Studies were undertaken to define the optimum dosage regimen for commercial loupingill virus vaccine, to investigate the susceptibility of vaccinated yet seronegative
sheep to challenge with live louping-ill virus and to determine the storage characteristics
of the vaccine at refrigerated and room temperatures.
Vaccination studies confirmed that two 1ml doses were required to provoke an
adequate immune response in the majority of animals and maximum antibody titres were
produced when an interval of two to eight weeks between injections was employed.
Vaccinated animals were clinically resistant to experimental challenge although in
animals exhibiting minimal or undetectable post-vaccinal antibody titres viraemias of
low intensity were demonstrated. Circulating virus was not detected in animals with
pre-challenge antibody titres greater than 1/10. Studies concerning vaccine stability
demonstrated that both the virus antigen concentrate and the emulsified final vaccine
were immunologically unstable following storage for periods of less than six months.
Production of louping-ill virus vaccine was therefore reappraised in an attempt
to improve vaccine potency and stability and to produce a one-dose vaccine capable
of consistently producing a satisfactory immune response in sheep.
Optimal conditions for the growth of louping-ill virus in BHK-21 cell monolayer
cultures were determined experimentally and using this technique under commercial
conditions titres of harvests ranged from 8.5 log₁₀pfu/ml to 6.1 log₁₀pfu/ml. A
direct correlation between the preinactivation infectivity titres of virus suspensions
and the immunogenicity of resulting vaccines was demonstrated and a threshold titre of
8.0 log₁₀pfu/ml determined. Harvests with preinactivation infectivity titres below
the threshold produced unacceptable vaccines. Concentration of virus antigen was
therefore required to consistently produce potent louping-ill virus vaccine.
The present commercial vaccine is manufactured by concentrating inactivated virus
harvests by methanol precipitation. A study of the efficacy of this concentration
technique demonstrated however that there was no increase in the immunogenicity of
concentrated virus harvests compared to unconcentrated virus harvests and in several
instances concentration resulted in vaccines of decreased potency. The low potency
characteristic of the commercial vaccine may therefore be partly attributable to the
ineffectiveness of this technique for concentrating viral antigen.
Concentration by methods other than methanol precipitation were examined; these
were ultrafiltration and precipitation with polyethylene glycol. Vaccines produced
from virus antigen concentrated by ultrafiltration and prepared under both experimental
and commercial conditions of production were consistently more immunogenic than
corresponding vaccines prepared by concentrating viral antigen by methanol precipitation
or polyethylene glycol precipitation. A single 1ml dose and two 1ml doses of vaccine
prepared by ultrafiltration produced a satisfactory immune response in sheep and cattle
respectively and antibody titres remained constant for at least six months.
Stability studies with vaccine prepared by ultrafiltration and stored at refrigerated
temperatures demonstrated that although the immunogenicity of the emulsified final vaccine
progressively declined over a six month period, immunogenicity of the virus antigen
concentrate remained stable. Further work therefore is required to improve stability of
the final vaccine and it is suggested that this be designed to evaluate the use of
alternate adjuvants, preservative agents and storage conditions.