MSc

A chicken embryo adapted vaccine strain of duck plague
virus was serially passaged in chicken embryo fibroblast cell
cultures and its immunogenicity was evaluated at different
passage levels.
The vaccine strain of DPV received from VBI, Pal ode was
revived in 11 day old chicken embryos by CAM route. The
infected embryos died in 70 to 120 hr PI with lesions of
congestion on the embryo and CAM and enlargement of liver and
spleen. This embryo passaged virus was propagated in CEF cell
culture, prepared from 12 day old embryonated chicken eggs.
The virus produced CPE, characterised by rounding and clumping
of cells, syncytium formation, vacuolation of cytoplasm and
eosinophilic intranuclear inclusion bodies.
The virus was adapted in CEF cultures by serial passage.
It was passaged for ten times and the various characters of
the fifth and 10th passaged viruses were studied. There was
no change in the CPE but the time required for the appearance
of CPE and total detachment of the cells decreased as the
passages increased. The CPE appeared at 48 hr, 30 hr and 24
hr for first, fifth and 10th passages respectively. Similarly
the time required for total detachment of cells also reduced

from 120 hr at first passage to 90 hr at fifth passage and 80
hr at 10th passage. The rapid onset of CPE and desquamation
of cells indicated the adaptation of the virus in CEF cell
culture.
The titres of fifth and 10th passage viruses in chicken
embryos were 104.75 and 105.77 ELD50/ml respectively. The titres
in CEF cultures were slightly higher. The values were 105.67
and 106.77 TCID50/ml respectively for the fifth and 10th
passaged samples.
The immunogenicity of the fifth and 10th passage viruses
were studied by vaccinating six weeks old ducklings. Each
duckling received 3.5 log10 TCID50 of either fifth or 10th
passaged virus intramuscularly. The birds remained normal
till the 20th day and when challenged with virulent virus.
Birds that received the fifth passaged virus showed mean
antibody titres of 64 and 32 by SNT and PHA respectively. All
the birds withstood challenge indicating the effectiveness of
fifth CEF passaged virus as a vaccine. In birds that received
the 10th passaged virus, the antibody titres were low both by
the SNT (1:54) and PHA (1:22). However all the ducks survived
without manifesting any clinical signs. All the control ducks
developed clinical signs of DP and died in seven to nine days
time.



The fifth and 10th CEF passaged viruses were sensitive to
pH 3 and 11, but stable at pH 7.2. They were completely
inactivated at 56°C in 30 min. These indicated that there was
no change in the above physical characters of the virus though
it was passaged in CEF cultures incubated at 38.5°C.
Though the efficacy of the 10th passage virus was
slightly lower as it was evident from the low antibody level,
a detailed study is required to establish the present findings
that an increase in the number of passages would result in
decreased immunogenicity of the DPV vaccine strain. However
from the results obtained during this study, it is evident
that cell culture adapted DP vaccine strain could be
recommended for production of vaccine against DP.