detection of brucella melitensis and chlamydophila abortus antibodies

Bull Vet Inst Pulawy 51, 493-495, 2007
DETECTION OF BRUCELLA MELITENSIS
AND CHLAMYDOPHILA ABORTUS ANTIBODIES
IN ABORTING SHEEP IN THE KARS PROVINCE OF TURKEY
SALİH OTLU, MİTAT ŞAHİN, AHMET UNVER, AND ÖZGÜR ÇELEBİ
Department of Microbiology, Faculty of Veterinary Medicine,
University of Kafkas, 36040 Kars, Turkey
salihotlu@hotmail.com
Received for publication March 08, 2007
Abstract
A total of 167 blood samples from aborting sheep
were collected from nine different villages and the serum
samples were tested for the presence of Brucella melitensis
antibodies by a serum agglutination test and Chlamydophila
abortus antibodies by ELISA. Seventy-one (40.11%) and nine
(5.38%) serum samples were found to be positive for B.
melitensis and C. abortus, respectively. These results may
indicate that both pathogens are the major infectious agents
responsible for abortions in sheep in the Kars province.
Key words: sheep, Brucella melitensis,
Chlamydophila abortus, antibodies, Turkey.
Ovine brucellosis, caused by Brucella
melitensis, is manifested by abortion, placentitis, and
mastitis in sheep and orchitis in rams (3, 12, 15). The
public’s health along with the economic impact of
brucellosis remains of particular concern in developing
countries throughout Africa, West Asia, and some parts
of Latin America. It is due to danger that infected
animals constitute a source of transmission of this severe
zoonosis to humans, as well as to the economic losses
associated with the disease in animals and the serious
constraints of the improvement of animal husbandry and
genetic resources in the affected areas (5). Brucellosis is
the primary cause of abortion in sheep raised in Turkey.
The prevalence of brucellosis was found to be 37.1%
among sheep population in the country (11, 14, 17, 18).
Another serious disease characterised by
abortion, placentitis, and decreased fertility is ovine
enzootic abortion (OEA), caused by Chlamydophila
abortus (Chlamidia psittaci serotype 1) (6, 7, 16). This
disease, common in Europe, North America, and some
parts of Africa, is a zoonosis; and therefore, the agent
must be dealt with great care and adequate
microbiological precautions and laboratory equipment.
The ELISA kits are commercially available for the
detection of Chlamydophila abortus antibodies in
animals and this assay is more sensitive and specific as
compared to complement fixation test (CFT) (19). The
studies from Turkey showed that OEA is relatively less
common than brucellosis; however, it causes significant
economic losses in this country (4, 8).
The present study aimed to determine the
prevalence of brucellosis and OEA among aborting
sheep in province of Kars by using serological methods.
Material and Methods
Serum samples. A total of 167 blood serum
samples collected from aborting sheep were used. The
sheep population was from 9 villages in Central Kars
and surrounding areas in Kars province. The sera were
stored at –20ºC until tested.
Serum agglutination test (SAT) for
brucellosis. The SAT Brucella antigen was obtained
from Pendik Veterinary Control and Research Institute,
Istanbul, Turkey. The assay was performed as described
by Alton et al. (2). Briefly, 0.5 ml of Brucella SAT
antigen was added to 0.5 ml of each serum sample,
diluted serially from 1:5 to 1: 640 in physiological saline
solution, and mixed thoroughly. The mixtures were
incubated at 37ºC overnight. The agglutination ++ and
stronger, observed in sera at dilution 1:20 and higher,
was considered to be positive.
ELISA for ovine enzootic abortion. ELISA
kit was obtained from Cypress Diagnostics Langdorp,
Belgium, and the assay was performed according to
manufacturer’s instructions. Briefly, appropriately
diluted serum samples with positive and negative control
sera were put into the wells of ELISA plate coated with
C. abortus antigen and incubated at 37ºC for 20 min.
The plate was washed with washing solution of the kit
and peroxidase conjugated anti-sheep IgG secondary
antibody was added to each well. After incubation at
37ºC for 20 min, the plate was washed three times. The
substrate was added to each well and incubated for 10
min. The reaction was stopped with stop solution and

494
the plate was read in ELISA reader (Metertech Σ 960) at
450 nm wavelength filter.
Results
Seventy-one (40.1%) serum samples from
sheep with a history of abortion were found to be
positive for brucellosis in the SAT, at the titre varying
between 20 and 1 280, and only 9 (5.4%) serum samples
were positive for OEA in the ELISA.
Discussion
Brucella melitensis infection and OEA are
leading causes of ovine abortions in many different
geographical areas. Brucellosis is an important zoonosis
in the world and has an economic impact on the
livestock industry. Ovine and caprine brucellosis, caused
by the mentioned microorganisms, are still endemic in
countries of the Mediterranean basin, the Middle East,
Central Asia, and Latin America (1). OEA is also
distributed worldwide and C. abortus is the main
causative agent of small ruminant abortion in numerous
countries, such as Spain, France, and United Kingdom
(7). It was also reported from other countries in the
continents of America, Asia, and Australia (8).
Turkey has a wide geographical variety and live
relations with many neighbouring countries in the terms
of animal movements. The independent serological
studies from Turkey revealed that the prevalence of
brucellosis in sheep was found to be as high as 37.1%.
OEA prevalence was reported with a lower percentage
compared to that of brucellosis. A serological study
from Karaman and Küçükayan (13) in the period of
1993-1997 in the Region of Central Turkey showed that
15.6% and 1.8% of aborting sheep were found positive
for brucellosis and OEA, respectively. Güler et al. (10)
reported the 17.6% brucellosis and 1.35% OEA in
aborting sheep raised in the same region between 1987
and 1998. Two studies carried out in Eastern Turkey
showed that the seroprevalence of brucellosis was
11.8% in Elazõğ and surrounding areas (14), 23.1% in
Van Region (11). The result of another study was 16.1%
in the Thrace Region located in the North-western
Turkey (9). The prevalence of brucellosis was higher
especially in north-eastern Turkey, including Kars, than
that in other parts of Turkey (17, 18). Duman and Durak
(8) found 45 out of 224 (20%) aborting sheep serum
samples positive for OEA by CFT with the titres
between 16 and 32 in Konya Region. In another
serological study carried out in Kars Region, 19.05%
and 17.95% of aborted sheep were found to be positive
for OEA by CFT and ELISA, respectively (4). In the
present study, while analysing serum samples from
aborting sheep, 71 (40.1%) and 9 (5.4%) out of 167
were found to be positive for brucellosis and OEA,
respectively. These results indicate that the prevalence is
higher compared with the other parts of Turkey and that
for OEA the difference is significant. The etiological
profile of ovine abortion in the Kars Region should be
investigated in further studies in aborting sheep
considering other diseases causing abortion
(campylobacteriosis, salmonellosis, leptospirosis,
listeriosis, bluetongue, border disease, toxoplasmosis,
etc.).
The results of the present study may show that
brucellosis and less commonly occurring OEA, are two
important diseases in aetiology of ovine abortions with a
higher prevalence in this region compared with other
areas of Turkey. Abortions are common in the Kars
Region causing significant economic loss in livestock.
The results of this study may reveal that the eradication
programme, or at least control measures and education
of farmers, need to be improved to prevent the
mentioned diseases.
References
1. Al-Talafhah A. H., Lafi S.Q., Al-Tarazi Y.:
Epidemiology of ovine brucellosis in Awassi sheep in
Northern Jordan. Prev Vet Med 2003, 60, 297-306.
2. Alton G. G., Jones L. M., Angus R.D., Verger J.M.:
Techniques for brucellosis laboratory. INRA, Paris,
1988, pp. 123-130.
3. Baum M., Zamir O., Bergman-Rios R., Katz E., Beider,
Z., Cohen A., Banai M.: Comparative evaluation of
microagglutination test and serum agglutination test as
supplementary diagnostic methods for brucellosis. J Clin
Microbiol 1995, 33, 2166-2170.
4. Baz E.: The detection of antibodies against Chlamydia
psittaci using complement fixation test (CFT) and
enzyme-linked immunosorbent assay (ELISA) in aborted
sheep in Kars district. PhD thesis, Institute of Health
Science, Kafkas University, Kars, Turkey, 2000.
5. Benkirane A.: Ovine and caprine brucellosis: World
distribution and control/eradication strategies in West
Asia/North Africa region. Small Rumin Res 2000, 62,
19-25.
6. Borel N., Doherr M. G., Vretou E., Psarrou E., Thoma
R., Pospischil A.: Seroprevalances for ovine enzootic
abortion in Switzerland. Prev Vet Med 2004, 65, 205-
216.
7. Buendia A. J., Cuello F., Del Rio L., Gallego M. C., Caro
M. R., Salinas J.: Field evaluation of a new commercially
available ELISA based on a recombinant antigen for
diagnosing Chlamydophila abortus (Chlamydia psittaci
serotype 1) infection. Vet Microbiol 2001, 78, 229-239.
8. Duman R., Durak Y.: Investigation on Chlamydia
psittaci infections causing abortion in sheep in Konya
district using complement fixation test. Turk J Vet Anim
Sci 1998, 22, 511-515.
9. Erdoğan İ., Gürel A., Tekin C., Uyanõk F., Bitgel A.:
Detection and distribution of bacterial abortion in sheep,
goats and cattle in the Thrace Region J Pendik Vet
Microbiol 1993, 24, 23-35.
10. Güler L., Gündüz K., Baysal T.: Bacteriological and
serological examination of material from sheep that
aborted at the Konya Veterinary Control and Resarch
Institute in 1998. Veterinarium 1998, 9, 3-10.
11. Gürtürk K., Solmaz H., Ekin İ. H., Aksakal A., Gülhan
T.: Bacteriological and serological examinations of
aborting sheep in Van Region. J Fac Vet Med Univ
Yuzuncu Yil 2000, 11, 19-22.

495
12. İyisan A. S., Akmaz Ö., Gökçen Düzgün S., Ersoy Y.,
Eskiizmirliler S., Güler L., Gündüz K., Işõk N.,
İçyerioğlu A. K., Kalender H., Karaman Z., Küçükayan
U., Özcan C., Seyitoğlu Ş., Tuna İ., Tunca T., Üstünakõn
K., Yurtalan S.: Seroepidemiology of brucellosis in cattle
and sheep in Turkey. J Pendik Vet Microbiol 2000, 30,
21-75.
13. Karaman Z., Küçükayan U.: The serological and
microbiologic examination results for the materials
consisted of aborted sheep sera that had been sent to Etlik
Central Veterinary Control and Research Institute
between 1993-1997. J Etlik Vet Microbiol 2000, 11, 23-
30.
14. Muz A., Ertaş H. B., Öngör H., Gülcü H. B., Özer H.,
Eröksüz H., Dabak M., Başbuğ O., Kalender H.:
Bacteriologic, serologic and pathologic studies on
abortus cases of goats and sheep in Elazõğ and its
vicinity. Turk J Vet Anim Sci 1999, 23, 177-188.
15. Öngör H., Muz A., Çetinkaya B.: Comparison of ELISA
with other serological tests in the diagnosis of ovine
brucellosis. Turk J Vet Anim Sci 2001, 25, 21-26.
16. Papp J. R., Shewen P. E., Gartley C. J.: Chlamydia
psittaci infection and associated infertility in sheep. Can J
Vet Res 1993, 57, 185-189.
17. Şahin M., Atabay H. İ., Otlu S., Ünver A., Çelebi Ö.:
Serological and cultural investigation of the prevalence
of brucellosis in cattle, sheep and humans in Kars and its
surrounding area. VI. National Congress of Veterinary
Microbiology Proceedings. Fõrat University, Veterinary
Faculty, Elazõğ-Turkey, September 14-16, 2004, p. 133.
18. Şeyda T., Güler M. A., Genç O.: Studies on diagnosis of
B. melitensis with microagglutination test (MAT) in
sheep. J Fac Vet Med Univ Kafkas 1997, 3, 67-72.
19. Travnicek M., Kovacova D., Bhide M. R., Zubricky P.,
Cislakova L.: Detection of IgG antibodies against
Chlamydophila abortus in sheep with reproductive
disorders. Acta Vet Brno 2003, 73, 95-99.