Singh, B.D. and Vijayalakshmi, R.M. 1977. Meiotic behavior
of spontaneous and mutagen induced partial desynaptic plants in
pearl millet. Cytologia, 42:41-48.
Singh, M.R. 2002. Cytogenetic studies of induced desynaptic mutants
in Barley (Hordeum vulgare L.). Cytologia, 67:129-133.
Swaminathan, M.S., Magoon, M.L. and Mehra, K.L. 1954. A simple
propionocarmine PMCs smear for plant with small chromosomes.
Indian Journal of Genetics and Plant Breeding, 14: 87-88.
Recieved on 16.2.2012 Accepted on 18.4.2012

Trends in Biosciences 5 (1): 17-19, 2012
Impact of Abiotic Factors on Population of Acridoid Fauna (Orthoptera) in Aligarh
Fort, Uttar Pradesh, India
MD. HUMAYOON AKHTAR, MOHD. KAMIL USMANI AND MD. RASHID NAYEEM
Section of Entomology, Department of Zoology, Aligarh Muslim University, Aligarh 202 002
email: usmanikamil94@gmail.com
ABSTRACT
A regular survey was carried out weekly during the period of
January to December 2011 from Aligarh Fort to record Acridoid
fauna. Their population have been compared with three abiotic
factors i.e., temperature, humidity and rainfall. Altogether 29
species representing 21 genera belonging to 3 tribes, 7
subfamilies and three families have been recorded from this
region. Out of 29 species Acrididae, Catantopidae and
Pyrgomorphidae are represented by 14, 10 and 5 species
respectively. Maximum number of specimens as well as species
have been recorded in the month of August, September and
October whereas minimum number of specimens and species
recorded in the month of December, January and February.
Acridoid population positively correlate with maximum
temperature, minimum temperature, minimum relative
humidity and rainfall but do not correlate with maximum
relative humidity.
Key words
Abiotic factors, Aligarh fort, Acrididae, Catantopidae,
Pyrgomorphidae
Aligarh Fort located in the city of Aligarh, Uttar Pradesh
situated on the Grand Trunk road and consists of a regular
polygon, surrounded by a very broad and deep ditch. Actually
the site is all natural but has maintained queue of hedges and
patches. There were grasses (Cynodon spp.) in the fort found
throughout the year. Aligarh a district of Uttar Pradesh, situated
at latitude 27.88°N, altitude 78.08°E having subtropical climate.
Average temperatures range between 28–33 °C. The monsoon
starts in late June continuing till early October, with high
humidity levels. Aligarh gets most of its annual rainfall of
800 milli metres during these months. Temperatures then
decrease, and winter starts in December which continues till
early February. Temperatures range between 12–16 °C. Winters
in Aligarh are generally mild, but fog and cold snaps may
occur.
Acrdoidea is one of the most important out of four
superfamilies of Caelifera which comprises five families out of
which Acrididae, Catantopidae and Pyrgomorphidae are
widely distributed in India. Members of Acridoidea are
commonly known as locusts and grasshoppers. Grasshoppers
are of great economic importance, because they constitute an
important group of pests and pose a constant threat to cereal
crops, pulses, vegetables, orchards, grassland and forest
plantations all over the world. Some grasshoppers cause
significant damage to tree seedlings and agricultural crops
(Joshi, et al., 1999). Most grasshoppers are oligophagous
and exhibit definite host preferences (Mulkern, 1967),
according to which grasshoppers are classified as grassfeeders
(graminivorous), forb-feeders (forbivorous) or a mix
of the two (ambivorous or mixed feeders).
MATERIALS AND METHODS
Weekly observations were recorded during the period
of January to December 2011 to assess the distribution and
diversity of grasshoppers in Aligarh fort. Grasshoppers were
collected through net sweeping and hand picking method
and killed using bottle having ethyl acetate. For the purpose
of correct identification and photography the grasshoppers
were stretched, pinned and labeled and examined under stereo
microscope and later kept in store boxes and cabinets for
further studies. For detailed study of genitalia permanent
slides were prepared and drawn it with the help of Camera
Lucida. Meterological records were obtained from Department
of Physics, Aligarh Muslim University, Aligarh. The number
of specimens collected from three sites has been shown in
Table 1, 2, and 3 respectively. Diversity was calculated by
using formula provided by Shannon - Wiener through
SPECDIV program.
H = pi log 2pi
Where, H = Diversity index, Pi = ni/N is the probability
of an individual to belong to a species.
ni = Number of individuals of each species in the sample.
N = Total number of individuals of all species in the samples.
RESULTS AND DISCUSSION
Twenty nine species representing 21 genera belonging
to 3 tribes, 7 subfamilies and 3 families have been recorded
from this region (Table 1). Maximum of 720 specimens have
been recorded in the month of September at 34.8°C and lowest
at 22.4°C while minimum number recorded is 122 in the month
of January at temperature 23.5°C and lowest at 3.4°C (Table 2).
With gradual increase of temperature and relative humidity,
their population also increases but the result shows that 35°C
is optimum temperature for grasshopper and locust population
in Aligarh. Mondal and Shishodia, 1982 also recorded maximum
population at 29°C from Calcutta. Out of total Acridoid
population family Acrididae constitutes maximum of 48%,

18 Trends in Biosciences 5 (1), 2012
Table 1. Population of Acridoid fauna in Aligarh Fort 2011
Months
Temperature (°C) Humidity (%)
Rainfall
No. of
Maximum Minimum Maximum Minimum (mm)
grasshoppers
January 23.5 3.4 97 53 0.00 122
February 29.6 7.5 98 51 57.0 125
March 35.6 10.0 93 30 3.6 140
April 43.0 15.8 74 19 16.0 369
May 44.0 20.8 73 22 45.0 215
June 44.4 23.4 98 26 98.0 140
July 37.4 24.8 98 46 365.8 480
August 35.8 24.4 98 58 223.4 720
September 34.8 22.4 97 52 85.6 628
October 34.6 15.4 81 36 0.00 602
November 32.0 11.6 98 51 0.00 275
December 28.8 4.6 97 49 0.00 206
followed by Catantopidae with 35% while least 17% by
Pyrgomorphidae (Figure 1.). This result correlates with Usmani,
et al., 2010 who described thirty three species of locusts and
grasshoppers from western Uttar Pradesh which constitutes
16 species of Acrididae, 13 species of Catantopidae and 4
species of Pyrgomorphidae. Grasshoppers of the family
Acrididae were recorded from Tamil Nadu. This result also
relates with Nayeem and Usmani, 2012 who have recorded 41
species of Acridoidea from Jharkhand. 17 species of acridids
were recorded by Julka, et al., 1982 from different sites of
Solan, Himachal Pradesh. Acrida exaltata was recorded
throughout the year which is in conformity of the result of
Susanta and Halder, 1998 from west Bengal but contrary to
the result of Azim, et al., 2010. Oedaleus abruptus not found
throughout the year which is contrary to the result of Khan
and Aziz, 1973 from Kashmir. Joshi, et al., 1999 collected total
seven species of Acrididae and two species of
Pyrgomorphidae from the moist deciduous forest in India.
Influence of three climatic factors i.e. maximum and
minimum temperature, relative humidity maximum and minimum
and rainfall on acridoid population fluctuation was analyzed
statistically. A study of coefficient of correlation shows that
population of acridoids positively correlate with maximum
temperature, minimum temperature, minimum relative humidity
Table 2. Relationship between Acridoid population and
climatic factors
Climatic factors
Cor-relation
coefficient (r)
Significance
(p)
Maximum temperature 0.149 ns
Minimum temperature 0.612 s
Relative humidity (Max.) -0.056 ns
Relative humidity (Min.) 0.287 ns
Rainfall 0.469 ns
Regression line.
Y= 36.86- 0.13x 1
- 9.49x 2
+ 18.30x 3
+1.09x 4
+0.86x 5
S= Significant at p (0.005), ns= Not significant, Y= Population of
Acridoids
x 1
= Maximum temperature, x 2
= Minimum temperature, x 3
= Relative
humidity (Max.),
x 4
= Relative humidity (Min.), x 5
= Rainfall
Figure 1. Composition of Acridoid fauna in Aligarh Fort
Figure 2. Species accumulation of Acridoids in Aligarh Fort
and rainfall but do not correlate with maximum relative
humidity. Maximum temperature, maximum & minimum relative
humidity and rainfall as individual factor have no signifance
positive or negative correlation with population of acridoids
but minimum temperature shows a significant positive
correlation (Table 3).
Maximum number of species have been recorded in the
month of August, September and October whereas minimum
number of specimens and species recorded in the month of
December, January and February (Figure 2.). Shannon
Diversity index (3.04) indicates richness of acridoid fauna in
Aligarh Fort. Fort is rich in grasses and bearing an undisturbed
ecosystem. Present findings indicate that undisturbed grass
ecosystem inside the fort supports a large number of
grasshopper species.

AKHTAR et al., Impact of Abiotic Factors on Population of Acridoid Fauna (Orthoptera) in Aligarh Fort, Uttar Pradesh 1 9
Table 1. Distribution of Acridoid fauna in Aligarh Fort during January - December 2011
Species January February March April May June July August September October November December
Acrida exaltata + + + + + + + + + + + +
Acrida gigentea + + + + + + + + + + + +
Phlaeoba infumata _ _ + + + + + + + _ _ _
Phlaeoba panteli _ _ + + + + + + + _ _ _
Orthoctha indica _ _ _ _ _ + + + + _ _
Ceracris nigricornis _ _ _ _ _ _ _ _ + + _ _
Trilophidia anuulata + + _ _ + + + + + + + +
Trilophidia repleta + + _ _ + + + + + + + +
Oedaleus abruptus _ _ + + + + + + + + + _
Oedaleus seneglensis _ _ + + + + + + + + + _
Aiolopus simulatrix _ _ _ + + + + + + + + _
Locusta migratoria _ _ _ _ + + + + + + _ _
Oedipoda miniata _ _ _ _ + + + + + _ _ _
Gastrimargus africanus _ _ _ _ + + + + + _ _ _
Chrotogonus trachypterus _ + + + + + + _ _ _ _ _
Chrotogonus armatus + + + + + + + _ _ _ _ _
Atractomorpha burri + + + + + + + + + + + +
Atractomorpha psittacina + + + + + + + + + + + +
Pyrgomorpha conica _ _ _ _ + + + + + _ _ _
Catantops karnyi + + _ _ _ _ _ _ + + + +
Catantops pinguis _ + _ _ _ _ _ _ + + + +
Schistocerca gregaria _ _ _ _ _ _ _ + + + + _
Eyprepocnemis alacris _ _ _ _ _ _ _ + + + + _
Heteracris littoralis _ _ _ _ _ _ _ + + + + _
Choroedocus robustus _ _ _ _ _ _ _ + + + + _
Spathosternum prasiniferum + + + + + + + + + + + +
Hieroglyphus nigrorepletus _ _ _ _ _ _ + + + + + _
Oxya velox _ _ _ _ _ _ _ + + + + _
Oxya fuscovittata _ _ _ _ _ _ _ + + + + _
ACKNOWLEDGEMENT
Authors are grateful to Prof. Irfan Ahmad, Chairman,
Department of Zoology, Aligarh Muslim University, Aligarh
for providing necessary facilities and also to Chairman,
Department of Physics, Aligarh Muslim University, Aligarh
for providing meteorological data of Aligarh.
LITERATURE CITED
Azim, M. Nayyar., Reshi, Shabir. A. and Rather. Ajaz Hassan. 2010.
Observation on the seasonal variation in population of three species
of grasshoppers (Orthoptera : Acrididae) of Kashmir Himalaya.
Journal of Entomological Research. 34 (4) : 259-264
Julka, J.M., Tandon, S.K., Halder, P. and Shishodia, M.S. 1982. Ecological
observation on grasshoppers (Orthoptera: Acridoidea) at Solan,
Himachal Pradesh, India. Oriental Insects, 16(1): 63-75.
Joshi, P.C., Lockwood, J.A., Vashishth, N. and Singh, A. 1999.
Grasshopper (Orthoptera: Acridoidea) community dynamics in a
moist deciduous forest in India. Journal of Orthoptera Research, 8:
17-23.
Khan, H.R. and Aziz, S.A. 1973. Observation on seasonal variation in
population of hoppers and adults of Oedaleus abruptus (Thunberg)
(Orthoptera : Acrididae). Indian Journal of Entomology. 35(4) :
300-305.
Mondal, S.K. and Shishodia, M. S. 1982. Population fluctuation of
grasshopper fauna in a field near Culcutta. Pro. Symp. Ecol. Anim.
Popul. Zool. Surv. India. 3: 127-132.
Mulkern, G.B. 1967. Food selection by grasshoppers. Annual Review
of Entomology, 12: 59-78.
Nayeem, M. R. and Usmani, M. K. 2012. Taxonomy and field
observations of grasshopper and locust fauna (Orthoptera:
Acridoidea) of Jharkhand, India. Munis Entomology & Zoology,
7(1): 391-417.
Prakash, C. Joshi., Jeffrey, A. Lockwood., N, Vashishth and A. Singh.
1999. Grasshopper (Orthoptera: Acridoidea) Community Dynamics
in a Moist Deciduous Forest in India. J. Orthoptera Res. 8: 17-23
Susanta, N. and Halder, P. 1998. Population dynamics of the grasshopper
Acrida exaltata (Walker) in the arid zone of West Bengal. Indian
journal of Interacademicia. 2(1-2) : 51-53.
Usmani, M.K, Khan, M.I. and Kumar, H. 2010. Studies on Acridoidea
(Orthoptera) of Western Uttar Pradesh. Biosystematica, 4(1): 39-
58.
Recieved on 16.2.2012 Accepted on 25.3.2012

Trends in Biosciences 5 (1): 20-21, 2012
In Vitro Evaluation of Anti Bacterial Properties of Anacardium occidentale L.
T.G. NAGARAJA 1* AND S.S. SHAIKH 2
1
Department of Agro Chemicals and Pest Management, Shivaji University, Kolhapur 416 004 (MS)
2
Department of Botany, Shivaji University, Kolhapur 416 004 (MS)
*e-mail: tgnagaraja2010@gmail.com
ABSTRACT
An attempt was made to study in vitro screening of anti bacterial
properties of leaf,stem and root of Anacardium occidentale L.in
acetone and ethyl acetate extract. Acetone and ethyl acetate
extract of leaf ,stem and root found to possess good bactericidal
properties against Bacillus megatarium, Micrococcus sp.,
Pseudomonas aeruginosa, Escherchia coli and Staphylococcus
aureus bacteria. While leaf,stem and root extracts in ethyl
acetate proves more bactericidal activity than acetone extracts.
Key words
Anti bacterial, Bacillus megatarium, Micrococcus sp.,
Pseudomonas aeruginosa, Escherchia coli
Anacardium occidentale Linn. a small much branched
evergreen tree, native to tropical America and cultivated in
the coastal parts of India, especially in the West Coast districts
of India, commonly called as East Indian Almond tree. The
liquid free nutshell contains several alkaloids such as
gallocatechin, narimgenin and pruning 6-o-p-coumarate. The
fruit contains anacardic acid, a phenol cardol, essential amino
acids, while the husk contains D-catechin, gallic, caffeic and
quinic acids, leucocyanidine and leucodelphinidin. The bark
and leaves are useful in ondontalgia andulitis, The fruit is
acrid, sweet, hot, antidiarrhoeal, aphrodisiac and anthelmintic,
cures vaata and kapha and is used as counter irritant, drugs
are prepared from shell oil.. Hence, the plant possesses multi
utility in the field of ayruveda, pharmaceutical and in traditional
medicine. Therefore, an attempt was made to study its anti
bacterial properties of leaf, stem and root in acetone and ethyl
acetate extracts in the laboratory.
MATERIALS AND METHODS
Freshly harvested leaf, stem and root of Anacardium
occidentale Lin were collected from Shivaji University campus
for experimental study. The collected samples were brought
to the laboratory, thoroughly washed in tap water and later by
distilled water, cut in to small pieces. Initially these small pieces
were shade dried for 48 hours, later dried in electric oven with
a temperature of 55-58 o C for consecutive 4 days. The dried
samples were than powdered with home grinder to fine powder.
This 20 gm of fine powder of leaf and stem were subjected for
extraction. Acetone and ethyl acetate used as solvents and
extraction was carried out by Soxhlet’s apparatus. Each of
these extracts were further evaporated by Rotary vacuum
evaporator till a gummy semi solid material was obtained. This
semi solid material is used for studying anti bacterial properties.
While studying anti bacterial properties the test bacteria
such as Bacillus megatarium, Micrococcus sp., Pseudomonas
aeruginosa, Escherchia coli and Staphylococcus aureus were
obtained from Department of Microbiology, Shivaji University,
Kolhapur and were maintained in Nutrient Agar and CDA
media. The bacterial suspension was prepared using saline
water and mixed with 100 ml of sterilized NA media with constant
shaking. This 20 ml of seeded medium was transferred to
sterile petriplates. After solidification, well or cup was scooped
with the help of cork borer 5 mm in diameter. The test solution
was poured in the wells with the help of sterilized pipettes.
Anti bacterial activity was carried out by agar well diffusion
method (Alice and Sivaprakasam, 1966). The cultures were
kept in incubator 28 o C for 48 hours and inhibition zone was
recorded in millimeters.
RESULTS AND DISCUSSION
The results were depicted in Table 1 among the five
bacterial species tested, a maximum zone of inhibition 20.1 mm
is recorded in acetone extract of leaf,stem and root of
Anacardium occidentale L followed by 15.2 mm against
Micrococcus sp. and Staphylococcus aureus. this indicates
that acetone extract of leaf,stem and root possess a good anti
bacterial potency. A parallel result was recorded by Shimpi, et
al., 2005 in Aristolechia bracteata and Vijayalakshmi Devi, et
al., 2011 in C.roseus. Simultaneously ethyl acetate extract of
stem, leaf and root also proves to possess good bactericidal
potency against Bacillus megatorium Micrococcus sp., and
Staphylococcus aureus (Table 1). A similar result was
documented by Nagaraja, et al., 2008 in Barringtonia
acutangula and Tejaswini, et al., 2011 in Tridax procumbens.
A negligible zone was recorded against Pseudomonoas
aeruginsa in leaf and stem extract prepared in acetone solvent.
Thus leaf and stem extract prepared in ethyl acetate reflects
powerful anti bacterial potency as compared to acetone extract
(Table 1). Therefore, the present study may helpful in preparing
different formulations of the plant product and may be used
as eco-friendly management of plant diseases.
ACKNOWLEDGEMENT
The author is very much thankful to authorities of UGC
(WRO) Pune for providing financial assistance and Coordinator
Department of Agro Chemical and Pest Management,

Table 1.
NAGARAJA and SHAIKH, In Vitro Evaluation of Anti Bacterial Properties of Anacardium occidentale L. 21
Acetone and ethyl acetate extracts of Anacardium occidentale L. against some bacteria
Sr. Test Bacteria
Zone of inhibition (Diameter in millimeter)
Acetone extract
Ethyl acetate extract
control leaf stem root control leaf stem root
1. Bacillus megatarium 5.1 10.0 15.1 09.1 8.1 22.1 17.1 14.5
2. Micrococcus sp. 5.1 15.2 20,1 21.1 8.1 20.1 30.2 15.2
3. Pseudomonas aeruginosa 5.1 11.0 13.1 10.4 8.1 21.3 19.1 16.2
4. Escherchia coli 5.1 12.1 08.1 15.2 8.1 16.1 17.2 23.3
5. Staphylococcus aureus 5.1 20.1 22.1 17.2 8.1 26.2 17.1 23.1
Shivaji University, Kolhapur for providing laboratory facilities.
LITERATURE CITED
Alice, D and Sivaprakasam, K. 1966. Fungicidal, Bactericidal and
Nematicidal Effect of Garlic-clove extract. Journal of Eco-Biology,
8(2): 99-103.
Nagaraja T. G., Sarang, S.V. and More, V.R. 2008. Anti microbial
activity of Barringtonia Acutangula, Bioinfolet, 5(4): 402-403.
Shimpi, S.R., Chaudhari, L.S., Bharambe, S.M., Kharche, A.T., Patil,
K.P., Bendre R.S. and Mahulikar, P.P. 2005. Evaluation of anti
microbial activity of organic extract of leaves of Aristolechia
bracteata, Pesticide Research Journal, 17(1): 16-18.
Tejaswini, K.B., Vishwanath Pradeep, K. Rudrama Devi, S. Shylaja and
K. Jyothsna. 2011. Phytochemical screening and anti microbial
activities of plant extract of Tridax procumbens, The Bioscan.,
6(2): 321-323.
Recieved on 18-2-2012 Accepted on 6-4-2012

Trends in Biosciences 5 (1): 22-24, 2012
Effect of Sowing Methods, Input and Varieties Levels on Growth, Yield Components,
Yield and Nutrient Uptake of Durum Wheat (Triticum durum Desf.)
PIYUSH JAIN
Directorate of Extension, MPUAT, Udaipur, Rajasthan
ABSTRACT
In order to evolve low cost, resource efficient production
technology for durum wheat, a field experiment entitled “Effect
of sowing methods, input and varieties levels on growth, yield
components, yield and nutrient uptake of durum wheat (Triticum
durum Desf.)was conducted on clay loam at the Instructional
Farm, Department of Agronomy, Rajasthan College of
Agriculture, Udaipur during the year 2002-03 and 2003-04.The
results showed that durum variety HI 8498 accumulated
maximum quantum of biomass m -2 and nutrients uptake (N, P
and K).These manifested in production of higher effective tillers
m -2 and improvement in yield attributes (test weight, grain
weight ear -1 and grain yield ear -1 ) with concomitant increase in
crop productivity in terms of grain and biological yield by 7.50
and 5.20 per cent over Raj 1555. Among sowing methods, cross
sowing significantly enhanced overall growth of the crop in
terms of biomass accumulation, RGR and CGR, effective tillers
and uptake of nutrients. Apart from increased operational cost,
adoption of cross sowing with higher inputs (125% SR + 125%
FD) gave highest productivity in term of grain (56.97 q ha -1 ) and
biological yield (125.80 qha -1 ) and also monetary returns (Rs
37904.80 ha -1 ) with B:C ratio of 2.50.Interaction effect between
sowing methods and input levels revealed that, cross sowing of
variety HI 8498 with 125% SR + 125% FD produced highest
yields (58.13 q grain and 66.94 q straw ha -1 ) and fetched net
returns of Rs. 38424 ha -1 with B:C ratio of 2.53. The adoption of
FIRB system did not provide any additional gain in productivity
and profitability over cross and line sowing.
Key words
Cross sowing, FIRBS, line sowing, input levels,
growth, yield, net returns
India is one of the major producers of durum wheat with
a total output of 2.5 million tones. The durum wheat accounts
for only eight per cent of total world wheat production. India
is one of the major producers of durum wheat with a total
output of 2.5 million tones. Generally durum growers are
growing their crop, as they adopt for aestivum. It has been
established that besides inherited genetic potentials,
productivity and quality of durum’s are markedly influenced
by agronomic environments. Thus, there is a need to develop
appropriate resource conservation technology, which can
increase production as well as improve quality of durum.
Among agronomic practices planting system plays a vital role
in increasing productivity per unit area, Several researchers
(Hussain, et al., 2003, Singh and Jat, 2002; Houssain, et al.,
2004) have reported benefits of cross sowing and Furrow
irrigated raised beds over conventional line sowing (22.5 cm
apart). Among agronomic variables, seed and fertilizer
accounts for higher cost. Hence, there is a need to evolve
cost effective technologies, which can increase efficiency of
these two inputs without reducing productivity.
MATERIALS AND METHODS
Performance of durum wheat (Triticum durum Desf.)
varieties under varying sowing methods and input levels was
conducted for two consecutive years during Rabi season of
the year 2002-03 and 2003-04 at the Instructional Farm,
Rajasthan College of Agriculture, Udaipur. The soils of the
experimental site were clay loam in texture, alkaline in reaction
(pH 8.18 to 8.21) and had bulk density of 1.46 to 1.45 Mg m -3 .
The soils were medium in available N and P and high in
available K. status (267 kg, 20.58 kg and 384 kg available N, P
and K ha -1 , respectively). The experiment consisted 2 durum
varieties (Raj 1555 and HI 8498), 3 sowing methods
(Conventional line sowing at 22.5 cm row distance, cross
sowing at spacing of 22.5 cm x 22.5 cm in both direction with
use of half seed rate in each direction, FIRB system having 2
wheat rows on each bed formed at a distance of 70 cm) and 5
input levels (Recommended 100% seed rate + 100% fertilizer
dose, 125% seed rate + 125% fertilizer dose, 100% seed rate +
75% fertilizer dose, 75% seed rate + 100% fertilizer dose and
75% seed rate + 75% fertilizer dose of recommendation). Thirty
treatment combinations comprising of two varieties and three
sowing methods in the main plots and five input levels in the
sub-plots were evaluated in split plot design with three
replications. The recommended seed rate was worked out on
the basis of 1000 grain weight of respective variety (Raj 1555
46 g and 50 g for HI 8498) keeping seed rate of 100 kg ha -1 for
test weight of 38 g and as per treatment seed rate was worked
out for sowing. Similarly, recommended dose of the region
was considered and doses were worked out as per treatments
(120 kg N + 60 kg P 2
O 5
ha -1 )
RESULTS AND DISCUSSION
Result of present investigation reveals that durum var.
HI 8498 accumulated significantly higher quantum of biomass
m -2 at each growth stage over var. Raj 1555. Thus when
compared to mean final dry weight of 1051.53 g m -2
accumulated by Raj 1555, Between the varieties, HI 8498 was
found more efficient, as it increased CGR and RGR between
maximum tillering–heading by 19.9 and 7.8 per cent. But later
on (heading – maturity), both the varieties did not show
perceptible variation in these parameters. Among yield
components, Raj 1555 produced higher number of spike lets
and grains ear -1 but effective tillers m -2 , test weight, ear weight
and grain weight ear -1 were significantly higher in HI 8498.

Table 1.
PIYUSH JAIN, Effect of Sowing Methods, Input and Varieties Levels on Growth, Yield Components, Yield and Nutrient 23
Effect of varieties, sowing methods and input levels on growth of durum wheat (Pooled for two years)
Treatments
Dry matter accumulation (g/m 2 ) CGR (g m –2 d –1 ) RGR (g g –1 d –1 )
Maximum
tilleging
At
heading
At
harvest
Max. tillering to
heading
Heading to
harvest
Max. tillering to
heading
Heading to
harvest
Varieites
Raj 1555 237.72 609.34 1051.53 11.07 12.26 0.0280 0.0152
HI 8498 262.78 675.33 1133.67 13.27 11.48 0.302 0.0131
CD (P=0.05) 6.81 17.33 27.54 0.64 0.70 0.0013 0.0011
Sowing methods
Line sowing 250.32 638.49 1074.80 12.07 11.45 0.0288 0.0140
Cross sowing 259.24 678.52 1151.10 12.85 12.55 0.0295 0.0141
FIRBS 241.20 610.00 1051.89 11.57 11.62 0.0291 0.0144
Input levels
100% SR + 100% FD 258.36 684.95 1126.53 13.35 11.59 0.0304 0.0132
125% SR + 125% FD 262.14 700.25 1167.97 13.71 12.23 0.0307 0.0134
75% SR + 100% FD 254.64 640.03 1094.61 11.70 12.01 0.0279 0.0143
100% SR + 75% FD 239.75 620.95 1076.89 11.70 12.26 0.0291 0.0149
75% SR + 75% FD 236.36 565.50 996.97 10.39 11.27 0.0273 0.0150
CD (P=0.05) 9.30 23.04 31.54 0.81 0.99 0.00016 0.0012
Table 2.
Effect of varieties, sowing methods and input levels on growth of durum wheat (Pooled for two years)
Treatments
Tiller Spiklets 1000 grain weight Grains Grain yield
Total uptake (kg ha –1 )
m –2 ear –1
(g)
ear –1 (q ha –1 ) Nitrogen Phosphorus Potassium
Varieites
Raj 1555 404 16.25 45.8 45.9 45.4 115.9 32.7 119.6
HI 8498 434 15.08 49.4 41.7 48.8 128.1 36.1 125.1
CD (P=0.05) 11 0.28 0.8 0.7 1.5 2.3 1.0 3.40
Sowing methods
Line sowing 411 15.57 47.6 43.8 46.3 118.5 33.5 119.8
Cross sowing 450 15.48 47.4 43.3 50.4 133.2 37.4 132.6
FIRBS 397 15.93 47.8 44.3 44.6 114.3 32.3 114.7
CD (P = 0.05) - - - - - - - -
Input levels
100% SR + 100% FD 426 16.06 48.2 45.1 48.9 128.1 36.2 126.8
125% SR + 125% FD 459 15.92 47.9 42.2 50.5 133.1 37.5 131.9
75% SR + 100% FD 409 15.92 48.6 45.0 46.9 124.4 35.0 123.2
100% SR + 75% FD 414 15.17 45.8 42.0 46.0 115.2 32.6 119.0
75% SR + 75% FD 388 15.25 47.5 44.0 43.3 109.2 30.7 110.9
CD (P=0.05) 14 0.4 1.1 1.1 1.8 4.0 1.2 4.5
Marked increase in growth and yield attributes in var. HI 8498
could be ascribed to overall improvement in crop growth as
evident from higher dry matter accumulation at successive
stages as well as concentration and uptake of nutrients. These
subscribe greater availability of photosynthates and nutrients
matching with demand for initiation and growth of each
reproductive structure. The results of present investigation
indicating increased overall growth of var. HI 8498 over Raj
1555 corroborate findings of the experiments conducted at
several locations of CZ (DWR, 1997; DWR, 1998).
Cross sowing improved dry matter production by 7.1
and 9.4% over line and FIRB sowings, while FIRB system
recorded reduction of 2.1% over line sowing. Cross sown
crop attained highest CGR(12.85 g m -2 d -1 between maximum
tillering-heading which was at par with line sowing but recorded
significant increase over FIRB system, while later sowing
methods were at par in this respect. Irrespective of crop
duration, RGR didn’t differ significantly between sowing
methods during either year of experimentation. Under cross
sowing better growth, implicated even distribution of seeds,
later on plants (after germination) over the cropped area, thus,
ensuing greater and uniform space to individual plant which
seems to have provided higher growth inputs (below and
above ground) without least competition under cross sowing
compared to line sowing and FIRB system. Several researchers
(Jat, 2001; Parashar, et al., 2004;) also reported reduction in
growth of wheat crop under FIRB system and ascribed this
on account of inter and intra plant competition, which was
not compensated by border effect
The biomass accumulation by plants showed significant
reduction to the extent of 4.4 and 11.5 per cent under reduced
input environments of 100% SR + 75% FD and 75% SR + 75%
FD over 100% SR + 100% FD, while input level of 125% SR +
125% FD failed to enhance it significantly over recommended
level of seed + fertilizer. .Between maximum tillering-heading
stage, CGR and RGR failed to show marked improvement with
higher seed + fertilizer (125% SR + 125% FD) over recommended
input level, while lesser inputs i.e., 100% SR + 75% FD and
75% SR + 75% FD significantly reduced these. The various
yield components were not improved significantly with
elevation in input level from 100% SR + 100% FD to 125% SR
+ 125% FD. However, these parameters showed significant

24 Trends in Biosciences 5 (1), 2012
reduction under lower input levels of 100% SR + 75% FD and
75% SR + 75% FD. Increased inter and intra plant competition
for various growth inputs under higher seed + fertilizers (125%
SR + 125% FD) might have restricted potentials of inputs
particularly increased fertilizers, to modify plant growth to the
fullest extent. On the other hand, reduction in fertilizers by 25
per cent with full seed rate (100% SR + 75%FD), as well as
reductions in both the inputs (75% SR + 75% FD) had a adverse
effect on overall growth of the crop by virtue of inadequate
mineral nutrition for development of various growth
parameters. These results are in conformity to the findings of
(Auti, et al., 1999; Behera and Pandey, 2002).
Var. HI 8498 was found efficient in nutrient uptake as it
accumulated significantly higher quantum of nutrients at
successive stages as well as at harvest, which manifested in
highest total uptake (128.18 kg N, 36.14 kg P, 125.17 kg K ha -1 )
by the crop compared to 115.92 kg N, 32.79 kg P and 119.63 kg
K ha -1 accumulated by Raj 1555. Marked variation in
concentration and uptake of nutrients between durum as well
as aestivum genotypes is in accordance with findings of
several research workers (Singhi and Jat, 2002).
The cross sown crop accumulated significantly higher
quantum of nutrients in plant parts at each stage, which
manifested in highest total uptake (133.20 kg N, 37.54 kg P and
132.61 kg K ha -1 ) representing increases of 12.3, 11.9 and 10.7%
over line sowing and 16.5, 16.1 and 15.5% over FIRB system.
The findings of present study corroborates results of the
experiments conducted by several workers (Jat, 2001)
The crop under higher input environment (125% SR +
125% FD) accumulated highest quantum of nutrients closely
followed by use of recommended seed and fertilizer, while
reduction in either fertilizer (100% SR + 75% FD) as well as
seed + fertilizer by 25 per cent (75% SR + 75% FD) significantly
reduced accumulation of nutrients by the crop)
Durum var. HI 8498 significantly out yielded Raj 1555
and recorded increase in mean grain and biological yield by
7.50% and 5.2%, respectively whereas varieties didn’t differ
significantly in straw production. The economic evaluation
revealed that variety HI 8498 (Malav Shakti) increased net
returns by Rs.2732 ha -1 over Rs.28794.50 ha -1 as compared to
Raj 1555 as well as enhanced net profit/rupee investment from
2.10 to 2.29.
Bidirectional sown crop produced significantly higher
grain, straw and biological yields (50.48, 61.33, 111.80 qha -1 )
representing increases of 9.0, 9.2 and 9.1% over conventional
line sowing. The corresponding increases over FIRB system
were to the extent of 13.0, 15.1 and 14.2%. Despite increased
operational cost, cross sowing gave additional returns of
Rs.3319.30 ha -1 and Rs.4971.30 ha -1 with B:C ratio of 2.33 as
compared to net returns and B:C ratio realized under
conventional line sowing (Rs.29604.70 ha -1 , 2.19) and FIRB
system (Rs. 27952.70 ha -1 , 2.07). The adoption of FIRB system
did not show economic viability as it reduced monetary gains
by Rs. 1652 ha -1 over conventional line sowing.
The interaction effect revealed marked variation in crop
response to input levels under each sowing method and viceversa.
Under line sowing, increase in inputs by 25% each of
seed rate and fertilizer over recommendation though had
positive impacts on overall performance of the crop but failed
to provide additional gains in crop productivity and
profitability. While reduction in inputs by 25% either fertilizer
and seed + fertilizer over recommended level adversely affected
overall productivity and profitability. Conversely, cross sowing
with higher input level of 125% seed + 125% fertilizer over
recommendation enhanced biomass and nutrients
accumulation, consequently production of higher effective
tillers and thereby realized highest productivity of the crop in
terms of grain (56.97 qha -1 ) and biological yields (125.80 q
ha -1 ). While under FIRB system, inputs levels failed to show
any significant variation in crop performance as well as
productivity and profitability. Among combination of studied
production factors, cross sowing of variety HI 8498 with 125%
SR + 125% FD produced highest yields (58.13 qha -1 grain and
66.94 qha -1 straw) and fetched net returns of Rs. 38424 ha -1
and B:C ratio of 2.53.
LITERATURE CITED
Auti, A.K., Wadile, S.C. and Pawar, V.S. 1999. Yield, quality and nutrient
removal of wheat (Triticum aestivum) as influenced by levels and
sources of fertilizer. Indian Journal of Agronomy 44: 119-122.
Behera, U.K. and Pandey, H.N. 2002. Improving productivity of durum
and aestivum wheat with limited irrigation in vertisols in Central
India. Extended summaries : 2 nd International Agronomy Congress,
Nov., 26-30 New Delhi, India, Vol. 2 pp. 1283-1284.
DWR. 1997. Results of the All India Coordinated Wheat and Triticale
Varietal Trials. AICWIP, Directorate of Wheat Research, Karnal,
India.
DWR. 1998. Results of the All India Coordinated Wheat and Triticale
Varietal Trials. AICWIP, Directorate of Wheat Research, Karnal,
India.
Hossain, M.I.M., Duxbury, C.M., Lauren, G., Rahman, M.M., Meer,
M.M. and RashidM.H.2004. Use of raised beds for increasing wheat
production in rice-wheat cropping systems. 4 th International Crop
Science Congress held at Australia
Hussain, I., Khan, A.K., Ahmad, K. 2003. Effect of row spacing on the
grain yield and yield components of wheat (Triticum aestivum).
Pakistan Journal of Agronomy, 2: 153-159.
Jat, L.N. 2001. Evaluation of FIRB planting system for development
of productive and sustainable wheat (Triticum aestivum) – based
cropping system. Ph.D. (Ag.) Thesis, Department of Agronomy,
MPUAT, Udaipur
Prashar, A., Thaman, S., Nayyar, A., Humphreys, E., Dhillon, S.S,
Singh, Y., Gajri, P.R. Timsina, J. and Smith, D.J. 2004. Performance
of wheat on beds and flats in Punjab, India 2 nd International Crop
Science Congress held at Australia.
Sharma R.K. and Randhir Singh 2002. “Furrow irrigated raised bed
planting system. An efficient input usage production technology.
Indian farming, 6:25-26.
Singhi, S.M. and Jat, L.N. 2002. Nitrogen uptake and water use efficiency
of wheat (Triticum aestivum) as influenced by planting system and
varieties. Extended summaries vol. 2 : 2 nd International Agronomy
Congress, Nov., 26-30 New Delhi, India, pp. 870-871.
Recieved on 04-01-2012 Accepted on 15-04-2012

Trends in Biosciences 5 (1): 25-30, 2012
Effect of Season on Helminth Parasitic Prevalence, Dominance, Means Intensity
and Abundance in Some Fresh Water Scaly Fish
KRISHNA SINGH AND ABHA MISHRA
Department of Applied Animal Sciences, Babasaheb Bhimrao Ambedkar University (A Central University),
Vidya Vihar, Raebareli Road, Lucknow 226 025
e-mail: drabhamishra@gmail.com, kri.singh1983@rediffmail.com
ABSTRACT
Five scaly fishes, Channa punctatus, Labeo rohita, Catla catla,
Tor tor and Anabas testudineaus were investigated for seasonal
density and infection intensity of helminths parasites of
Lucknow region during October 2007 to September 2009. Total
fourteen species of helminths were recovered: nine species of
ectoparasites (Dactylogyrus and Gyrodactylus); twelve species of
endoparasites (Clinistomum, Euclinostomum, Gastrohtylax,
Paramphistomum, Opisthorchis, Aspidogaster, Fasciola, Teania,
Trichinella and Pallisentis). The overall prevalence of helminth
parasites infection was recorded as 66.15 % out of 73.84 %
infection in examined scaly fishes. The infestation exhibited
significant monthly or seasonal fluctuation. The maximum
prevalence and mean intensity of helminth parasite was
recorded in summer season, moderate in monsoon and
minimum in winter season. The prevalence, dominance, mean
intensity and abundance of the helminth parasite were also
found to be different between scaly fishes and were varied with
the seasons. Among helminth parasites trematode showed
significantly higher infection intensity as compared to cestode,
nematode and acanthocephalan parasites.
Key words
Helminth parasites, scaly fishes, prevalence,
dominance, mean intensity, abundance.
Climatic change effects on parasites and diseases of
key species may cascade through food webs, with
consequences for entire ecosystems (Marcogliese, 2004,
2008). The seasonal fluctuation affected the parasitic
population (Paulin, 2007; Jørgensen, et al., 2008) and their
infection rate (Ozturk and Altunel, 2006). Optimum temperature
supports all life cycle stages of helminth parasite including
their infectivity, reproduction and hatching rate (Ondrackova,
et al., 2004; Shalaby, et al., 2004). Higher temperature results
in increasing outbreaks of helminth parasite (Ondracji kova,
et al., 2004; Shalaby, et al., 2004).
The prevalence, intensity and abundance of the
infection with helminth parasites species richness increases
with host body size and shows fluctuation in the months of
the year (Bayoumy, et al., 2008; Owolabi, 2008). The medium
sized and weight fish were more infected and their prevalence,
intensity and abundance were highest (Tasawar, et al., 2007;
Bhuiyan, et al., 2008; Owolabi, 2008; Tekin-ozan, et al., 2008).
Carp fishes are reported to show the severe infection
with helminth parasites (Bhuiyan, et al., 2008; Tekin-Özan, et
al., 2008). Fresh water fish, both scaly and scale less, show
the infestation with helminth parasite (Owolabi, 2008;
Morenikeji and Adepeju, 2009) but low temperature only
support the parasitization in scaly fish (Gelnar, 1987).
Helminth parasitic infection not only deteriorate the fish
nourishing quality but also makes wound on their flesh that
leads to secondary infection and cause disease and ultimately
death (Bhuiyan et al., 2008). The present study was done to
investigate the prevalence, dominance, means intensity and
abundance of helminth parasite species on some fresh water
scaly fish of Lucknow region. Study will represent a
comparative assessment of infestation of different helminth
community on five common edible fresh water scaly fish. In
this work we will also focus on the effect of seasons on
helminth parasite infection in scaly fish.
MATERIALS AND METHODS
Lucknow has a warm subtropical climate with cool dry
winters from December to February and dry hot summers from
April to June. The rainy season is from mid-June to mid-
September. In winter the maximum temperature is around 24
degrees Celsius and the minimum is in the 3-4 degrees Celsius
range. Lucknow gets an average rainfall of 101 cms mostly
from the south-west monsoon winds between June and
September.
The survey covers the relative prevalence of helminth
parasites in some fresh water scaly fish during the period
October 2007- September 2009. Fish specimens were procured
from local fish markets. The fishes examined for study were
Channa punctatus (Bloch.) Ophiocephalidae; Labeo rohita
(Ham.) Cyprinidae; Catla catla (Ham.) Cyprinidae; Tor tor
(Ham.) Cyprinidae and Anabas testudineus (Bloch.)
Anabantidae. Fish specimens of each genus were of medium
and approximately same size, same weight to avoid infection
versus fish health factor.
The collected fish were examined externally as well as
anatomically to collect the external and internal helminth
parasites. For collection of external parasites fish were placed
in containers of 4% formalin, shaken and the sediment
examined for parasites. For internal parasites dissected fish
were examined paying special attention to the color and
consistency of the organs. Parasitic helminths were removed
from the fish carcass. These could be cysts, juveniles or larval

26 Trends in Biosciences 5 (1), 2012
forms but only adult one were collected, identified and
numbered for the study to avoid confusion and mistake. For
better observation, hand lens and microscope were used. The
gall bladder was removed and the content examined on a slide.
The wall of stomach and intestine were carefully examined for
the presence of ulcers, cysts and burrowing parasites. The
content of stomach and intestine were flushed and solid
contents being collected for examination.
Nematodes were relaxed for 3 minutes in physiological
saline and washed with water, examined alive, fixed in F. A. A.
(formalin-acetic-alcohol) and transferred in a solution of 5 parts
of glycerin and 95 parts of ethyl alcohol. A few drops of
saturated aqueous picric acid are added to stain and mount in
DPX.
Cestodes were placed in ice water in vials and left
overnight in a refrigerator. This makes the worms relaxed so
their scolex, which is of taxonomic importance. Then flattened
worm between glass and transferred in 70% alcohol 10 minutes.
Wash it again with 70% alcohol and stained with Borax Carmine
for 5 minutes. Afterward, washed it with 90% alcohol and then
give two changes for 5 minutes in absolute alcohol and mount
in DPX.
Trematodes were pressed between two slides with glacial
acetic acid (GAA). This makes the worms transparent so
internal organs can be seen. All parasites were preserved in
70% alcohol for 2-3 hours. Remove the glass and transfer
worms to fresh fixative. After 2-3 hours stain the worms with
Borax carmine, differentiate it with acid water and transferred
in 90% alcohol 10 minutes. Now give two changes in 100%
alcohol for 5 minutes and then clear it with xylene or benzene
and mount with DPX.
Acanthocephalan were first relaxed in 0.6% physiological
saline for 1 hour and then fixed in F. A. A. solution for 24 hours
and stained with Mayer’s Carmalum, dehydrated in a graded
series of alcohol and mount in DPX.
The collected adult helminth parasites were identified
according to Kabata 1985, Yamaguti 1963, 1985 and Verma
2000 on the basis of their internal and external features.
Standard statistical computations viz; prevalence,
dominance, mean intensity and abundance were carried out
according to Margolis, et al., 1982. Study data presented as
season wise according to Pokale, et al., 2002 viz; winter
(October to January), summer (February to May) and Monsoon
(June to September).
The complex chi-square and contingency test were used
for data significance and relationship between parasites
intensity and different seasons (viz., winter, summer and
monsoon).
RESULTS AND DISCUSSION
The prevalence of infection during the study period was
73.84 % in scaly fish. In this study, 3610 parasites of 14
helminth species, including seven digenean trematode
(Clinostomum spp., Euclinostomum spp., Gastrothylax spp.,
Paramphistomum spp., Ophisthorchis spp., Aspidogaster
spp. and Fasciola spp.) and two monogenean trematodes
(Gastrothylax spp. and Dactylogyrus spp.), three cestodes
(Teania spp.), one nematode (Trichinella spp.), and one
acanthocephalus (Pallisentis spp.) were found. Helminth
infection caused deterioration in fish external as well as internal
tissue intactness. During the study, presence of cloudiness
of skin (grey and white), reddening, ragged or torn fins, raised
scales, white spots or parasites visible to naked eyes were
among the common morphological features. The common
changes in internal body parts i.e., yellowish color of liver,
congestion and reddening of liver, pale color of kidney,
transparency and inflammatory reddening of intestine,
hardening of bile and gall bladder, adipose tissues between
the intestine and liver, fluid in the peritoneal cavity, white
nodule on liver, pancreas and kidney were also observed.
Occurrence wise, among different helminth community,
trematode showed maximum prevalence percentage followed
by acanthocephalan, cestode and nematode; however last
two showed fluctuating infection season wise (Fig. 1). Seasonal
effect in % prevalence, % dominance, mean intensity and
abundance of helminth parasite in local fresh water scaly fish
during two successive years was recorded. Data represents a
comparative statement of seasonal effect on different helminth
parasitic group in five common edible fresh water scaly fish. It
was found statistically that the occurrence of all four helminth
parasitic groups (trematode, cestodes, nematode and
acanthocephalan) were not influence each other (÷ 2 < 1, below
the significant value) but their availability in scaly fish groups
was affected by different seasons with varied significant level
among helminth community (Fig. 1; for winter ÷ 2 = 6.48, P <
0.9; for summer ÷ 2 = 8.50, P < 0.75; for monsoon ÷ 2 = 21.95, P <
0.05, complex chi-square and contingency test). A single fish
may be the habitat for a single helminth parasitic community
or may be the host for two or three or all the four helminth
parasitic community with visible preference. The most common
Fig. 1.
Infection rate (%) of helminthes parasites in some
scaly fishes during different seasons.
W= winter; S = summer; M = monsoon

SINGH AND MISHRA, Effect of Season on Helminth Parasitic Prevalence, Dominance, Means Intensity and Abundance 27
combination of helminth parasites was trematode and
acanthocephalan and rare combination was cestode and
nematodes.
With respect to annual helminth infection, minimum
infection intensity for overall community was detected in C.
punctatus whereas maximum infection was recorded in case
of L. rohita, most common edible major carp, as compared
with other scaly fish (Fig. 1). Data presents a clear significant
affinity between helminth community and fish species as well
as they show the seasonal dependence also. During winter
infestation of helminth community was minimum and maximum
was recorded during summer period in all five examined fresh
water fish (Fig.1, Table 1-5).
In winter season, over all infection with helminth
community was highest in T. tor and minimum in C. punctatus
(Fig. 1). The pattern of per cent prevalence of helminth
parasites in all checked scaly fish were same viz. higher with
trematode followed by acanthocephalan, cestode and minimum
with nematode. The prevalence of helminth community was
comparatively highest in T. tor than others (Table 1-5). The
dominance of helminth parasites was also differing in scaly
fishes and follows the same trend of per cent prevalence during
the study period. Per cent dominance of trematode in winter
was high in L. rohita and C. catla; acanthocephalan was high
in A. testudineus and T. tor; cestode was more in L. rohita and
A. testudineus; nematode was maximum in A. testudineus
Table 1.
Prevalence, dominance, mean intensity and
abundance of Helminthes parasites in Channa
punctatus under different climatic conditions.
Trematode
Seasons Prevalence Dominance Mean Abundance
(%) (%) intensity
Winter 15.89 50.91 4.72 0.76
Summer 53.70 46.36 5.07 2.16
Monsoon 38.47 53.62 4.67 1.80
χ 2 =15.24, p

28 Trends in Biosciences 5 (1), 2012
Table 3.
Seasons
Prevalence, dominance, mean intensity and
abundance of Helminth parasites in Catla catla
under different climatic conditions.
Prevalence
(%)
Trematode
Dominance
(%)
Mean
intensity Abundance
Winter 24.70 52.01 1.78 0.41
Summer 78.08 51.89 2.84 2.22
Monsoon 43.91 58.16 3.38 1.46
χ 2 =24.78, p

SINGH AND MISHRA, Effect of Season on Helminth Parasitic Prevalence, Dominance, Means Intensity and Abundance 29
Table 5.
Prevalence, dominance, mean intensity and
abundance of Helminth parasites in Anabas
testudineus different climatic conditions.
Trematode
Seasons Prevalence Dominance Mean
(%) (%) intensity Abundance
Winter 19.51 41.62 1.64 0.21
Summer 76.25 47.43 3.14 2.39
Monsoon 43.18 45.16 1.83 0.92
χ 2 =16.28, p

30 Trends in Biosciences 5 (1), 2012
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Trypanosoma spp. From the Fishes of Aurangabad, India. Uttar
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Ecology meets Geography and Climate. Journal of Parasitology,
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Occurance of Helminth Parasites Infecting Anabas testudineus in
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Received on 8-6-2011 Accepted on 10-12-2011

Trends in Biosciences 5 (1): 31-34, 2012
Life cycle, Population Index and Feeding Activities of the Lime Butterfly, Papilio
demoleus (Lepidoptera: Rhopalocera: Papilionidae)
DURGESH NANDNI, ARUN RAGHUWANSHI* AND VINOY KUMAR SHRIVASTAVA*
Government College, Ashta, District Sehore, M.P.
*Department of Biosciences, Barkatullah University, Bhopal 462 026 (M.P)
email: vinujks2001@yahoo.com
ABSTRACT
The present paper deals about the life history stages of Lime
butterfly which includes larval development period, pupal
period, measurements of body length of caterpillars, population
index and feeding time. The larval instars i.e. Ist, IInd, IIIrd,
IVth and Vth of Lime butterfly lasts for an average of 2.50 ±
0.32 , 4.07 ± 0.61 , 4.10 ± 0.62 , 3.60 ± 0.28 and 3.71 ± 0.25 days
respectively and their body length measured 4.06 ± 0.43 mm,
8.98 ± 0.62 mm, 18.61 ± 0.94 mm, 25.37 ± 0.99 mm and 35.09
±1.45 mm respectively during different larval stages. The total
larval period of Lime butterfly was observed to be 17.98 ± 2.08
days. The pupal period was 11.57 ± 0.92 days. The total
development period was recorded 29.55 ± 3.06 days. During 4 th
and 5 th instar feeding activities was rapid in comparison to 1 st
and 3 rd instar stages. The Lime butterflies were active in
January, February, June, July and October months of the year.
The percentage of survival from egg to adult was maximum in
the month of June (80%) and July (80%) and was minimum
(60%) in month of February.
Key words
Autecology, life history, papilionidae, papilio demoleus,
instars, population index.
Lepidoptera, one of the highly specialized insect order,
includes scale- winged insects of the holometabolous
endoperygote series which includes butterflies and moths
that show total metamorphosis and pass through egg, larva/
caterpillar, pupa and adult stages. Several entomologists were
studied the life cycle of different butterflies and moths
(Ramkrishna-Ayyar,1984; Metcalf and Flint, 1973). Atluri, et
al., 2002 and Vasait, 2002 studied the autecology and life cycle
of Papillio polytes Lepidoptera: Papillionidae. Apart from
this several workers have also revealed the life cycle of many
others butterflies (Singh and Shrivastava, 2008; Dharshey and
Shrivastava, 2010; Nandni, et al. 2011).
MATERIALS AND METHODS
The present study has been made in the garden and
nurseries of Bhopal city. Five sampling stations were opted
and they were representing approximately the four directionsnorth,
east, south and west and the central part of the Bhopal
city. The sites selected for study represent all different
vegetational groups and topography of Bhopal. The eastern
sampling station was “Madhuban Garden” which is situated
in Bharat Heavy Electrical Limited (BHEL) area and is named
as sampling station ‘A’ (Fig. 1). Similarly the sampling stations
in the northern, southern, western and central part of the
Bhopal city were named as sampling stations B, C, D and E
(Fig. 1) respectively; ‘B’ sampling station was “Sundarban
Nursery” situated at Bairagarh, which is about 8km away from
Bhopal on Sehore-Astha-Indore road, while ‘C’ station was
“Mohan/Agra Nursery”, which lies at Misrod road, about 12
km. from Bhopal. ‘D’ station was “Shahpura Garden” located
nearby Maneesha market . Apart from these, ‘E’ station was
“Govt.Park 1”, situated on the link road near the New Market.
‘A’, ‘B’, ‘C’ and ‘E’ sampling stations are relatively dry and
do not have any pond or water body in close proximity. ‘D’
sampling station has a big lake which remains filled through
out the year. In connection to this, the vegetations i.e. herbs,
shrubs, trees and of course grasses were abundant in all the
sampling stations, but A, B, and ‘E’ stations were have more
abundant vegetations than C and D stations (Fig. 1). The life
cycle of Lime butterflies were observed from Sundarban and
Misrod sampling sites, because the greater appearance was
more at these stations in comparison to other sampling
stations. Twigs with leaves carrying eggs of butterfly were
identified on Citrus limon plant. The eggs were observed till
they hatched and along with this, the feeding behavior of
caterpillars was also observed. The caterpillars start feeding
from the periphery of the leaves and reach to midrib. The
caterpillar moves from one leaf to another for feeding. At the
time before pupa formation, the caterpillar stops eating and
forming pupa. During pupation the caterpillar attaches itself
to the walls of Citrus limon’s leave and form pupa.
The present paper deals about the life history stages,
larval development period, pupal period, measurements of
body length of caterpillars, population index and feeding time
of the Lime butterfly Papilio demoleus. In population index
studies, readings were taken for number of eggs, pupae formed
and number of adults emerged. For feeding activity the
observations were recorded of one larva out of all hatched in
that particular month of rearing. Observations made on feeding
activity for a 4 h period. Feeding activity and population index
studies were conducted in the months of January, February,
June, July and October.
RESULTS AND DISCUSSION
The eggs were yellowish in color and arranged on the
leaf surface in a group [Fig: 2(a)]. There were 5 larval instars

32 Trends in Biosciences 5 (1), 2012
observed. The larval period was completed in 17.98 ± 2.08
days. Instar wise development period varies from 2.50 ± 0.32
days (I instar) to 4.10 ± 0.62 days (III instar) [Table 1]. The
pupal period was 11.57 ± 0.92 days. Total developmental period
from Ist instar to adult emergence was completed in 29.55 ±
3.06 days. Ist instar larva was pale brown in color, 4.06 ± 0.43
mm in length. IInd instar larva was slightly dark brown and
feeding on leaves. The fully developed IInd instar larva
measured 8.98 ± 0.62 mm in length. In second and third instar,
body is rough in texture, snuff colored with white markings
Fig. 1
on the dorsal profile of abdomen and anal region. The third
one was 18.61 ± 0.94 mm in length [Table1, Fig. 2(b and c)]. In
fourth and fifth instar, caterpillar turns into pale green. White
and brown band developed on 8 th and 9 th segments. Black
band developed on 4 th and 5 th segments with two black spots.
The fourth and fifth instar measures 25.37 ± 0.99 mm and 35.09
± 1.45 mm long [Table1, Fig. 2(d)]. The pupal colour was light
green. Its length was 30.11 ± 0.74 mm. Posterior end is pointed,
attached to the substratum while anterior end is V shaped
[Table 1; Fig. 2(e)].

NANDNI et al., Life cycle, Population Index and Feeding Activities of the Lime Butterfly, Papilio demoleus 33
Fig. 2. Showing the different stages of life cycle of lime butterfly (Papilio demoleus)
Fig. 2(a): Eggs, Fig. 2 (b&c): Different Instars of caterpillar, Fig. 2(d): Caterpillar preparing to pupate, Fig. 2(e): Pupa, Fig. 2(f): Adult
Butterfly
Table 1. Developmental period and body measurement of Lime Butterfly.
Species
Developmental period in days
Body measurements (mm)
Larval instars
I II III IV V
Larval
period
Pupal
period
Total dev.
period
(Larva to
adult)
Larval instars
I II III IV V
Pupa
Lime
Butterfly
2.50
±
0.32
4.07
±
0.61
4.10
±
0.62
3.60
±
0.28
3.71
±
0.25
17.98
±
2.08
11.57
±
0.92
29.55
±
3.06
4.06
±
0.43
8.98
±
0.62
18.61
±
0.94
25.37
±
0.99
35.09
±
1.45
30.11
±
0.74
Table 2. Population index of Lime butterfly (Papilio demoleus)
Month
Number of eggs
incubated
Number of
larvae hatched
Number of
pupae formed
Number of
adults emerged
Number of
adults emerged in %
Jan 8 6 5 5 62.6
Feb 5 4 3 3 60.0
Jun 10 9 8 8 80.0
Jul 10 8 8 8 80.0
Oct 8 7 6 6 75.0
During population index, it has been observed that, June
(80%) and July (80%) months were the most congenial period
for the development of Lime butterfly and February month
was least favorable month [Table 2]. Observations made on
feeding activity for a 4 hour period. During this period June
and July months were observed, when the feeding time was
maximum. In these months the time was 17.07 ± 6.40 min and
16.69 ± 4.79 min with 3.53 ± 0.84 bouts and 3.46 ± 0.63 bouts
respectively [Table-3]. It was found that in 4 th and 5 th instar
feeding was rapid and in first to third instar feeding was slow.

34 Trends in Biosciences 5 (1), 2012
Table 3.
Month
Feeding activity of Papilio demoleus larvae on
Citrus limon leaves over 4h period
Instar
No.
Day of
Observation
No. of
feeding
bouts
Total length of
larval feeding
time(min)
I 1 - -
2 2 9
Jan II 1 3 13
2 3 13
3 2 14
III 1 3 12
2 4 15
3 4 15
IV 1 3 17
2 3 17
3 3 19
V 1 3 22
2 4 22
3 4 27
3.15 ± 0.66 16.53 ± 4.71
Feb I 1 - -
2 2 9
II 1 2 10
2 4 13
3 4 14
III 1 3 10
2 4 15
3 4 16
IV 1 3 17
2 3 17
3 4 20
V 1 4 24
2 4 25
3 3 27
3.38 ± 0.73 16.69 ± 5.63
Jun I 1 - -
2 2 9
II 1 3 10
2 3 10
3 3 12
III 1 3 10
2 4 16
3 4 16
IV 1 3 17
2 5 22
3 4 22
Breeding of Lime butterfly was observed in January, February,
June, July and October months. The mean temperature of these
months were 18.81 ºC, 24.34 ºC, 31.74 ºC, 26.35 ºC and 26.01 ºC
respectively.
The mean percentage relative humidity of these specific
months was 51.81, 40.55, 56.73, 87.46 and 61.62 respectively.
Eggs and larvae found on various host plants revealed this
period to be most favourable period of this butterfly as it was
also reflected by the higher population index and feeding
activity of the larvae.
ACKNOWLEDGEMENT
We wish to express sincere deep sense of gratitude to
Prof. M. Banerjee, HOD Bioscience Dept., B.U. Bhopal, Dr.
V 1 4 25
2 5 26
3 3 27
3.53±0.84 17.07 ± 6.40
Jul I 1 - -
2 3 10
II 1 2 10
2 4 13
3 3 12
III 1 3 14
2 4 16
3 4 16
IV 1 3 17
2 4 19
3 4 20
V 1 4 20
2 4 25
3 3 25
3.46 ± 0.63 16.69 ± 4.79
Oct I 1 - -
2 2 6
II 1 3 11
2 3 14
3 4 15
III 1 3 10
2 4 14
3 4 15
IV 1 3 18
2 4 19
3 5 20
V 1 4 22
2 5 24
3 3 26
3.61 ±
0.83
16.46 ±
5.52
Rajesh Verma, Principal scientist, Entomology, FRS, Intkhedi,
Bhopal (M.P.) and Dr. K. Chandra, Office Incharge, Central
Regional Station, Zoological Survey of India, Jabalpur (M.P.)
for the valuable guidance for the study.
LITERATURE CITED
Atluri, J. B., Venkata Ramana, S. P., and Subba Reddi, C. 2002. Autecology
of the Common Mormon butterfly, Papilio polytes (Lepidoptera:
Rhopalocera: Papilionidae). Journal of Environmental Biology,
23(2):199-204.
Dharshey, D.N. and Shrivastava, V.K. 2010. Autecology of the heverfly
Allograpta obliqua say (Diptera : Syrphidae) at Shahpura lake (Chuna
bhatti), Bhopal, M.P. Biospectra, 5(1): 01-04.
Kalyanam, N. P. 1967. Common Insects of India, Asia publishing house,
P.1.
Metcalf, C.L. and Flint, W.P. 1973. Destructive and useful insects. 4 th
ed, Tata McGraw Hill Publishing Co Ltd. New Delhi, pp.1087.
Nandni, D., Raghuwanshi, A. and Shrivastava, V. K. 2011. Autecology
of the Common Mormon butterfly, Papilio polytes (Lepidoptera:
Rhophalocera, Papilionidae). J. Ent. Res., 35(2) : 1-4.
Ramkrishna Ayyar, T.V. 1984. Hand book of Economic Entomology
for South India. Government Publications, Madras.
Singh, H. and Shrivastava, V.K. 2008. Autecology of the lime butterfly,
Papilio demoleus demoleus (Lepidoptera : Rhophalocera,
Papilionidae). J. Exp. Zoology, India, 11(2): 331-333.
Vasait, J.D. 2002. Study of the life cycle of Papilio polytes Lepidoptera:
Papilionidae, Ecology. Environment. and Conservation, 8(1): 79-
84.
Recieved on 2-8-2011 Accepted on 15-12-2011

Trends in Biosciences 5 (1): 35-37, 2012
In Vitro Quantification of B-Cell Proliferation in Kidney Culture of Channa striatus
Exposed to Different Doses of KOV Antigen of Aeromonas hydrophila, using
Lymphocyte Transformation Test (LTT)
S.A. MASTAN
Post-Graduate Department of Biotechnology, D.N.R. College, P.G. Courses and Research Centre,
Bhimavaram-534 202, W.G. Dist, A.P.
email: shaikmastan2000@yahoo.com
ABSTRACT
The present work aims to study the specific immune response
shown by the fish against various doses of KOV antigen of
Aeromonas hydrophila. Tissue culture techniques were used
for the culture of lymphoidal cells of head kidney. The cell
number was primarily calculated by using Nicobar’s Chamber
and the cells were infused with various doses of antigen. The
highest number of lymphocytes transformation was observed
in the well treated with 5.25µl antigen while the lowest number
of lymphocyte transformation was observed in the treated with
0.002µl of antigen.
Key words Aeromonas hydrophila, lymphocyte transformation
test, Channa striatus
Immune mechanism of the fish have not been as
extensively studied as those of mammals but they appear to
share a number of structural and functional characteristics
important in the humoral, cell – mediate and non specific
immune response. Immune cells of fish, similar to those of
mammals include : (a) antibody producing cells thought to be
analogous to B – lymphocytes (Anderson, et al., 1982): (b) T
– lymphocyte – like antigen sensitive cells, which participate
in reactions important in cell – medicated immunity such as
delayed type hypersensitivity: (c) non – specific cytotoxic
cells (NCC), a proposed equivalent to mammalian natural killer
cells (NK), which are cytotoxic to a variety of allogenic and
xenigenic tumour cells in vitro (Graves, et al., 1982); and (d)
phagocytic cells including polymophonuclear like leucocytes
and macrophages, which are important in both non specific
and cell mediated immunity (McKinne, et al., 1961).
Lymphocyte transformation is conventionally measured by
incorporation of radioactive thymidine into the DNA of
dividing cell. The rate and amount of isotope incorporated,
expressed as count per minute (CPM), is a reflection of
response and is directly related to the number of cells
responding. Other parameters of lymphocytes response can
be measured e.g. incorporation of amino acids into proteins,
amino acid transport, enzyme synthesis, RNA synthesis and
membrane changes. Lymphocyte transformation may also lead
to secretion of lymphokinase e.g. blastogenic factor, migration
– inhibition factor and interferon; however elaboration of
lymphoid products may occurs in the absence of proliferation
(Lamers, et al., 1992). The substances that can induce
lymphocyte transformation have been classified as specific
or non-specific depending upon whether or not response to
the stimuli requires prior exposure of the cell donor to the
stimulant. In experimental animal model system, this is referred
to as the requirement for pre sensitization. Generally, the
procedure for preparation and maintenance of fish cell cultures
do not differ from those adopted for higher vertebrates.
However, the appropriate physiological and culture condition
depend upon the type of tissue under cultivation, which
include growth responses, culture media, temperature and
osmolarity of the culture environment. The present paper
reports the in vitro quantification of B-cell proliferation in
kidney culture of Channa striatus exposed to various doses
of KOV antigen of Aeromonas hydrophila, using Lymphocyte
Transformation Test (LTT).
MATERIALS AND METHODS
For the purpose of present study, the cell cultures were
developed from kidney of healthy Channa striatus, as per
procedure described by Freshney, 1960. Fish tissues were
taken aseptically at field sites, placed in growth medium and
transported at ambient temperature to the laboratory. They
were processed further and seeded on culture medium. The
cultures thus obtained successfully were used for further use.
The supernatant was centrifuged at 1500 rpm; the pellets were
suspended in PBS. After Pippeting out the non-adherent cells
from the flask, the adherent cell layer was washed with PBS.
Dislodging of the adherent cell was avoided during the
washing steps. The lymphocytes were poured in multi well
plates and the cells were counted in special chamber called
the Nicobar’s Chamber. The haemo-cytometer was also used
to count the number of lymphocytes. Cell suspension was
diluted before counting and carefully added to the area under
the cover slip so that the grid area was just full and not
overwhelming into the over flowing well. The cells were allowed
to settle. The cells were counted in each of the four large
squares. Only those cells were counted touching two outside
borders. The average of the cells per large square was
determined. This was the number of cells per 10 -4 ml. Thus
cells per ml = (Av. No. per large square) X 10 -4 X 1/dilution.
Specific antigen was added (LPS of A. hydrophila) in different
concentrations. The cell number was counted after one week’s
time.

36 Trends in Biosciences 5 (1), 2012
RESULTS AND DISCUSSION
In the present study, lymphocyte transformation test
was used in vitro quantification of lymphocytes. The cell
number increases from 24,600 to 24,670 in the control well.
This may be due to the stress posed by cell during culture.
The cell number increases significantly in LSP treated wells.
The highest transformation of cells (36,866) was observed in
well treated with 6.25µl of antigen. While lowest transformation
of cells 24,850) was observed in the well treated with 0.002 ul
of antigen (Table 1).
It is generally agreed that lymphocytes carry specific
receptors that recognize antigen on the surface of the cell.
Since the only molecule known to combine with antigen is
antibody, the speculation was that the receptor of B-
lymphocyte is a kind of immunoglobulin which appears
structurally related to IgM. It has been shown that cell
expressing IgM can also express IgD from the base of cell
Table 1. In vitro qualification of lymphocytes
S.No
Initial value
(Number of cells)
Amount of
LPS Added
Transformed
value (No. of cells)
1 24,750 6.25 36,860
2 24,750 1.25 33,230
3 24,750 0.25 27,480
4 24,750 0.05 27,050
5 24,750 0.01 25,950
6 24,750 0.002 24,850
7 24,750 Control (No
LSP added)
24,670
receptor. The production of each class of immunoglobulin is
otherwise determined by the presence of class specific Ig on
B- lymphocytes as determined by the genes. When an antigen,
which may be a foreign organism or compound, enters into
the tissue of the animal, lymphocytes are stimulated to
differentiate and divide. Similar to the bone marrow derivative
lymphocytes, an interaction of antigen with thymus derive
lymphocytes can also give rise to primary and secondary
responses in which no specific antibody molecule is involved.
The specific changes from virgin lymphocyte into effector
cell demonstrate, for example ,by specific cytotoxic action to
target cell in response to antigenic determinant indicate that
T- lymphocytes are also antigen sensitive. It also indicate
that the lymphocytes derive from bone marrow and thymus
respectively and activated by appropriate antigen and finally
expanded into short lasting effector cells and long lasting
memory cells. While the former response protects the host
against acute phase of infection as results of first contact and
eliminates the infective agents, the latter provides protection
against infection by an effective interaction on subsequent
contact with primary antigen survived from the microbes.
Activation and proliferation along with differentiation
into effector cells are important response of immune system
to challenge by invading pathogen. Cellular expansion is
meant to increase a number of effector cells, capable of
generating both the necessary inflammatory milieu and an
increased frequency of pathogen specific responder cells.
Depending on the nature of the pathogen and challenge to
the organisms, the response may differ considerably in term
of cellular component involved and appropriate effector
function, chosen by immune system for instance, CTL usually
appears during viral infection. The cell responses are induced
by intracellular bacteria such as mycobacterium, while Th2
pattern can be detected in connection with helminthes
infection and humoral (B-cell) immune response frequently
observed following immunization with protein. Moreover,
cooperative action is characteristics but still not fully
understood aspects of immune system, involving the various
components during the maturation of immune response to
varying degrees. Thus, proliferation is a frequent response of
T-Lymphocytes to an antigenic stimulation. In vivo
experiments are an important supplement of studying the
infectious diseases. Proliferation studies performed in vitro
using isolated and highly purified lymphocytes subsets in
order to detect cellular complexity and to focus on particular
role of individual subset In teleosts, ability to respond to T-
cell and B- cell mitogenes are established some 20 years ago
(Eltinger, et al., 1976) and a number of studies followed
(reviewed by Rowley, et al., 1988). The requirement for
accessory cell activation of T- lymphocytes by mitigen was
confirmed by channel catfish peripheral blood lymphocyte
(Sizemore, et al.,1984) it now seems likely that these accessory
cells (monocytes) were acting by secretion of cytokinesis,
precisely IL1 (Millev, et al.,1985, Clem, et al.,1985). Sizemore,
et al., 1984 separated SIg+ cells from SIg- cells by panning
and demonstrating that SIg+cells responded to LSP regardless
the presence or absence of monocyte, while the cells remain
unresponsive to either LPS or Con A, unless the accessory
cells were present in which respond to mitogens. In Pearman
et al., 1963 reported that lymphocytes of tuberculin – positive
individual Transform in vitro in presence of purified protein
derivatives obtained from tuberculin (PPD) whereas the
lymphocytes of tuberculin negative individual fail to transform.
The strength of skin test responses has been found to correlate
well with the degree of lymphocyte transformation
(Miller and Jones, 1973). High dose of stimulation with
tuberculin, however, may cause lymphocytes from skin test
negative donor to respond (Nillson). Oppenhiem, et. al., 1967
similarly demonstrated transformation to PPD and to guinea
pig albumin orthanilic acid and with lymphocytes that had

MASTAN, In vitro Quantification of B-Cell Proliferation in Kidney Culture of Channa striatus 37
been taken from guinea pig pre immunized with same antigen.
Teleosts fishers possess T and B lymphocytes as found in
mammals (Clem. 1991). Monoclonal antibodies have been
used to separate the SIg+ from SIg- in experiments Labb et al
., found that SIg+ are 40 % while SIg- 2-5%. In ultra structural
study Ig occurs mainly in clusters on membrane of B cells. In
mammals certain substances ( Mitogens) are specific for a
particular array of carbohydrates moieties occur on T cells
but not of B cells e.g PHA and Con A. The other mitogens like
LPS activate the proliferation of B cells but not the T cells.
Marsden, 1995 suggested that SIg- cells were activated by
PHA and vice versa for SIg+ population peripheral
lymphocytes of carp. Some SIg+ cells respond only dull in
immunoflorescent technique. The lymphocyte transformation
test can be done according to procedure given by Scheffold
and Radbruch 1998 and Pechhold and Kabeltz 1998.
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Recieved on 3.8.2011 Accepted on 5.3.2012

Trends in Biosciences 5 (1): 38-40, 2012
Pathogenicity and Mass Production of Entomopathogenic Nematode, Heterohabditis
indica on Major Insects of Agricultural Importance
RISHI PAL*, G.N. TIWARI AND C.S. PRASAD
Sardar Vallabhbhai Patel University of Agriculture & Technology, Meerut, U.P.
Biocontrol Laboratory, Department of Entomology
email: rishipal.biocontrol@gmail.com
ABSTRACT
A study on pathoge nicity and mass pro duction of
entomopathogenic nematode, Heterohabditis indica was carried
out in Meerut, India. Observed data showed that the larval
stages of Plutella xylostella, Bombyx mori, Leucinodes orbonalis,
Corcyra cephalonica, Galleria mellonella and Helicoverpa
armigera were found highly susceptible to the IJs of H. indica
causing 73.3 to 100% mortality at 48 h of exposure. Other
insects, viz., Spodoptera litura, Spilosoma obliqua, Pieris brassicae,
Earias vittella and Holotrichia consanguinea were less susceptible
(3.3 to 73.5% mortality). The maximum IJs were produced from
the infective larvae of G. mellonella (50,000-2,00,000) followed
by H. armigera (50,000-1,50,000). Whereas, the minimum IJs
were produced from infective larvae of P. xylostella (100-2000)
followed by B. mori (2000-3000).
Key word
Pathogenicity, mass production, entomopathogens,
heterohabditis indica
A number of insect pests cause serious damage to cash
crops i.e. cotton, sugarcane, fruits and vegetables. Insect pest
management continues to be one of the major constrains to
the productivity. According to an estimate, the yield losses
caused by insect pests in different crops were about 12-15%
valued at Rs. 3615 crores.
Entomopathogenic nematodes associated with insects
causing disease in susceptible insects and killing them. They
are lethal obligatory parasites of insects (Kaya and Gaugler,
1993), yet pose no threat to plants, vertebrates and many
invertebrates. Entomopathogenic nematodes possess many
attributes of an excellent biological control agent. They have
a broad host range and are highly virulent, killing the host
rapidly. The present information generated will support further
investigation to understand biological and ecological
characteristics of these nematodes, which are important factors
to consider while using them in Bio-intesive Integrated Pest
Management (BIPM) programme.
MATERIALS AND METHODS
Entomopathogenic nematode, Heterorhabditis indica
was cultured on Galleria mellonella larvae as described by
Kaya and Stock, 1997. Infective juveniles emerging within
first 2 weeks were collected from modified White traps (Kaya
and Stock, 1997) and stored at 10 o C. Only fresh cultures were
used in the pathogenicity.
Larvae of Leucinodes orbonalis (shoot and fruit borer),
Earias vittella (spotted bollworm), Spodoptera litura
(tobacco cutworm), Plutella xylostella (diamondback moth),
Pieris brassicae (cabbage butterfly), Spilosoma obliqua
(Bihar hairy caterpillar), Helicoverpa armigera (cotton
bollworm) and Holotrichia consanguinea (white grub) were
collected from infested brinjal shoots/fruits, okra fruits,
cabbage leaves, chickpea field and upper soil layer (up to 30
cm depth) of sugarcane fields, respectively. Whereas, the larvae
of Galleria mellonella (greater wax moth), Corcyra
cephalonica (rice meal moth) and Bombyx mori (silkworm)
were reared in laboratory and used in laboratory pathogenicity
experiment (Table 1).
Filter paper impregnation method was used by exposing
host insects (except white grubs) on filter paper at the bottom
of petri plate (dia. 100 mm) or specimen tube (dia. 37 mm;
height 100 mm). The nematode concentrations at a rate of 0,
10, 25, 50, 75, 100, 200, 300, 400 and 500 IJs/larva was distributed
over the filter paper and 10 healthy larvae were introduced in
each petri plate, covered with lid and kept at room temperature
(30±2 o C). The lethal effect of nematode concentration was
checked and per cent larval mortality was recorded at every
24 h interval up to 3 to 7 days. H. armigera larva was treated
singly in specimen tube. Cannibalism among P. brassicae was
observed when larvae were introduced in bigger petri plates
and therefore, pathogenicity was repeated in small Petri dish
(dia. 40 mm) in which single larva of P. brassicae was confined
and treated with a predefined nematode concentration. In case
of white grub, soil column assay was conducted by placing a
single grub in specimen tube filled with sterile moist soil (15%
moisture w/w) to a depth of 40 mm. The grubs were allowed to
acclimatize at room temperature for 24 h prior to exposure of
nematode concentrations. The pathogenicity experiment on
H. consanguinea was repeated twice as highly variable results
were observed in earlier experiment. There were 10 treatments
with four replicates per treatment whereas in P. brassicae and
H. consanguinea pathogenicity, each treatment had 40
replications.
Further, dead cadavers were placed on modified White
traps, emerging progeny were harvested within 2 weeks of
infection.
The per cent mortality of insect larvae was first arcsine
transformed before analysis of ANOVA. The means were

PAL et al., Pathogenicity and Mass Production of Entomopathogenic Nematode, Heterohabditis indica 39
separated using Least Significant Difference (LSD). The
differences between treatments were considered significant
at P < 0.05.
RESULTS AND DISCUSSION
The results showed that most susceptible host insects
were C. cephalonica and G. mellonella while the poorest one
was Holotrichia sp. (Table 2). Absolute mortality of C.
cephalonica/L. orbonalis and G. mellonella/E. vittella were
obtained at 72 and 120 h of treatment, respectively at different
concentration of IJs/larva. More than 95% larval mortality
was observed in C. cephalonica and H. indica was very
efficient and caused absolute mortality of C. cephalonica at a
dose of 100 IJs/larva at 48 h whereas Karunakar, et al., 1999
and Zaki, et al., 2000 achieved the same level of kill at 37.6 and
96 h, respectively. At 48 h of treatment application more than
95% larval mortality was observed in L. orbonalis, and = 85%
mortality in G. mellonella. Hussaini et al. (2002) reported
maximum mortality of L. orbonalis at an inoculum level of 50
IJs/larva within 48 to 72 h of exposure of H. indica. However,
86.7% mortality was obtained in E. vittella (except dosage 10,
25 and 50 IJs/larva) at 96 h. while, no larval mortality was
recorded in control. In case of Holotrichia consanginea,
erratic results were observed but 26.7 to 66.7% mortality was
noted down within first 48 h of treatment which increased to
60 to 100% at 168 h. Kumar, et al., 2003 reported significant
mortality (17 to 89%) by inoculation of Heterorhabditis indica
³ 50 IJs/100 g soil within 48 h whereas 50% mortality was
observed at 72 h of exposure to 100 IJs whereas, Hussaini, et
al., 2005b found 30-40% mortality of white grub by S.
carpocapsae and S. abbasi whereas H. indica and H.
bacteriophora caused 20 to 25% mortality at 10 days of
nematode treatment.
In S. litura, 12.5 to 60% mortality was recorded at 48 h of
exposure of different concentrations of H. indica. However at
96 h, 100% mortality was obtained at dosage of 100 and more
than 400 IJs/larva whereas 87.5% mortality was obtained at a
dose of 25 IJs/larva.
Bombyx mori and Plutella xylostella were found highly
susceptible, in which absolute mortality was observed at
dosages ³ 25 and ³ 50 IJs/larva, respectively at 48 h. Whereas
at 24 h, 17.5 to 87.5% and 27.5 to 90% mortalities were recorded
at all dosages, respectively. Singh and Shinde, 2002 and
Saravanapriya and Subramanian, 2007 reported final instar
larvae of P. xylostella as most susceptible stage with an LC 50
value of 9 and 2 IJs/larva, respectively. While, Zaki, et al.,
2000 obtained 100% mortality of third instar larvae of B. mori
at 72 h.
Highly variable mortalities were observed in S. obliqua
and P. brassicae to which nematode dosages appeared to
have less effect on per cent mortality. Highest mortality of
82.5% was recorded in S. obliqua at 500 IJs/larva whereas
56.7% mortality occurred at dosages of 300 and 500 IJs/larva
at 96 h. At 48 h, mortality of 25 to 72.5% and 3.3 to 33.33% were
caused at all dosages. Ali, et al., 2005d reported S. obliqua as
poor host for nematode
In case of H. armigera, absolute mortality was recorded
at dose of ³ 75 IJs/larva at 72 h. More than 73% mortality was
noted at dose ³ 50 IJs/larva at 48 h. The result corroborates
the report of Jothi and Mehta, 2006 who achieved 100%
mortality of H. armigera by H. indica at an inoculum of 80 IJs/
larva.
The results showed that G. mellonella (50,000–200,000
IJ/Larva) produced maximum IJ followed by H. armigera
(50,000–150,000 IJ/Larva) could be a potential host for H.
indica production. The next effective host for the maximum IJ
production was H. consanguinea (20,000–100,000 IJ/Larva) >
S. litura and C. cephalonica (10,000–30,000 IJ/Larva) > E.
vittella (3,000-20,000 IJ/Larva) > P. brassicae (10,000-15,000
IJ/Larva). Comparatively, rearing of C. cephalonica (10,000–
30,000 IJ/Larva) and are easy and require less labour and
contamination chances are less. Nevertheless, proper care
and regular monitoring are crucial in rearing these insects.
The minimum IJ production was recorded in P. xylostella larvae
(100-2000 IJ/Larva) followed by B. mori (2000-3000 IJ/Larva).
This is in contradictory reports of Pribhuraj, et al., 2003 who
reported B. mori as a suitable host for production of an
unidentified Heterorhabditis sp. However, present study
corroborates the report of Zaki, et al., 2000 about unsuitability
of fifth instar B. mori for in vivo mass culturing; however a
maximum of 2,750 juveniles of H. bacteriophora was harvested
from third instar larva. The above results are corroborate with
the findings of Ali, et al., 2005c; Hussaini, 2003.
Table 1.
Insect pests of various crops against which pathogenicity of Heterorhabditis indica was tested.
Scientific name Common name Order: Family Commodity
Leucinodes orbonalis Early shoot and fruit borer Lep.: Crambidae Eggplant/brinjal
Earias vittella Spotted bollworm Lep.: Noctuidae Bhindi, cotton
Spodoptera litura Tobacco cutworm Lep.: Noctuidae Tobacco, tomato
Helicoverpa armigera Cotton bollworm / legume pod borer Lep.: Noctuidae Cotton, vegetables, pulses, cereals
Plutella xylostella Diamondback moth Lep.: Plutellidae Crucifers
Pieris brassicae Cabbage butterfly Lep.: Pieridae Cabbage
Galleria mellonella Greater wax moth Lep.: Pyralidae Honey comb
Corcyra cephalonica Rice meal moth Lep.: Pyralidae Stored grains
Bombyx mori Mulberry silkworm Lep.: Bombycidae Mulberry
Holotrichia consanguinea White grub Col.: Scarabaeidae Sugarcane
Spilosoma obliqua Bihar hairy caterpillar Lep.: Arctiidae Vegetables; crucifers

40 Trends in Biosciences 5 (1), 2012
Table 2. Pathogenicity of Heterorhabditis indica against lepidopteran and coleopteran insects at different dosages (10, 25,
50, 75, 100 to 500 IJs/larva) and progeny produced (mass production of IJs)
Insects
Per cent mortality of insect larva/grub hours after inoculation
(instar tested) 24 48 72 96
Corcyra cephalonica (L5) 10 to 80% mortality at 95% mortality at 100% mortality at 50
all dosages
50 IJs/larva)
IJs/larva
Galleria mellonella (L5) 10 to 95% mortality at
all dosages
Leucinodes orbonalis (L4 and
L5)
Earias vittella (L4 and L5)
Holotrichia consanguinea (G4
and G5)
Spodoptera litura (L4)
Helicoverpa armigera (L4)
Spilosoma obliqua (L4)
10 to 92.5% mortality
at all dosages
60% mortality at
300 IJs/larva
Bombyx mori (L5) 17.5 to 87.5%
mortality at all dosages
Plutella xylostella (L5)
Pieris brassicae (L4)
27.5 to 90% mortality
at all dosages
85% mortality at 50
IJs/larva and above
dosage
95% mortality at
75 IJs/larva
50% mortality at all
dosages
26.7 to 66.7%
mortality at all dosages
12.5 to 60% mortality
at all dosages
73.3% mortality at
50 IJs/larva
25 to 77.5% mortality
at all dosages
92.5% mortality at 10
IJs/larva; 100%
mortality at 25
IJs/larva
85 and 95% mortality
at 10 and 25 IJs/larva,
resp.; 100% mortality
at 50 IJs/larva
3.3 to 33.3% mortality
at all dosages
100% mortality at 25
IJs/larva
47.5% mortality at
50 IJs/larva
100% mortality at 75
IJs/larva
80 to 95% at dosage 10,
25 and 75 IJs/larva;
100% mortality at other
dosages
86.7 to 96.7% mortality
at 10 to 75 IJs/larva;
100% mortality at other
dosages
53.3 to 80% mortality at
all dosages
87.5% mortality at
25 IJs/larva; 100%
mortality at 100 and
400 IJs/larva
Highest mortality of
82.5% at 500 IJs/larva
Highest mortality of
56.7% at dosage of 300
and 500 IJs/larva
Progeny produced
(IJs/larva)
10,000 – 30,000
50,000 – 200,000
5,000 – 15,000
3,000 – 20,000
20,000 – 100,000
10,000 – 30,000
50,000 – 150,000
5,000 – 10,000
2,000 – 3,000
100 – 2,000
10,000 – 15,000
ACKNOWLEDGEMENT
The authors are very thankful to Dr. Abid Hussain and
Dr. Milan Prasad for their valuable time and suggestion during
research work. Authors also thankful to ICAR funded Research
Project “Niche Area Excellence Programme” for their financial
support.
LITERATURE CITED
Ali, S.S. Ahmad, R, Hussain, M.A. and Pervez, R. 2005d. Pest
Management in Pulses through Entomopathogenic Nematodes.
Indian Institute of Pulses Research, Kanpur, India, pp. 58.
Hussaini, S.S. 2003. Progress of research work on entomopathogenic
nematodes in India, pp. 27-68. In: Current Status of Research on
Entomopathogenic Nematodes in India (eds. S.S. Hussaini, R.J.
Rabindra and M. Nagesh). Project Directorate of Biological Control,
Bangalore, India.
Jothi, B.D., Mehta, U.K. 2006. Pathogenicity of three species of EPN
against cotton bollworm Helicoverpa armigera Hub. Entomon, 31:
259-266.
Karunakar, G., Easwaramoorthy, S., David, H. 1999. Susceptibility of
nine lepidopteran insects to Steinernema glaseri, S. feltiae and
Heterorhabditis indicus infection. International Journal of
Nematology, 9: 68-71.
Kaya, H.K. and Stock, S.P. 1997. Techniques in insect nematology, In:
Manual of Techniques in Insect Pathology (ed. LA Lacey). Academic
Press, San Diego, CA, pp. 281-324.
Kumar, M.R., Parihar, A., Siddiqui, A.U. 2003. Effects of
entomopathogenic nematode, Heterorhabditis sp., on Spodoptera
litura. Annals of Plant Protection Sciences, 11: 406-407.
Prabhuraj, A., Viraktamath, C.A., Kumar, A.R.V. 2003. Evaluation of
silkworm larva and root grub for the in vivo mass production of
entomopathogenic nematodes. Indian Journal of Entomology, 65:
34-37.
Saravanapriya, B., Subramanian, S. 2007. Pathogenicity of EPN to
certain foliar insect pests. Annals of Plant Protection Sciences, 15:
219-222.
Singh, N.P., Shinde, S. 2002. Relative susceptibility of different life
stages of Plutella xylostella (L.) to entomopathogenic nematode,
Heterorhabditis bacteriophora Poinar. Entomon, 27: 281-285.
Zaki, F.A., Mantoo, M.A., Gul, S. 2000. In: vivo culturing of
entomopathogenic nematodes Heterorhabditis bacteriophora and
Steinernema carpocapsae on silkworm (Bombyx mori) and their
effect on some lepidopterous insects. Indian Journal of Nematology,
30: 1-4.
Recieved on 29-2-2012 Accepted on 30.3.2012

Trends in Biosciences 5 (1): 41-44, 2012
Bioefficacy of Cow Urine Decoction (CUDs) of Different Plants on Population Growth
of Lipaphis erysimi (Kalt.) on B. juncea under Field Conditions
WAJID HASAN* AND M.S. ANSARI
Department of Plant Protection, Faculty of Agricultural Sciences, Aligarh Muslim University,
Aligarh-202002 (U.P.)
*e-mail: entowajid@gmail.com
ABSTRACT
The per cent population growth of L. erysimi after two sprays of
CUDs was lowest in neem oil 0.005% (-56.2) followed by A.
indica (-20.9), neem oil 0.003% (-18.6), A. squamosa (3.1), C.
citriodora (8.9), P. hysterophorus (10.7), A. cepa (15.4), A. sativum
(24.4), C. papaya (27.5), C. gigantea (66.5) and highest in
untreated control (76.3) in timely sown crop. While in late
sown crop it was lowest in Neem oil 0.005% (-15.1) followed by
neem oil 0.003% (-14.5), A. cepa (-10), A. indica (-7.8), A. sativum
(-6.7), A. squamosa (-1.9), C. gigantea (6.2), C. papaya (7.7), C.
citriodora (7.7), P. hysterophorus (7.8) and highest in untreated
control (24.7). On the basis of percent population growth, A.
indica was the best among all CUDs used.
Key words
Cow urine decoctions, lipaphis erysimi, bioefficacy
Out of various insect-pests associated with Rapeseed-
Mustard, Lipaphis erysimi Kalt. is the key pest which feeds
by sucking sap from its host and damage to the crop ranging
from 9 to 96% and reach up to 100% (Bakhetia, 1986; Singh
and Sachan, 1999; Parmar, et al., 2007; Singh and Sharma,
2002). An estimate revealed that 2, 121 plant species possess
pest control properties, out of which 1,005 species have
insecticidal, 384 antifeedant, 297 repellents, 27 attractants and
31 growth inhibitory properties (Reheja, 1998). In the era of
environment awareness, more emphasis is given to the natural
insecticides, as they are biodegradable and less harmful to
environment. Considering the economic importance of the
pest and to reduce the poisonous effect of chemical
insecticides to natural enemies, cow urine decoctions (CUD)
of botanicals were tried for its efficacy against mustard aphid,
L. erysimi by the keeping in view the finding of Gupta, 2005;
Purwar and Yadav, 2004; Hasan and Singh, 2008, 2009 and
Manglik, 2002. Biopesticide applications that conserve
biological activity post-treatment, provide additional
suppression of pest populations. Use of biopesticides that
can be used when pollinators are present, would allow
improved IPM practices by replacing prophylactic and more
harmful cover sprays with a program that emphasizes treatment
based on economic thresholds and conserves beneficial
biological activity (Hasan and Singh, 2009). The present
investigation was undertaken to study the bioefficacy of cow
urine decoctions (CUD) @5% of plants viz., Allium cepa,
Allium sativum, Annona squamosa, Azadirachta indica,
Cariaca papaya, Parthenium hysterophorus, Calotropis
gigantea, Cymbopogon citriodora and neem oil (@ 0.005
and 0.003%) were evaluated against nymphs of L. erysimi
under filed conditions.
MATERIALS AND METHODS
The experiments were conducted during 2010-11 at
Ramgarh Panjeepur village of Aligarh district of U.P. to test
the bioefficacy of cow urine decoctions (CUD) of plants viz.,
A. cepa, A. sativum, A. squamosa, A. indica, C. papaya, P.
hysterophorus, C. gigantea, C. citriodora @5% and neem oil
(@ 0.005 & 0.003%) were evaluated against per cent population
growth of L. erysimi under field conditions. Seed of Indian
mustard (B. juncea cv. chinkara NV-4102) was sown on 18
October, 2010 (timely sown) and 24 November, 2010 (late sown).
Experiments were laid out in randomized block design (RBD)
with five replications. Standard recommended cultural
operations were followed to raise the healthy crop of Indian
mustard except the plant protection measurers. To prepare
the cow urine decoctions (CUD) of 250 gm leaves were added
in 500 ml of cow urine and allow it to boil slowly up to half
original volume was evaporated than it was cooled. After that
material was filtered with muslin cloth than filled in bottle and
stored into the cool and dry place. Two sprays were conducted
at 12 days intervals with knapsack sprayer. The number of
aphids was counted on 10 cm apical central shoot of
inflorescence on 10 tagged plants in each replication before
spray, one, three, five and seven days after spray.
RESULTS AND DISCUSSION
Tables 1 and 2 revealed that the per cent population
growth of L. erysimi after two sprays of CUDs was lowest in
neem oil 0.005% (-56.2) followed by A. indica (-20.9), neem oil
0.003% (-18.6), A. squamosa (3.1), C. citriodora (8.9), P.
hysterophorus (10.7), A. cepa (15.4), A. sativum (24.4), C.
papaya (27.5), C. gigantea (66.5) and highest in untreated
control (76.3) in timely sown crop. While in late sown crop it
was lowest in neem oil 0.005% (-15.1) followed by neem oil
0.003% (-14.5), A. cepa (-10), A. indica (-7.8), A. sativum (-6.7),
A. squamosa (-1.9), C. gigantea (6.2), C. papaya (7.7), C.
citriodora (7.7), P. hysterophorus (7.8) and highest in untreated
control (24.7). On the basis of per cent population growth, A.
indica was the best among all CUDs used.
The highest mean yield i.e. 1925.83 Kg./ha was recorded

42 Trends in Biosciences 5 (1), 2012
Table 1.
Bioefficacy of cow urine decoction (CUDs) of different plants on percent population growth of L. erysimi under field
conditions (Timely sown)
Treatments
First spray
Second spray
Population
Before
spray
One
DAS
Three
DAS
Five
DAS
Seven
DAS
Percent
increase
Before
spray
One
DAS
Three
DAS
Five
DAS
Seven
DAS
Percent
increase
Growth
(%)
1 2 3 4 5 6 7 8 9 10 11 12 13 14
Allium sativum
72 72 68 78 92
134 126 152 150 162
27.8
(Garlic)
(8.43) (8.43) (8.19) (8.79) (9.46)
(11.40) (11.01) (12.08) (12.03) (12.56)
20.9 24.4
Cariaca papaya 68 73 81 88 102
200 190 206 202 210
50
(Papaya)
(8.23) (8.53) (8.90) (9.35) (10.19)
(14.11) (13.72) (14.29) (14.19) (14.46)
5 27.5
Calotropis gigantean 66 71 88 126 142
192 188 194 198 226
115.2
(Milk weed)
(8.09) (8.39) (9.36) (11.21) (11.91)
(13.81) (13.69) (13.86) (13.95) (14.95)
17.7 66.5
Parthenium
hysterophorus
62 70 72 72 78 25.8 176 154 134 132 168 -4.5 10.7
(Congress grass) (7.85) (8.36) (8.48) (8.46) (8.76)
(13.24) (12.40) (11.57) (11.48) (12.94)
Azadirachta indica 70 66 50 46 52
200 208 178 168 168
-25.7
(Neem)
(8.24) (8.04) (6.96) (6.71) (7.15)
(14.10) (14.39) (13.31) (12.94) (12.95)
-16 -20.9
Allium cepa
62 64 68 74 84
214 226 216 198 204
35.5
(Onion)
(7.79) (8.0) (8.18) (8.58) (9.15)
(14.59) (15.00) (14.63) (14.03) (14.21)
-4.7 15.4
Annona squamosa 80 78 80 84 92
246 232 212 208 224
15
(Sharifa)
(8.85) (7.96) (8.91) (9.15) (9.56)
(15.64) (15.19) (14.52) (14.35) (14.88)
-8.9 3.1
Cymbopogon citriodora 68 75 71 82 92
238 222 214 200 196
35.3
(Lamon grass) (8.23) (8.72) (8.41) (9.03) (9.57)
(15.33) (14.77) (14.55) (14.05) (13.95)
-17.6 8.9
Neem oil (0.005%) 80 73 63 16 26
176 163 108 62 97
-67.5
(Neemerin)
(8.92) (8.64) (7.93) (3.95) (5.08)
(13.25) (12.74) (10.38) (7.84) (9.86)
-44.9 -56.2
Neem oil (0.003%) 70 60 42 12 74
196 148 102 104 112
5.7
(Neemerin)
(8.36) (7.73) (6.31) (3.38) (7.77)
(13.98) (12.16) (10.08) (10.19) (10.58)
-42.9 -18.6
Untreated
70 86 108 142 154
202 213 234 240 268
120
(8.31) (9.25) (10.33) (11.91) (12.39)
(14.09) (14.47) (15.17) (15.42) (16.29)
32.67 76.3
CD (P?0.01) 1.681 1.568 1.761 1.334 2.698 2.34 2.372 2.481 2.494 2.459
CD (P?0.05) 1.256 1.172 1.316 0.997 2.016 1.75 1.773 1.854 1.864 1.837
SE±m 0.439 0.410 0.460 0.349 0.705 0.61 0.620 0.649 0.652 0.643
*Figures in parenthesis indicate square root transformation, DAS= Days After Spray,
Population Growth (%)= (column 7 + column 13)/2, Percent increase =percent of (column 6- column 2)
in plots treated with neem oil (0.005) followed with neem oil
(0.003) 1885.00, 1825.16 Kg./ha with CUDs of A. indica, 1816.16
Kg./ha with CUDs of A. squamosa, 1782.66 Kg./ha with CUDs
of A. sativum, 1742.83 Kg./ha with CUDs of A. cepa, 1742.33
Kg./ha with CUDs of C. citriodora, 1738.16 Kg./ha with CUDs
of C. gigantea, 1722.50 Kg./ha with CUDs of P. hysterophorus,
1705.50 Kg./ha with CUDs of C. papaya and lowest 1654.16
Kg./ha in untreated control (Table 3). The mean pooled yield
was higher in timely sown crop (1928.18 Kg./ha) and lower in
late sown crop (1624.60 Kg./ha).
Similar results were recorded by Hasan and Singh, 2008.
Gupta, et al., 2005 reported the efficacy of neem in combination
with cow urine against mustard aphid. Sachan and Bansal,
1975 reported that the first, second, third and fourth nymphal
stages last for 1-2, 2, 2, 3 days respectively and wingless
females produce 70-87 nymphs in the lifetime while winged
females produce 31-40 nymphs. Rai and Gupta, 2002 studied
the effect of neem [Azdirachta indica] kernel extract in cow
urine (NSKE; 30 ml/litre), neem oil (1.0%), dimethoate (0.045%),
NSKE + dimethoate (0.03%), NSKE (3.0%) + dimethoate
(0.03%), neem oil (1.0%) + dimethoate (0.03%), and neem
kernel extract in cow butter milk (NCBM; 20 ml/litre) against L.
erysimi on Brassica juncea (cv. Pusa Bold). Purwar, 2001
reported that in soybean that cow urine in 5%, 10%, 25% and
75% has some suppressing effect on larval population of
defoliators (Manglik, 2002). Abebe, 1999 observed that cow
urine 10% and cow dung ash is effective against tobacco
caterpillar (S. litura), caster semilooper (P. orichalcea) of
soybean. Hasan and Singh, 2008 and 2009 reported the
aphidicidal activity of cow urine decoctions of botanicals
against mustard aphid, Lipaphis erysimi Kalt. and coriander
aphid, Hyadaphis coriandri (Das.). This information would
support growers wanting to transition efforts to reduce their
use of higher risk pesticides. Thus it was concluded that all
the above CUDs were found to be less suitable for growth
and development of aphid with respect to the control.
ACKNOWLEDGEMENT
The research work has been sponsored by the
University Grant Commission under the UGC DR. D.S. Kothari
Post Doctoral Fellowship Scheme.

Table 2.
HASAN AND ANSARI, Bioefficacy of Cow Urine Decoction (CUDs) of Different Plants on Population Growth 43
Bioefficacy of cow urine decoction (CUDs) of different plants on percent population growth of L. erysimi under field
conditions (Late Sown)
Treatments
First spray
Second spray
Population
Before
spray
One
DAS
Three
DAS
Five
DAS
Seven
DAS
Percent
increase
Before
spray
One
DAS
Three
DAS
Five
DAS
Seven
DAS
Percent
increase
Growth
(%)
1 2 3 4 5 6 7 8 9 10 11 12 13 14
Allium sativum
256 244 232 245 240
480 442 430 404 446
-6.2
(Garlic)
(15.97) (15.58) (15.18) (15.65) (15.49)
(21.84) (20.96) (20.68) (20.05) (21.07)
-7.1 -6.7
Cariaca papaya 240 252 240 262 282
500 476 458 362 391
17.5
(Papaya)
(15.45) (15.83) (15.44) (16.14) (16.75)
(22.13) (21.56) (21.21) (18.75) (19.54)
-2.1 7.7
Calotropis gigantean 250 252 244 238 282
446 426 396 418 444
12.8
(Milk weed)
(15.78) (15.81) (15.57) (15.37) (16.79)
(21.08) (20.60) (19.85) (20.39) (21.00)
-0.4 6.2
Parthenium
hysterophorus
200 218 232 230 230 15 370 351 334 354 372 0.5 7.8
(Congress grass) (14.09) (14.69) (15.17) (15.05) (15.07)
(19.14) (18.64) (18.20) (18.72) (19.21)
Azadirachta indica 164 182 190 168 152
300 270 240 266 275
-7.3
(Neem)
(12.70) (13.49) (13.76) (12.88) (12.32)
(17.25) (16.38) (15.48) (16.31) (16.58)
-8.3 -7.8
Allium cepa
202 178 190 152 176
310 285 246 228 188
-12.9
(Onion)
(14.19) (13.30) (13.76) (12.25) (13.20)
(17.53) (16.80) (15.65) (15.07) (13.57)
-7.1 -10
Annona squamosa 210 196 186 176 204
300 296 292 272 297
-2.8
(Sharifa)
(14.45) (13.97) (13.62) (13.25) (14.21)
(17.32) (17.15) (17.05) (16.45) (17.23)
-1 -1.9
Cymbopogon
citriodora (Lamon 191 163 184 196 208 8.9 250 228 248 254 266 6.4 7.7
grass)
(13.78) (12.68) (13.54) (13.95) (14.37)
(15.78) (15.06) (12.38) (15.90) (16.28)
Neem oil (0.005%) 170 150 158 132 142
220 196 156 196 190
-16.5
(Neemerin)
(13.04) (12.24) (12.57) (11.46) (11.89)
(14.81) (13.95) (17.70) (14.00) (13.78)
-13.6 -15.1
Neem oil (0.003%) 230 210 182 158 195
190 184 150 104 164
-15.2
(Neemerin)
(15.14) (14.47) (13.45) (12.52) (13.00)
(13.78) (13.51) (12.18) (10.19) (12.81)
-13.7 -14.5
Untreated
240 254 260 290 310
305 322 330 326 382
29.2
(15.45) (15.90) (16.08) (17.03) (17.6)
(17.39) (17.87) (18.02) (17.98) (19.49)
20.2 24.7
CD (P?0.01) 2.079 2.068 1.901 2.525 2.835 3.292 3.422 3.237 2.908 3.329
CD (P?0.05) 1.554 1.545 1.434 1.887 2.119 2.459 2.558 2.419 2.173 2.488
SE±m 0.544 0.541 0.502 0.660 0.741 0.861 0.895 0.846 0.760 0.870
*Figures in parenthesis indicate square root transformation, DAS= Days After Spray,
Population Growth (%)= (column 7 + column 13)/2, Percent increase =percent of (column 6- column 2)
Table 3.
Effect of cow urine decoction (CUDs) of different plants and date of sowing on yield of B. juncea
Treatments
Yield (Kg./ha)
Timely Sown Late Sown Mean
Allium sativum (Garlic) 1935.33 (44.00) 1630.00 (40.39) 1782.66 (42.19)
Cariaca papaya (Papaya) 1840.33 (42.91) 1570.66 (39.64) 1705.50 (41.28)
Calotropis gigantean (Milk weed) 1870.66 (43.26) 1605.66 (40.08) 1738.16 (41.67)
Parthenium hysterophorus (Congress grass) 1855.00 (43.08) 1590.00 (39.89) 1722.50 (41.49)
Azadirachta indica (Neem) 1989.66 (44.62) 1660.66 (40.76) 1825.16 (42.73)
Allium cepa (Onion) 1875.33 (43.32) 1610.33 (40.14) 1742.83(41.74)
Annona squamosa (Sharifa) 1981.33 (44.52) 1651.00 (40.64) 1816.16 (42.61)
Cymbopogon citriodora (Lamon grass) 1881.00 (43.38) 1603.66 (40.06) 1742.33 (41.73)
Neem oil (0.005%) (Neemerin) 2122.33 (46.08) 1729.33 (41.59) 1925.83 (43.88)
Neem oil (0.003%) (Neemerin) 2079.66 (45.61) 1690.33 (41.12) 1885.00 (43.42)
Untreated 1779.33 (42.19) 1529.00 (39.11) 1654.16 (40.67)
Mean 1928.18 (43.91) 1624.60 (40.31) 1776.39 (42.14)
Treatments (a) Date of sown (b) axb
CD (P?0.01) 0.048 0.021 0.068
CD (P?0.05) 0.036 0.015 0.051
SE±m 0.013 0.005 0.018
*Figures in parenthesis indicate square root transformation
LITERATURE CITED
Abede, M. 1999. Comparison of newer and traditional methods to
control the insect pest of soybean. M. Sc. Thesis, G.B.P.U.A.&T.,
Pantnagar, p.89.
Bakhetia, D.R.C. 1986. Control of insect pest of toria, sarson & rai.
Indian Farming, 36: 43-44.
Gupta, M.P. 2005. Efficacy of neem in combination with cow urine
against mustard aphid and its effect on coccinellid predators. Natural
Product Radiance, 4(2): 102-106.

4 4 Trends in Biosciences 5 (1), 2012
Hasan Wajid and Singh, C. P. 2008. Bioeffecacy of cow urine decoctions
of botanicals against mustard aphid, Lipaphis erysimi (Kalt.) and
coriander aphid, Hyadaphis coriandri (Das.) (Homoptera:
Aphididae). Journal of Aphidology, 22(1&2): 41-46.
Hasan Wajid and Singh, C.P. 2009. Efficacy of cow urine decoctions of
botanicals against mustard saw fly. Annals of Plant Protection
Sciences, 17(1): 234-235.
Manglik, G. 2002. Field efficacy of Cow urine in comparison to chemical
and Bio pesticide against insect pests of soybean. M.SC. Thesis,
G.B.P.U.A. & T., Pantnagar. pp. 86.
Parmar, G.M., Kapadia, M.N., Jadav, N.B. and Zizala, V.J. 2007. Avoidable
losses due to Lipaphis erysimi (Kalt.) in mustard. Asian Journal of
Bio Science, 2(1/2): 73-75
Purwar, J.P. and Yadav, S.K. 2004. Evaluation of age related response
of Spilarctia oblique (Walker) to biorational insecticides. Annals
of Plant Protection Sciences, 12(20): 271-273.
Purwar, J.P. 2001. Evaluation of bioregional pesticide against major
insect pests of soybean. M.SC. Thesis, G.B.P.U.A. & T., Pantnagar.
p.108.
Raheja, A.K. 1998. Role of crop protection chemicals in IPM. Pestology,
22 (4): 6-11.
Rai, H.S. and Gupta, M.P. 2002. Integrated management of mustard
aphid, Lipaphis erysimi Kalt. Annals of Plant Protection Sciences,
14(1): 76-79.
Sachan, J.N. and Bansal, O.P. 1975. Influence of Different Host Plants
on the Biology of Mustard Aphid, Lipaphis erysimi Kalt. Indian
Journal of Entomology, 37(4): 420-424.
Singh, C.P. and Sachan, G.C. 1999. Ecofriendly management of Lipaphis
erysimi kalt. in Brassica carinata. Proceeding of 10 th International
Rapeseed Conference Canberra, Australia.
Singh, Y.P. and Sharma, K.C. 2002. Integrated approach to manage the
mustard aphid, Lipaphis erysimi (Kaltenbach) (Homoptera:
Aphididae) in oilseed Brassica crops- A review. Journal of
Amphibiology, 16:77-88.
Vaibhav Mathur, Singh, C. P. and Hasan Wajid. 2011. Effect of cow
urine decoction of some plants on the biology of Lipaphis erysimi
(Kalt.). Proceedings of National Conference on 3 rd Congress on
Insect Science (April 18-20, 2011) held at Department of
Entomology, Punjab Agricultural University, Ludhiana. pp. 77.
Recieved on 17-3-2012 Accepted on 05.4.2012

Trends in Biosciences 5 (1): 45-46, 2012
Effect of Different Formulations of Bacillus thuringiensis on Larval Mortality of
Diacrisia obliqua
ZEENAT WARSI AND AJAY CAPOOR
Department of Zoology, Agra College, Agra, U.P.
ABSTRACT
This study was carried out to determine the effect of commercial
preparation of Bacillus thuringiensis (biocontrol agent) namely
Dipel, Thuricide HP and Bactospeine against the Bihar hairy
caterpillar that is Diacrisia oblique walker of the family
Arctiidae belong to order Lepidoptera .Five concentrations i.e.
0.05, 0.10,0.50, 0.75, and !.0% of different formulations are
used against the third instar larvae of D. obliqua. All these
three microbial prepration that is Dipel, Thuricide HP and
Bactospeine were toxic to the test insect but Dipel proved to
be most effective.
Key words
Mortalit, diacrisia obliqua, formulation, efficacy
Many conventional insecticides such as cholorinated
hydrocarbons, organophosphates, carbonates etc., were used
in the controlling insect pest but, many of these insecticides
are harmul to man and beneficial organism which causes
ecological disturbense. Newer compound were developed
which affecting mortality, growth and developmental
processes in insects, such as chitin synthesis inhibitors and
different formulation of Bacillus thuringiensis. Progress has
been made to introduce iimproved biocontrol agents such as
B.T. for controlling Lepidopteran, Coleopteran and Dipteran
pest. B.T. kills insects primarily through the action of delta
indotorxins which is a protenieous constituent produced
during sporulation. It affects the insect midgut epithelium upon
ingestion (Narayan and Jayraj, 1975).
Most conventional B.T. products are based on the sub
species kurstaki HD-1 introduced by Abbott in laboratory
followed by Sandoz, Nova, Ecogen, Monsato for controlling
lepidopteran pest. Various formulation of B.T. kurstaki sucah
as Dipel ‘HD-1 Abbott) Thuricide, (HD-1 Sandoz), Biobit (HD-
1 Nova), Javelin (NRD-12, Sandoz) are available for controlling
the lepidoteran pest. Diacrisia obliqua Walker belonging the
family Arctiidae and order Lepidoptera and commonly known
as Bihar Hairy caterpillar. The population of D. obliqua started
growing faster and reached an explosive stage in August 2002.
A perusal of literature divulge that work done in India related
to effect of B.T. formulation on D. obliqua is very secured
and fragmentary. However there are some important aspects
like effect of exposure period on insects mortality, effect of
lethal and sub-lethal concentration of B.T. on the mortality
growth and development of insects.
MATERIALS AND METHODS
Three commercial preparations of B.T. (Bacillus
thuringiensis) namely Dipel, Thuricide HP and
Bactospeine.These are commercial preparations of B.T used
against the third instar larvae of D. obliqua taken from the
laboratory stock. Five concentrations i.e. 0.05, 0.10, 0.50, 0.75
and 1.0% of different formulations are used. Uniform size of
castor plants were used and these plants were treated with
each concentration of bacterial preparations by leaf dip
method. In this method of treatment small and uniform size of
leaves of host plant (castor plant) were treated with each
concentration of particular bacterial preparations by dipping
the leaf in microbial preparation of B.T.
Twenty larvae were taken from laboratory stock already
starved for 12 hours. After 12 hours starvation larvae were
released on the treated leaves for 24 hours for feeding and
after that normal food was offered to them for the rest period.
Observatinos on moratlity of larvae were recorded after 24
hours and continued for the period of 10 days. Moribund
(dying condition) larvae were also counted as dead. In control
larvae were treated with water containing 2% skimmed milk
solution.
The data thus recorded were subjected to probit analysis.
for calculating the LC50 and LT50 values. The LT50 was
calculated log days in place of log concentration on the basis
of screening different bacterial preparations of B.T.
RESULTS AND DISCUSSION
Data pertaining to influence of bacterial preparations
against the larvae of D. obliqua shows that all the three
microbial prparations i.e. Dipel, Thuricide HP, Bactospeine
were toxic to the larvae of test insect but Dipel proved to be
the most effective followed by Thuricide HP and Bactospeine
at all the concentrations tested. It can be further seen that
LC50 value has negative corelation with the toxicity of
microbial preparations tested, the lowest (0.0930) was observed
in case of Dipel while it was highest against Bactospiene
Table 1.
Effect of Dipel, Thuricide HP and Bactospeine
against 3 rd instar larvae of D. obliqua
S. Name of Average per cent mortality LC50
No formulation Concentration of bacterial
preparations
0.025 0.05 0.10 0.25 0.50
1. Dipel 24.13 31.03 44.82 82.75 89.65 0.930
2. Thuricide 20.00 30.00 46.67 80.00 86.66 0.1039
HP
3. Bactospeine 13.33 23.33 30.00 66.67 80.67 0.1540

46 Trends in Biosciences 5 (1), 2012
Table 2.
Toxicity index of different bacterial preparations against 3 rd instar larvae of D. obliqua
S.No Name of formulation Spore count/gram of
active product
Regression
equation
Lietero
genecity
Fiducial limit
1. Dipel 25*10^9 Y=1.905+1.57 3.605 0.1312 (U)
0.066 (L)
2. Thuricide HP 30*10^6 Y=1.860+1.556 4.97 0.144 (U)
0.740 (L)
3. Bactospeine 10*10^8 Y=1.584+1.562V 2.28 0.2157 (U)
0.1096 (L)
Toxicity
1.00
0.89
0.60
(0.1540). Thuricide HP behaved intermediary (0.1039). Log
probit curves of these microbial preparations clearly shows
the values ranging from 1.556-1.571 (Table 1).
Low LC50 value for Dipel against other Lepidopteran
insects have also been reported by Srivastava and Nayak,
1978; Caturvedi, 2002.
As regards the toxicity index of different varieties of
microbial preparations used in this investigation, it can clearly
be seen from Table 2 that Dipel has the highest toxicity index
1.0 whereas it was lowest 0.60 in case of Bactospeine. Thuricide
HP had toxicity index (0.89).
Dipel took the minimum time (48 hrs) to to while kill of
bring the insect and maximum time (120 hrs) to 89.99%
mortality at one % concentration, whereas at the same
concentration Bactospeine consumed as much as 168 hrs to
start and 240 hrs for giving the highest (80%) larval mortality.
Results shown that all the formulations tested offered
effective control of the test insect but Dipel manifested it
superiority over the other microbial preparations. It is
interesting to note that larval mortality caused by Dipel was
89.99% and it was closely followed by Thuricide HP 86.66%
ACKNOWLEDGEMENT
Authors are grateful to Dr. A.V. Singh Principal, Agra
College, Agra and Dr. R.S. Rawat, Head, Department of
Zoology, Agra College, Agra for providing necessary facilities
and moral support during the course of the present
investigations.
LITERATURE CITED
Chaturvedi, R.K. 2003. Studies on the effectiveness of Bacillus
thuriengiensis against Utetheisa pulchella (Lipidoptera; Arctiidae),
Indian Society of Life Sciences. pp. 68-69.
Srivastava, R.P and Nayak, P. 1978. Laboratory evaluation of four
formulations of B. thuriengiensis against Cnaphalocrosis medinalisgn
(Pyrilidae;Lepidoptera) The rice roller, zeitschrift fiir
pflanzenkarankheitem un pflanzenschuz, 85(11): 641-644.
Narayan, K. and Jayraj, S. 1975. Effect of B. lhurengiensis on size,
weight and fat content of citrus leaf caterpillar, Papilio demdeus.
Madras Agric. J., 62: 367-370.
Recieved on 15-7-2011 Accepted on 20-12-2011

Trends in Biosciences 5 (1): 47-50, 2012
An Empirical Study of Land Degradation its Magnitudes and Casualities
ANIL KUMAR 1 , B.S. SACHAN 2 AND KESHAV PRASAD 2
1
Seed and Farm, C.S.A. University of Agriculture Technology Kanpur 02
2
Deptt. of Agril. Eco., C.S.A. University of Agriculture Technology Kanpur 02
ABSTRACT
A modest attempt has been made to study the impact of water
and fertilizer intensive agriculture on land degradation, in the
specific context of the situation in D.P., where such a technology
is known to be the predominant one while the increases in crop
production in this region have received considerable attention,
possible negative effects of the new technology have by and
large, gone unnoticed. Alternatively, it has led to exaggeration
of the possible dreading effect of this technology in some circles.
The present paper is an attempt in this direction in the context
of D.P. It shall postulate that the steady decrease in water
logging can be attributed to the changes in level of water flow
and use in the region, and salinity to the simultaneous use of
heavy doses of fertilizer. The variable, volume of canal water
per hectare of cultivated area, number of tube wells per
thousand hectares of cultivated area and rainfall capture the
moisture related factors and level of fertilizer use per hectare
of cropped area captures the fertilizer effect. Two models shall
be postulated to explain the variations in water logging and
salinity, both over time and between districts. These shall be
the ordinary least squares and the weighted least squares based
on linear probability model for the year cross-section analysis
and pool data for three sets of years (1986-87 to 88-89, 1994-95
to 1996-97 and 2003-04 to 2005-06 respectively) after estimating
district level data, on the following four sets’ of variables. Land
degradation includes waterlogged and saline land. Recent
studies indicate that high levels of salinity are found to occur
in regions where fertilizer consumption is high. Consumption
of fertilizer per hectare of cropped area is included as one of
the explanatory variables in the model to capture this impact.
In order to see what impact this increase had on the water
balance and hence on the problems of water logging and salinity.
The results of both the ordinary least squares and the weighted
least squares specifications are reasonably acceptable. All the
explanatory variables appear with the expected signs. Which
the variables, rainfall volume of canal water and level of
fertilizer use, land to increases in land degradations, the
number of tube-wells per thousand and hectares decreases land
degradation due to water logging and salinity. In the ordinary
least squares specification, the variables, volume of canal water
per hectare ,and fertilizer use, per hectare are significant at
the 10 per cent level. Our results show that the new agricultural
technology characterised, by water and fertilizer intensive
agriculture have had second round effects. Remedial measures
that help to limit the negative environmental impact to the
minimum should be planned for ideally, water resource
planning in a river basin should be based on long run estimates
of availability of water and land.
Key words
Land degradation, canal water, tubebell, fertilizers
The degradation of land that irrigation leads to takes
the form of water-logging and salinity. Large tracts of land go
out of cultivation due to waterlogging. Alternatively, the
productivity of cultivated areas decreases, with the
consequence that an ecological imbalance is initiated that
may result in a considerable dent being made in the benefits
resulting from land augmentation. Salinity is another related
from of land degradation.
Soils become saline as a result of concentration of salts
due to evaporation of groundwater moving upwards. At cases,
this is the outcome of a rise of groundwater from a shallow
water-table. In such cases, salinity appears together with
waterlogging. Alternatively, it could also appear independently
of any occurance of waterlogging (Karanth, 1996) in part, and
weather salinity together as elements of land degradation and
do not make an attempt to divide the total into its two
component parts. The crux of the problem appears to be a
distortion of the natural equilibrium between groundwater
discharge and recharge stemming from percolation losses in
the irrigated area, a phenomenon that hydrologists study with
the help of water-balance equations.
Simultaneously, however, this phenomenon has resulted
in both lower yields from land and larger amounts of land
going out of cultivation. In the Sharda Sahayak command
area, yields of paddy and wheat decreased by about 40 to 70
per cent due to waterlogging and by 50 to 60% on account of
salinity decreased paddy and wheat decreased by about 40 to
70% due to waterlogging and by 50 to 60% on account of
salinity. In Demol village of Kheda district in Gujarat it was
estimated that different levels of salinity decreased paddy
and wheat yields by 10 to 80% Further, in a study of four
Haryana villages it was found that farms having salinity had
to leave 25% of their as fallow as compared to only 4% on
farms without salinity.
Later, with the addition of other canal systems, in
particulars the Bhakra Canal system, the volume of water
flowing through the region increased and this led to changes
in the hydrology of the region. The increase in water flows
trough the region could be expected to result in waterlogging
and salinity. Even tough the soils in this part of the country
are mainly comprised of sand and silt mixed with clay, the
excessive use of surface water without adequate groundwater
pumping and sub-surface per year from 1997 to 1998-99. It has
been estimated that around 1996-97 there existed an annual
surplus groundwater-balance equivalent to a rise of 1.40
meters.

48 Trends in Biosciences 5 (1), 2012
In particular, Lakhimpur, Hathras ben tile chronically
waterlogged areas. Of late, waterlogging has increased in
Sitapur, Barabanki, Pilibhit districts also. Together these five
districts account for per cent of the waterlogged area in the
State. Simultaneously, the levels of fertilizer use have also
increased. Joshi, 2003, among others, has postulated that this
has resulted in increased levels of salinity, one of the
components of land degradation.
By 2006-07 60.90, thousand hectares were affected by
waterlogging and salinity. Up to 2000-01, there is an increase
in the magnitude of saline and waterlogged land. After that
year , there is a steady fall. By 2006-07 the area affected by
waterlogging and salinity had decreased to less than half its
extent in 2001-02. The factor landing to this fluctuation in the
levels of waterlogging need to be examined both on their own
account and because of the such an examination would
through on policy aspects of irrigation management.
It is postulatel that the steady decrease in waterlogging
can be attributed to the changes in the level of water flow and
use in the region, and salinity to the simultaneous use of
heavy doses of fertilizer. The variable, volume of canal water
per hectare of cultivated area, and rainfall capture the moisture
related face and level of fertilizer use per hectare of cropped
area captures the fertilizer effect.
Two models shall be postulated to explain the variations
in waterlogged and salinity, both over time and between
districts. These shall be the ordinary least squares (OLS) and
the weighed least squares (WLS) based on linear probability
model. For the cross-section analysis, we pool together data
for three sets of years (Table 1) respectively after estimating
district level data, on the following four dets of variables. For
the purpose of this analysis, data for the following set of
variables are compiled.
(a) Saline and waterlogged Land as a Percentage of
Geographical Area. The data on and waterlogged land are
taken from a publication of the Directorate of Economics and
Statistics of the Punjab Government entitled “Floods, Irrigation
and Waterlogging in Punjab. The dependent variable is
defined as saline and waterlogged land as percentage of
geographical area in the district. An alternative formulation
could be to take degraded land as a proportion of cultivated
area or net area sown. If the object of the exercise is to examine
the impact of irrigation on extending or limiting the land
resources available to agriculture, the latter is the better
specification to use. This is because irrigation would have led
to an expansion of sown area and the decreases in sown area
due to the accompanying degradation must be examined to
get the net effect. In a model that seeks to determine the factors
leading to degradation, the geographical area is a better base
as it will capture the total effect of loss irrespective of whether
the land is cultivated or uncultivated. Further, if the net sown
area does not change significantly in a certain period of time
(as it does not in Uttar Pradesh, see Table 2), either definition
is acceptable.
It may be said that the intensity of waterlogging and
salinity should be taken into account. As hydrologists will
vouch for, however, this intensity may vary locally, even within
a small region. It is, therefore, not possible to account for it at
this level of aggregation. It is our contention, however, that
some insight into degradation of land can be attained even by
the less aggregated analysis.
The presence or otherwise of irrigation is captured better
by the volumetric supply of canal water reaching the land
than by the area irrigated. For explaining waterlogging and
salinity it is all the more relevant to examine the volume of
water as only its physical presence can result in land being
waterlogged. A detailed estimate of volumetric supply of canal
water from the different canal systems is available for the
entire time period from the unpublished data of the irrigation
department. The districtwise estimates of the volumetric
supply of canal water are arrived at on the basis of length of
the canal system in the district.’ This provides an
approximation to the magnitude of canal water reaching the
district. Table 2 gives the estimates.
On waterlogging maintain that rainfall has an important
impact on water-balance in the region. Rainfall at the district
level is, therefore, taken as one of the explanatory variables.
The effect of rainfall would depend partly on the soil
characteristics. However, as we are taking district level data
for the analysis, we expect that the level of aggregation will
capture differences in the soil conditions and we need not
take a separate soil variable.
The three variables, rainfall, volume of water supplied
and volume of water extracted through tubewells together
stand as a proxy, for the excess of water supply over demand.
This is because it is only water that has not been used by the
cropped fields or that has not evaporated, which seeps into
the ground and adds to the level of groundwater, thereby
making H possible to increase extraction through tubewells.
In an indirect fashion, therefore, demand for water has been
accounted for.
One component of the water-balance equation that
affects the magnitude of land degradation is the amount of
water that gets drawn from the water-table through pumping.
Over the period 2005 to 2006 there has been a rapid increase in
the tubewells in the State. In order to see what impact this
increase had on the waterbalance and hence on the problems
of water logging and salinity, we take the number of tube
wells per hectare of net sown area as one of the explanatory
variables in the equation.
Definition of land degradation includes waterlogged and
saline land. Recent studies indicate that high levels of salinity
are found to occur in regions where fertilizer consumption is
high. Consumption of fertilizer per hectare of cropped area is
included as one of the explanatory variables in the model to

KUMAR et al., An Empirical Study of Land Degradation its Magnitudes and Casualities 49
capture this impact. The actual quantity of unabsorbed fertilizer
would perhaps have been a better explanatory variable. Even
where agronomic estimates of unabsorbed fertilizer are
available, they are region- and crop-specific. In the absence
of district level, time-series data on this agronomic variable,
fertilizer consumption is taken as the proxy variable.
RESULTS AND DISCUSSION
Two linear regression models are now set up to test for
the hypothesis that the levels of land degradation depend on
the variables listed above. In other words, the alternative
sources of water availability together with the use of fertilizer
determine land degradation caused by water logging and
salinity. The models pool time-series data for three sets of
years, i.e., 1986-87, 1988-89, 1994-95, 1996-97 to 2003-04 and
1982-83 to 1984-85. In the first instance, an OLS model is setup
for each set and an F -test is conducted to ensure that the
pooling is in order.
Alternatively, the dependent variable, being a fraction,
i.e., proportion of land degraded, can be interpreted as the
probability that degradation will occur given the total
geographical area and the water availability from different
sources. The estimated value of the dependent variable will
then give the estimated probability that degradation will occur.
The OLS estimates may not be efficient and it may be better to
use weighted least squares defining.
The WLS estimates so obtained for the two sets of years
are tabulated together’ with the OLS results in Table 2. The
results of both the OLS and the WLS specifications are
reasonably acceptable.
All the explanatory variables appear with the expected
signs. While the variables, rainfall, volume of canal water and
level of fertilizer use, lead to increases in land degradation,
the number of tube wells per thousand hectares (by reducing
the water-table) decreases land degradation due to water
logging and salinity. In the OLS specification, the variables,
volume of canal water per hectare and fertilizer use per hectare
are significant at the 5 per cent level whereas the variable tube
wells per thousand hectares is significant at the 10 per cent
level. The WLS specification changes the level of significance
somewhat but the direction of the results remains the same.
The model explains 19 to 46 per cent (for different time periods)
of the variation in the dependent variable, which, for pooled
data, is not unacceptable. It is correct to conclude, therefore,
that the adoption of the canal water- fertilizer based technology
contributed to degradation of land and, but for the offsetting
Table 1.
Year
Saline and waterlogged area, sown area and
geographical area in Uttar Pradesh
Land saline and
waterlogged
(2)
Net sown
area
(3)
(1)
1987-88 72.13 4602
1988-89 70.72 4687
1989-90 68.40 4782
1990-91 65.33 4826
1991-92 63.86 5036
1992-93 82.21 5082
1993-94 61.87 5137
1994-95 60.39 5223
1995-96 58.21 5382
1996-97 57.26 5476
1997-98 55.46 5482
1998-99 53.62 5531
1999-2000 51.62 5636
2000-01 50.21 5682
2001-02 49.32 5733
2002-03 48.46 5872
2003-04 45.32 5921
2004-05 44.36 6023
2005-06 43.12 6134
2006-07 42.41 6272
Table 2.
The models pool time series data for 3 sets of years
Specification Year Intercept Canal water
per hectare
Rainfall Tube wells per
thousand hectare
Ordinary least squares 1984-85 to 1986-87 0.436
0.454
0.0029
-0.0089
(0.267) (2.267)
(0.777)
(-1.403)
R 2 =0.193 F-ratio=3.01 n=33
1997-98 to 1999-2000 -0.703 0.575
0.0129
-0.0058
(-0.893) (4.291)
(0.201)
(-1.329)
R 2 =0.46 F-ratio = 6.508 n=33
2003-04 to 2005-06 0.06
0.675
0.23
-0.0069
(0.07) (3.29)
(0.314)
(-1.429)
R 2 =0.42 F-ratio = 5.617 n = 33
Linear probability model 1986-87 to 1988-89 -0.860 0.611
0.0054
-0.013
(-1.860) (3.777)
(1.179)
(-1.2999)
R 2 =0.41 F-ratio=6.58 n=33
1997-98 to 1999-2000 0.675
0.232
0.00161
5.491
(1.269) (2.839)
(-2.345)
(0.937)
R 2 =0.19 F-ratio = 2.887 n=33
2003-04 to 2005-06 0.725
0.324 0.0018 (0.512) -0.0078
(1.369) (3.749)
(-3.24)
R 2 =20.24 F-ratio = 3.88 n=33
Fertiliser user
per hactare
9.609
(1.635)
14.085
(2.179)
12.085
(2.28)
19.146
(2.116)
6.491
(0.84)

50 Trends in Biosciences 5 (1), 2012
effect of tube well expansion in the late sixties and seventies,
the land degrading effect of the technology would have been
far more.
Results show that the new agricultural technology
characterised by water and fertilizer intensive agriculture have
had second round effects. It was just circumstantial that the
accelerated investment in tube wells offset some of these
effects. For the Uttar Pradesh region, then these ecological
effects of irrigation are not substantial and have been
decreasing over time as a consequence of the conjunction of
a number of factors. One cannot, however, in the planning for
large surface irrigation systems depend on such fortuitous
happenings. Detailed projections of the likely environmental
impact (Central Board of Irrigation and Power, 1985) should
form a part of the planning exercise.
It is important to realise that the kind of planning
mentioned above is significant, because in its absence,
measures to correct for land degradation may turn out to be
costly. Joshi et al., 2001, for instance, find that sub-surface
drainage to reclaim degraded land in Haryana would cost
Rs.21,913 per hectare under one set of technical parameters.
In another estimate the production loss from land degradation
for eleven irrigation projects studied is estimated at Rs.l,803
million (Joshi and Agnihotri, 1984). Moreover, this loss not
accounted for the loss of forest land in catchments areas and
the cost of abilitation of displaced persons. The significance
of a proper pre-implementation evaluation of all aspects of
large irrigation projects cannot, therefore, be over-estimated
m if in some cases the adverse effects may turn out to be
insignificant. Following a strategy of the above kind would
ensure also that estimates of these phenomena are made re
firm. This development would then provide a link in the study
of the inter-relation between ecology and development.
LITERATURE CITED
Central Board of Irrigation and Power. 1985. Indo-Soviet Workshop
on Evaluation and Modeling of Impacts on Environment of Water-
Resources Projects, Government of India, New Delhi.
Centra: Ground Water Board. 1983. Proceedings of the Seminar on
Assessment, Development and Management of Ground Water
Resources, Vol. II, Ministry of Water Resources, Government of
India, New Delhi.
Central Ground Water Board. 1986. Proceedings of the Seminar on
Conjunctive Use of Surface and Ground Water, Ministry of Water
Resources, Government of India, New Delhi ..
Chopra, Kanchan. 1982. “Alternative Sources of Irrigation and Land
Use Patterns m the Uttar Pradesh”, Indian Journal of Agricultural
Economics, Vol. 37, 0.2, April-June.
Chow, G.C. 1960. “Tests of Equality between Sets of Coefficients m
Two Imear Regressions”, Econometrica Vol. 28, No.3, July, pp.
591-605.
Government of India 1976. Report of the National Commission on
Agriculture 1976, Part V: Resource Development, Ministry of
Agriculture and Irrigation, New Delhi.
Joshi, P.K. 1987. “Effect of Surface Irrigation on Land Degradation-
Problems and Strategies”, Indian Journal of Agricultural Economics,
Vol. 42, No.3, July September .
Joshi, P.K. and A.K. Agnihotri 1984. “An Assessment of the Adverse
Effects of Canal Irrigation in India”, Indian Journal of Agricultural
Economics, Vol. 34, No.3, July-September.
Joshi, P.K., O.P. Singh, K. V.G.K. Rao and K.N. Singh 1987. “Sub-
Surface Drainage for Salinity Control: An Economic Analysis”,
Indian Journal of Agricultural Economics, Vol. 42, No.2, April-
June.
Kanwar, J.S. 1977. Soil Bulletin No. 34, Food and Agriculture
Organization of the United Nations, Rome.
Karanth, K.R. 1986. “Water logging: Causes, Prevention and Control”,
in Central Ground Water Board : Proceedings of the Seminar on
Conjunctive Use of Surface and Ground Water, Ministry of Water
sources, Government of India, New Delhi.
Recieved on 16.12.2011 Accepted on 25.2.2012

Trends in Biosciences 5 (1): 51-53, 2012
Comparative Efficacy of Three Chemical Products on the Root-Knot Development
and Plant Growth of Green Gram, (Vigna radiata L.)
MOHD. YAQUB BHAT 1 , A.H. WANI 1 AND NASEER HUSSAIN SHAH 2
1
P.G. Department of Botany, University of Kashmir, Srinagar 190 006
2
Department of Botany, Govt. Degree College Kupwara, Srinagar
email: myaqub35@gmail, ahamidwani@yahoo.com
ABSTRACT
A green house study using artificial inoculations in earthen
pots was made to ascertain the efficacy of three different
chemicals viz. Carbofuran, Oncol,and Posse against root-knot
nematode infecting green gram. The study revealed that all
the chemicals applied either as soil drench or seed soaking
were effective and superior to untreated control in reducing
nematode population and gall index and in improving plant
growth. The chemical Carbofuran showed better performance
than that of Oncol and Posse. Seed soaking treatment
registered slightly better plant growth in comparison to soil
drench application. However, soil drench application of
chemicals showed much efficacy in restricting nematode
population and gall development. Higher concentration (0.1%)
of chemicals appeared as the most effective than the lower
concentration 0.05%). The number of bacterial nodules
increased in all chemical treatments with a more stimulating
effect at lower dosages.
Key words Chemicals, root-knot nematode, Vigna radiata L.
growth, gall index, nodules.
The root-knot nematode Meloidogyne incognita
(Kofoid and White, 1919) Chitwood 1949, is an important pest
of most grain legumes including green gram Vigna radiata
(L.) Wilczek and causes severe economic losses to these crops
every year. Effectiveness of nematicides in controlling plant
parasitic nematodes is well recognised and considerable work
has been done on the effect of various chemicals on the
nematode population in order to find suitable control measures
(Azam, et al., 1978; Sharma and Trivedi, 1985, Haq, et. al.,
1987, Sethi and Maher, 1989, Dareker, et.al., 1990 Giannakou,
et. al., 2002). Oxamyl application reduced root-galling severity
and increased marketable yield however the cost-effectiveness
of Oxamyl application was related to the level of soil infestation
with nematode (Gugino, et.al., 2006). It is concluded that
there will most probably be not a single nematode controlling
agent but combinations different control methods will be
required according to specific needs (Mc Sorley, 1998, Abou-
Jawdah, et.al., 2000). Now it seems that chemical control of
nematodes is the most desirable component to be involved in
the nematode management system (Fazal, et.al., 2011).
Therefore it order to chose a chemical for effective integrated
nematode management system it was felt desirable to carry
out the green house study to evaluate the effect of three
chemical products viz., Carbofuran, Oncol (Benfurocarb) and
Posse, as seed soaking and soil drench treatment on the
nematode development and plant growth of green gram in
two consecutive seasons.
MATERIALS AND METHODS
Two concentrations, 0.05% and 0.1 % of Carbofuran
(10G) 2,3- Dihydro-2,2 dimethyl-7-benzofuranylmethyl
carbamat), Oncol (50SP) (Benfurocarb)(ethyl N-[2,3-dihydro-
2,2-dimethyl benzofuran—7-yloxycarbonyl (methylaminothio]
= N- isopropyl-B-alaminate) and Posse (25 STP)(2,3-dihydro-
2,2-dimethyl benzofuran-7-yl(dibutylamino thio) methyl
carbamate), prepared from granular formulation on active
ingredient basis, were used as seed soaking or as soil drench.
For seed soaking, seeds of green gram were soaked in both
the concentrations of each chemical for 2 hr separately. For
soil drench, 25 ml of both the concentrations of each chemical
was applied around the root zone of each seedling, individually.
Soaked seed were treated with Rhizobium (green gram strain)
and sown in 15 cm earthen pots filled with 1 kg steam sterilized
soil and farm yard manure in the ratio of 2:1. On germination,
one healthy seedling was retained per pot and each seedling
inoculated with 1000 J2 of Meloidogyne incognita and 1g of
Rhizobium. For soil drench treatment, green gram seedling
germinated in earthen pots, as mentioned previously were
applied with 25 ml of each concentrations (0.05% and 0.1%)
of each chemicals, separately. One day after chemical
application, the seedling were inoculated with 1000 J2 of M.
incognita. Each treatment was replicated thrice and pots were
arranged in randomised block design on green house benches.
Sixty days after inoculation, the plants were harvested
and roots were washed thoroughly to free them of adhered
soil particles. Data on shoot and root length was recorded in
cm, Number of bacterial nodules and root knot were counted
and number of nematode populations in root and soil were
determined ( Southey, 1988). Nematode reproductive factors
was calculated for each treatment (Oostenbrink, 1966). For
rating gall index (G 1) Taylor and Sasser, Scale was used. (Taylor
and Sasser, 1978). The data obtained was subjected to
statistical analysis and critical difference (CD) was calculated
at P= 0.05

52 Trends in Biosciences 5 (1), 2012
RESULTS AND DISCUSSION
Data on the performance of three nematicides as seed
soaking treatment against root knot disease of green gram
Vigna radiata L. in pots showed that all the nematicides
tested, decreased the nematode reproductive factor and rootknot
index and increased the plant growth significantly (P=
0.05) compared to plants green gram inoculated with M
.incognita alone(Table 1). In general, treatments were most
effective at higher concentration (0.1%) and least at lower
concentration (0.05%) with non significant differences.
Carbofuran was most effective among the different chemicals
in reducing nematode reproductive factor and root-knot index
and increasing plant dry weight followed by Posse and Oncol
(benfuracarb). However among these nematicides carbofuran
and posse showed significant increase in plant dry weight in
comparison to Oncol at both concentrations. These
nematicides at tested concentrations had variable effect on
bacterial nodules. Posse at both the concentrations was most
effective in increasing the bacterial nodules significantly (P=
0.05) followed by 0.05% carbofuran. However, Oncol at both
the concentrations enhanced the number of bacterial nodules,
non significantly. Results on the efficacy of three nematicides
as soil drench treatment for the management of root knot
disease of green gram in pot conditions revealed that
Carbofuran was most effective in reducing the nematode
reproductive factor and gall index and increasing the plant
growth, followed by and Posse and Oncol. All these
nematicides at tested concentration increased the bacterial
nodules significantly (P= 0.05) compared to green gram plants
inoculated with M. incognita alone (control). In seed soaking
treatment Oncol failed to enhance the bacterial nodule number
significantly in comparison to control but increased the nodule
number significantly (P= 0.05) in comparison to control in
drench treatment at the same concentrations. However, the
differences among Carbofuran, Posse and Oncol, in
suppressing nematode reproductive factor and gall index and
Table 1.
Treatments
Effect of three chemical products on the root-knot developnent and plant growth of green gram (Vigna radiata (L.)
Wilzeck) (Seed soaking treatment).
Carbofuran.05%
Carbofuran.1%.
Oncol .05%
Oncol .1%
Posse .05
Posse .1%
Untreated inoculated control
DF (6,14)
Bacterial
nodules
/plants
56.20
50.20
52.26
50.06
65.26
63.60
48.33
Each value is mean of three replicates
*Reproductive factor (R) = Final population / Initial populations
Dry weight of plant
Nematode population
Wt. (g)
Root Shoot Total Soil Root Total
A B A+B A B A+B
1·40 3.13 4.53 1496 100 1596
1.50 3.45 5.25 1460 88 1549
1.50 3.26 4.76 1583 108 1691
1.76 3.16 4.92 1545 89 1650
1.00 2.26 3·26 1752 136 1888
1.21 2.36 3.56 1702 132 1834
0.43 1.36 1.80 3306 290 3596
* R =
Pf/Rf
Root-knot
index
F 13.97 8.93 21.53 29.34 742.5 254.9 1171 1183 34.30
St.error of mean 1.73 0.15 0.16 0.22 24.13 4.3 21.24 0.02 0.19
P=0.5 4.59 0.38 0.39 0.55 60.10 10.8 52.91 0.05 0.49
1.59
1.54
1.69
1.65
1.88
1.83
3.59
1.80
1.80
2.20
2.20
3.50
2.80
5.0
Table 2.
Treatments
Effect of three chemicals products on the root-knot development and plant growth of green gram (Vigna radiata (L.)
Wilzeck.) (Soil drench treatment).
Carbofuran.05%
Carbofuran.1%.
Oncol .05%
Oncol .1%
Posse .05
Posse .1%
Untreated inoculated control
Bacterial
nodules
64.20
63.90
62.93
59.03
66.20
66.13
49.88
• Each value is mean of three replicates
*Reproductive factor (R) = Final population / Initial populationss
Dry weight of plant
Nematode population
Wt. (g)
Root Shoot Total Soil Root Total
A B A+B A B A+B
1.60 3.20 4.80 1490 116 1606
1.60 3.50 5.18 1362 112 1447
1.16 2.93 4.10 1402 120 1522
1.40 3.86 4.26 1535 125 1660
0.81 2.40 3.21 1698 128 1826
1.10 2.20 3.30 1638 142 1780
0.40 1.30 1.70 3308 290 3598
* R =
Pf/Rf
1.65
1.47
1.52
1.66
1.82
1.78
3.59
Root-knot
index
F 18.74 12.84 11.37 29.25 1.5±0.4 379.10 8239.27 48.77 7360.35
St.error of mean 1.32 0.12 0.21 0.21 5.69 3.26 8.25 0.19 0.008
P=0.5 3.31 0.29 0.54 0.53 14.17 8.12 20.56 0.02 0.48
1.93
1.43
2.06
1.55
3.3
2.6
5.0

MOHD. YAQUB BHAT, et al., Comparative Efficacy of Three Chemical Products on the Root-Knot Development 5 3
increasing plant growth and nodule number were nonsignificant.
Seed soaking and soil drench evaluation data on
the efficacy of nematicides were almost similar. However, soil
drench treatment was comparatively most effective in reducing
the root knot disease as evident from reproduction factor and
gall index.
Present studies revealed that most of the chemicals
tested, either as soil drench or seed soaking treatment,
provided protection to green gram plants against M. incognita
race-1 to maximum extent. The variation in effectiveness of
chemicals can be attributed to the property of chemicals by
which these chemicals breakdown and are metabolized to
hydrolytic products in the form of plant conjugates. Reduced
nematode population and gall indices in seed soaking
treatment can be attributed to the fact that chemical absorbed
by seeds in sufficient quantity was probably responsible for
nematostatic action at the initial stage (Haq, et al., 1987). In
soil drench application, both contact and systemic action of
chemicals probably resulted in reduced nematode reproductive
factor and gall indices. This assumption was reflected in less
number of nematode population in soil and root and are in
consistency with studies of Gugino, et al., 2006, Giannakou,
et al., 2002) Increase in bacterial nodules with chemical
treatments shows their less phytotoxicity and confirm the
findings of Shakuja and Singh, 1976. The results of present
investigation highlight the importance of seed soaking and
soil drench application of chemicals in crop protection
strategies against nematodes species to enhance the crop
productivity. These studies also help in knowing the
importance of particular chemical to be involved in integrated
nematode management strategies as has been studied by
Kamra, et al., 2008.
ACKNOWLEDGEMENT
The authors are thankful to Head, Department of Botany,
University of Kashmir, Srinagar, for providing necessary
facilities to complete this work.
LITERATURE CITED
Abou-Jawdah,Y., Melki, K., Hafez, S.L., Sofa, H., El.Masri, Y. and
Sundararaj, P. 2000. Alternatives to methyl bromide for root knot
nematode management on cucumber in Lebanon: Nematropica,
30:41 -45
Azam, M.F., Khan, A. M. and Saxena, S.K. 1978. Population of plant
parasitic nematodes as influenced by certain nematicides around
some ornamental plants: Indian J. Nematol., 8: 69-73.
Darekar, K. S., Mhase, N. L. and Shalke, S. S. 1990. Effect of green
gram seed treatment with certain nematicides on root-knot
nematodes and crop yields: International Nematology Network
Newsletter, 7: 4-5.
Fazal, M., Bhat, M. Y. and Ashaq, M. 2011. Combined application of
Paecilomyces lilacinus and Carbosulfan for management of
Meloidogyne incognita and Rotylenchulus reniformis. Annals of
Plant Protection Sciences, 19(1) In press.
Giannakou, I. O., Sidiropoulos, A. and Athanasiadou, D. P. 2002.
Chemical alternatives to methyl bromide for thecontrol of rootknot
nematodes in greenhouses. Pest Management Science, 58:
290-296
Gugino, B.K., Abawi, G.S. and Ludwig , J. W. 2006. Damage and
Management of Meloidogyne hapla Using Oxamyl on Carrot in
New York J. Nematol., 38 (4): 483-490
Haq, S., Khan, M.W. and Saxena, S.K.1987. An evaluation of different
modes of application of systemic nematicides for the control of
root-knot nematodes in tomato: Phytoprotection, 68: 57-63.
Kamra, A. Sharma, H. K. and Pankaj 2008.Effect of seed treatment
with pseudomonas flurescens alone and in combination with soil
application of carbofuran and neem seed. Pesticide Research
Journal, 20(1): 79-82.
Mc Kenry, M Buzo, T., Kretsch, J., Kalu, S. Otomo, E. Ashcroft, R.,
Lange, A. and Kelley, K. 1994. Soil fumigants provide multiple
benefits: Alternatives give mixed results: Calif. Agric., 48:22-28.
Mc Sorley, R., Stansly, P. A., Noling, J.W., Obreza, T.A. and Conner,
J.M. 1998. Impact of organic soil amendments and fumigation on
plant parasitic nematode in a southwest Florida vegetable fields:
Nematropica, 27:181-189.
Oostenbrink, M. 1966. Major characteristic of the relation between
nematodes and plants: Medel Hogesh, wageningen, 66: 3-46.
Sakhuja, P.K. and Singh, I. 1980. Effect of nematicidal treatment
against M. incognita on mung (P. aureus) and bacterial nodulation:
Nematol. Medit., 8: 201-203.
Sethi, C.L. and Meher, H.C. 1989. Effect of Phenamiphos on soil
nematodes and yield of cowpea, pea and okra: Indian J. Nematol.,
19(2): 89-94.
Sharma, R.K. and Trivedi P.C. 1985.Nematotoxicity of some
organophosphates and carbamates against Meloidogyne incognita
and their effect on plant growth and rhizobial nodulation in pea :
International Nematology. Network Newsletter, 2(4): 10-12.
Southey, J.F. 1970. Laboratory methods for work with plant and soil
nematodes, Ministry of Agriculture, Fisheries and Food. Tech. Bull.
2. HerMajesty’s Stationary Office, London.
Taylor, A.L. and Sasser J.N. 1978. Biology, identification and control
of root-knot nematodes (Meloidogyne species):Coop. Publ. Dept.
Plant Pathology, NCSU and USAID, Raleigh, N.C. 111.
Recieved on 3.10.2011 Accepted on 2.4.2012

Trends in Biosciences 5 (1): 54-56, 2012
Effect of Some Aqueous Plant Extracts on Egg Hatching of Meloidogyne incognita
(Kofoid and White) Chitwood, the Root-Knot Nematode
HARPREET KAUR* AND ANU KATOCH
Department of Zoology & Environmental Sciences, Punjabi University, Patiala 147 002
*email: harpreet_bimbra@yahoo.com
ABSTRACT
Root-knot nematode disease had been effectively and mostly
controlled by synthetic nematicides which have been confirmed
to be the source of food, water and environmental pollution. A
study was conducted to evaluate potential of aqueous extracts
from mint, ginger, garlic, marigold and eucalyptus in controlling
the hatch of eggs of the RKNs, Meloidogyne incognita. Five
concentrations of water soluble extracts from the five plant
species were filtered, added to petridishes and infested with
eggs of M. incognita. The AMinE was the most effective in
delaying and suppressing hatching. Lowest concentration i.e.
0.5% AMinE caused delay upto 19 hours with 2.3% of hatch in
comparison to 17 hours delay in 0.5% AGarE and AGinE with
3% and 3.8% of hatch. AMarE was least effective as the delay
was 14 hours in 0.5% AMarE with 5.2% egg hatch.
Key words
Meloidogyne, egg hatch, plant extracts, mint, garlic
Most crop plants including vegetables, cereals,
ornamental plants and fruits are attacked by one or more
species of nematodes. It has been reported that average yield
loss of world’s crops due to plant parasitic nematodes is 12.3%
(Sasser, 1998). Nematodes are microscopic, eel-like round
worms. The most troublesome species in the field are the ones
that live and feed within plants than others which live freely
in the soil and feed on plant roots. Although there are different
species of root feeding nematodes, most important are
Meloidogyne spp. which are difficult to control. It is impossible
to precise the loss caused by them. However, some estimates
of individual crop losses have been made by various authors.
In tomato plants, when M. incognita occurs, 50% of fruit
yield is suppressed (Lamberti, 1979). In tropical areas root
knot nematodes cause losses in potato crops estimated to be
24% (Sasser, 1979). In addition, these nematodes have ability
to interact synergistically with other plant pathogens and
cause upto 5-34% yield loss in vegetables in tropical climates.
The presence of root knot nematode, Meloidogyne spp. is
known to increase the severity of Rhizoctonia solani, the
causative agent of root rot in many plants.
The present research was undertaken to access the
effect of aqueous extracts of five plants in controlling the
hatching of eggs of M. incognita, the root knot nematode in
laboratory conditions.
MATERIALS AND METHODS
The effect of different concentrations of five selected
plant extracts on egg hatching of M. incognita was evaluated
as follows. The plants selected for the present study were
mint (Mentha erecta), garlic (Allium sativum), ginger (Zingiber
officinale), marigold (Tagetes erecta) and eucalyptus
(Eucalyptus). 25 gms of dry leaves of mint, fresh leaves of
marigold and eucalyptus, garlic bulbs and ginger rhizome were
mixed separately in an electric blender in 100 ml of distilled
water for 10 minutes and kept in water bath for 8 hours at
65°C. These extracts filtered through muslin cloth and each
filtrate was considered as standard solution of 100%
concentration. Each extract was autoclaved at 15 lb pressure
at 121°C temperature, allowed to cool and stored in refrigerator
for further experimentation. (Ahmed, et al., 2004). Root samples
of infected banana plant were collected from selected localities.
Root samples were collected by digging the soil around the
roots of the plants. Samples were collected in polythene bags
and stored in refrigerator at 4°C. Fresh and uniform egg masses
of M. incognita were collected using forceps. One egg mass
of M. incognita was added to each petridish containing the
extracts. The effect of different concentrations levels on the
hatching of nematodes was determined at four intervals i.e 12,
24, 48 and 72 hours. Data was recorded and comparison of
hatching percentage were made on the basis of average 600
eggs per egg mass.
RESULTS AND DISCUSSION
During the present study significant effect of different
concentrations of five plant extracts were observed on the
egg hatching of M. incognita.
Aqueous extracts of all the selected plants delayed the
egg hatching of M. incognita. The effect of different
concentrations and exposure time on hatching of eggs varied
with plant material and incubation period. In the control, egg
hatching started after 4 hours, whereas for other treatments
there was no hatching till 14 to 19 hours. The percentage of
hatching increased from 24.8% - 31.2% at 12 hours to 63.1-
70.1 at 72 hours in the control. In 0.5 % AMinE the egg hatching
was delayed to 19 hours with 2.3 % egg hatching rate in
comparison to 12 % hatching at 4 hours in control. The
hatching increased to 16.2% at 24 hours and then decreased
to 12.4 at 72 hours in 0.5 % concentration of AMinE. 100 %
inhibition of egg hatching was observed in higher
concentration i.e. 3 % and 4% of AMinE.
The average time taken to start egg hatching was 17
hours (3.8%) in 0.5% AGarE. The hatching increased to 17.3

KAUR and KATOCH, Effect of some Aqueous Plant Extracts on Egg Hatching of Meloidogyne 55
% at 24 hours and then decreased to 10.2 % at 72 hours in 1%
concentration of AGarE. In concentrations, 1%, 2% and 3%
of AGarE egg hatching was delayed upto 24 hours. In AGinE,
average time taken to start egg hatching was 17 hours with
hatching rate 3% at 0.5 %. The hatching increased to 17.4% at
24 hours and then decreased to 11.3% at 72 hours in 0.5%. In
concentrations, 1% to 3% hatching started at 24 hours with
rate of hatching 29.2% to 70.1% upto 72 hours. No hatching
occurred in higher concentration i.e. 3% AGinE (72 hours).
The egg hatching of M. incognita was delayed upto 15
hours with egg hatching rate of 4% at 0.5% AEucE. In 1%, 2%
and 3% AEucE delay was upto 24 hours with egg hatching
rate 17.4%, 10.4% and 8% respectively. 100% inhibition was
observed in higher concentrations 3% at 72 hours and 4%.
In 0.5% concentration of aqueous marigold extract, the
egg hatching of M .incognita was delayed upto 14 hours with
egg hatching rate 5.2%. The hatching percentage increased
to 19.8% at 24 hours and then decreased to 14.2% at 72 hours.
In the present study, all the concentrations of aqueous
extracts of various botanicals caused delay in hatching.
According to the in vitro experiments conducted the average
time taken to start hatching of eggs was 19 hours in 0.5%
AMinE with only 2.3% of hatching followed by 17 hours delay
in 0.5% AGarE and AGinE with 3% and 3.8% of hatch; 15
hours delay in 0.5% AEucE with 4% of egg hatch and 14
hours delay in 0.5% AMarE with 5.2% of egg hatch. Therefore,
indicating AMinE to be the most effective in delaying and
suppressing of hatching of eggs of M. incognita. Pandey
and Kalra, 2010 reported the inhibition in hatching of eggs in
the aqueous extracts of vermicompost produced from wastes
of menthol mint (Mentha arvensis) and garden mint (Mentha
viridis) i.e. 90.6% and 73.2% respectively. Oka, et al., 2000
also reported essential oils of Mentha species as highly
effective in causing hatching inhibition at concentration of
600 µl/Lt in an in vitro and in pot experiment. According to
them, isomers of 1, 2, epoxy methylacetate had highest
nematicidal activity as compared to limonene and peperitone
of M. retundifolia.
Table 1.
In the present study mint was found to be the most
Effect of aqueous mint extract (AMinE) on egg
hatching of M. incognita
Treatment Concentration
of extract
Mint
extract
(leaves)
Hatching (%)
Average time
taken to start
hatching of
eggs (%)
12 24 48 72
hours hours hours hours
0.5% Nil 16.2 14.6 12.4 19 hours (2.3)
1% Nil 14.2 11.8 10.1 24 hours
(14.2)
2% Nil 12.9 10.6 8.4 24hours
(12.9)
3% Nil Nil Nil Nil Nil
4% Nil Nil Nil Nil Nil
Control Distilled water 27.7 35.1 49.6 63.1 4 hours
(13)
effective botanicals among all the selected plants in inhibiting
and controlling the hatching of eggs of M. incognita. In 3%
AMinE at 48 hours reduced hatching to 0% followed by 4%
AGarE 5% in AGinE; 5.3% in AEucE; 9.8% in AMarE. Abo-
Elyousr, et al., 2009 found that garlic extract at a very low
concentration (S/100) suppressed the nematode activity of
M. javanica and M. incognita as was also evident with our
finding. The present findings confirm nematicidal potential of
ginger, Zingiber spp. as also indicated by Zareen, et al., 2003
where aqueous extract of ginger significantly (P

56 Trends in Biosciences 5 (1), 2012
Table 4.
Effect of aqueous eucalyptus extract (AEucE) on
egg hatching of M. incognita
Treatment Concentration
of extract
Eucalyptus
extract
(leaves)
12
hours
Hatching
(%)
24
hours
48
hours
72
hours
Average
time
taken to
start
hatching
of eggs
(%)
0.5% Nil 18.4 15.4 14.8 15 hours
(4%)
1% Nil 17.4 14.3 12.1 24 hours
(17.4)
2% Nil 10.4 9.8 9.4 24hours
(10.4)
3% Nil 8 5.3 Nil Nil
4% Nil Nil Nil Nil Nil
Control Distilled water 27.9 35.8 52.7 67.3 4 hours
(13)
Table 5.
Effect of aqueous marigold extract (AMarE) on
egg hatching of M. incognita
Treatment Concentration
of extract
Marigold
extract
(leaves)
12
hours
Hatching (%)
24
hours
48
hours
72
hours
Average
time taken
to start
hatching of
eggs (%)
0.5% Nil 19.8 16.4 14.2 14 hours
(5.2)
1% Nil 16.2 14.1 12.9 17 hours
(13.3)
2% Nil 13.8 12.7 11.8 24hours
(13.8)
3% Nil 11.2 9.8 Nil 24hours
(11.2)
4% Nil Nil Nil Nil Nil
Control Distilled water 24.8 34.7 57.9 69.1 4 hours
(13)
aqueous leaf extract of eucalyptus was effective and stopped
hatching of eggs of M. javanica after 72 hours.
The present study indicate the AMarE was least effective
in causing delay and inhibition of hatching of eggs of M.
incognita even at higher concentration i.e. 3%. Percentage of
hatched eggs were 9.8% at 48 hours in comparison to 4–5.3%
in rest of other extracts (AGarE, AGinE and AEucE) and no
hatching in AMinE. These observations in contrast with
Bhardwaj and Sharma, 2007 according to which all
concentrations of Tagetes patula leaf extract inhibited the
hatching except the lowest concentration 6.6% where in
hatching was found to be 0.11% at 24 hour and it increased up
to 0.33% at 48 hours. Hot water extract of T. patula was found
to be even better in comparison to T. patula leaves in delaying
the hatching of eggs as there was only 0.11% hatching at 48
hours. Swarup and Sharma, 1967 reported T. erecta and T.
halisciencis root extract were lethal or inhibited egg hatching
of M. javanica, M. arenaria and M. incognita respectively.
Pandey and Kalra, 2010 evaluated aqueous vermicompost
extract having egg hatching inhibitory activity causing upto
81.1% of inhibition. Toida, 1972 had also reported the inhibition
of M. incognita juvenile hatch caused by root exudates of T.
minuta.
LITERATURE CITED
Dawar, S., Younus, S.M. and Zaki, M.J. 2007. Use of Eucalyptus sp., in
the control of M. javanica root knot nematode. , Pakistan Journal
of Nematology, 39(6): 220, 9-2214.
Hassan, S.M.E., Rahman, M. Sq., Amin, M. R., Majumdar, U.K. and
Taj, H.F.El. 2001. Study of ginger on root knot disease of brinjal.
Online journal of biological sciences, 1(7): 560-562.
Lamberti, F. 1979. Economic of Meloidogyne spp. in subtropical and
mediterran climates, in root knot nematodes (Meloidogyne species).
In: Systematic, Biology and control (eds. Lamberti, F. and Taylor,
C.F). Academic press, New York. pp. 341-357.
Oka, Y., Nacar, S., Putievsky, E., Ravid, U., Yaniv, Z., and Spiegel, Y.
2000. Nematicidal avtivity of essential oils their components against
the root knot nematode. Phytopathology, 90: 710-715.
Pandey, R and Kalra, A. 2010. Inhibitory effects of vermicompost
produced from agrowaste of medicinal and aromatic plants on
hatching in Meloidogyne (Kofoid and White) Chitwood. Current
science, 98(6).
Sasser, J. N. 1998. A perspective on nematode problems worldwide. In:
Nematode Parasitic to cereals and legumes in temperate semi arid
regions. (eds. Sasena, M. C., Sikora, R. A. and Srivastava). ICARDA.
Syria, pp. 8-12.
Sasser, J.N. 1979. Economic importance of Meloidogyne in tropical
countries. In: Root knot nematodes (Meloidogyne spp.). Systemetics,
Biology and Control (eds. Lamberti, F. and Taylor, C.E.). Academic
Press, Newyork. pp. 359-374.
Swarup, G. and Sharma, R.D. 1967. Effect of root extract of Asparagus
racemosus and Tagetes erecta on hatch of eggs on M. javanica and
M. arenaria. Indian Journal of Experimental Biology, pp. 5-59.
Toida, Y. 1972. Nematicidal effect of Maxican marigold against
nematodes associated with mulberry. Japanese Journal of
Nematology, 1: 18-21.
Zareen, A., Zaki, M.J. and Javed, N. 2003. Nematicidal activity of
ginger and its effect on the efficacy of Pasteuria penetrans for the
control of root knot nematodes on tomato. Asian journal of plant
sciences, 2(11): 858-860.
Recieved on 18-01-2012 Accepted on 20.4.2012

Trends in Biosciences 5 (1): 57-60, 2012
Effect of Bacterial Preparations on Development of Diacrisia obliqua
ZEENAT WARSI AND AJAY CAPOOR
Department of Zoology, Agra College, Agra, U.P.
ABSTRACT
This study was carried out to determined the effectiveness of
different concentrations of all the bacterial preparations of
B.thuringiensis like Dipel, Thuricide HP, Bactospeine against
the development of test insect Bihar hairy caterpillar i.e. D.
obliqua. The biocontrol agents such as Bacillus thuringiensis
used for controlling Lepidopteran, coleopteran and dipteran
pests. These insecticides are considered safe to the environment
and natural enemies. It is evident from the result of the
experiments that lethal concentration of dipel had marked effect
on the larval development and it is also significant that the
surviving larvae of D. oblique were shorter in length, and
lighter in weight when compared with control.
Key words
Diacrisia obliqua, efficacy insecticide, bacillus
thuringiensis
The worldwide attention is to develope to compounds
acting selectively on same groups of insects by inhibiting the
activity of bio chemical sides such as respiration
(diafenthiuron), the nicotinyl acetyl chlorine receptor, the
salivary glands of sucking pests (pymetrozine). Progress has
been made to introduce improved bio control agents such as
Bacillus thuringiensis (Bt) for controlling lepidopteran,
coleopteran and dipteran pests. Bt kills insects primarily
through the ation of delta-endotoxins, a proteinous
constituents produced during sporulation it effect the insects
midgut epithelium. Many insecticides such as cholorinated
hydrocarbon, organophosphates, carbonates etc. where used
in the controlloing insects pest but many of these insecticides
are harmful to man and beneficial organism which causes
ecological disturbances.
MATERIALS AND METHODS
The effect of different concentrations was studied on
all the bacterial preparations viz., Dipel, Thuricide HP,
Bactospeine on the development of D.obliqua was studied in
response to their application to Moth by Leaf dip method and
Topical method (Table 1).
Leaf dip method was studied with immediately hatched
larvae obtained from females fed on a strength of a bacterial
preparation were employed. This was tested by one experiment
designed separately for each strength of bacterial preparation.
Twenty such larvae per replicate were reared on tender leaves
of host plants until their pupation, number of larvae pupated,
their developmental duration and survival were recorded.
These pupae were kept to obtain adults on emergence of
adults, the number of adults emerged and their pupal period
were recorded. Besides these, the sex ratio of adults was also
recorded. The experiment was further extended to record the
life duration of males and females. For this purpose males and
females were maintained individually in glass chimneys on
daily supply of 20% of sugar solution till their natural death
and on their expiry of longevity was recorded.
Topical method of treatment was studied experimentally
with newly hatched larvae of adults which were already forced
to contact their residue film of a strength of each bacterial
preparation. Twenty such larvae were reared on leaves of C.
juncea till their pupation and when they pupated the number
of pupae obtained and the larval period were recorded. The
pupae were kept in glass chimneys date wise and when moths
emerged from them, their number and pupal periods were
noted. Besides this the sex ratio was also recorded. The
experiment was further extended for recording the life span of
males and female moths. For this purpose males and females
were maintained individually date wise in glass chimneys on
daily supply of 20% sugar solution till their natural death and
on expiry of their longevity was recorded.
RESULTS AND DISCUSSION
The larva of untreated adults with different
concentrations were reporte 89.43% survival. In response to
treatment by leaf dip method with different concentrations of
the Dipel, result showed that the larval survival , varying from
22.14%-64.36% and decreasing with the advancing
concentration depended on the concentration of the Dipel.
Further the larvae of the untreated adults grew faster than of
the adults treated with any concentration of the Dipel. The
leaf dip treatment with any concentration of the Dipel curtailed
the longevity of both male and female adults significantly as
compared to non-treatment at early stage. The female live
longer than the male in response to treatment with any
concentration of the Dipel. The life span in both sexes
decreasing with the increasing concentration. In response to
treatment by leaf dip method with different concentrations of
Dipel from 0.05%-1.0% the pupal period varying from 14.52-
29.18 days and increasing with the increasing concentration.
The percentage of the emergence varying from 22.28% to
70.62% and decreasing with the increasing concentration but
there was no significant difference observed in the pupal period
in both treatments and 0.05%.
In response to parents treatment with Dipel under
Topical method, the larval period varied from 24.42 to 40.43
days among different concentrations.
The treatment of parent adults with residue film of all
concentration of the Dipel reduced the longevity in adults of

58 Trends in Biosciences 5 (1), 2012
Table 1.
Mode of treatment Concentration(%) Pupation(%) Larval period(days) Emergence(%) Pupal
period(days)
L.D.M.
T.M.
Effect of Dipel on post embryonic development in D. oblique at different concentrations under different modes of
treatment.
0.05 65.35 22.65±0.68 70.62 14.52±0.56
0.10 57.38 24.85±0.26 60.54 16.76±0.75
0.50 44.44 28.47±0.25 40.34 20.64±0.31
0.75 35.26 34.24±0.26 35.27 24.44±0.56
1.00 22.14 36.42±0.44 22.28 29.18±0.91
0.05 70.65 24.42±0.54 70.74 16.17±0.88
0.10 58.37 25.42±0.84 60.83 17.72±0.94
0.50 45.26 26.72±0.72 41.37 20.84±0.44
0.75 36.68 30.57±0.77 38.33 24.52±0.14
1.00 25.42 40.43±0.67 25.74 30.41±0.48
Control 89.42 16.35±0.42 100.00 12.76±0.32
both sexes as compared to the untreated parent adults. The
life span of progeny adult in either sex verifying from 3.45 to
7.80 days in male and from 3.55 to 9.76 days in females in
response to residue films of different concentrations and
tending to be in indirectly proportional to the residue film
concentration and differed significantly with the strength of
the residue film of the Dipel (P

Table 3.
WARSI AND CAPOOR, Effect of bacterial preparations on development of diacrisia obliqua 59
Effect of Thuricide HP on post embryonic development in D. obliqua at different concentrations under different
modes of treatment. (Values are mean ± S.E.)
Mode of treatment Concentration (%) Pupation (%) Larval period (days) Emergence (%) Pupal period (days)
L.D.M.
0.05 69.42 22.76±0.68 71.72 14.82±0.42
0.10 58.48 26.52±0.66 65.65 15.76±0.44
0.50 48.24 28.27±0.28 54.72 19.76±0.42
0.75 36.37 33.32±0.37 42.65 23.86±0.56
1.00 25.26 36.46±0.72 22.12 29.76±0.40
T.M.
0.05 69.85 19.64±0.75 72.82 14.94±0.22
0.10 58.40 24.78±0.69 68.47 15.78±0.34
0.50 48.32 26.85±0.42 56.32 20.87±0.46
0.75 36.72 30.75±0.17 48.11 23.60±0.06
1.00 27.12 36.30±0.12 29.75 28.70±0.82
Control 89.43 16.35±0.42 100.00 12.76±0.32
Table 4.
Mode of
treatment
L.D.M.
T.M.
Effect of Thuricide HP on longevity in male and
female D. obliqua at Different modes of treatment
(Values are mean ±S.E.)
Concentration
(%)
Longevity (in days)
Male
Female
0.05 7.92±0.22 8.24±0.42
0.10 7.86±0.44 7.84±0.44
0.50 6.06±0.42 6.06±0.28
0.75 5.04±0.22 5.50±0.20
1.00 3.76±0.15 3.86±0.25
0.05 8.60±0.22 8.90±0.42
0.10 7.02±0.42 7.00±0.46
0.50 6.06±0.45 6.87±0.37
0.75 5.66±0.14 5.80±0.88
1.00 3.68±0.32 3.94±0.36
Control 11.35±0.22 13.76±0.24
applied through leaf dip method prolonged the larval period
varying from 20.86 to 33.35 days and prolonged the pupal
period as compared with the control experiment. The pupal
stage varied from 14.75 to 27.12 days among different
concentration (Table 3). The Leaf dip treatment with any
concentration of the Bactospeine reduce the life span of
progeny adults of the both sexes as compared to the untreated
once. In male moth, the longevity varies from 4.82 to 7.56 days
and in female from 4.05 to 7.66 days differed with the
concentration of the Bactospeine (Table 6).
All concentration of the Bactospeine applied under
Topical method to adults reduced the larval survival and
delayed the pupation as compared to the untreated condition
of parents. The larval period varying from 19.08 to 32.42 days
and prolonging with the advancing concentration of
Bactospeine differed from concentration to concentration
applied earlier by Topical method (Table 5) and the larval
survival varying from 26.52% to 76.25% among different
concentrations from 0.05% to 1.0%
Table 5.
Mode of treatment Concentration (%) Pupation (%) Larval period (days) Emergence (%) Pupal period (days)
L.D.M.
T.M.
Effect of Bactospeine on post embryonic development in D. obliqua at different concentrations under different
modes of treatment. (Values are mean ± S.E.)
0.05 72.76 20.86±0.12 71.82 14.75±0.12
0.10 59.72 22.24±0.14 67.35 15.76±0.22
0.50 48.46 26.18±0.44 54.36 18.52±0.24
0.75 38.57 30.74±0.28 40.21 23.22±0.46
1.00 25.16 33.35±0.79 22.28 27.12±0.22
0.05 76.25 19.08±0.64 75.72 15.25±0.20
0.10 60.72 22.02±1.14 70.25 16.60±0.24
0.50 49.48 24.25±2.12 60.35 18.72±0.46
0.75 38.84 28.42±0.12 42.11 22.15±0.48
1.00 26.52 32.42±0.14 23.82 26.26±0.68
Control 89.43 16.35±0.42 100.00 12.76±0.32

60 Trends in Biosciences 5 (1), 2012
Table 6.
Mode of
treatment
L.D.M.
T.M.
Effect of Bactospeine on longevity in male and
female, D. obliqua at different modes of treatment.
(Values are mean ± S.E.)
Concentration
(%)
In the concentration range of 0.05% to 1.0% the
emergence varying from 23.82% to 75.72% and tending to
decrease with the increasing concentration. The male adult
obtained from the control experiment had the more life span
(11.35 days) as compared to the adult obtained from the parents
treated with any concentration of the Bactospeine and this
fact was applicable to the female adult also. The longevity of
the male adult varying from 4.96 to 7.52 days and that of the
female adult varying from 3.36 to 7.90 days tended to decrease
with the increase in the concentration of the Bactospeine.
(Table 6).
It is evident from the results of the experiments that
lethal concentration of Dipel had marked effect on the larval
development. It would be worthy to mention that surviving
larval of D.obliqua were significantly shorter in the length
and lighter in weight when compared with control. Larvae
also manifested prolongation in larval duration besides
pupation of D.obliqua larvae over and above this moth larval
casualty was also noted in treated moths.
ACKNOWLEDGEMENT
Longevity (in days)
Male
Female
0.05 7.56±0.12 7.66±0.14
0.10 6.80±0.14 7.20±0.13
0.50 6.42±0.43 6.09±0.82
0.75 6.02±0.03 6.80±0.60
1.00 4.82±0.16 4.05±0.26
0.05 7.52±0.12 7.90±0.24
0.10 7.82±0.14 6.85±0.58
0.50 6.48±0.14 6.60±0.36
0.75 6.26±0.18 5.92±0.48
1.00 4.96±0.26 3.36±0.32
Control 11.35±0.22 13.76±0.24
Authors are grateful to Dr. A.V. Singh, Principal, Agra
College, Agra and Dr. R. S. Rawat, Head Department of
Zoology, Agra College, Agra, for providing facilities and
constant encouragement.
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(Lepidoptera : Noctuidae) and persistence in cotton terminals. J.
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Bajpai, Anju. 2003. Studies on the effects of Bacillus thuringiensis
Ber.on the growth and development of Leucinodes orbonalis.Indian
society of Life Science. pp.75-77.
Cantwell, G.E., William, W.C.and Cantwell, M.A. 1986. Effect of ß-
toxin of B.thuriengiensis on the development of Mexican bean
beetle (Epilachna varivosis) (Coleoptera : Coccinellidae). Great
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Chandra, A. Kaushik, N.C. and Gupta, G.P. 1996. Studies of B.
thuringiensis on growth and development of Helicorverpa armigera.
Hubner Annl. Pl. Prot. Sci., 7(2):154-158.
Chaturvedi, R.K. 2003. Studies on the effectiveness of Bacillus
thuringiensis Ber. Against Utetheisa pulchella Linn. (Lepidoptera :
Arctiidae). Indian society of life sciences. pp. 68-69.
Dulmage, H.T. and Martinez, E. 1973. The effect of continuous exposure
to low concentration of delta- endotoxin of B. thuringiensis on
the development of tobacco bud worm, Heliothis virescens. J.
Invertebr. Patho., 22 : 14-22.
Huang, F.N. Buschman, L.L; Higgins, R.A. 2005. Larval survival and
development of susceptible and resistant -Ostrinia nubilalis
(Lepidoptera : Pyralidae ) on diet containing B. thuringiensis.
Agric and Forest, Ent. 7(1): 45-52.
Salama, H.S., Foda, M.S., El-Sharaby, A., Matter, M. and Khalafallah,
M. 1981. Development of some lepidopterous cotton pests as
affected by exposure to sub-lethal level of endotoxin of
B.thuringiensis for different period. J. Invertebr. Pathol., 38(2):220
-227.
Sharma, M. 1993. Effect of certain insect growth regulators on the
growth and development of U. pulchella Linn. (Lep. : Arctiidae)
A. Thesis submitted to Kanpur university for the award of Ph.D.
Degree in Zoology.
Sreenivas, A.G.; Ashoka, j.; Patil, B.V. 2002. Evaluation of the effects
of Bacillus thuringiensis (Berlin er) commercial products against
mulberry silkworm, Bombyx mori (L.). Indian Journal of Sericulture,
41(1): 54-56.
Srivastava, K.L. and Ramkrishnan, N. 1980. Potency of Bactospeine
and Dipel, two commercial formulations of B. thuringiensis Berliner
against Castor semilooper, Achea janata Linn. Indian J. Ent., 42(2):
769-771.
Recieved on 15-9-2011 Accepted on 30-12-2011

Trends in Biosciences 5 (1): 61-63, 2012
Determination of Antimycobacterial Activity of Polyenzyme Preparation Immunoseb-
S using In Vitro Methods
ANITA JOSHI 2 , SHILPA RISBUD 2 , BHAVANA PRADHAN 2 , SMITA DESHPANDE 1
AND RENU BHARADWAJ 1
1
Medical College, Pune. 2 Advanced Enzyme Technologies Ltd. Thane
email : renu.bharadwaj@gmail.com
ABSTRACT
The polyenzyme formulations developed by Advanced Enzyme
Technologies Ltd., Thane, Exclzyme and Immunoseb-S, were
tested for their ability to inhibit the growth of the tubercle
bacilli. For this, the liquid medium method (REMA) was used.
It was found that both the formulations have antitubercular
properties and can inhibit the growth of M tuberculosis at a
Minimum Inhibitory Concentration of 5 and 10 mg/ml
respectively. Although it was not possible to show any
potentiation of standard drugs by in vitro methods, a detailed
study using MDR strains and sensitive strains showed that the
novel formulations are equally effective against both sensitive
and MDR strains. Exclzyme and Immunoseb-S may hence be a
major breakthrough in the management of MDR tuberculosis.
Key words
Polyenzyme formulation, novel antimicrobial, broad
spectrum antimicrobial, MIC, synergy
Currently, tuberculosis is one of the greatest contributors
to adult mortality among infectious diseases. The disease is
responsible for approximately 2 million deaths every year.
According to the WHO, India has the largest no. of TB cases
in the world, accounting for nearly one third of the global
burden. Additionally the HIV epidemic has had a dramatic
impact on the global epidemiology of TB since it is the most
commonly occurring secondary infection in HIV patients.
Tuberculosis is also a major cause for concern due to
the emergence of multi drug resistant strains of
Mycobacterium tuberculosis (MDR-TB). These strains show
resistance to Isoniazid and Rifampicin, the two most powerful
anti-TB drugs, with or without resistance to other drugs
(Paramasivan, 1998). The problem of drug resistance is most
severe in developing countries such as India and China
(Martin, et al., 2003) where additionally poor socioeconomic
conditions, population, HIV and poorly implemented health
measures prevail. Consequently, there is a need for new drugs
for the treatment of TB.
Several plant extracts and other natural bioactive
molecules are being screened as alternatives to these
conventional medicines (Ramaswamy, et al., 2000). Thus to
control the spread of TB it is imperative that cases be detected
and treated effectively in a time bound manner.
Conventionally, antimycobacterial drug susceptibility
testing is done by the agar proportion method using the
inexpensive Lowenstein Jensen medium or Ogawa medium or
Middlebrook’s medium in which results are obtained only after
6-8 weeks. Owing to this, attempts are being made to develop
novel methods in which the test is done using liquid media
(Luz Caviedes, et al., 2000). These methods have been found
to be simple, rapid, quantitative and inexpensive and are done
in microtiter plates. They employ an oxidation-reduction
indicator such as Alamar blue which changes colour from
blue to pink during growth (Varma et al., 2000, Yajko, et al.,
1995, Franzblau, et al., 1998) or MTT which is a tetrazolium
dye. A good correlation has been obtained between the
method and the conventionally used agar proportion method
(Varma, et al., 2002). Further modification by Martin et al.,
2003 suggests the use of the indicator Resazurin which has
been found to be the main component of Alamar blue and is
readily available.
In this study we have used the Resazurin method of
Martin et al., 2003 to screen for antituberculosis properties of
polyenzyme formulations, Exclzyme and Immunoseb-S. We
further modified the method by replacing Middlebrook’s
medium with Kirchner’s medium without the phenol red
indicator. The Mininmum Inhibitory Concentrations of the
formulations were determined. Lastly, the possibility of the
effect of these polyenzymes as potentiators of
antituberculosis drugs was also explored.
MATERIALS AND METHODS
The medium used for culturing organisms was the
Kirchner’s medium without phenol red indicator. The medium
was supplemented with filtered 10% horse serum, Penicillin
100 U/ml of medium and Cycloheximide.
Cultures used in the study were maintained on
Lowenstein Jensen medium. The standard strain of
Mycobacterium tuberculosis: H37RV for screening for
antimycobacterial activity and determination of MIC values.
For the potentiation studies, five known sensitive clinical
isolates and five known MDR isolates were used.
Standard antibiotics used were Rifampicin and INH. The
polyenzymes were dissolved in Tris buffer pH 7.0. Screening
for anti-TB activity and MIC determination were done as per
the method described by Martin et al, 2003 in microtiter plates.
Briefly, serial dilutions of standard drugs and test formulations
were done in tubes. From these, 100 ml was added to

62 Trends in Biosciences 5 (1), 2012
appropriate wells. To these, 100 ml of standardized suspension
of H37RV was added and the plates were incubated at 37ºC for
7 days. Resazurin (0.01% w/v in distilled water; 30 ml) was
added to each well on the seventh day and the plates were
incubated further for 48 hours. Change in colour for every
well was recorded and MIC was calculated.
Change in colour to pink indicated that growth /
respiratory metabolism had taken place. Thus MIC was the
highest dilution which prevented change in colour from blue
to pink.
After determination of MIC values of Exclzyme and
Immunoseb-S, a study was undertaken to study the effect of
the two formulations on 5 sensitive and 5 MDR strains and to
determine whether it was possible to decrease the MIC values
of standard drugs such as Rifampicin and INH by using fixed
concentrations of polyenzymes along with different dilutions
of the standard drug. Thus a test was designed to study
whether any combination therapy was possible with Exclzyme
and Immunoseb-S.
The method used was a combination assay in microtitre
plates with Kirchner’s medium in which Resazurin was added
as a growth indicator on the seventh day. Briefly, serial dilutions
of standard drugs and test formulations were done in tubes.
To wells 50 ml of polyenzyme (MIC concentration) and 50 ml
of dilution of standard antibiotic wee added. Next, 100 ml of
standardized inoculum of the above mentioned clinical isolates
was added to each well. Appropriate controls were included
in each test. Incubation period for each test was 7 days at 37
0
C. Plates were sealed. On the seventh day, Resazurin was
added to each well and the plates were incubated further for
48 hours. The plates were interpreted as before.
RESULTS AND DISCUSSION
Based on these observations, MIC value for MIC value
for Exclzyme was 5 mg/ml and for Immunoseb-S was 10 mg/ml.
The MIC of INH = 0.25 mg/ml and that of Rifampicin = 1 mg/ml
reflecting the accuracy of the assay. The above results were
confirmed using standard staining methods: acid fast bacilli.
Controls for enzyme, antibiotics were blue. Growth control
was pink.
It was observed that it was possible to grow
Mycobacterium tuberculosis using Kirchner’s liquid medium
in seven days instead of the usual 3-4 weeks required on solid
media such as LJ medium. Furthermore, MIC could be
determined in 9 days. MIC values of standard drugs
corresponds to values documented in literature
Both novel formulations Exclzyme and Immunoseb-S
have anti-tubercular properties. The MIC of Exclzyme was 5
mg/ml and MIC of Immunoseb-S was 10 mg/ml.
In the potentiation studies, the MIC values of Rifampicin
and INH did not drop due to addition of Exclzyme and
Immunoseb-S. Thus, the polyenzymes do not have any
potentiation effect on standard antibiotics in vitro
However, it must be noted that Exclzyme and
Immunoseb-S are equally effective at the same concentrations
against both sensitive as well as resistant strains.
Table 2.
MIC values for Exclzyme in combination and alone
Sensitive strains Excl INH INH+ Excl Rifa Rifa+ Excl
A 5 0.125 0.25 0.5 0.5
B 10 0.125 0.25 0.5 0.5
C 5 0.125 0.125 0.25 0.25
D 5 0.062 0.125 0.5 0.25
E 5 0.125 0.125 0.5 0.5
Resistant strains
A 5 0.5 0.25 0.5 0.5
B 5 0.25 0.25 10 0.5
C 10 0.125 0.125 0.25 0.25
D 5 0.25 0.25 1 0.5
E 10 0.25 1 1 0.5
Table 3.
MIC values for Immunoseb-S in combination and
alone
Sensitive strains SEB-S
Mg/ml
INH
g/ml
INH+ SEB-s
g/ml
Rifa
g/ml
Rifa + SEB-s
g/ml
A 5 0.125 0.125 0.5 0.5
B 10 0.125 0.062 0.5 0.5
C 10 0.125 0.25 0.25 0.25
D 10 0.062 0.25 0.5 0.5
E 5 0.125 0.125 0.5 0.5
Resistant strains
A 10 0.5 0.5 0.5 0.5
B 5 0.25 0.25 10 1
C 10 0.125 0.125 0.25 0.25
D 10 0.25 0.25 1 0.5
E 10 0.25 0.5 1 0.25
Controls: Growth control: pink indicating growth
Enzyme / antibiotic controls: blue indicating inhibition
Table 1. Exclzyme
Dilution (mg/ml) 40 20 10 5 2.5 1.25 0.62 0.31 0.15 0.07
Colour of medium Blue Blue Blue Blue Pink Pink Pink Pink Pink Pink
Immunoseb-S:
Dilution (mg/ml) 40 20 10 5 2.5 1.25 0.62 0.31 0.15 0.07
Colour of medium Blue Blue Blue Pink Pink Pink Pink Pink Pink Pink
Std drugs:
INH g/ml 4 2 1 0.5 0.25 0.125
Colour of medium Blue Blue Blue Blue Blue Pink
Rifampicin g/ml 4 2 1 0.5 0.25 0.125
Colour of medium Blue Blue Blue Pink Pink Pink

JOSHI, et al., Determination of Antimycobacterial Activity of Polyenzyme Preparation Immunoseb-S 63
The in vitro results are definitely encouraging and in
vivo studies in appropriate models need to be undertaken.
ACKNOWLEDGEMENT
The authors wish to thank Mr. V. L. Rathi and Mr. C. L.
Rathi, of Advanced Enzyme Technologies Ltd., Thane, and
Specialty Enzymes Co. USA, not only for funding this study
at the B. J. Medical College, Pune but also for their enthusiastic
encouragement, valuable inputs and support throughout these
studies. The authors also wish to thank Dr. A. Martin
Mycobacteriology Unit, Institute of Tropical Medicine,
Nationalestraat, 155, Antwerp, B-2000 Belgium for his patient
support and inputs.
Fig.1.
The Resazurin Microtiter Assay for determination of
MIC values of antimycobacterial drugs.
Pink : growth or no inhibition Blue : inhibition
The following study was undertaken to determine
whether the novel polyenzyme formulations Exclzyme and
Immunoseb-S developed by Advanced Enzyme Technologies
Ltd., Thane have any antitubercular properties. Furthermore,
the possibility of using the formulations in combination with
the currently used drugs was also explored.
It was found that the two formulations have
antitubercular properties and the Minimum Inhibitory
Concentration (MIC) values were established using the
Resazurin Microtiter Assay (REMA) for M. tuberculosis. The
MIC value was 5 and 10 mg/ml respectively.
However, the use of the polyenzyme formulations in
conjunction with Rifampicin or INH did not lower the
established MIC values of Rifampicin and INH. This shows
that it was not possible to demonstrate potentiation of standard
fdrugs by using polyenzyme formulations by in vitro methods.
Nevertheless, an interesting finding of this study was
that MDR strains and sensitive strains showed a similar
response to the polyenzyme formulation. Therefore the use
of the formulations could be a breakthrough in the management
of MDR strains.
It should also be noted that since more than one enzyme
has been included in the formulation, development of
resistance to the formulation may be a complicated and unlikely
phenomenon. This may help restrict circulation of MDR
strains.
LITERATURE CITED
Franzblau, S. G., R. S. Witzig, J. C. McLaughlin, P. Torres, G. Madico, A.
Hernandez, M. T. Degnan, M. B. Cook, V. K. Quenzer, R. M.
Ferguson, and R. H. Gilman. 1998. Rapid, low-technology MIC
determination with clinical Mycobacterium tuberculosis by using
the Microplate Alamar Blue Assay. J. Clin. Microbiol., 36:362-
366.
Luz Caviedes, Tien-Shun Lee, Robert, H. Gilman, Patricia Sheen, Emily
Spellman, Ellen H. Lee, Douglas E. Berg, Sonia Montenegro-James,
and The Tuberculosis Working Group in Peru. 2000 Rapid, Efficient
Detection and Drug Susceptibility Testing of Mycobacterium
tuberculosis in Sputum by Microscopic
Observation of Broth Cultures. Journal of Clinical Microbiology,
38(3):1203-1208.
Martin, A., Camacho, M., Portaels, F. and Palomino, J.C. 2003. Resazurin
Microtiter Assay Plate testing of Mycobacterium tuberculosis
Susceptibilities to second-line drugs: rapid, simple and inexpensive
method. Antimicrobial agents and Chemotherapy, 47(11): 3616-
3619.
Paramasivan, C.N. 1998. An overview on drug resistant tuberculosis in
India. Ind. J. Tub., 45: 73-81.
Ramaswamy, A.S. and Sirsi, M., 2000. Antitubercular activity of some
chemical constituents from higher plants. Ind. J. of Pharmacy,
22(2): 34-35.
Varma, M., Kumar, S., Kumar, A. and Bose, M. 2002. Comparison of
Etest and Agar proportion method of testing drug susceptibility of
M tuberculosis. Ind. J. Tub., 490: 217-220.
Yajko, D.M., Madej, J.J., Lancaster, M. V., Sanders, C. A., Cawthon, V.
L., Gee, B., Babst, A. and Hadley, W. K. 1995. Colorimetric method
for determining MICs of antimicrobial agents for Mycobacterium
tuberculosis. J. Clin. Microbiol. 33:2324-2327.
Recieved on 7-2-2012 Accepted on 12.4.2012

Trends in Biosciences 5 (1): 64-67, 2012
Genetic Divergence Analysis in Faba Bean (Vicia faba L.)
B.K. CHAUBEY 1 , C.B. YADAV 1 , V.K. MISHRA 2* AND K. KUMAR 1
2
Department of Genetics and Plant Breeding, 1 Department of Plant Molecular Biology and Genetic
Engineering, N. D. University of Agriculture and Technology, Kumarganj, Faizabad 224 229 (U.P.)
*e-mail: vinay.mishra111@gmail.com
ABSTRACT
A ?eld experiment was conducted at N.D.U.A. & T., Kumarganj,
Faizabad, to study genetic diversity in faba bean (Vicia faba L.)
landraces. Seventy random germplasm accessions were grown
in an augmented block design. The genetic divergence existing
in 70 faba bean germplasm collection was studied by employing
non-hierarchical Euclidean cluster analysis for 11 quantitative
characters. The 70 genotypes were grouped into eight distinct
clusters, this indicated existence of high degree of genetic
diversity among germplasm collection in the present study.
The eight clusters in divergence analysis contained genotypes
of heterogeneous origin, thereby indicating no parallelism
between genetic and geographic diversity. Therefore, crosses
between the members of clusters separated by high inter-cluster
distance are likely to throw desirable segregants. The different
clusters showed considerable differences in intra-cluster group
means of eleven characters and genotypes having distinctly
different mean performance for various characters were
separated into different clusters.
Key words
Fababean, genetic divergence, segregants, clusters.
Faba bean ( Vicia faba L.) is one of the most important
legumes in the world. Due to its partial cross-pollination, faba
bean is highly heterogeneous and consequently contains
broad genetic variation within varieties and landraces, which
make the conservation of germplasm resources more expensive
and difficult (Duc, et al., 2010, Gemechu, et al., 2005). Genetic
diversity arises either due to geographical separation or due
to genetic barriers to crossability (Singh, 1990). Whether
differences in geographic origin (source) imply genetic
distance in parental selection for hybridization is still a matter
of some controversy. Joshi and Dhawan, 1966 suggested the
concept that geographic diversity may serve as an index of
genetic diversity in parental selection. Others argue that
genetic divergence was not apparently related to geographic
diversity in some crops (Durga Prasad, et al., 1985, Sindhu,
1985, Nadaf, et al., 1986, Rezai and Frey, 1990, Katule, et al.,
1992).
MATERIALS AND METHODS
The experiment was conducted at the Student’s
Instructional Farm of Narendra Deva University of Agriculture
and Technology, Narendra Nagar (Kumarganj), Faizabad (U.P.),
India, which is geographically situated between 26.47 0 N
latitude, 82.12 0 E longitudes and at an altitude of 113 meters
above the mean sea level in the gangetic plains of eastern U.P.
The climate of district Faizabad is semi-arid with hot summer
and cold winter. Nearly 80 per cent of total rainfall is received
during the monsoon (only upto September) with a few showers
in the winter. The experimental material for the present
investigation consisted of 70 germplasm lines of faba bean
and three check varieties namely, PRT 7, PRT 12 and Vikrant
were planted in augmented block design with three checks
repeated after every 10 lines of the test entries. The checks
were distributed systematically in each block. Each entry and
checks were grown in double row of 4 m length; spaced 30 cm
apart and distance of plants within rows was maintained at
about 10 cm by thinning. Observations on days to 50%
flowering, days to maturity, plant height (cm), number of
branches per plant, number of pods per plant, number of
seeds per pod, biological yield per plant (g), seed yield per
plant (g) , harvest index (%), 100-seed weight and protein
content (%) were recorded on the basis of five plants randomly
selected and tagged from each row except for days to 50%
flowering and days to maturity, which were recorded on the
plot basis. All the recommended cultural practices were
followed to raise a good crop (Table 4).
RESULTS AND DISCUSSION
The genetic divergence existing in seventy faba bean
germplasm collection was studied by employing nonhierarchical
Euclidean cluster analysis for eleven quantitative
characters. The pseudo F-test revealed that eight cluster
arrangements were the most appropriate for this material.
Therefore, the 70 genotypes were accepted to be grouped
into eight different non-overlapping clusters. The distribution
of 70 faba bean lines in eight clusters is given Table 1.
The highest number of genotypes appeared in cluster V
which contained 17 entries followed by cluster VIII having 15
genotypes. Cluster III, cluster VI, cluster VII and cluster I
having 12, 12, 7 and 3 respectively. Cluster II and cluster IV
both were represented by 2 entries each and had minimum
number of genotypes among all the clusters. The estimates of
average intra and inter cluster distance for the eight cluster is
presented in Table 2. The highest intra-cluster distance was
recorded for cluster VIII (2.491) followed by cluster II (2.327),
while the lowest value was recorded in case of cluster IV
(1.808). The maximum inter cluster distance was recorded
between cluster IV and cluster I (7.604) followed by cluster

Table 1.
CHAUBEY, et al., Genetic Divergence Analysis in Faba Bean (Vicia faba L.) 65
Clustering pattern of 70 Faba bean genotypes on the basis of non-hirarchical Euclidean cluster analysis.
Cluster No. of
Genotypes
Number genotypes
I 3 EC 251014, EC 324677, EC 361427.
II 2 EC 331587, EC 382423.
III 12 EC 243626, EC 25192, EC 243637, EC 329605, EC 329627, EC 374731, EC 329673, EC 329715, EC 243860,
IC 361498, IC 348948, EC 329700.
IV 2 EC 243709, EC 117739.
V 17 EC 361470, EC 331549, EC 117734, EC 117744, EC 323731, EC 117749, EC 243772, EC 329683, EC 243786,
EC 117842, EC 267648, EC 361496, EC 243525, IC 331571, EC 2558, EC 322967, EC 329677.
VI 12 EC 293713, EC 243755, EC 249947, EC 329003, EC 243784, EC 243791, EC 329812, EC 243782, EC 243781,
EC 117753, EC 243743, EC 329707.
VII 7 EC 117745, EC 267641 EC 117726, EC 248945, EC 329668, EC 329662, EC 329714.
VIII 15 EC 354951, EC 249851, EC 343808, EC 329724, EC 329675, EC 329725, EC 25085, EC 243596, EC 243770,
EC 243529, EC 351919, EC 247696, EC 243895, EC 343696, EC 329701.
Table 2.
Estimates of average intra and inter-cluster distances for the eight clusters
Clusters I II III IV V VI VII VIII
I 2.194 4.245 3.548 7604 3.080 4.734 3.356 4.466
II 2.327 5.779 7.008 4.936 4.894 6.613 4.645
III 2.028 6.546 2.331 3.105 2.609 3.525
IV 1.808 7.112 5.614 7.305 4.257
V 2.104 3039 2.836 3.255
VI 2.265 4.446 2.463
VII 1.859 4.023
VIII 2.491
Bold figures represent intra-cluster distances.
VII and cluster II (7.305). The inter cluster distance between
cluster IV and cluster II (7.008), cluster VII and cluster II (6.613)
cluster IV and cluster III (6.546) and cluster III and cluster II
(5.779), were also in high orders. The minimum inter cluster
distance was observed between cluster V and cluster III (2.331)
followed by cluster III and cluster VI (2.463). The cluster
means, standard deviation and coefficient of variability for
eleven characters are presented in Table 3.
The genotypes of cluster VII took maximum days to
50% flowering (x = 79.43, cv=3.62) followed by cluster V (x=
76.82, cv=5.25). The genotypes took minimum days to 50%
flowering were concentrated cluster III (x=64.83, cv=6.57)
followed by cluster VI (x= 67.75, cv=10.69). The entries
representing cluster VII showed highest cluster mean for days
to maturity (x=168.00, cv= 1.38) followed by cluster VI (x=
166.92, cv= 1.95), while the lowest mean for the characters is
showed by cluster II (x=157.00, cv= 3.61) followed by cluster
I (x= 161.00, cv= 1.86). The entries representing cluster IV
showed highest cluster mean for plant height (x= 132.50,
cv=2.67) followed by cluster VII (x= 129.14, cv = 3.94), while
the lowest cluster mean for plant height was observed in
cluster V (x= 114.78, cv= 6.72) followed by cluster II (x=116.70,
cv= 2.31). The genotypes of cluster IV (x = 8.80, cv= 16.02)
had highest cluster mean for number of branches per plant
followed by cluster II (x= 7.10, cv = 5.92), while lowest cluster
mean for characters was observed in cluster VII (x=4.73, cv=
9.72). The cluster IV (x = 53.40, cv= 36.75) had highest cluster
mean for number of pods per plant followed by cluster II (x=
44.30, cv= 11.81), while the cluster VII (x= 19.63, cv = 20.17)
had lowest cluster mean for number of pods per plant followed
by cluster I (x= 21.60, cv= 4.90). The highest cluster mean for
number of seeds per pod showed by genotypes of cluster VI
(x= 3.12, cv= 6.41) followed by cluster IV (x= 3.08, cv = 1.95)
and lowest cluster mean for number of seeds per pod showed
by genotypes of cluster III (x= 2.70, cv= 5.93) followed by
cluster V (x= 2.76, cv= 7.25). 100-seed weight was highest in
the genotypes of cluster VI (x= 34.30, cv = 8.60), followed by
cluster II (x= 34.19, cv= 9.940), while it was lowest for entries
occurring in cluster I (x= 28.70, cv= 23.38) followed by cluster
IV (x= 29.28, cv= 16.19). The highest cluster means for biological
yield per plant exhibited by cluster VIII (x = 99.62, cv= 17.47)
followed by cluster IV (x= 82.50, cv= 19.70), while the lowest
cluster mean for biological yield per plant showed by
genotypes of cluster I (x= 51.67, cv = 21.58) followed by cluster
VII (x = 52.63, cv= 22.25). The highest cluster mean for seed
yield per plant was observed in case of cluster IV (x= 34.41,
cv= 4.01), which indicates that lines having very high seed
yield were concentrated in this cluster followed by cluster
VIII (x= 27.91, cv= 16.12). The genotypes with low seed yield
were found to be grouped in cluster VII (x= 13.39, cv= 30.47)
followed by cluster I (x= 14.37, cv= 17.12), cluster III (x= 16.97,
cv= 20.74) and cluster V (x = 18.32, cv= 22.54). Remaining
cluster had mean seed yield per plant of medium range. The
cluster II (x = 38.67%, cv= 35.51) comprised of entries which
produced highest mean for harvest index followed by cluster
VI (x = 36.595, cv = 15.91). The lower value was observed for
cluster IV (x= 18.97%, cv= 12.86) followed by cluster VII (x=
25.51%, cv= 20.07). Rest of the clusters showed moderate
mean value for this character. The protein content was the
highest in the genotypes of cluster III (x= 26.00%, cv= 0.58)

66 Trends in Biosciences 5 (1), 2012
Table 3.
Clusters
I
II
III
IV
V
VI
VII
VIII
Cluster mean, standard deviation and coefficient of variability for different characters for 8 clusters
Days to
50%
flowering
Days to
maturity
Plant
height
(cm)
No. of
branches/
plant
No. of
pods/plant
No. of
seeds/pod.
100-seed
weight
(g)
Biological
yield/ plant
(g)
Seed
yield/
plant (g)
Harvest
index (%)
Protein
content
(%)
1 2 3 4 5 6 7 8 9 10 11
Mean 72.67 161.00 120.40 4.73 21.60 2.92 28.70 51.67 14.37 28.02 25.03
S.D. 2.08 3.00 7.81 0.46 1.06 0.33 6.71 11.15 2.46 1.44 0.25
C.V. % 2.86 1.86 6.48 9.72 4.90 11.30 23.38 21.58 17.12 5.14 1.00
Mean 69.00 157.00 116.70 7.10 44.30 2.92 34.19 72.20 27.16 38.67 24.65
S.D. 1.41 5.66 2.69 0.42 5.23 0.51 3.40 11.03 5.65 13.73 0.07
C.V. % 2.04 3.61 2.31 5.92 11.81 17.47 9.94 15.28 20.80 35.51 0.28
Mean 64.83 166.08 121.90 5.20 24.22 2.70 30.33 63.18 16.97 28.34 26.00
S.D. 4.26 2.64 7.65 0.92 5.70 0.16 2.56 17.76 3.52 6.98 0.15
C.V. % 6.57 1.59 6.28 17.69 23.53 5.93 8.44 28.11 20.74 24.63 0.58
Mean 69.00 166.50 132.50 8.80 55.40 3.08 29.28 182.50 34.41 18.97 25.85
S.D. 1.41 3.54 3.54 1.41 20.36 0.06 4.74 16.26 1.38 2.44 0.21
C.V. % 2.04 2.13 2.67 16.02 36.75 1.95 16.19 8.909 4.01 12.86 0.81
Mean 76.82 162.53 114.78 4.93 26.71 2.76 32.43 56.54 18.32 32.97 25.85
S.D. 4.03 2.62 7.71 1.03 5.84 0.20 3.07 13.86 4.13 5.59 0.18
C.V. % 5.25 1.61 6.717 20.89 21.86 7.25 9.47 24.51 22.54 16.95 0.70
Mean 67.75 166.92 118.07 5.88 40.12 3.12 34.30 75.04 26.90 36.59 25.99
S.D. 7.24 3.26 5.48 1.29 8.48 0.20 2.95 13.56 3.21 5.82 0.23
C.V. % 10.69 1.95 4.64 21.94 21.14 6.41 8.60 18.07 11.93 15.91 0.88
Mean 79.43 168.00 129.14 4.29 19.63 2.79 28.70 52.63 13.39 25.51 25.84
S.D. 2.88 2.31 5.09 0.56 3.96 0.29 3.49 11.71 4.08 5.12 0.15
C.V. % 3.62 1.38 3.94 13.05 20.17 10.39 12.16 22.25 30.47 20.07 0.58
Mean 76.60 164.60 127.91 6.44 41.03 2.95 33.65 99.61 27.91 28.79 25.80
S.D. 5.89 4.87 5.21 0.98 8.72 0.26 3.10 17.40 4.50 6.29 0.20
C.V. % 7.69 2.96 4.07 15.22 21.25 8.81 9.21 17.47 16.12 21.85 0.04
followed by cluster VI (x= 25.99%, cv= 0.88), while it was lowest
for entries occurring in cluster II (x= 24.65%, cv= 0.28) followed
by cluster I (x= 25.03%, cv= 1.00). Remaining clusters had
moderate means for protein content.
Cluster I had 3 genotypes which were characterized by
lowest cluster mean for 100- seed weight and biological yield
per plant and average cluster mean for remaining traits. Two
genotypes presents in cluster II resulted in lowest cluster
mean for days to maturity, protein content and average cluster
mean for rest of the characters except harvest index that has
highest cluster mean performance. The cluster III had 12
genotypes resulting in lowest cluster mean for days to 50%
flowering and number of seeds per pod and highest cluster
mean for protein content, having average cluster mean for all
other characters. Two genotypes falling in the cluster IV
resulted in better and highest cluster mean for all the characters
except, harvest index that had lowest cluster mean performance.
Cluster V comprising 17 genotypes had lowest cluster mean
performance for plant height and average cluster mean
performance for remaining traits. Cluster VI having 12
genotypes possessed highest cluster mean for number of
seeds per pod and 100 seed weight and medium cluster mean
for rest of the other characters. The seven genotypes of cluster
VII produced highest cluster mean for days to 50% flowering
and days to maturity and lowest cluster mean for number of
branches per plant, number of pods per plant, 100- seed weight
and seed yield per plant and average cluster mean for all other
characters. The cluster VIII contained 15 genotypes having
highest cluster mean for biological yield per plant and average
mean for remaining all characters. The above observation
confirm wide variation from one cluster to another in respect
of cluster mean, which indicated that genotypes having
distinct different mean performance for various characters
were separated into different clusters. The eight clusters in
the aforesaid divergence analysis contained frequently the
genotypes of heterogeneous origin (Table 1), although the
genotype originated in same place or geographic region were
also found to be grouped together in same cluster, the instance
of grouping of genotypes of different origin or geographical
region also in same cluster were observed in case of all the
eight clusters. This suggests lack of parallelism between
genetic and geographic diversity. Therefore, the selection of
parental material for hybridization programme simply based
on geographic diversity may not be a successful exercise for
the choice of suitable divergent parents. Selection on the basis
of genetic divergence analysis would be more rewarding than
the choice made on the basis of geographic diversity. This
finding is in agreement with the report advocating lack of
definite relationship between genetic and geographic diversity
in faba bean.
An examination of the estimates of within and between
cluster diversity presented by intra and inter cluster D 2 values
revealed that the genotypes of the same cluster had little
divergence from each other with respect to aggregate effect
of 11 characters under study (Table 4). Therefore, the chances
of obtaining good recombinants in segregating generation

Table 4.
CHAUBEY, et al., Genetic Divergence Analysis in Faba Bean (Vicia faba L.) 67
The most desirable genotypes identified for eleven characters studied
S. No. Characters Genotypes
1. Days to 50% flowering EC 293713, EC 243791, EC 243860, EC 25192, EC 243626, EC 117753
2. Days to maturity EC 267648, EC 25085, EC 247696, EC 243525, EC 243860, EC 243791, EC 322967
3. Plant height (cm EC 247696, EC 329714, EC 243709, EC 329673, EC 243626, EC 343696
4. No. of branches/ plant EC 243709, EC 243782, EC 382423, EC 243860, EC 117739, EC 243525
5. No. of pods/plant EC 243709, EC 243529, EC 329701, EC 243755, EC 25085, EC 247696
6. No. of seeds /plant EC 382423, EC 243784, EC 249851, EC 247696, EC 361427, EC 293713
7. 100-seed weight EC 329707, EC 243781, EC 243808, EC 243895, EC 329701, EC 329673
8. Biological yield/ plant (g) EC 243709, EC 117739, EC 243696, EC 329701, EC 25085, EC 117753
9. Seed yield/ plant (g) EC 243529, EC 243709, EC 117753, EC 329701, EC 243784, EC 25085, EC 329724, EC 329725,
EC 243525, EC 243781
10. Harvest index (%) EC 382423, EC 267648, EC 329003, EC 243781, EC 329812, EC 249947, EC 243529
11. Protein content (%) EC 243784, EC 243755, EC 323731, EC 248945, EC 329707, EC 243696, EC 243626
by crossing the members of the same cluster are very low. It
is, therefore, suggested that crosses should be attempted
between the genotypes belonging to clusters separated by
large inter-cluster distance in this respect. The highest inter
cluster distance was observed between cluster IV and I.
Cluster I also had high inter cluster distance from cluster V,
VIII, II and III. Thus the crossing between the genotypes
belonging to cluster I with those of cluster IV, V, VII, II and III
may throw desirable transgressive segregants. The lowest
inter- cluster distance was observed between cluster III and V
followed by cluster VI and VIII, cluster III and VII, cluster V
and VII, and cluster V and VI which indicated that genotypes
present in these clusters were genetically close to each other.
The crosses between genotypes belonging to the cluster
separated by low inter cluster distances are unlikely to
generate promising recombinants in segregating generations.
The intra-cluster group mean for eleven characters revealed
considerable differences between the clusters in respect of
cluster means (Table-2). The crosses between the entries
belonging to cluster pairs separated by large inter- cluster
distances and having cluster means for one or other characters
to be improved is likely to be more useful.
The concept that difference in geographic origin should
be used as an index of genetic diversity in parental selection
could not reliably be used as a strategy and parental selection
should rather be made based on systematic assessment of
genetic distance in a specific population. Breeding programs
should also focus on effective and efficient exploitation of
not only inter-regional diversity in the country as a whole but
also intra-regional diversity in India. Inter-cluster gene
recombination of sample accessions drawn from the
significantly distant clusters followed by selection should
prove to generate agronomically desirable progenies as
expected.
LITERATURE CITED
Duc , G. , S. Y. Bao , M. Baum , B. Redden , M. Sadiki , M. J. Suso , M.
Vishniakova , and Zong, X. X. 2010 . Diversity maintenance and
use of Vicia faba L. genetic resource. Field Crops Research, 115:
270-278 .
Durga Prasad, M.M.K., Arunachalam, V. and Bandyopadyay, A. 1985.
Diversity pattern elucidating parents for hybridization in varieties
of groundnut, Arachis hypogaea L. Trop. Agric., 62: 237–242.
Gemechu Keneni, Belay Simane and Getinet Gebeyehu 1997. Genetic
diversity of groundnut germplasm in Ethiopia. Ethiop. J. Agri. Sci.,
16(1&2): 1–13.
Gemechu Keneni, Mussa Jarso, Tezera Wolabu and Getnet Dino. 2005.
Extent and pattern of genetic diversity for morpho-agronomic
traits in Ethiopian highland pulse landraces II. Faba bean (Vicia faba
L.). Genetic Resources and Crop Evolution, 52: 551–561.
Joshi, A.B. and Dhawan, N.L. 1966. Genetic improvement in yield with
special reference to self-fertilizing crops. Indian J. Genet., 26A:
101–103.
Katule, B.K., Thombare, M.V., Dumbre, A.D. and Pawar, B.B. 1992.
Genetic diversity in bunch groundnut. J. Maharashtra Agri. Univ.,
17: 302–303.
Nadaf, H.L., Habia, A.F. and Goud, J.V. 1986. Analysis of diversity in
bunch groundnut. J. Oilseeds Res., 3: 37–45.
Rezai, A. and Frey, K.J. 1990. Multivariate analysis of variation among
wild oat accessions-seed traits. Euphytica, 49: 111– 119.
Sindhu, J.S. 1985. Multivariate analysis in faba bean (Vicia faba L.).
FABIS, 12: 5–7.
Singh, B.D. 1990. Plant Breeding: Principles and Methods. Kalyani
Publishers, New Delhi-Ludhiana.
Recieved on 15.1.2012 Accepted on 18.4.2012

Trends in Biosciences 5 (1): 68-70, 2012
Evaluation of Different Substrate for Mass Multiplication of Trichoderma spp.
ANURADHA SINGH, MOHD SHAHID, VIPUL KUMAR AND MUKESH SRIVASTAVA
Department of Plant Pathology, C.S. Azad University of Agriculture & Technology, Kanpur
e.mail: singhanu@gmail.com,mo.shahid@sify.com
ABSTRACT
Trichoderma viride and T. atroviride are two most widely used
bio-control agents. They are applied to soil, foliage or roots. In
spite of their high efficacy under controlled condition, their
performance at farmers field is not consistent. Unlike chemicals
these bio-control agents need support even after their
application to get established in targeted niche. Present study
was undertaken to achieve major issue involved in mass
production and utilization of bio-control agent by selection of
effective strains for mass multiplication and its formulation.
Which deals with use of cow dung, neem cake, wheat straw,
sorghum grains, rice bran and spent compost of mushroom
either alone or in certain combinations, with or without
additives such as jaggery and wheat flour and having differential
moisture levels were evaluated as substrates for mass production
of T. viride and T. atroviride. Jaggery and wheat flour served as
nutritional supplements, which enhanced the conidial yield.
An increase in the number of viable propagules up to 30 days
was noted regardless of the substrates and its moisture levels.
Although highest initial population of Trichoderma viride was
observed in sorghum grains (132 x 10 7 cfu g -1) , propagule viability
was low in spent compost of mushroom (11.9 x 10 7 cfu g -1) that
compared to other substrates. Neem cake + wheat flour mixture
at 35% moisture also gave longer shelf life(114.3 x 10 7 cfu g -1)
for Trichoderma progagules.
Key words Biocontrol, organic substrates, substrate moisture levels.
Among several groups of plant diseases major amount
of work has been done on the biological/integrated control of
soil borne fungal plant pathogens by using fungal antagonist
like, Trichoderma spp. Trichoderma is one of the common
fungal biocontrol agent, is being used world wide for suitable
management of various foliar and soil borne plant pathogens.
Biological agents like Trichoderma viride and Trichoderma
atroviride are acclaimed as effective, ecofriendly and cheap,
nullifying the ill effectives of chemicals. Therefore, of late,
these biocontrol agents are identified to act against on array
of important soil borne plant pathogens causing serious
diseases of crops. Therefore considering the cost of chemical
pesticides and hazardous involves, biological control of plant
disease appears to be an effective and eco-friendly approach
being practice world over. Further biological control strategy
is highly compatible with sustainable agriculture and has a
major role to play as a component of integrated pest
management (IPM) programme (Chaudhari, et. al.,. 2011).
Trichoderma as a potent fungal biocontrol agent against
a range of plant pathogens has attracted considerable scientific
attention (Tewari and Mukhopadhyay, 2001). Different organic
media like neem cake, coir path, farmyard manure, and
decomposed coffee pulp also have been suggested for its
multiplication (Saju, et al., 2002). Yet reports on the optimum
moisture levels of these substrates for high inoculum
production of Trichoderma spp. are inadequate. Therefore, a
study was conducted to evaluate some locally available
organic substrates and to standardize the optimum moisture
levels for mass multiplication and long-term survival of
Trichoderma spp.( Rini and Sulochana, 2007). Large scale
production, along with shelf life and establishment of bioagents
in targeted niche, determine the success of biological
control. Therefore cost effective large scale production, shelf
life of formulation, establishment of bio-agent in to targeted
niche and consistency in disease control are the primary
concern with augmentative biological control.
For mass multiplication of bioagent through solid state
fermentation technology an enormous quantity of spore
biomass is needed. Various substrates like Cow dung, neem
cake, wheat straw, sorghum grains, rice bran ,paddy straw
and spent compost of mushrooms are being used for mass
multiplication of Tricoderma spp. with various degree of
success.
MATERIALS AND METHODS
Isolation and Identification of Trichoderma spp. : Soil
samples collected from various rhizosphere soil of infected
field of different places of Uttar Pradesh and Trichoderma
spp. were isolated on PDA medium by following serial dilution
plate technique as described by (Johnson and Curl, 1972).
Ten gram soil sample from well pulverized, air dried soil was
added into 90 ml sterile water in a flask to make 1:100 dilution
(10 -1 ).The mixture was vigorously shaken on a magnetic shaker
for 20-30 minutes to obtain uniform suspension. One ml of
suspension from flask was transferred into a test tube
containing 9 ml sterile water under aseptic condition to make
1:10 (10 -2 ) dilution. Further dilution 10 -7 was made by pipetting
1ml suspension into additional water as prepared above. One
ml each liquids of 10 -7 dilution was transferred into 10 sterile
Petri plates which was previously poured by 15 ml sterile PDA
medium and spread uniformly, PDA is the best solid media for
growth and sporulation of Trichoderma spp ( Singh, et. al.,
2011). The Petri plates were incubated at 25±2 0 C for 7 days in
an incubator, because the most favourable temperature for
growth of Trichoderma spp was found in between 25-30 ºC
(Shahid, et. al. 2011). As soon as the mycelial growth were

SINGH, et al., Evaluation of Different Substrate for Mass Multiplication of Trichoderma spp. 69
visible in the PDA culture medium. The hyphal tips from the
advancing mycelium were cut and transferred into the culture
slants containing PDA medium for further purification and
identification of pathogen. The pure culture was obtained by
adopting single spore technique. Green conidia forming fungal
bodies were selected and microscopic observation was
identified to be Tricoderma viride and Trichoderma atroviride
The culture was maintained on PDA slants. The purified
culture was then confirmed by ITCC, Division of Plant
Pathology IARI, New Delhi-12 for further experiment and
investigation.
Processing of substrates: The substrates included
preboiled sorghum grains, cow dung, Neem cake, rice bran,
paddy straw and spent compost of mushroom either alone or
uncertain combinations (Table 1). Jaggary (3%) and wheat
flours (10%) were used as nutritional supplements for
enhancing conidial yield (Prasad, et.al., 2002) and having
differential moisture levels were evaluated as substrates for
mass multiplication of T. viride and T. atroviride. The waste
substrate cuts in the form of pieces and shade dried it. The
dried substrate measure and add jaggery and wheat flour for
nutritional support the moisture level of that mixture was
maintained up to 30-35%. The media got sterilized by atuoclaved.
Each solid substrate was taken in petri-plates,
inoculated with fungal mycelia and incubated at 25±2°C in
incubator for 7-10 days. The treatments were arranged in
completely randomized design with three replications and the
data analyzed using the analysis of variance technique
(Chaudhari, et. al., 2011). Colony forming units (cfu) were
counted after 2 days of incubated. The study was conducted
during the period from September to December 2011.
Mass multiplication of Trichoderma spp: Trichoderma
spp. have been grown on wide range of grains viz. maize,
sorghum, pearl millet, wheat, Jhangora weed (Echinocloa
frumantaceae), wheat bran, wheat straw, waste tea leaves,
banana fruit bark, coffee husk, paddy-straw, Diatomaceous,
earth granule impregnated with molasses (Zaidi and Singh,
2004). Sugarcane pressmud have been widely utilized for mass
multiplication and delivery of the Trichoderma biopesticides
in soil (Singh and Joshi, 2007).Mass cultures of Trichoderma
isolates were made on sorghum grains. Half boiled sorghum
grains (250gm) were filled in polypropylene bags (25 × 30 cm)
and autoclaved at 15 psi pressure for half an hour. After cooling
the grains were inoculated with seven days old culture of
Trichoderma, tied and incubated at 25±2°C in incubator for 7-
8 days.After seven days colonized grains were air-dried in
open shade, grounded with the help of wily mill to get fine
powder and passed through 50 and 80 mesh size sieves,
simultaneously to obtained pure spore powder.
RESULTS AND DISCUSSION
Among the different substrates tested, sorghum grains
with jaggery showed the highest growth of T. viride. White
mycelial growth was observed on sorghum grains on the 5 th
day of incubation and it covered the entire surface of the
sorghum grain substrate with green sporulation in 8-10 days.
Maximum biomass production of T. viride (132.93 x 10 7 cfug -1 )
at 10 days of incubation which was significantly superior to
others (Table 1). In case of Neem cake+ wheat flour mixture
mycelial growth was visible over the surface on 5 th day and it
took 10 days to cover the whole substrate. After incubation
of 10 th days the population of 114.30 x 10 7 cfug -1 ) was recorded
on this substrate. On cowdung and spent compost of
Table 1.
Effect of different substrates on the population of
Trichoderma viride and T. atroviride
Treatments
Mean population (x 10 7 cfu g -1 )
at 10 th day of incubation (n=3)
T. viride T. atroviride
Cowdung 19.600 16.636
Neem cake + wheat flour 10% 114.300 107.566
Sorghum grains + Jaggery 3% 132.93 123.400
Paddy straw 25.533 19.800
Rice brans 31.033 19.400
Spent compost of mushroom 11.933 10.7667
CD (0.05) 5.9052 6.530
Neem cake + wheat(10%) flour colonized by T. viride
Sorghum grain powder + (5%) Jaggery colonized by T. viride.

7 0 Trends in Biosciences 5 (1), 2012
mushroom when used along scanty growth of the fungus
was noted. Rice bran recorded a population of 31.03 x 10 7
cfug -1 which was statistically on per with paddy straw
substrate.
Growth rate of T. atroviride on different organic
substrates was similar to that of T. viride. Sorghum grains
maintained the maximum growth and spore count of 123.4x10 7
cfug -1 followed by neem cake + wheat flour mixture 107.5x10 7
cfug -1 . Cowdung and spent compost of mushroom recorded
lower spore counts (16.6 x 10 7 cfu and 10.7 x 10 7 cfug -1
respectively) compared to other substrates (Table 1). Rini, et
al., 2007 also concluded that sorghum grain with jaggery
served as nutritional supplements and enhanced the conidial
yield of Trichoderma viride. Sorghum grain was found as
superior grain substrate as it gave maximum population and
proved very useful and effective for mass multiplication.
ACKNOWLEDGEMENT
The authors are grateful for the financial support granted
by the ICAR, under Niche Area of Excellence of IPM
Programme running in Department of Plant Pathology, C.S.
Azad University of Agriculture & Technology, Kanpur
LITERATURE CITED
Chaudhari, P.J., Srivastava, P. and Khadse, A.C. 211. Substrate evaluation
for mass cultivation of Trichoderma viride. Asiatic J. of Biotech.
Res., 2 (04): 441-446.
Johnson, L.F. and curl, A.E., 1972. Method for the research on ecology
of soil borne plant pathogen. Burgess publishing company,
minnepolis, MN, pp. 247.
KAU. 2002. Package of Practices Recommendations; Crops. Twelth
edition. Directorate of Extension, Kerala Agricultural University,
Thrissur, 278p.
Prasad, R.D., Rangeshwaran R. and Sunanda, C.R. 2002. Jaggery- an
easily available alternative to molasses for mass production of
Trichoderma harziaum. Pl. Dis., Res., 17:363-365.
Rini, C.R. and Solochana, K.K. 2007. Substrate evaluation for
multiplication of Trichoderma spp. J. Trop. Agric., 45 (1-2): 58-
60.
Rini, C.R. and Sulochana, K.K. 2007. Usefulness of Trichoderma and
Pseudomonas against Rhizoctonia solani and Tusarium oxysporum
infecting tomato. J. Trop. Agric., 45: 21-28.
Saju, K.A., Anandaraj, M. and Sarma, Y.R. 2002. On farm production
of Trichoderma harzianum using organic matter. Indian Phytopath.,
55: 227-281.
Shahid, M., Singh, A., Srivastava, M., Mishra, R.P. and Biswas, S.K.,
2011 Effect of temperature, pH and media for growth and
sporulation of Trichoderma longibrachiatum and self life study in
carrier based formulations. Ann. Pl. Protec. Sci., 19(1): 147-149.
Singh, A., Shahid. M., Pandey, N.K., Kumar,S., Srivastava, M. and
Biswas S.K., 2011. Influence of temperature, pH and media for
growth and sporulation of Trichoderma atroviride and its shelf life
study in different carrier based formulations. J. Pl. Dis. Sci 6:32-
34.
Singh, V. and Joshi B.B. 2007. Mass multiplication of Trichoderma
harzianum on sugarcane press mud. Ind. Phytoath., 60: 530-531
(2007).
Tewari, A.K. and Mukhopadhyay, A.N. 2001. Testing of different
formulations of Gliocladium virens against chickpea wilt complex.
Indian Phytopath., 54: 67-71.
Zaidi, N. W. and Singh U.S. 2004. Use of farm yard manure for mass
multiplication and delivery of biocontrol agents. Trichoderma
harzianum and Pseudomonas fluorescens. Asian Agric. Hist., 52:
165-172.
Received on 10.3.2012 Accepted on 15.3.2012

Trends in Biosciences 5 (1): 71-73, 2012
Compatibility of Entomopathogenic Nematodes (Nematoda: Rhabditida) with
Pesticides and their Infectivity against Lepidopteran Insect Pest
RASHID PERVEZ* AND S.S. ALI
Indian Institute of Pulses Research, Kanpur, 208 024
*Present add.: Indian Institute of Spices Research, Calicut 673 012
email: rashid_pervez@rediffmail.com, ss_ali@rediffmail.com
ABSTRACT
Studies on the effect of aqueous suspension of the insecticides
(Endosulfan and Monocrotophos), fungicide (Mancozeb),
weedicide (Pendimenthilene) and botanical (Nemmarin) on
the activity of infective juveniles of Steinernema masoodi, S.
seemae, S. carpocapsae and S. mushtaqi and infectivity of IJs pre
exposed to these pesticides against Corcyra cephalonica larva
were carried out under laboratory condition. The aqueous
suspensions of these pesticides were prepared as per the field
recommended doses along with control (exposed to water).
Results show that, S. mushtaqi was found more compatible with
all tested pesticides followed by S. masoodi and S. seemae.
However, S. carpocapsae was found least compatible with tested
pesticides. Immobility of IJs increased when increase the
exposure period. Maximum number of inactive juveniles was
recorded after 72 h exposure. Among the tested pesticides,
Endosulfan is more affective on the activity of the juveniles
followed by Monocrotophos. While, Nemmarin showed less
effective on the activity of the IJs. The infectivity of 24 h
pesticides exposed IJs was not much affected against C.
cephalonica larva as compare to control. Among the tested
pesticide, all tested IJs of EPN shows less infectivity when
exposed with Endosulfan followed by Monocrotophos. However,
Pendimenthlin and Nemmarin exposed infective juveniles
found more pathogenic to rice moth larva.
Key words Entomopathogenic nematodes, pesticides, compatibility
Recently, entomopathogenic nematodes (EPNs) are
emerging as biological control agents of insect pests of crops
due to their wide host range, ease to handle, short life cycle
and environmental safety (Gaugler and Kaya, 1990, Ali, et al.,
2005a and 2008, Pervez, et al., 2007, Shapiro, et al., 2002).
It has been suggested that combining low-impact
insecticides or reduced rates of insecticides with biological
control could achieve adequate control while reducing the
adverse effects of insecticides (Forschler, et al., 1990; Alfred
and Grewal, 2004). Several studies have demonstrated additive
or synergistic relationships between the combined use of lowimpact
insecticides and biological control agents
(Koppenhofer and Kaya, 1998; Nishimatsu and Jackson, 1998).
Koppenhofer and Kaya, 1998 described a strong
synergistic effect on mortality of 2 scarab species,
Cyclocephala hirta LeConte and C. pasadenae Casey with
combinations of imidacloprid, another reduced risk insecticide
and entomopathogenic nematodes. Use of chemical pesticides
along with different species of EPNs in integrated pest
management (Hussaini, et al., 2001).
Present study carried out, compatibility of S. masoodi,
(Ali, et al., 2005b), S. seemae (Ali, et al., 2005b), S. carpocapsae
(Weiser, 1955) Wouts, et al., 1982 and S. mushtaqi Pervez, et
al., 2009 with insecticides (Endosulphan and
Monocrotophos), fungicide (Mancozeb), weedicide
(Pendimenthilene) and botanical (Nemmarin) has been
examined and effect of the exposed infective juveniles
infectivity against Corcyra cephalonica also tested.
MATERIALS AND METHODS
The infective juveniles of test nematodes were obtained
from nucleus culture of the nematodes maintained in the
Nematology laboratory, Indian Institute of Pulses Research,
Kanpur. All of these EPNs species were cultured on fully grown
Galleria mellonella larvae as per the procedure described by
Woodring and Kaya 1988. Emerged infective juveniles (IJs)
were surface sterilised in 0.01% Hyamine solution, stored in
distilled water in tissue culture flasks for study. While, larvae
of the rice moth, C. cephalonica were collected from the culture,
which maintained on sorghum grain.
Third stage juveniles of tested species of EPNs (50)
were inoculated into 5.5 cm petri plates containing 2 ml of
each concentrations of Endosulfan (1.6 ppm), Monocrotophos
(1.2 ppm), Mancozeb (2600 ppm), Pendimenthilene (0.75 ppm)
and Nemmarin (1.5 ppm) solution along with control (exposed
to distill water) separately and kept in BOD incubator at 28 ºC.
All treatments were replicated ten times. The immobility of
juveniles was observed after 24, 48 and 72 h exposure under a
stereoscopic microscope. Inactive juveniles, mobile responded
after the pricking the tail considered being immobile. The per
cent of immobility was calculated.
Infectivity of 24 h test pesticides exposed to IJs of test
species of EPNs against rice moth, Corcyra cephalonica was
kept in petriplates. For this, ten larva of tested insect was kept
in each petri plate and 500 infective juveniles (IJs) of each test
species of EPNs were inoculated and their mortality was
recorded after 72 h. Each species of EPN as well as pesticides
were tested separately and singly. The experiment was
conducted at 28 0 C in a BOD incubator and replicated five
times along with control. The per cent mortality was calculated

72 Trends in Biosciences 5 (1), 2012
according to following formula.
Mortality (%) = D x 100 / N
Where:
D- Number of dead larvae; N – Total number of larvae
RESULTS AND DISCUSSION
Results showed that, S. mushtaqi was found more
compatible with all test pesticides followed by S. masoodi
and S. seemae. However, S. carpocapsae was found least
compatible with test pesticides. Immobility of IJs increased
when increase the exposure period. Maximum number of
inactive juveniles of EPNs was recorded after 72 h exposure.
Among the test pesticides, Endosulfan is more effective on
the activity of the infective juveniles than to Monocrotophos.
While, Nemmarin shows very little effect on the activity of
the IJs or recoded on par as similar to control (Fig. 1).
Fig. 1.
The infectivity of 24 h exposed pesticides IJs was not
Inactivity of the infec tive juveniles o f the EP Ns
exposure to pesticides in aqueous solutions.
much affected against C. cephalonica larva as compared to
control. Among the test pesticides all tested IJs of EPN shows
less infectivity when exposed with Endosulfan followed by
Monocrotophos. However, Pendimenthlin and Nemmarin
exposed infective juveniles found more pathogenic to rice
moth larva (Fig. 2).
Rovesti and Deseo, 1990 reported that
organophosphorus compounds and Endosulfan caused a
reduction in the movement of the infective juveniles of S.
carpocapsae and S. feltiae but infectivity of them was
negligible. However, earlier studies have shown that some
pesticides can reduce entomopathogenic nematode viability
and infectivity (Zhang, et al ., 1994; Krishnayya and Grewal
2002). Chemicals insecticides and EPN offer different but
potential compatible approaches to suppress insect
populations (Nishimatsu and Jackson 1998).
EPNs are often applied in combination with pesticides,
Fig. 2.
Infectivity of pesticides exposed infective juveniles of
EPNs against C. cephalonica
soil amendments and fertilizers. IJs of EPN have been found
to be tolerant to short exposures (2 to/6 h) of most acaricides,
fungicides, herbicides, and insecticides (Rovesti, et al ., 1988;
Rovesti and Deseo 1990) and can therefore be applied
simultaneously with many pesticides. Moreover,
entomopathogenic nematode species differ in their
susceptibility and sensitivity to different formulations of the
same chemical pesticide (Grewal 2002).
This study revealed that, few pesticides had adverse
effect on the activity of infective juveniles, however, the other
were not affected and are hence compatible. The incompatible
species can be improved by exposing IJs to lower
concentration of pesticides and increasing the concentration
for exposure of subsequent generations of tolerated species.
Further study on the residual effects of the pesticides in the
field, which directly affects the EPNs populations are to be
explored.
ACKNOWLEDGEMENT
The authors express their gratitude to Director, Indian
Institute of Pulses Research, Kanpur, for providing all the
facilities for this study. First author also thankful to Department
of Science and Technology (DST), Ministry of Science &
Technology, Government of India, New Delhi, for providing
financial support.
LITERATURE CITED
Alfred, A. and Grewal, P. S. 2004. Tank mix compatibility of the
entomopathogenic nematodes, Heterorhabditids bacteriophora and
Steinernema carpocapsae, with selected chemicals pesticides used
in Turfgrass. Biocontrol Science and Technology, 14 (7): 725-730.
Ali, S. S., Ahmad, R., Hussain, M. A. and Pervez, R., 2005a. Pest
management of pulses through entomopathogenic nematodes.
Indian Institute of Pulses Research, Kanpur, Army press, Lucknow
(India), pp. 59.
Ali, S. S., Pervez, R., Hussain, M.A. and Ahmad, R., 2008. Susceptibility
of three lepidopteran pest to five entomopathogenic nematodes
and in vivo mass production of these nematodes. Archieves of

PERVEZ AND ALI, Compatibility of Entomopathogenic Nematodes (Nematoda: Rhabditida) with Pesticides 73
Phytopathology and Plant Protection, 41 (4): 300 – 304.
Ali, S. S., Shaheen, A., Pervez, R. and Hussain, M.A. 2005b. Steinernema
masoodi sp. n. and Steinernema seemae sp. n. (Rhabditida:
Steinernematidae) from Uttar Pradesh, India. International Journal
of Nematology, 15 (1) : 89–99.
Forschler, B. T., All, J. N. and Gardner, W. A. 1990. Steinernema feltiae
activity and infectivity in response to herbicide in aqous and soil
environment. J. Inv. Pathology, 55: 375-379.
Gaugler, R. and Kaya, H. K. 1990. Entomopathogenic nematodes in
Biological control. CRC Press, Boca Raton, Florida, pp. 365.
Grewal, P.S. 2002. Formulation and Application technology, In :
Entomopathogenic Nematology (ed: Gaugler, R.). CABI Publishing.
CAB International, Wallingford. pp. 311-/332.
Hussaini, S.S., Singh, S. P. and Shakeela, V. 2001. Compatibility of
entomopathogenic nematodes (Steinernematidae,
Heterorhabditidae : Rhabditida) with selected pesticides and their
influence on some biological traits. Entomon, 26 (1): 37- 44.
Koppenhofer, A. M., and H. K. Kaya. 1998. Synergism of imidacloprid
and an entomopathogenic nematode: a novel approach to white
grub (Coleoptera: Scarabaeidae) control in turfgrass. J. Econ.
Entomol, 91: 618-623.
Krishnayya, P.V. and Grewal, P.S. 2002. Effect of neem and selected
fungicides on viability and virulence of the entomopathogenic
nematode Steinernema feltiae. Biocontrol Science and Technology,
12: 259-/266.
Nishimatsu, T., and Jackson, J. J. 1998. Interaction of insecticides,
entomopathogenic nematodes, and larvae of the western corn
rootworm (Coleoptera: Chrysomelidae). J. Econ. Entomol, 91:
410- 418.
Pervez, R., Ali, S. S. and Ahmad, R. 2007. Efficacy of some
entomopathogenic nematodes against mustard saw fly and in vivo
production of these nematodes. International Journal of
Nematology., 17 (1): 55-58.
Pervez, R., Ali, S. S. and Asif, M. 2009. A new species of
entomopathogenic nematodes Steinernema mushtaqi sp. n.
(Nematoda : Rhabditidae : Steinernematidae) from chickpea
rhizosphere. In: International Conference on Legumes (ICGL), at
Indian Institute of Pulses Research, Kanpur, Feb., pp. 14-16.
Rovesti, I., Heinzpeter, E.W., Tagliente, F. and Deseo, K.V. 1988.
Compatibility of pesticides with the entomopathogenic nematode
Heterorhabditis bacteriophora Poinar (Nematoda:
Heterorhabditidae). Nematology, 34: 462- 476.
Rovesti, L. and K. V. Deseo. 1990. Compatibility of chemical pesticides
with the entomopathogenic nematodes, Steinernema carpocapsae
Weiser and S. feltiae Filipjev (Nematoda: Steinernematidae).
Nematology, 36: 237-245.
Shapiro, D. I., Mizell, R. F. III., Cambell, J. F., 2002. Susceptibility of
the plum curculio, Conotrachelus nenuphar, to entomopathogenic
nematodes. Journal of Nematology, 34 (3) : 246-249.
Weiser, J. 1955. Neoaplectana carpocapsae n. sp. (Anguillulata,
Steinernematidae) novy cizopasnic housenik obalece jablecneho,
Carpocapsa pomonella L. Vestnik Cesk. Zool. Spolecnosti, 19:
44-52.
Woodring, J. L. and Kaya, H. K. 1988. Steinernematid and
heterorhabditid nematodes: In: a handbook of biology and
techniques. Southern Cooperative Series Bulletin 331, Arkansas
Agricultural Experiment Station, Arkansas, Fayetteville, pp 28.
Wouts, W. M., Mracek, Z., Gerdin, S. and Bedding, R. A. 1982.
Neoaplectana Steiner, 1929 a junior synonym of Steinernema
Travassos, 1927 (Nematoda: Rhabditida). Systematic Parasitology,
4 (2): 147-154.
Zhang, I., Shonu, T., Yamanaka, S. and Tanabe, H. 1994. Effects of
insecticides on the entomopathogenic nematode Steinernema
carpocapsae. Applied Entomology and Zoology, 29 (4): 539-/
547.
Recieved on 5-12-2011 Accepted on 5-3-2012

Trends in Biosciences 5 (1): 74-76, 2012
Prevalence of Grain Moulds in Sorghum Growing Areas of Northern Karnataka
Y.S. MAHESH 1 , S. LINGARAJU 2 , B. RAVI KUMAR 1 , B.MANJUNATH 1 AND M.G. PALAKSHAPPA 2
1
Department of Plant pathology, University of Agricultural Sciences, Bangalore-560 065, Karnataka
2
Department of Plant pathology, University of Agricultural Sciences, Dharwad-580 005, Karnataka
e-mail: maheshsgowda@gmail.com
ABSTRACT
Sorghum is known to suffer from various diseases in the field.
Among them Grain mold is a major disease wherever sorghum
is grown if moist weather conditions prevail after flowering
until grain maturity and before harvest. Survey revealed the
presence of disease in all three districts viz., Belgaum, Dharwad
and Haveri. The per cent disease index ranged from 61.25 to
77.25. Per cent disease index was high in Belgaum district
followed by Haveri and Dharwad district. Among different
cultivars under cultivation in these districts, Maldandi
(M 35-1) was more susceptible with per cent disease index of
77.25 followed by CSH 5 which recorded a per cent disease
index of 77.0.
Key words
Sorghum, grain mould, survey, north Karnataka
Sorghum (Sorghum bicolor (L.) ranks fourth among the
world’s cereals after wheat, maize and rice. Karnataka state
accounts for 1790 thousand hectares area with the production
of 1440 thousand tonnes of grain. The cultivation of sorghum
is concentrated more in northern districts of Karnataka viz.,
Bijapur, Dharwad, Belgaum, Raichur, Gulbarga and Bellary.
Globally grain mould causes an estimated loss of $130 million
annually (Anon., 1992). More than forty fungal genera are
associated with moulded grains and most of them are
facultative parasites or saprophytes, only a few fungi infect
sorghum flower tissue during early stages of grain
development. These include Fusarium moniliforme,
Curvularia lunata, Alternaria alternata, Phoma sorghina
and Exserohilum sp. Grain moulds have become increasingly
important particularly after the introduction of early maturing
hybrids and varieties. In kharif, the maturity of grains
invariably coincides with the torrential late rains seen in
September and October months in states of Karnataka, Andhra
Pradesh, Maharashtra and Tamil Nadu, resulting in high grain
moulds incidence. Therefore the present study was undertaken
to assess the disease incidence in northern Karnataka.
MATERIALS AND METHODS
An intensive roving survey for grain mould disease was
conducted during kharif 2003 to know the disease incidence
in sorghum growing areas of Dharwad, Belgaum and Haveri
districts of north Karnataka. In each district, three taluks were
selected. In each field 10 plants were randomly selected and
grain mould severity was recorded by following 0-5 scale.
Then the per cent disease index (PDI) was calculated by using
the formula given by Wheeler, 1969.
Sum of individual disease 100
ratings
Per cent disease index= —————————— × —————
Number of samples Maximum
disease grade
RESULTS AND DISCUSSION
A roving survey to know the incidence of sorghum grain
moulds was carried out in three districts viz., Belgaum, Dharwad
and Haveri. A total of 45 villages, 15 from each district were
surveyed, data obtained are presented in Table 1.
Incidence of grain moulds of sorghum was noticed in all
the places surveyed on two commonly grown sorghum
cultivars CSH 5 and Maldandi. In Belgaum district, the per
cent disease index (PDI) ranged from 64.00 to 77.25, the highest
PDI of 77.25 was recorded in Hulikatti village of Saundatti
taluk followed by 77.0 in Yakkundi and 76.00 in Hanchinal
village of same taluk, which is higher than district average
(71.38%). The lowest PDI of 64.00 was recorded in agarage
village of Belgaum taluk followed by 65.20 in Peeranawadi of
Belgaum taluk and Hosur village of Saundatti taluk.
In Dharwad district, highest per cent disease index of
73.76 was recorded in Dastikoppa village of Kalaghatagi taluk
followed by 72.27 in Hirchanalli village of same taluk and
72.25 in Kundagola of Hubli taluk. The lowest PDI of 61.25
was recorded in Haralikatte village of Dharwad taluk. Overall
among three taluks, highest PDI was recorded in Kalaghatagi
taluk with 68.86 which is higher than district average of 66.80%.
In Haveri district, the PDI ranged from 62.00 to 73.40.
Highest percent disease index was recorded in Bankapur
village with 73.40 of Shiggaon taluk followed by 73.25 in
Shiggaon village of same taluk and 72.25 in Negalur village of
Haveri taluk, which is higher than the district average (68.74).
Lowest PDI of 62.00 was recorded in Hanumanatti village of
Haveri taluk. Among three districts, highest disease index
was recorded in Belgaum district with 71.38% followed by
68.74% in Haveri district and lowest disease index of 66.80%
was recorded in Dharwad district.
Variation in disease incidence was observed variety wise
and location wise. Among different cultivars grown in three
districts, variety Maldandi (M 35-1) was the most popular
one, which was seen in 29 fields followed by CSH 5 in 16
fields. PDI was more on Maldandi (77.25) in Saundatti taluk
but it was very low in Dharwad taluk (61.25). However CSH 5
recorded a high disease index of 77% in Saundatti taluk and a

MAHESH et al., Prevalence of Grain Moulds in Sorghum growing areas of Northern Karnataka 75
Table 1.
Survey for grain mould incidence in different districts of north Karnataka
District Taluk Village Variety/hybrid PDI
Belgaum
Belgaum Agarage CSH 5 64.00
Hirebagewadi CSH 5 69.00
Japarwade Maldandi 75.00
Peeranawadi CSH 5 65.20
Kaduli Maldandi 68.27
Mean 68.29
Saundatti Hanchinal Maldandi 76.00
Hosur Maldandi 65.20
Munavalli CSH 5 71.20
Hulikatli Maldandi 77.25
Yakkundi CSH 5 77.00
Mean 73.29
Bailhongal Sangolli Maldandi 72.50
Nesaragi Maldandi 75.20
Halaki Maldandi 72.70
Tigadi CSH 5 73.00
Kitlur CSH 5 70.00
Mean 72.58
District average 71.38
Dharwad
Dharwad Narendra Maldandi 68.00
Chabbi Maldandi 64.00
Haralikatle Maldandi 61.25
Navalur Maldandi 62.25
Yarikoppa Maldandi 71.25
Mean 65.35
Hubli Shiraguppi Maldandi 68.00
Ingalahalli CSH 5 64.26
Kundagola CSH 5 72.25
Navanagar Maldandi 63.27
Noolvi Maldandi 64.26
Mean 66.40
District
Taluk Village Variety/hybrid POI
Kalaghatagi Haliyala CSH 5 64.00
Kalaghatagi CSH 5 62.25
Nirsagar Maldandi 71.05
Hirchanalli Maldandi 72.27

76 Trends in Biosciences 5 (1), 2012
Haveri
Dastikoppa Maldandi 73.76
Mean 68.86
District average 66.80
Haveri Hanumanatti CSH 5 62.00
Sangur Maldandi 68.00
Bilavagi Maldandi 72.00
Negalur Maldandi 72.25
Devihosur CSH 5 65.78
Mean 68.00
Shiggaon Shiggaon CSH 5 73.25
Bankapur CSH 5 73.40
Hirebendigeri Maldandi 63.52
Dhundashi Maldandi 72.00
Konankeri Maldandi 72.00
Mean 70.83
Ranebennur Chalageri Maldandi 63.00
Honnatti CSH 5 68.70
Ranebennur Maldandi 65.25
Sunkabidari Maldandi 70.25
Halageri Maldandi 69.75
Mean 67.39
District average 68.74
low of 62.25% in Kalaghatagi taluk.
Hiremath and Palakshappa 1992 also found severe grain
mould incidence on all genotypes of sorghum in Karnataka.
Similarly, Padaganur and Palakshappa, 1998 also reported
severe incidence of grain moulds in transitional zone of north
Karnataka.
The severity of the disease may be due to prolonged
rainy season or coincidence of continuous rains and high
degree of relative humidity during grain formation stage (From
September to first week of October).
LITERATURE CITED
Anonymous. 1992. Medium Term Plan 1994-98. Research theme
datasets Volume 3, International Crops Research Institute for Semi
Arid Tropics, Patancheru, Andhra Pradesh, India, pp.229.
Hiremath, R.V. and Palakshappa, M.G. 1992 Annual Report, All India
Coordinated Research Project on Sorghum Improvement, pp.1-3.
Padaganur, G.M. and Palakshappa, M.G., 1998, Survey of diseases of
sorghum in North Karnataka. Annual Report on Sorghum, All India
Coordinated Sorghum Improvement Project, University of
Agricultural Sciences, Dharwad, pp.106-108.
Wheeler, B.E.J. 1969. An Introduction to Plant Disease, John Willey
and Sons Limited, London, pp.301.
Received on 4-7-2011 Accepted on 12-12-2011

Trends in Biosciences 5 (1): 77-78, 2012
SHORT COMMUNICATION
Genetic Diversity in Bread Wheat [Triticum aestivum (L.) Em. Thell]
HASAN TANVEER * , ARVIND KUMAR AND HAMVEER SINGH *
Krishi Vigyan Kendra, Ruastamnagar, Bilari, Moradabad 202 415, Uttar Pradesh
e-mail: htdania@yahoo.com
Wheat is a major edible cereal crop of India. There is
need to develop better genotypes suitable for higher yield
with consumer acceptability. Thus there is an urgent need to
improve grain yield productivity. In the present study genetic
divergence (Mahalanobis, 1936; Rao, 1952 ) among available
genotypes of wheat were investigated through D 2 statistics.
The materials of present study comprised improved varieties
of wheat. The genotypes were grown during rabi, 2008-2009
in randomized block design with three replications at Krishi
Vigyan Kendra, Bilari, Moradabad. The plot size 3.0 x 2.5 m
comprised 11 rows and the sowing was done in rows 25 cm
spacing. Observations were recorded on eight characters
including yield.
Data on yield components viz., plant height, number of
tillers per plant, flag leaf area (Lazerao, 1965), spike length,
days to maturity, number of grains per spike, 1000-grain weight
were recorded on five randomly selected plants. Analysis of
variance was performed on mean data. The genotypes were
grouped into three different clusters based on D 2 values. The
Table 1.
Sl.
No.
Variation of different morpho-physiological, yield
and yield component traits in bread wheat.
Characters Range Mean S.E. (m)
1 Plant height (cm) 77.00-106.33 89.37 0.99
2 Number of tillers per
plant
9.33-13.00 11.00 0.84
3 Flag leaf area (cm 2 ) (3) 16.55-35.48 21.62 0.51
4 Spike length (cm) 9.33-13.00 10.87 0.70
5 Days to maturity 90.00-131.00 115.96 3.21
6 Number of grains per
spike
45.00-66.00 55.89 1.72
7 1000-grain weight (g) 38.33-47.00 42.00 0.94
8 Grain yield (q/ha) 38.07-60.45 45.59 1.16
Table 2.
Average intra and inter cluster D 2 values in bread
wheat
Clusters I II III
I 2.097 3.40 3.59
II 1.84 2.80
III 1.26
correlation among different traits were performed as per Dewey
and Lu, 1959.
The average performance of bread wheat cultivars in
respect of different characters, yield and yield components is
presented in Table 1. Highly significant differences (P=0.01)
for all traits were recorded, indicating substantial divergence
in the collection. The mean grain yield of trial was 45.59 q ha -
1
with the range of 38.07 to 60.45 q ha -1 . The flag leaf area (cm 2 )
mean was 21.62 with the range of 16.55 to 35.48 cm 2 .
Number of grains per spike ranges between 45.00 to
66.00 with average of 55.89. While range between 90 to 131
days for days to maturity with the mean of 115.89.
Nine genotypes could be grouped in three clusters
based on D 2 values and their intra and inter cluster distance
are presented in Table 2. Inter cluster D 2 is a measure of genetic
distance between the two clusters and was observed to be
the highest for cluster number (I) and cluster number (III) and
the lowest between cluster number (II) and cluster number
(III).
Three genotypes were clustered in cluster I, four
genotypes in cluster II while two genotypes in cluster III. The
genotype K 9423, K 9644 and K 7903 (all three from C.S.A.U.A
& T, Kanpur) were found together in cluster I. It shows that
these genotypes are linked together in respect of their
development origin. The four genotypes, K 9351, K 0307,
DBW 16 and PBW 502 were in cluster II suggests that the
geographical isolation was not the only factor causing genetic
diversity. Same criteria were also observed in cluster III
because genotype K 9107 was developed from C.S.A.U.A &
T, Kanpur and variety PBW 373 from P.A.U., Ludhiana.
The wheat breeding for higher grain yield with duration
is important, variety K 7903 represented by cluster I embodies
the genes for short duration as identified by different
physiological indices, however, it is poor in yield. On contrary,
genotype K 0307 in cluster II embodies high grain yield
potential. K 7903 and K 0307 can be undertaken for further
hybridization programme to select short duration and high
yielding segregants.
Correlation analysis (Table 3) revealed that grain yield
positively and highly significant genotypic correlation with
number of tillers per plant. It is considered that selection for
number of tillers per plant can improve grain yield while
selecting plants in the segregating generations.

78 Trends in Biosciences 5 (1), 2012
Table 3. Correlation among different traits in bread wheat
p
g
Plant height
(cm)
Number of
tillers per
plant
Flag leaf area
(cm 2 ) (3)
Spike length
(cm)
Days to
maturity
Number of
grains per
spike
1000-
Grain
weight (g)
Grain
yield
(q/ha)
Plant height (cm) 1.00 0.403 -0.028 0.535 0.581 -0.144 0.072 0.625
Number of tillers per plant 0.525 1.00 -0.307 0.235 0.614 -0.235 -0.031 0.674 *
Flag leaf area (cm 2 ) -0.037 -0.480 1.00 -0.014 -0.393 0.205 -0.148 -0.149
Spike length (cm) 0.668 * 0.015 -0.047 1.00 0.380 0.049 0.200 0.213
Days to maturity 0.612 0.910 ** -0.414 0.526 1.00 -0.267 0.448 0.731
Number of grains per spike -0.138 -0.298 0.225 0.172 -0.285 1.00 0.231 -0.407
1000-grain weight (g) 0.095 -0.012 -0.143 0.259 0.524 0.283 1.00 0.097
Grain yield (q/ha) 0.649 * 0.930 ** -0.152 0.223 0.755 * -0.405 0.109 1.00
LITERATURE CITED
Dewey, D. R. and Lu K. H. 1959. A correlation and path coefficient
analysis of components of crested wheat grass seed population.
Agron. J., 51: 515-518.
Lazerao, R. 1965. Coefficient for determining the leaf area in certain
agricultural crops. Rast. Nauki, 2: 27-37.
Mahalanobis, P.C. 1936. On the generalized distance in statistics. Proc.
Nat. Inst. Sci., 2: 49-55.
Rao, C.R. 1952. Advanced Statistical Methods in Biometrical Research,
ed. I, John Willey and Sons, New York.
Recieved on 12-11-2011 Accepted on 15-2-2012

Trends in Biosciences 5 (1): 79-80, 2012
SHORT COMMUNICATION
Relation of Temperature and Humidity to Postharvest Rot of Chickpea Caused by
Fungi
SARITA JOSHI AND KAMLESH GUPTA
Botany Department, Dayanand Girls P. G. College Kanpur
email : jhimanshu1983@gmail.com
Effect of temperature and humidity on the decay of
mango, banana, guava, litchi, grapes and certain other fruits
has been investigated by several workers (Bhargava, et al.,
1965; Tandon and Singh, 1969; Bilgrami, 1973; Grewal, 1988;
Krishnaiah, 1989 and Haware, 1998). However no significant
information appears to to be available about the effect of
temperature and humidity on the decay of chickpea (Cicer
arietinum). The present work attempts to investigate the
influence of above said factors on their post harvest fruit rot
disease.
For determining the effect of different temperature,
healthy mature seeds of chickpea were obtained from Indian
Institute of Pulse Research Kanpur. These seeds were surface
sterilized with 0.1% Sodium hypochlorite solution. Seeds were
inoculated with spore suspension of different fungi by pinprick
injury method of inoculation. Inoculated seeds were incubated
at different temperatures up to 12 days in open sterile
polythene bags.
Different levels of relative humidities were maintained
in sterilized desicators according to method of Buxton and
Mellanby, 1934. The percentage of rot was calculated by the
formula:
W - w
Percentage rot = ——— * 100
W
Where W = Weight of the fruit before infection, and
w = Weight of the fruits after removal of the infected
tissue.
Low temperature showed poor fungal advancement in
seeds tissue, but this temperature increased the susceptibility
of seeds. The fungal infection was more between 20 0 -30 0 C
(Table 1) showing severe rotting.
The disease was most severe when the RH was between
90-100% at decreased humidity the severity of rot also
decreased. Probably this is the reason that these seeds get
spoiled more during wet season when humidity ranges
between 90-100%. It has been generally observed that high
humidity is most favourable for spore germination and
penetration in the seeds tissues (Table 2).
The rotting of chickpea by Ascochyta rabiei,
Phytopthora megasperma and Sclerotium rolfsii was high
when they were incubated at 25 0 C and 30 0 C. Phoma glomerata
showed slower rate of rotting at different temperatures. It
appears from the results of the present study that temperature
has a pronounced effect on the rate of development of decay
Table 1.
Percentage rot of chickpea seeds inoculated with
respective pathogens during storage at different
temperature
Pathogen
Days of Temperature in 0 C
Incubation 15 25 30
Phoma glomerata 3 - 4.5 6.3
6 - 4.8 6.5
9 2.5 5.0 7.5
12 3.8 5.5 8.0
Ascochyta rabiei 3 1.5 4.5 5.7
6 2.7 6.8 7.8
9 3.0 7.6 8.9
12 3.5 9.5 10.5
Phytopthora megasperma 3 1.0 4.0 5.5
6 2.8 5.5 7.8
9 3.6 5.9 9.7
12 4.1 7.8 11.5
Sclerotium rolfsii 3 2.3 3.0 6.5
6 3.0 4.1 7.9
9 3.5 5.6 8.7
12 4.1 7.9 12.6
Table 2.
Percentage rot of chickpea seeds incubated with
respective pathogens during storage at different
relative humidities
Pathogen
Days of Relative Humidity in %
Incubation 30 50 70 90 100
Phoma glomerata 3 2.5 3.1 10.5 14.1 16.8
6 3.1 3.9 11.2 15.8 17.9
9 3.9 4.2 11.7 17.9 21.3
12 4.2 4.6 12.2 20.1 24.6
Ascochyta rabiei 3 2.2 3.4 9.5 15.1 18.3
6 3.4 4.5 10.6 17.5 19.9
9 3.6 5.2 11.0 18.9 22.0
12 4.1 5.5 12.1 20.9 24.6
Phytopthora megasperma 3 2.5 3.5 7.9 14.4 18.3
6 3.1 4.1 8.3 16.8 22.4
9 3.7 5.2 9.9 19.7 24.1
12 4.6 6.3 11.1 21.6 26.7
Sclerotium rolfsii 3 3.1 3.4 7.5 17.1 20.1
6 3.6 3.9 8.2 19.2 21.7
9 4.3 4.8 9.5 20.7 24.1
12 5.1 5.3 11.4 22.0 26.3

80 Trends in Biosciences 5 (1), 2012
and consequently on the magnitude of the loss due to fungal
infection.
Relative humidity appears to have pronounced influence
on the development of fungal disease as fairy infection was
noticed at higher levels of humidity. In general it can be said
that higher the humidity there is high percentage of rotting
and at lower humidity rotting was poor.
LITERATURE CITED
Bhargava, S.N., Ghosh, A.K., Srivastava, M.P., Singh, R.H. and Tandon,
R.N. 1965. Studies on fungal disease of tropical fruits VII. Effect of
temperature on the decay on mango, banana, guava caused by some
important pathogens. Proc. Natl. Acad. Sci. India, 35 :393-398.
Buxton, P.A., Mellanby, K. 1934. The measurment and control of
humidity. Bull. Entam. Res. 25:171-175.
Grewal, J.S. 1988. Disease of pulse crops: An overview. Phytopath. 41:
1-14.
Haware, M.P. 1998. Disease of chickpea. The pathology of food and
Pasture Legumes (eds. D.J. Allen and J.M.Lenne) CAB International
ICARDA Wallingford U.K. pp.473-506.
Krishnaiah, J. and Thirupathaiah, V. 1989. Environmental factors in
relation to post harvest rot of grapes caused by fungi. Res. J. Pl.
Environ., 5(1): 59-62.
Prasad, S.S. and Bilgrami, R.S. 1973. Investigation on disease of Litchi
II: influence of temperature and humidity on the decay of fruits
caused by nine virulent pathogens. Indian Phytopath, 26:517-522.
Tandon, I.N. and Singh, B.B. 1969. Studies on anthracnose of guava
and its control. Indian Phytopath, 22:322-326.
Recieved on 12.1.2012 Accepted on 3.3.2012
1. ERRATUM
Research paper Sida cordifolia: Morphological Characterization of Plants Growth in a Variant Habitat of Kanpur Nagar
ALPANA TEWARI* AND NAINA SRIVASTAVA published in Trends in Biosciences 4(2): 233-234, 2011 should be read as
Sida cordifolia: Morphological Characterization of Plants Growth in a Variant Habitat of Kanpur Nagar
ARCHANA SRIVASTAVA, ALPANA TEWARI* AND NAINA SRIVASTAVA, Trends in Biosciences 4(2): 233-234, 2011.
2. ERRATUM
Research paper Efficacy of Herbal Flavoured Sterilized Milk-A Dairy Based Nutraceutical RITU P. DUBEY, POOJA
KUMARI AND MEENU SINGH published in Trends in Biosciences 4(2): 198-200, 2011 should be read as Efficacy of Herbal
Flavoured Sterilized Milk-A Dairy Based Nutraceutical RITU P. DUBEY, MEENU SINGH AND POOJA KUMARI, Trends
in Biosciences 4(2): 198-200, 2011.

Trends in Biosciences 5 (1): 81-83, 2012
Author Index
(Vol. 4, No. 1&2, 2011)
A
Ahmad Javed 169
Ahmad Tabrez 180
Ahmed Mohammed Osman 138
Ali, S.S. 31,82,98,103,123,140
Ashok Rathore 1
Asif Mohammad 35,82,98, 140
Attri Rajni 219
Awasthi Nimisha 230,235
Azad, B.S. 75
Azra Shaheen 98
Ahmad Javed 169
Ahmad Tabrez 180
Attri Rajni 219
B
Basharat, N. 95
Bhagat, I. 56
Bhaskar, R.S. 79
Burungale,S.V. 120
C
Channa Ashok 19
Chatterjee Sayan 157
Chauhan, R.M. 120
Chauhan, S.K. 68
Chavan, Sagar 86
D
Dar, S.A. 44
Devasahayam Mercy 88,175
Dubey Deepak Kumar 38,210
Dubey P. Ritu 198,208, 228
Dubey Rajni 75
Dubey Ritu Prakash 182
Dwivedi Pratibha 77
E
Elangovan Vadamalai 8, 53
G
Gami, R.A. 120
George Rachel 230
Gupta Astha 63
H
Haider Jamal 175
I
Idowu,A.A. 126
Irulan, A. 8
J
Jakhmola, S.S. 25
Johnson William 130
K
Kamaluddin 172
Katna Sapna 51
Kaur Harpreet 219
Khan Khalil 230,235
Khan Khalil 109
Koli,N.R. 224
Kumar Anil 41, 106
Kumar Ashok 38, 210
Kumar Rajesh 53,134
Kumari Pooja 182,198
L
Lodam, V.A. 91
Lok Nath 12
M
Makhdoomi, M.I. 44
Malik Mohd. Nadim 169
Mall, T.P. 47, 58, 128, 132,185
Mandhan Rishi Pal 161

82 Trends in Biosciences 5 (1), 2012
Manjunath, B. 31,71
Marimuthu, Y. S. 8
Masih, Sam A 175
Mastan, S.A. 138
Mecarty Samuel D 1, 88
Meena Girraj Singh 77
Mercy Devasahayam 1
Mir Imtiyaz Hussain 19
Misra Atul Kumar 61
Mohan Jitendra 118
Mohan, J. 101
Mohan, N. 101
Munjal Neera 157
Murugesh, K.A. 79
N
Nabi Sumaira 19
Nagaraja, T.G. 86, 112
Nagpal Neeraj 157
Naik,B. Gangadhara 71
Narayan Shyam 101
Narayanaprasad, G.B. 114
Nargund, V.B. 31
Nathan, P. T. 8
O
Osunlola, O.S. 126
P
Pandey Alok 75
Pandey, H.P. 68
PandeyAvinash 95
Pankaj, T. 146
Parameswaran, P. 130
Patel, I.S. 116,136
Patel, J.K. 116, 136
Patel, P.S. 116, 136
Patel, P.T. 120
Patel, S.A. 116, 136
Patil Shridhar 23
Patil, P.P. 91
Patil, S.R. 91
Paul Newton 180
Pervez Rashid 103
Pooja Kumari 228
Prabhu,Aditi M. 194
Prakash Chandra 224
Prasad Keshav 41
Prasad, C.S. 12
Prasad, G.K. 114
Prasad, H. N. 5
Prasad, P.S. 71
Pravin Kumar Singh 5
Priya, G. 8
R
Rajnish Kumar 75
Rana Kiran 51
Rana, B.S. 51
Randhawa, H.S. 56
Rani Sandhya 161
Rathore Ashok 88
Raut,S.V. 194
Raza, A.K. 68
Razvi, S.M. 172
S
Sachan, B.S. 41,106
Sachan,C.P. 205
Sagar Alka 66
Sahui Anjali 235
Saifulla Muhammad 71
Samundeeswari, S. 130
Shaheen Azra 82, 123
Shahid Mohammad 21, 205
Shakya, Narendra B. 101
Shalini Mishra 175

Author Index (Vol. 4, 1&2, 2011) 83
Shankar, P. 140
Sharma,A.K. 180
Sharma Parvati 215
Sharma Sanjay 148
Sharma, H.K. 51
Sharma, M.K. 66
Sharma, Suresh K. 142
Shivakumar, P.S. 114
Shivran,R.K. 224
Shukla, P.K. 21
Shukla,C.P. 189
Siddiqui, A.U. 148
Sihag, R.C. 215
Simon Sobita 95
Singh Ajai 235
Singh Anuradha 21, 205
Singh Archana 118
Singh Bijendra 109
Singh Dileep Kumar 180
Singh Meenu 198
Singh Neeraj 230
Singh Pradyumn 25
Singh, A. K. 63
Singh, D.P. 47, 58, 132
Singh, S.K. 75
Sirohi Anita 146
Sirvastava, S.P. 140
Sofi, P.A. 172
Srivastava Amit 134
Srivastava Archana 201
Srivastava Mukesh 205
Srivastava Naina 233
Srivastava, S.P. 75
Sudarshan 114
Sudarshan, G.K. 31
Sultan, M. Sarwat 142
Sunanda, B.S. 148
Sundaresha 114
T
Taiwo, B.R. Aminu 126
Tewari Alpana 201, 233
Tewari Seema 165
Tewari Vijay 118
Thakor, D.M. 120
Tiwari Sugandha 222
Tiwari, G.N. 12
Tripathi Madhu 165
Trivedi Parul (Mishra) 232
U
Umer Shahid 169
V
Varpe, P.G. 91
Vashi, R.D. 91
Vyas Priti 23
Y
Yadav Swapanil 66

Trends in Biosciences 5 (1): 84-87, 2012
Subject Index
(Vol. 4, No. 1&2, 2011)
A
Acute exposure 165
Agriculture 5
Algae 118
Alizarin reds 19
Allelopathy 68
Alternaria alternate 112
Alternaria 47,58
Anti fungal 112
Antimicrobial activity 23
Aphid 56,116
Arthropod diversity 12
Aspergillus niger 112
Azolobaetor 109,169
B
Bacillus sp. 8
Basic chromium sulphate 38
Bioagents 116
Biocontrol 71
Biodiversity 189
Biofertilizer 101,194
Biomarkers 210
Biomarkers 38
Biomethanation 66
Bioremediation 175
Blorilized milk 198
Botanicals 25, 79
Bread wheat 120
Brinjal 12
Bruchid 25
Bt 35
Bumble bee 51
C
Calcium 19
Callosobruchus maculates 25
Callus 21
Catfish 165
Cereals 68
Channa punctatus 38,210
Character association 172
Characterization 219
Chickpea 35,82,205
Chitin 8
Chlorophylls 86
Ciliated protozoa 180
Citrobacter sp. 8
Cladocerans 180
Cladosporum 112
Co-efficient of variation 44
Colony forming unit 71
Colour 157
Columnar epithelial cells 19
Combining ability 120
Common bean 44, 172
Composts 66
Conopid fly 51
Coturnix coromandelica 61
Coulombic efficiency 175
Cross bred cattle 1
D
Daphnia 180
Diabetes 161
Diallel analysis 120
Disease index 31
Diseases 215
E
Eco-races 53
Ecosystem 82

Subject Index (Vol. 4, No. 1&2, 2011) 85
Efficiency 224
Electronic resources 5
Employment 106
Enterobacter sp. 8
Entomopathogenic nematodes 35, 103, 123
Enzymatic activity 95
Epizootical ulcerative syndrome 215
Eri silkworm 53
Essential oils 23
Exorista bombycis 79
Explant 21
F
Fecundity 75
Fennel 116
Fertizer 41
Fish gills 165
Fish 215
Flavor 198
Fluoride toxicity 165
Foliar fungi 58
Foliicolous fungi 47, 185
Frozen yoghurt 182
Fructated DNA 161
Fungicides 71
Fusarium oxysporum 112
G
Gamma rays 77
Ganga 118
Genetic divergence 63
Genetic diversity 63
Genetic grades 1
Genotypes 114
Germination 205
Grain damage 75
Gram-negative 23
H
H. armigera 35
Hemorrhagic septicemia 215
Hepatotoxicity 38
Heritability 44
Heterosis 91
Hipposiderous fulvus 8
Holstein friesian 1,88
Hormonal secretions 61
Horticultural 106
Human diet 208
Hyadaphis coriandri 116
I
Indian major carp 215
Indirect selection 172
Infectivity 103
Information technology 5
Inland water 175
Internet 5
Intestinal tract 19
K
Key stone sp. 189
L
Lactation yield 88
Larval period 75
Larvicidal effect 95
Lathyrus sativus 77
LC 50
38, 210
Line x tester 91
Lipid 86
Liquid formulation 35
Lucknow city 180
M
M. javanica 219
Maggoticides 79

86 Trends in Biosciences 5 (1), 2012
Malaria 222
Margosa 25
Market survey 201
Maruca vitrata 103
Mass production 103
Mdr pathogens 23
Medicinal plants 33,201
Meloidogne incognita 95
Mfc 175
Microorganism 157
Milking management system 1, 88
Morphologically 219
Morphometrically 213, 219
Morphometrics 98
Morphotaxonomy 47
Mycorrhiza 169
N
Natural colorant 157
Natural day length 61
New species 98,123
Nigrocine black 210
Nitrogen 109,169
NMU 77
North central tarai forest 185
NPU 35
NSKE 35
Nutrient 109, 208
O
O. columbiana 82
O. hussainii 82
Oscheius ciceri 82
Oscheius nadarajani 98
P
Paddy 41
Paecilomyces lilacinus 95
Papaya pulp 182
Parasitization 51
Parthenium hysterophorus 68
Pesticide resistant 194
Petroleum ether 25
Phosphorous 169
Photoperiodic schedule 61
Physico-chemical quality 118
Phytostimulation 194
Pigeonpea 82
Plant age 222
Plant density 222
Plant height 222
Plant products 75
Plantago ovate 169
Plasmid profile 194
Ponds 180
Population 56, 219
Potassium 169
Productivity 41
Protein profile bioprotection 194
R
Reactive oxygen species 161
Rearing performance 51, 53
Resistance 114
Resources 5
Rice weed control 224
Rice 91,114
S
Sca 120
Scenario 106
Schizaphis graminum 56
Schizothorax curvifrons 19
Seasonal abundance 116
Seed yield 169
Seedling length 205
Selection index 172

Subject Index (Vol. 4, No. 1&2, 2011) 87
Sensory quality 182
Sewage water 175
Sheath rot 114
Slow release water soluble micronutrient brick 86
Slow rusting 31
Somaclonal variations 21
Species novel 58
Species novum 47
Spirulina 182
Springseason 53
Star fruit 208
Steinernema sayeedae 123
Steinernema seemae 35
Stem rot 71
Structural alterations 165
Sunflower genotypes 31
T
Taxonomy 98, 123
Testicular activity 61
Tomato 101
Topical application 79
Traditional knowledge 189
Traditional medicine 201
Trigonella foenum graceum 86
Triticum aestivum 56
U
Uasb reactors 66
Ultra-structural 210
Urban environment 201
Uredospores 31
V
Variability 44
Vigour 205
Vindhyan region 189
W
Weather factors 12
Web-based 5
Weed dynamics 224
Wheat 41, 56
Y
Yield components 91
Yield 41
Z
Zea mays 63

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