Investigation Into the Comparative Efficacy of Three West Nile Virus Vaccines in Experimentally Induced West Nile Virus Clinical Disease in Horses

This is the first report of a comparative vaccine-efficacy trial in horses where West Nile Virus-induced encephalomyelitis was shown in all of the control animals. These studies show differences in the abilities of two commercial vaccines and a live-chimera vaccine to protect against West Nile Virus-induced disease at 28 days post-vaccination in horses.

1. Introduction

Since its emergence into the United States in 1999, West Nile virus (WNV) has killed a total number of 785 humans, 6000 horses, and an estimated hundreds of thousands of birds. This mosquito-borne encephalitic flavivirus has caused clinical disease in 19,444 humans and 22,908 horses [1]. Using a reproducible model of WNV-induced encephalomyelitis [2], the efficacy of a new live-chimera vaccine and two commercially available vaccines for prevention of clinical signs of WNV-induced disease was tested in horses.

2. Materials and Methods

Animals

Twenty-four healthy, WNV-seronegative horses of varying ages and gender were randomized into three blinded vaccination/challenge trials. Horses were maintained according to University of Florida Animal Care Services and IACUC (Institutional Animal Care and Use Committee) guidelines.

Efficacy Trials

Horses were placed into three randomized, blinded trial groups consisting of eight horses with two horses in each group receiving (1) a killed WNV vaccine, (2) a modified-live vaccine containing WNV prM and E proteins expressed by a canarypox vector, (3) a live-chimera vaccine containing WNV prM and E proteins expressed by a YF17D vector, or (4) a diluent. For the commercial inactivated and modified-live vaccines, horses received an initial primary injection followed in 28 days with a second injection according to manufacturer’s label. For the chimera vaccine, each horse received a primary injection dose of diluent followed in 28 days by a single vaccine dose. All horses were challenged intrathecally with virulent WNV 28 days after vaccination.

Monitoring of Animals

Complete physical and neurological evaluations were performed for 21 days post-challenge (PC). Horses were observed for clinical and neurological signs of mentation, fasciculations, paresis, ataxia, and cranial-nerve abnormalities [3]. Horses were euthanized for humane reasons because of overall severe health condition as a result of WNV infection and/or the inability to locomote without assistance. Twenty-one days PC, all remaining horses were euthanized, and a full gross and histopathological evaluation was performed. Cross sections of the brain and spinal cord were examined and quantified for the presence of gliosis and perivascular cuffing. Serum and plasma samples were collected before and after challenge for virological evaluation.

Statistical Analysis

Neurologic signs were analyzed as 0 = none and 1 = present. Data from all three trials were combined as 2×2 contingency tables and analyzed using χ2 and Fischer Exact. Level of significance was set at p < 0.05. Statistical analysis was performed by use of computer spreadsheet [a] statistical-analysis package [b].

3. Results

Three of six control horses had increased rectal temperature (≥102.5°C). Six of six control horses developed PC viremia and grave neurological disease, and they were all euthanized for humane reasons before the end of the trial. Gross and histopathological changes consistent with WNV polioencephalomyelitis were observed in all control horses. None of the vaccinates, irrespective of the vaccine administered, developed PC viremia, and all survived to the end of the trial period at which time a full gross and histopathological evaluation was performed. For the chimera group, none of the vaccinates developed increased rectal temperatures, and no neurological signs were observed. One of the horses given the modified-live vaccine was determined to have pre-existing neutralizing antibody to WNV and was removed from the study. For the modified-live vaccinates, one of five horses developed increased rectal temperatures, and one of five had consistent mild neurological signs in several categories. Another had mild malaise and anorexia after challenge. For horses given the inactivated vaccine, one of six developed increased rectal temperature, and four of six developed mild to moderate neurologic signs that occurred relatively late in the challenge period. Chimera vaccinates showed significantly fewer clinical signs than horses vaccinated with the inactivated vaccine (p ≤ 0.035) or control animals (p ≤ 0.01). Histopathological analysis showed moderate to severe changes in the controls consistent with WNV encephalitis and mild changes in some vaccinated horses.

4. Discussion

Using a severe challenge model of WNV that induced encephalomyelitis, the efficacy of a new live-chimera vaccine and two commercially available vaccines for prevention of clinical signs of WNV-induced disease was tested in horses. Challenge of horses by this model caused grave and reproducible neurological signs in all six horses that were not vaccinated. In contrast, vaccination with one dose of the chimera vaccine or two doses of the commercial inactivated or modified-live vaccines resulted in 100% survivorship. Horses in both the inactivated and modified-live vaccine groups had a higher occurrence of clinical signs PC when compared with the chimera group. The efficacy of these vaccines against WNV-induced disease in horses challenged >28 days post-vaccination is not known. Additional studies with larger sample sizes and at longer durations after initial vaccination are warranted.

Funding was provided by Intervet, Inc. (Millsboro, DE). Technical assistance was provided by R. A. Bowen (Colorado State University, Fort Collins, CO) and T. Monath (Acambis, Cambridge, MA). In addition, the authors acknowledge the contributions of the following undergraduate and veterinary students: David Gosche, Brett Weldon, Jeremy Camp-field, and Tamara Weber. Dr. Gibbs has been a consultant for Akzo Nobel.

Footnotes

[a] Excel, Microsoft, Redmond, WA 98052.
[b] SysStat, Point Richmond, CA 94801.