Abstract

West Nile (WN) virus caused the death of more than 15 birds in the avian collection at the Wildlife Conservation Society (WCS)—Bronx Zoo within a 3-wk period associated with high mosquito activity in late summer 1999. On preliminary evaluation, more than 50 birds in the WCS collection had serologic evidence of exposure to WN virus. Multiple and varied avian species appeared to be susceptible, with both mortality and morbidity, to this virus. The return of mosquito vectors after the winter quiescent period was considered a risk factor for initiation of a second outbreak of WN virus for collection birds, as well as local indigenous birds.

Japanese encephalitis (JE) virus is in the same group of flaviviruses as WN virus and shares significant nucleotide sequence homology with WN virus; previous studies in non-human primates have shown cross-protection between JE and WN virus vaccines after challenge.¹ We were aware of no studies that had been conducted in birds and considered that JE virus immunization could prove useful to protect endangered species and collection birds, if safe and efficacious. We asked two research questions: (1) Is the inactivated JE virus vaccine (JE-VAX) safe for avian species with considerations of mortality, adverse systemic reactions, local complications, and general behavior? and (2) Does JE-VAX induce a serologic antibody response (neutralizing) to WN virus in vaccinates?

Tragopans (Tragopan caboti x temminckii), ring neck doves (Streptopelia risoria), and laughing gulls (Larus atricilla), as well as previously free-ranging pigeons (Columba livia) were utilized for the study. Birds were selected as model avian species for those avian species with high conservation value and for avian species which suffered high morbidity or mortality associated with WN virus in WCS collection birds.

Briefly, on days 0, 7, and 30, all birds in the study were manually restrained for physical examination, observation of the pectoral muscles (inoculation site), blood collection, and subcutaneous injection of saline or the JE virus vaccine (JE-VAX® aka BIKEN®, manufactured by The Research Foundation for Microbial Diseases, Osaka University, Suita, Osaka, Japan; distributed by Pasteur Mérieux Connaught, Swiftwater, PA, USA). Birds in the experimental group were inoculated subcutaneously with 0.25 ml JE-VAX (tragopans and pigeons) or 0.10 ml JE-VAX (gulls and doves), while birds in the control group were administered a 0.25 ml or 0.10 ml saline placebo subcutaneously, respectively. On day 60, all birds in the study and control groups were manually restrained for physical examination, observation of the inoculation site, and blood collection. Blood samples were processed for complete blood count, biochemical evaluation, and storage for batch processing for WN serology. Frozen plasma and serum samples were delivered to the United States Army Medical Research Institute of Infectious Diseases (USAMRIID) for evaluation of neutralizing antibody titers to JE and WN viruses.

The inactivated JE virus vaccine (JE-VAX) was safe for all avian species in the project. No mortality or morbidity was noted throughout the study period. Statistical analysis and trend identification for the complete blood counts and plasma biochemical panels are in progress.

JE-VAX did not induce a serologic antibody response (neutralizing) to WN virus in previously WN-negative vaccinates. JE-VAX appeared to induce a serologic antibody response (neutralizing) to WN virus in pigeons that already had low WN serologic titers when vaccinated (Table 1). JE-VAX did induce serologic antibody response (neutralizing) to JE virus in some birds (Table 1). Collection birds at WCS were not vaccinated with JE-VAX during the 2001 mosquito season for protection against WN virus, based upon the results of the project.

Further vaccine trials and research are warranted. JE-VAX protection from lethal infection with WNV in American crows (Corvus brachyrhynchos) is under evaluation at the National Wildlife Health Center, Madison, Wisconsin (RG McLean, personal communication). Pending financial investment, other investigators plan avian trials of a recently developed WN vaccine that has proved effective in horses (T.S. McNamara, personal communication).

Table 1. Serologic antibody titers in selected birds to Japanese encephalitis (JE) virus and West Nile (WN) virus at 0, 7, 30, and 60 days post-inoculation with JE virus vaccine or saline placebo.

Acknowledgments

The authors thank the veterinary technicians, avian care staff, veterinary student preceptors, and veterinary pathologists at the Wildlife Conservation Society for their efforts that made this project possible. A Small Grants Award from the Species Survival Fund at the Wildlife Conservation Society funded portions of this project.

Literature Cited

1.  Goverdhan M.K., A.B. Kulkarni, A.K. Gupta, C.D. Tupe, and J.J. Rodrigues. 1992. Two-way cross-protection between West Nile and Japanese encephalitis viruses in bonnet macaques. Acta Virol. 36:277–283.