Pathological investigation and protective immunity of recombinant vaccine candidates of equine influenza virus in BALB/c mice
Gupta, R.P.
Equine influenza is an important viral respiratory disease of equines caused by Influenza A virus. The antigenic drift in H3N8 virus necessitates regular update and harmonization of vaccine strain with the circulating virus. Thus, recombinant HA subunit vaccine/ RG based EIV vaccine could be potential tools wherein vaccine virus strain can be quickly updated in state of emergency. The present study has been undertaken to evaluate the protective efficacy of recombinant Haemagglutin-1 (rHA-1), rHA-1 conjugated with bacteriophage (VTCCBPA6) and recombinant equine influenza virus adjuvanted with alhydrogel in murine model. For the protection efficacy studies, mice were divided into 4 groups. Mice from group A were immunized with rHA1, group B with rHA1 conjugated with bacteriophage and group C with inactivated recombinant EIV adjuvanted with alhydrogel followed by two booster doses. In group D mice immunization has not been done. On day 42 of primary immunization, mice from each group were subdivided into A1, A2, B1, B2, C1, C2, D1, D2 from which groups A1, B1, C1 and D1 challenged with wild EIV (H3N8). Mice from group D2 were mock infected with sterile PBS, served as negative control. Mice from each subgroup were sacrificed on 0, 1, 3, 5, 7, 10 and 14 days post challenge. The mice were monitored for clinical signs, body weight loss, gross and histopathological lesions, ultrastructural changes, immunohistochemical localization of EIV antigen in tissues, humoral immune responses, CMI response by cytokine estimation using qRT-PCR, residual virus titration in eggs and virus genome quantification by qRT-PCR. Immunization of mice results in antibody titres which was highest in group C1 which further boosted after challenge. CMI response was highest in group B followed by group A and C. Unvaccinated mice showed severe respiratory signs and body weight loss after challenge while vaccinated groups showed mild signs. Gross and histopathological lesions were severe in positive control group whereas vaccinated groups particularly group C1 suffered only mild disease. Group B1 and A1 had mild to moderate intensity of lesions but comparatively less than group D1 and the same were validated by the EIV antigen detection by IHC. Virions ranging from 80-120nm size visualized intra/intercellularly by TEM. Live EIV in nasopharynx and lung tissue was detected in group D1 till 5 dpc with high titre whereas it was not detected in group C1 and with lesser titres in group A1 and B1. Virus titres were further validated with EIV RNA copy numbers in nasal wash and lung tissue. Thus it can be concluded that inactivated recombinant virus (RG-EIV) conferred best protection followed by rHA1 conjugated with bacteriophage and rHA alone.
Veterinary Pathology
Pathological investigation and protective immunity of recombinant vaccine candidates of equine influenza virus in BALB/c mice
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LUVAS-375023- Ramesh Kumar.pdf