Abstract

Since the arrival of West Nile virus (WNV) in North America in 1999, preventive measures have been taken to control the spread of the disease in bird collections in the zoo community. One approach is the use of a killed vaccine approved for equines. A currently accepted vaccine protocol consists of a three-vaccine series administered 3–4 weeks apart. Blood samples are collected each time the bird is inoculated. The Houston Zoo has vaccinated over 350 birds and tested over 550 blood samples for WNV.

Introduction

West Nile virus (WNV) has been well studied, both by the human and veterinary medicine sector, since it first entered the USA in fall 1999.¹ One of the very first available WNV vaccines to be used was a killed vaccine (West Nile-Innovator, Ft. Dodge Animal Health, IA, USA) licensed for horses. This vaccine has been used in vaccine trials in selected birds. The first case of WNV confirmed in a bird in Houston occurred 17 June 2002.⁴

Materials and Methods

In anticipation of the arrival of WNV, the Houston Zoo launched a vaccination program for the bird collection in August 2001, primarily those more susceptible species such as the corvids (crows, magpies and jays) and those housed outdoors.

The WNV vaccine being used in this project was a killed vaccine (West Nile-Innovator, Ft. Dodge Animal Health, IA, USA) licensed for equines. Initially, the birds were given between 0.25 ml to 1.0 ml IM in the pectoral muscles, depending upon bird size. However, this protocol was later changed to 1.0 ml, regardless of weight. Birds weighing less than 1.0 kg received the vaccine subcutaneously in the flank web. The vaccine was given at day 0 and then repeated at 3–4-week intervals for a total of three inoculations. Blood samples were collected at pre-vaccination, then at each following vaccination, and finally at 4 weeks after the third and final vaccination. Blood collections were performed mostly under manual restraint using a 25-ga needle with a 1.0- or 3.0-ml syringe. Venipuncture sites included the jugular vein (most frequent), ulnar (wing) vein and metatarsal (leg) vein. The blood collected was transferred into a heparinized collection tube and transported to the hospital laboratory. After centrifugation for 5–10 minutes, plasma was separated, transferred into a cryovial, and stored in an ultracold freezer at -70°C before shipment to the Animal Health Diagnostic Lab (New York State College of Veterinary Medicine, Cornell University, Ithaca, NY) for processing using the Plaque Reduction Neutralization Test (PRNT). This project was part of the American Zoo and Aquarium Association (AZA) ongoing study. Additionally, an initial batch of pre-vaccination samples was submitted to Zoonosis Control Division (Public Health Region 6, Texas Department of Health (TDH), Houston, TX) for processing using the hemagglutination inhibition (HI) test.

The bird collection consisted of approximately 800 specimens representing 250 species. The vaccination program commenced in August 2001 and continues to the present time. To date, the zoo has vaccinated well over 350 birds and tested more than 550 samples for WNV (Table 1).

Table 1. West Nile virus (WNV) vaccination serology results for Houston Zoo, 2001–2003^a

^aTotal number birds vaccinated; Pre-vax, serology prior to vaccination; Post-vax, serology after vaccination series with plaque reduction neutralization test (PRNT) seroconversion noted after inoculation #1, #2, or #3
^bNegative PRNT titer 1:20
^cNo blood drawn post-vaccination
^dPositive titer seroconversion variable, refer to Table 2
^ePositive PRNT titer 1:20 3 weeks after 2nd vaccination
^fPositive PRNT titer 1:20 4 weeks after 3rd vaccination

Results and Discussion

The batch of pre-vaccination samples (primarily Attwater’s prairie chicken Tympanuchus cupido attwateri and Chilean flamingo Phoenicopterus chilensis) that was sent to TDH tested negative for WNV. This result was expected since it was a naïve population prior to the arrival of WNV in Houston.

As a result of the lack of seroconversion seen in the Attwater’s prairie chicken in the initial phase of the study,^2,3 all birds regardless of weight thereafter received 1.0 ml of the vaccine. It was believed that the recommendation of the manufacturer to use a dose of 1.0 ml for horses also applied to the birds tested, as this was the minimal volume required to invoke an immune response.

Of the 132 birds tested for WNV pre-vaccination, none were positive. On that same number of birds tested for WNV post-vaccination, only three species demonstrated seroconversion: the white-necked raven (Corvus albicollis), the green jay (Cyanocorax yncas) and the Chilean flamingos (Phoenicopterus chilensis) (Tables 1 and 2).

Table 2. Plaque reduction neutralization test (PRNT) serology results for West Nile virus (WNV) vaccination in Chilean flamingos (Phoenicopterus chilensis), for Houston Zoo, 2001. Interpretation of negative (Neg) or positive (Pos) and PRNT titer (1: “dilution level”) is listed for samples collected 3–4 weeks after each WNV vaccination.

^aSample showed 50% plaque reduction at negative 1:20.
^bSample showed 60% plaque reduction at negative 1:20.

At this point, it is uncertain whether vaccination with the WNV vaccine has conferred protective immunity. None of these birds were clinically challenged, as many of them are rare and endangered species. However, we suggest that for several of the species, immunity might have been conferred based on the relatively few numbers of WNV-related cases seen after WNV arrived in Houston. These five cases included the Palawan peacock-pheasant (Polyplectron emphanum), the Marianas/Guam crow (Corvus kubaryi), the Mauritius pink pigeon (Columba mayeri), and two nenes (Branta sandvicensis). Only the crow received the three WNV vaccine series, while the others did not. Except for the nenes, the remaining cases involved other medical complications.²

In all the vaccine trials, no adverse side effects have been observed. A possible exception existed in a corvid (plush-crested jay, Cyanocorax chrysops) wherein minor feather loss occurred at the injection site.

In summary, until another vaccine is proven to be more efficacious, the Houston Zoo will continue to use this particular WNV vaccine to vaccinate susceptible bird species and will administer boosters on an annual basis, since the vaccine does not appear to be harmful and may afford some immunity.

Acknowledgments

I thank the veterinary and bird staff involved in this project; Lorna Schnase, zoo volunteer, for tabulation of data; Dr. Amy Glaser at NY State Diagnostic Lab for processing plasma samples; and AZA for funding of the study.

Literature Cited

1.  Layton, M. 2000. The epidemiology of West Nile virus in New York, 1999. Proc. West Nile Virus Action Workshop 1999: 5.

2.  Llizo, S.Y. Management of West Nile virus in zoo birds. Proc. Assoc. Avian Vet. Conf. 2003: 18.

3.  Okeson, D.M., S.Y. Llizo, C.M. Miller and A.L. Glaser. Antibody response of four bird species after vaccination with a killed West Nile virus vaccine. Proc. Am. Assoc. Zoo Vet. Conf. 2000; addendum provided at conference.

4.  Texas Department of Health. 2002. News release: Mosquito precautions issued after WNV detected in Texas. Available at http://www.tdh.state.tx.us/news/b_new421.htm. (VIN editor: Link was not accessible as of 2-4-21).