Bluetongue.

Summary Bluetongue (BT) is an arthropod-transmitted viral disease of non-African ungulates, principally sheep. The disease results from vascular injury analogous to that of human haemorrhagic viral fevers, with characteristic tissue infarction, haemorrhage, vascular leakage, oedema, and hypovolaemic shock. Importantly, BT is not zoonotic. Bluetongue virus (BTV) infection of ruminants and vector Culicoides midges is endemic throughout many tropical and temperate regions of the world; however, within this global range the virus exists within relatively discrete ecosystems (syn. episystems) where specific constellations of BTV serotypes are spread by different species of biting Culicoides midges. Recently discovered goat-associated BTVs, notably BTV serotype 25 (BTV-25) in central Europe, appear to have distinctive biological properties and an epidemiology that is not reliant on Culicoides midges as vectors for virus transmission. Bluetongue virus infection of ruminants is often subclinical, but outbreaks of severe disease occur regularly at the upper and lower limits of the virus's global range, where infection is distinctly seasonal. There have been recent regional alterations in the global distribution of BTV infection, particularly in Europe. It is proposed that climate change is responsible for these events through its impact on vector midges. However, the role of anthropogenic factors in mediating emergence of BTV into new areas remains poorly defined; for example, it is not clear to what extent anthropogenic factors were responsible for the recent translocation to northern and eastern Europe of live attenuated vaccine viruses and an especially virulent strain of BTV-8 with distinctive properties. Without thorough characterisation of all environmental and anthropogenic drivers of the recent emergence of BT in northern Europe and elsewhere, it is difficult to predict what the future holds in terms of global emergence of BTV infection. Accurate and convenient laboratory tests are available for the sensitive and specific serological and virological diagnosis of BTV infection and confirmation of BT in animals. Prevention and control strategies for BT are largely reactive in nature, and typically are reliant on vaccination of susceptible livestock and restrictions on animal trade and movement.

Serological evidence of H3N8 canine influenza-like virus circulation in USA dogs prior to 2004.

H3N8 canine influenza virus (H3N8 CIV) was first reported as a novel canine respiratory pathogen in racing greyhounds and shelter dogs in the U.S.A. in 2004. Phylogenetic analyses determined that this host-adapted pathogen originated from interspecies transmission of an equine influenza virus (EIV), but it is unknown when the transmission occurred prior to discovery in 2004. The objective of this study was to determine if racing greyhound and shelter dog sera collected from 1984 to 2004 had serological evidence of exposure to H3N8 CIV or EIV. Archived sera from 702 racing greyhounds and 1568 shelter dogs were tested for H3 antibodies to the original 2004 CIV isolate, as well as EIV isolates from 1991 to 1999. None of the racing greyhounds from 1984 and 1985 had detectable H3 antibodies. One of the shelter dogs, which entered a north Florida shelter in 2004, was seropositive. For racing greyhounds sampled from 1999 to 2004, 133/520 (26%) dogs had antibodies to both CIV and EIV H3 proteins. The annual seroprevalence was 27% in 1999, 28% in 2000, 10% in 2001, 1% in 2002, 41% in 2003, and 28% in 2004. The odds of H3 seropositivity were greater among dogs that raced > or =6 months, raced on > or =2 tracks, and raced in 1998, 2002, and 2003. Many of the seropositive dogs raced at tracks that were involved in 'kennel cough' epidemics in 1998-1999 and 2002-2003. Based on serological evidence, a H3N8 canine influenza-like virus was circulating in racing greyhounds in the U.S.A. as early as 1999.

Canine H3N8 influenza virus infection in dogs and mice.

An H3N8 influenza virus closely related to equine influenza virus was identified in racing greyhound dogs with respiratory disease in 2004 and subsequently identified in shelter and pet dogs. Pathologic findings in dogs spontaneously infected with canine influenza virus were compared with lesions induced in beagle and mongrel dogs following experimental inoculation with influenza A/canine/Florida/43/2004. BALB/c mice were inoculated with canine influenza virus to assess their suitability as an experimental model for viral pathogenesis studies. All dogs inoculated with virus developed necrotizing and hyperplastic tracheitis and bronchitis with involvement of submucosal glands as well as mild bronchiolitis and pneumonia. Viral antigen was identified in bronchial and tracheal epithelial cells of all dogs and in alveolar macrophages of several dogs. Many dogs that were spontaneously infected with virus also developed bacterial pneumonia, and greyhound dogs with fatal spontaneous infection developed severe pulmonary hemorrhage with hemothorax. Virus-inoculated BALB/c mice developed tracheitis, bronchitis, bronchiolitis, and mild pneumonia in association with viral antigen in airway epithelial cells and in type 2 alveolar epithelial cells. Virus was not detected in extrarespiratory sites in any animals. The results indicate that canine influenza virus infection consistently induces acute tracheitis and bronchitis in dogs. Mice may be a useful model for some pathogenesis studies on canine influenza virus infection.

Essential veterinary education: equipping students with an understanding of the need for research in global veterinary public health.

Research is the foundation of health advancement; therefore, it is imperative that all health professionals are well versed in its importance during their formal training. Since veterinary education in most countries is now focused on preparing clinicians rather than public health practitioners or research scientists, educators should recognise the importance of research by emphasising the principles and key methodologies that are generic in the life sciences. This exposure will provide a baseline understanding for all students, may encourage some to complete research projects and research-focused externships during school, and will ultimately inspire others to pursue research training after graduation. All aspects of veterinary research would benefit from this approach, including veterinary public health. This paper discusses the essential understanding of research that should be gained through veterinary education, particularly within the evolving nature of veterinary public health education.

Safety of an attenuated West Nile virus vaccine, live Flavivirus chimera in horses.

REASON FOR PERFORMING STUDY: West Nile virus (WNV) infection is endemic and able to cause disease in naive hosts. It is necessary therefore to evaluate the safety of new vaccines. OBJECTIVES: To establish: 1) the safety of a modified live Flavivirus/West Nile virus (WN-FV) chimera by administration of an overdose and testing for shed of vaccine virus and spread to uninoculated sentinel horses; 2) that this vaccine did not become pathogenic once passaged in horses; and 3) vaccine safety under field conditions. METHODS: There were 3 protocols: 1) In the overdose/shed and spread study, horses were vaccinated with a 100x immunogenicity overdose of WN-FV chimera vaccine and housed with sentinel horses. 2) A reversion to virulence study, where horses were vaccinated with a 20x immunogenicity overdose of WN-FV chimera vaccine. Horses in both studies were evaluated for abnormal health conditions and samples obtained to detect virus, seroconversion and dissemination into tissues. 3) In a field safety test 919 healthy horses of various ages, breeds and sex were used. RESULTS: Vaccination did not result in site or systemic reactions in either experimental or field-injected horses. There was no shed of vaccine virus, no detection of vaccine virus into tissue and no reversion to virulence with passage. CONCLUSIONS: WN-FV chimera vaccine is safe to use in horses with no evidence of ill effects from very high doses of vaccine. There was no evidence of reversion to virulence. In addition, administration of this vaccine to several hundred horses that may have been previously exposed to WNV or WNV vaccine resulted in no untoward reactions. POTENTIAL RELEVANCE: These studies establish that this live attenuated Flavivirus chimera is safe to use for immunoprophylaxis against WNV disease in horses.

Efficacy, duration, and onset of immunogenicity of a West Nile virus vaccine, live Flavivirus chimera, in horses with a clinical disease challenge model.

REASON FOR PERFORMING STUDY: West Nile virus (WNF) is a Flavivirus responsible for a life-threatening neurological disease in man and horses. Development of improved vaccines against Flavivirus infections is therefore important. OBJECTIVES: To establish that a single immunogenicity dose of live Flavivirus chimera (WN-FV) vaccine protects horses from the disease and it induces a protective immune response, and to determine the duration of the protective immunity. METHODS: Clinical signs were compared between vaccinated (VACC) and control (CTRL) horses after an intrathecal WNV challenge given at 10 or 28 days, or 12 months post vaccination. RESULTS: Challenge of horses in the immunogenicity study at Day 28 post vaccination resulted in severe clinical signs of WNV infection in 10/10 control (CTRL) compared to 1/20 vaccinated (VACC) horses (P<0.01). None of the VACC horses developed viraemia and minimal histopathology was noted. Duration of immunity (DPI) was established at 12 months post vaccination. Eight of 10 CTRL exhibited severe clinical signs of infection compared to 1 of 9 VACC horses (P<0.05). There was a significant reduction in the occurrence of viraemia and histopathology lesion in VACC horses relative to CTRL horses. Horses challenged at Day 10 post vaccination experienced moderate or severe clinical signs of WNV infection in 3/3 CTRL compared to 5/6 VACC horses (P<0.05). CONCLUSIONS: This novel WN-FV chimera vaccine generates a protective immune response to WNV infection in horses that is demonstrated 10 days after a single vaccination and lasts for up to one year. POTENTIAL RELEVANCE: This is the first USDA licensed equine WNV vaccine to utilise a severe challenge model that produces the same WNV disease observed under field conditions to obtain a label claim for prevention of viraemia and aid in the prevention of WNV disease and encephalitis with a duration of immunity of 12 months.

Comparative efficacies of three commercially available vaccines against West Nile Virus (WNV) in a short-duration challenge trial involving an equine WNV encephalitis model.

We used a severe challenge model that produces clinical West Nile virus (WNV) disease to test the efficacy of three commercially available equine WNV vaccines in horses. Twenty-four healthy, WNV-seronegative horses of varying ages and genders were placed, in random and blind manner, into three trial groups consisting of eight horses each; two horses in each group received (i) an inactivated WNV vaccine (K-WN), (ii) a modified-live vaccine (CP-WN) containing the WNV prM and E proteins expressed by a canarypox vector, (iii) a live-chimera vaccine (WN-FV) containing WNV prM and E proteins expressed in a YF17D vector, or (iv) a diluent control. Challenge by this model caused grave neurological signs, viremia, moderate to severe histopathologic lesions in the brain and spinal cord, and an outcome of 0% survivorship in all six control horses. In contrast, challenge in horses at between 28 days postvaccination with the chimera vaccine and 56 days postvaccination with the commercial inactivated or modified-live vaccine resulted in 100% survivorship (protection from the onset of WNV encephalitis and viremia). Horses vaccinated with the live-chimera vaccine showed significantly fewer clinical signs than did the control horses (P

Emerging zoonotic epidemics in the interconnected global community.

The rate at which epidemics of zoonotic disease in humans have surfaced over the past 25 years has shaken--some would say shattered--the assumption that zoonotic diseases are under control, says Paul Gibbs. In this review he analyses the global factors that have led to the increased emergence of zoonotic diseases, sketches several recent epidemics (and where relevant, their relationship to bioterrorism), discusses the lessons learned, and concludes by outlining an agenda for action.

Transmission of equine influenza virus to dogs.

Molecular and antigenic analyses of three influenza viruses isolated from outbreaks of severe respiratory disease in racing greyhounds revealed that they are closely related to H3N8 equine influenza virus. Phylogenetic analysis indicated that the canine influenza virus genomes form a monophyletic group, consistent with a single interspecies virus transfer. Molecular changes in the hemagglutinin suggested adaptive evolution in the new host. The etiologic role of this virus in respiratory disease was supported by the temporal association of rising antibody titers with disease and by experimental inoculation studies. The geographic expansion of the infection and its persistence for several years indicate efficient transmission of canine influenza virus among greyhounds. Evidence of infection in pet dogs suggests that this infection may also become enzootic in this population.

Characterization of surface interleukin-2 receptor expression on gated populations of peripheral blood mononuclear cells from manatees, Trichechus manatus latirostris.

An in vitro system to determine surface interleukin-2 receptor (IL-2R) expression on mitogen-stimulated peripheral blood mononuclear cells (PBMC) from free-ranging manatees, Trichechus manatus latirostris was developed. Human recombinant IL-2, conjugated with a fluorescein dye was used in conjunction with flow cytometric analysis to determine changes in surface expression of IL-2R at sequential times over a 48-h period of in vitro stimulation. Surface expression of IL-2R was detected on manatee PBMC, which also cross-reacted with an anti-feline pan T-cell marker. An expression index (EI) was calculated by comparing mitogen-activated and non-activated PBMC. Based on side- and forward-scatter properties, flow cytometric analysis showed an increase in the number of larger, more granular "lymphoblasts" following concanavalin A (Con A) stimulation. The appearance of lymphoblasts was correlated with an increase in their surface expression of IL-2 receptors. Surface IL-2R expression, in Con A-stimulated PBMC, was detected at 16 h, peaked at 24-36 h, and began to decrease by 48 h. Characterization of the IL-2R expression should provide additional information on the health status of manatees, and the effect of their sub lethal exposure to brevetoxin.

Venereal transmission of pseudorabies viruses indigenous to feral swine.

Between 1995 and 1998, we designed a series of studies in which we attempted to determine the main routes of transmission involved in the natural infection of pseudorabies virus (PRV) indigenous to free-ranging feral swine (Sus scrofa). Naturally infected feral sows transmitted the infection to uninfected feral boars, with which they had been commingled for a 6-wk period. Pseudorabies virus was isolated from boar preputial swabs, but not from nasal swabs. Three of the same PRV-infected feral sows did not transmit the infection to domestic boars during a 16 wk commingling period, despite the fact that they became pregnant. Feral boars, naturally infected with PRV transmitted the virus to domestic gilts while penned together during 6 wk. Pseudorabies virus was isolated from vaginal swabs, but not from nasal swabs of gilts, after 2 and 3 wk of commingling. When the same infected boars were commingled with either feral or domestic boars for 13 wk, PRV transmission did not occur. None of the exposed boars developed neutralizing antibodies or yielded virus from their preputial or nasal swabs. Our results indicate that PRV indigenous to feral swine is preferentially transmitted to feral or domestic swine of the opposite sex by the venereal route. This mode of transmission differs from that seen in the natural transmission of PRV prevalent in domestic swine, where contaminated secretions, excretions and aerosols are responsible for the spread of the virus. Based on these results, we feel that as long as feral swine do not come into direct contact with domestic swine, PRV-infected feral swine probably pose only a limited risk to the success of the National Pseudorabies Eradication Program. The fact that PRV is usually transmitted from feral to domestic swine at the time of mating would indicate that the isolation of domestic herds by the use of a "double fence," should be adequate protection against reinfection with PRV.

Safety testing of needle free, jet injection devices to detect contamination with blood and other tissue fluids.

Needle free jet injection guns have been used extensively in both veterinary and human health to deliver both vaccine and drugs, but in recent years, concerns have mounted for their potential to transmit blood borne disease agents among consecutive vaccinates. A Ped-O-Jet type jet injection device was used to deliver serial subcutaneous injections of 0.5 mL saline (as a surrogate for vaccine) into calves and pigs, with intervening ejectates collected in vials to represent what the next vaccinate would have received. An enzyme linked immunosorbant assay was developed to detect species specific albumin as a marker for blood, using calibration standards from known dilutions of bovine or porcine blood. Assay sensitivity of 20 pL/mL corresponded to the estimated minimal chimpanzee infectious dose of 10 pL for hepatitis B virus. The methodology and available results for evaluating the safety of jet injector devices are reported.

The public health risks associated with wild and feral swine.

Wild swine populations (Sus scrofa) are present in many regions of the world. Large feral populations in North America and Australia are principally derived from introduced domestic pigs. In Europe, most wild boar are found in Germany and Poland. While wild swine are certainly a significant reservoir of infection for domestic swine diseases (for example, African swine fever virus in wild boar in Sardinia), these swine generally do not constitute a major public health risk. Brucella suis infections and trichinellosis are the most important zoonotic threats to public health.

Mechanisms of transmission of Aujeszky's disease virus originating from feral swine in the USA.

To understand the possible mechanisms of transmission of Aujeszky's disease virus (pseudorabies or PRV) from a feral pig reservoir, intranasal infections were initiated in domestic pigs and in pigs from a herd derived from captured feral pigs. Virus strains originating from feral pigs and from domestic pigs were compared. Similar shedding patterns were obtained in both feral-derived and domestic pigs, however, virus strains from feral pigs were markedly attenuated. Virus could be isolated after acute infection from nasal secretions, tonsils and occasionally from genital organs. In studies of transmission of PRV by cannibalism, either latently infected or acutely infected tissue was fed to both domestic and feral-derived pigs. In two similar experiments, latently infected tissue did not transmit virus, but tissues from acutely infected pigs did transmit infection. Cannibalism was observed typically in both types of pigs older than 6 weeks of age. It was concluded that transmission of PRV originating from feral pigs can occur by several mechanisms including the respiratory route and by cannibalism of pigs that die of acute infection. Transmission of PRV from feral swine may, however, result in sub-clinical infection.

Genital infection and transmission of pseudorabies virus in feral swine in Florida, USA.

Seventeen feral swine (FS) naturally infected with pseudorabies virus (PRV) and treated with dexamethasone (4 mg/kg body wt) on five consecutive days shed virus primarily from the genital tract and less frequently from the upper respiratory tract. The FS isolates were identified as PRV by virus neutralization with specific polyclonal antiserum and by direct immunofluorescence. Restriction endonuclease analysis with BamHI showed that representative samples from a total of 62 isolates were identical to each other, but differed in at least 5 DNA bands from the PRV Shope reference strain profile. DNA purified from FS isolates propagated in Vero cells or DNA extracted directly from genital swabs were amplified in the polymerase chain reaction using primers specific for the gpII (gB) gene of PRV. This amplification yielded a product of the expected size (200 bp), which specifically hybridized to a digoxigenin-labelled 30-mer probe complementary to an area within the region defined by the primers. In a transmission experiment, PRV was recovered from the vagina at 1 and 6 weeks after uninfected feral gilts were mixed with infected feral boars. PRV was not isolated from the upper respiratory tract of either gilts or boars. At eight weeks, 4 of the 5 gilts had developed low titer neutralizing antibodies to PRV. Our results indicate that PRV in FS is transmitted through sexual contact.

The epidemiology of bluetongue.

The perception that bluetongue virus (BTV), once introduced to a country, would decimate its sheep industry, grew from the acceptance in the late 1950s that it was an emerging virus with Africa as its source. Epidemiological studies in the 1960s and early 1970s confirmed that the geographic distribution of BTV infections included regions of the world, outside the traditionally defined areas where BT was observed. This was interpreted at the time as representing serious and rapid spread of the virus. This review provides evidence to rebut this earlier view. What has emerged through the 1980s is: (a) the recognition that BTV is a common infection of ruminant livestock throughout the tropics and sub-tropics apparently within several separate ecosystems; (b) in most areas of the world, infection is sub-clinical; (c) incursions of virus (with accompanying disease) into temperate climates do occur at certain key locations, but "die out" usually within the same year; (d) recognition of the vector competence of Culicoides spp in the different ecosystems of the world is critical for understanding the epidemiology of disease; (e) persistent infection in ruminants is no longer considered important in the long term perpetuation of the virus.

Evaluation of swinepox virus as a vaccine vector in pigs using an Aujeszky's disease (pseudorabies) virus gene insert coding for glycoproteins gp50 and gp63.

Pigs were vaccinated by scarification or intramuscular injection with a swinepox virus-Aujeszky's disease (pseudorabies) recombinant (rSPV-AD) constructed by inserting the linked Aujeszky's disease virus genes coding for glycoproteins gp50 and gp63, attached to a vaccinia virus p7.5 promoter, into the thymidine kinase gene of swinepox virus. By 21 days after vaccination, 90 and 100 per cent of the animals vaccinated by scarification or intramuscular injection, respectively, had developed serum neutralising antibodies to Aujeszky's disease virus. Upon challenge with virulent virus, significantly fewer vaccinated pigs developed clinical Aujeszky's disease, nasal shedding of challenge virus was markedly reduced, and the vaccinated groups of pigs maintained or gained weight during the week after challenge whereas the unvaccinated control group lost weight. No transmission of rSPV-AD to in-contact controls was detected during the three weeks before challenge. In a second experiment, serum neutralising antibodies to Aujeszky's disease virus persisted for 150 days after the pigs were vaccinated with rSPV-AD by scarification or intramuscular injection and all the pigs showed an anamnestic response when they were revaccinated.

Epidemiology of bluetongue in Central America and the Caribbean: initial entomological findings. Regional Bluetongue Team.

Forty-four species of Culicoides (Diptera: Ceratopogonidae) were caught in insect light traps during the first 2 years of studies on the epidemiology of bluetongue virus in the Caribbean and Central America. Traps were operated near sentinel ruminants which were bled monthly for serologic evaluation and then virus isolation. More than 570,000 individuals were identified. Culicoides insignis Lutz accounted for 90% of the catch, C. filarifer Hoffman/C. ocumarensis Ortiz 5%, C. furens Poey 3% and C. pusillus Lutz 2%. Other species accounted for less than 1% of the total catch. Sentinel ruminants became seropositive when C. insignis populations were high at many study sites. At a few sites C. pusillus and C. filarifer/C. ocumarensis were predominant or were present in large numbers during seroconversions of sentinels. Virus isolations were obtained from sentinel ruminants during times when these same species were present in large populations.

Prevalence of pseudorabies (Aujeszky's disease) virus antibodies in feral swine in Florida.

Serum samples collected from feral swine (Sus scrofa) throughout Florida (USA) from 1980 to 1989 were tested for antibodies to pseudorabies virus (PRV) by the serum neutralization test, the latex agglutination test, or by the enzyme-linked immunosorbent assay. Seropositive swine were detected at 11 of 13 sites with a composite seroprevalence of 34.8% (579 of 1,662 samples; range = 5.9% to 58.2%) for sites with seropositive swine. Data on age and sex of the swine were available from three sites. Seroprevalence in males and females did not differ significantly (P = 0.62 for the combined data). Seroprevalence in adult (> or = 8 mo) and juvenile swine (< 8 mo) was significantly different at all sites (P < 0.05 for the combined data). From these data, PRV infections appear to occur widely in populations of Florida feral swine and may seriously undermine efforts to eradicate this virus from the domestic swine population of the USA.