ANNUAL REPORT - Department of Biotechnology

ody hepatitis (IBH) in domestic fowl were studied
with an aim to develop a cell culture based vaccine.
Both the viruses were successfully isolated in
chicken embryo liver (CEL) and chicken kidney (CK)
cell lines. Cytopathic effects (CPE) produced by HPS
virus were characterized, which was evident from the
first passage itself. All three HPS isolates were
serotyped. Viral DNA of all 3 HPS isolates extracted
from infected CEL revealed the presence of 1223 bp
and 1190 bp PCR products.
In an effort to develop a candidate vaccine against
Haemonchus contortus, at IVRI, Izatnagar, a 66 kDa
antigen (p66) secreted by adult worms was
characterized as monocyte inhibiting factor.
Challenge experiment in goats showed reduction in
worm burden that ranged between 34% (lowest
reduction) to 70% (highest protection). The
protection reported to be mediated by antibody as
immunized animals had elevated levels of anti-p66
antibodies whereas no cell mediated immune
response was observed. Characterization of
Haemonchus CalR indicates that it may act as an
anti-coagulant. A 55 kDa protein that inhibits
neutrophil and monocytes functions also appeared to
down regulate T lymphocyte but not B cells.
At IVRI, Bangalore, attempts were made to develop a
DBT Annual Report 2006-07
82
self replicating gene vaccine for foot and mouth
disease virus(FMDV). Two vectors viz. one for
cellular response carries a CMV promoter and
polyadenylation signal and another vector for
humoral immune response contains eEF1 promoter,
Sindbis virus polymerase gene and secretory and
anchoring signals were constructed. The linked
polyvalent protein genes of FMDV serotype A, O and
Asia 1 were cloned into the vectors and the presence
of the insert was confirmed by restriction enzyme
digestion. The constructed DNA vaccine was injected
into guinea pigs and the immune response was
compared with the naked DNA vaccine. This
approach of constructing self replicating DNA
vaccine for humoral response is the first report in
FMD.
At NII, New Delhi, attempts were made to develop
recombinant ε-toxin and DNA based vaccine against
Clostridium perfringens. Gene encoding epsilon
toxin of C.perfringens type D has been cloned in
pQE60 expression vector. High level expression and
single step purification of recombinant epsilon toxin
have been achieved. Toxicity of recombinant epsilon
toxin was tested in MDCK cells and has been found to
be toxic. The gene for the toxin has been cloned in
order to produce epsilon toxin independently or in
fusion with GFP aimed at the development of a DNA
based vaccine. Recombinant constructs have been
confirmed by DNA sequencing and authenticated by
the fluorescence analysis of GFP in CHO-K1 cells
transfected with the recombinant constructs.
At IVRI, Izatnagar, a recombinant Brucella antigen
(L7/L12 ribosomal protein) was developed and
found to provide considerable protection to
immunized mice. To increase the efficiency of this
DNA vaccine, two additional immunodominant
antigens viz. Cu-Zn Superoxide dismutase (SOD)
and P39 antigen were also cloned and expressed.
DNA vaccine constructs of these genes have been
made and three antigens (L7/L12, SOD and P39)
were fused to generate a chimeric antigen.
Recombinant proteins as well as the DNA vaccine
constructs were purified and produced in large scale.
Antisera were also raised against the recombinant
antigens and their protective efficacy tested in
laboratory animals.
In a joint programme implemented at MVC, Chennai