Distribution of bluetongue in the United States of America, 1991-2002.

Bluetongue virus (BTV) distribution in the United States of America (USA) is limited by the range of the vector Culicoides spp. Regional differences exist with the north-eastern states being free of BTV, while the central and north-western states are seasonally free of virus. Activity of the virus can be observed throughout the year in the southern USA. Serological evidence defining the distribution of BTV in selected regions of the USA is gathered regularly through serological surveys conducted on samples from slaughter cattle. From 1991 to 2002, ten serological surveys were completed. Results from Alaska, Hawaii, Michigan, Minnesota, New York, Wisconsin and New England consistently demonstrated a seropositive rate of less than 2%, confirming BTV-free status. Antibody against BTV was sporadically detected in cattle originating from states contiguous to the BTV-free regions. Additional information on BTV distribution in the USA is obtained through identification of BTV or BTV RNA in diagnostic, surveillance and export specimens submitted to the National Veterinary Services Laboratories. Results confirm that BTV serotypes 2, 10, 11, 13 and 17 are present in the USA.

Veterinary diagnostic laboratories and their support role for Veterinary Services.

National Veterinary Services administer a number of regulatory programmes, such as foreign animal disease (FAD) surveillance and exclusion of FADs, and certification of regions as free from disease. Laboratory testing is an important part of any surveillance or control programme. Most countries have a national laboratory which performs testing for FADs and provides support for national disease eradication and control programmes. State laboratories provide testing for surveillance programmes and export purposes, in addition to diagnosis of clinical cases. Many national and state laboratories are developing quality assurance programmes to assure the reliability of testing results. Veterinary Services are reliant on the diagnostic expertise of the laboratory system of that country to be able to respond to FAD introductions and to provide the surveillance programmes needed to detect the introduction of diseases and to certify freedom from disease.

Equine West Nile encephalitis, United States.

After the 1999 outbreak of West Nile (WN) encephalitis in New York horses, a case definition was developed that specified the clinical signs, coupled with laboratory test results, required to classify cases of WN encephalitis in equines as either probable or confirmed. In 2000, 60 horses from seven states met the criteria for a confirmed case. The cumulative experience from clinical observations and diagnostic testing during the 1999 and 2000 outbreaks of WN encephalitis in horses will contribute to further refinement of diagnostic criteria.

Detection of North American West Nile virus in animal tissue by a reverse transcription-nested polymerase chain reaction assay.

A traditional single-stage reverse transcription-polymerase chain reaction (RT-PCR) procedure is effective in determining West Nile (WN) virus in avian tissue and infected cell cultures. However, the procedure lacks the sensitivity to detect WN virus in equine tissue. We describe an RT-nested PCR (RT-nPCR) procedure that identifies the North American strain of WN virus directly in equine and avian tissues.

A comparison of diagnostic assays for the detection of type A swine influenza virus from nasal swabs and lungs.

Nasal swabs and lung samples from pigs experimentally infected with H1N1 swine influenza virus (SIV) were examined for the presence of SIV by the indirect fluorescent antibody assay, immunohistochemistry, cell culture virus isolation, egg inoculation, and 2 human enzyme immunoassays (membrane enzyme immunoassay, microwell enzyme immunoassay). Egg inoculation was considered to be the gold standard for assay evaluation. The 2 human enzyme immunoassays (EIA) and egg inoculation agreed 100% for the prechallenge nasal swabs. Agreement on SIV identification in nasal swabs with egg inoculation following challenge was considered to be good to excellent for membrane EIA (kappa = 0.85) and microwell EIA (kappa = 0.86). Agreement on SIV identification in lung tissue with egg inoculation following challenge was good to excellent for membrane EIA (kappa = 0.75), fair for microwell EIA, fluorescent antibody, and cell culture virus isolation (kappa = 0.48, 0.64, 0.62, respectively), and poor for immunohistochemistry (kappa = 0.36). No assay was 100% accurate, including the "gold standard," egg inoculation. In light of this information, it is important to consider clinical signs of disease and a thorough herd history in conjunction with diagnostic results to make a diagnosis of SIV infection.

Pathogenicity of West Nile virus in chickens.

In the fall of 1999, West Nile virus (WNV) was isolated for the first time in the Western Hemisphere during an outbreak of neurologic disease in humans, horses, and wild and zoo birds in the northeastern United States. Chickens are a potential reservoir for WNV, and little is known about the pathogenicity of WNV in domestic chickens. Seven-week-old chickens derived from a specific-pathogen-free flock were inoculated subcutaneously with 1.8 x 10(3) 50% tissue culture infectious dose of a crow isolate of WNV in order to observe clinical signs and evaluate the viremic phase, gross and microscopic lesions, contact transmission, and immunologic response. There were no observable clinical signs in the WNV-inoculated chickens during the 21-day observation period. However, histopathologic examination of tissues revealed myocardial necrosis, nephritis, and pneumonitis at 5 and 10 days postinoculation (DPI); moderate to severe nonsuppurative encephalitis also was observed in brain tissue from one of four inoculated birds examined at 21 DPI. WNV was recovered from blood plasma for up to 8 DPI. Virus titers as high as 10(5)/ml in plasma were observed at 4 DPI. Fecal shedding of virus was detected in cloacal swabs on 4 and 5 DPI only. The WNV also was isolated from myocardium, spleen, kidney, lung, and intestine collected from chickens euthanatized at 3, 5, and 10 DPI. No virus was isolated from inoculated chickens after 10 DPI. Antibodies specific to WNV were detected in inoculated chickens as early as 5 DPI by the plaque reduction neutralization test and 7 DPI by the indirect fluorescent antibody test. Chickens placed in contact with inoculated chickens at 1 DPI lacked WNV-specific antibodies, and no WNV was isolated from their blood plasma or cloacal swabs throughout the 21 days of the experiment.

Recombination between North American strains of porcine reproductive and respiratory syndrome virus.

Porcine reproductive and respiratory syndrome virus (PRRSV), a recently discovered arterivirus swine pathogen, was shown to undergo homologous recombination. Co-infection of MA-104 cells with two culture-adapted North American PRRSV strains resulted in recombinant viral particles containing chimeric ORF 3 and ORF 4 proteins. Nucleotide sequence analysis of cloned recombinant PCR products, encompassing 1182 bases of the 15.4 kb viral genome, revealed six independent recombination events. Recombinant products persisted in culture for at least three passages, indicating continuous formation of recombinant viruses, growth of recombinant viruses in competition with parental viruses, or both. The frequency of recombination was estimated from <2% up to 10% in the 1182 b fragment analyzed, which is similar to recombination frequencies observed in coronaviruses. An apparent example of natural ORF 5 recombination between naturally occurring wild type viruses was also found, indicating that recombination is likely an important genetic mechanism contributing to PRRSV evolution.

Public veterinary medicine: public health. Serologic evaluation of vesicular stomatitis virus exposure in horses and cattle in 1996.

OBJECTIVE: To determine potential risk factors for vesicular stomatitis (VS) in Colorado livestock in 1995 and evaluate VS virus (VSV) exposure of Colorado livestock in 1996. DESIGN: Retrospective case-control study of VS risk factors and seroprevalence evaluation. SAMPLE POPULATION: Premises included 52 that had VS-positive animals and 33 that did not have VS-positive animals during the 1995 epidemic, and 8 in the vicinity of premises that had VS-positive animals during the 1995 epidemic. PROCEDURE: Layout and management data for premises were collected during site visits in 1996. Signalment and management data were collected for animals from which samples were obtained, and samples were tested by serologic examination and virus isolation. The VSV seroprevalence rate was estimated for Colorado, using serum obtained for equine infectious anemia testing and from the Market Cattle Identification program in Colorado. RESULTS: At least 1 animal was seropositive for VSV on 35 of 52 (67%) premises, and 71 of 228 (31%) animals tested were seropositive for VSV. Seroprevalence was 63 of 170 (37%) for horses and 8 of 54 (15%) for cattle. Seroprevalence of VSV in animals from non-study premises in Colorado in 1996 was estimated to be 1.1% in cattle and 0.8% in horses. CLINICAL IMPLICATIONS: Overall VSV seroprevalence in Colorado livestock was less than seroprevalence in epidemic areas, and seroprevalence rates in epidemic areas were greater for horses than cattle. Results may indicate that some animals had subclinical VSV infection during epidemics and that animals may be exposed to VSV between epidemics.

Evaluation of PCR for diagnosis of bovine viral diarrhea virus in tissue homogenates.

Tissue homogenates from 60 specimens submitted to the Veterinary Diagnostic Center were evaluated by polymerase chain reaction (PCR) for detection of bovine viral diarrhea virus (BVDV). Conventional virus isolation procedures showed the specimens contained BVDV. The BVDV RNA was extracted from the homogenates and subjected to a reverse transcription reaction followed by PCR amplification. The PCR product was blotted onto a nylon membrane and hybridized with a 30-base pair oligonucleotide probe labeled with 32P. One set of PCR primers detected BVDV in 46/60 (77%) of the tissue homogenates. An additional set of primers was used to detect 10/11 samples that had escaped detection with the first set of primers. The results indicate that BVDV can be detected by PCR directly out of tissue homogenates generated in a diagnostic setting.