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.

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.

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.

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.

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.

Epidemiology of bluetongue viruses in the American tropics. Regional Bluetongue Team.

A study of the epidemiology of bluetongue viruses is in progress with the collaboration of 11 Central American and Caribbean countries. To date, over 200 bluetongue virus isolates have been obtained from cattle and sheep in sentinel groups distributed in the participating countries. Bluetongue serotypes identified include 1, 3, 6, and 12, virus types not previously recorded in the Western Hemisphere. Although the clinical impact of bluetongue virus infections in this hyperendemic environment appears to be minimal, the ubiquity of infection causes restrictions on the export of ruminant livestock and germ plasm. The stability of the Caribbean region ecosystem and the long-range implications of the interface with the northern temperate bluetongue virus ecosystem are reviewed.

Seroepidemiology of Bivens Arm virus infections of cattle in Florida, St. Croix and Puerto Rico.

Bivens Arm virus (BAV) is a newly discovered rhabdovirus infecting cattle and water buffalo in Florida. The virus is classified as a member of the Tibrogargan group, members of which have hitherto been found only in Australasia. They are considered to be transmitted by Culicoides species. Bivens Arm virus was first isolated from Culicoides insignis which suggests that BAV is also transmitted by this genus. A serological survey of two small groups of cattle raised in St. Croix and Puerto Rico, in the Caribbean, established that antibody to BAV, or a closely related virus, exists on both island. A retrospective analysis of seroconversions to BAV in sentinel calves in Florida, relative to populations of potential Culicoides vectors, failed to demonstrate any statistically significant correlation.

Evaluation of Culicoides insignis (Diptera: Ceratopogonidae) as a vector of bluetongue virus.

Based upon epidemiological evidence, Culicoides insignis Lutz is a probable biological vector of bluetongue viruses (BTV) in South Florida, the Caribbean Region and Central America. The vector potential of this species for BTV was evaluated in the laboratory in a series of experiments using insects caught in the field. Although there was great variation in the percentage of flies that fed from any one catch, it was demonstrated that C. insignis became infected after membrane feeding on a mixture of blood and virus. The infection rates ranged from 20 to 62.5%. Following intrathoracic inoculation, BTV replicated to high titres in C. insignis. Such flies were also shown to be capable of transmitting BTV to susceptible sheep and embryonated chicken eggs. This series of experiments provides the first conclusive evidence that C. insignis is a biological vector of bluetongue virus. This is the first proven vector of BTV in the neotropics.

Bivens arm virus: a new rhabdovirus isolated from Culicoides insignis in Florida and related to Tibrogargan virus of Australia.

During field studies in 1981 on the transmission of bluetongue viruses in ruminants in Florida, a virus was isolated from Culicoides insignis collected near water buffalo (Bubalus bubalis) recently imported from Trinidad. Electron microscopy showed that this isolate, for which the name Bivens Arm virus is proposed, has rhabdovirus morphology. Serologic comparisons were made with recognized rhabdoviruses from terrestrial vertebrates and hematophagous arthropods. Indirect fluorescent antibody, complement fixation and neutralization tests indicated antigenic reactivity between Bivens Arm virus and two rhabdoviruses found only in Australia, Tibrogargan and Coastal Plains viruses. The Australian isolates cause subclinical infections in cattle and water buffalo and are believed to be transmitted by Culicoides. Initially, it was thought that Bivens Arm virus may have been introduced to Florida with the water buffalo from Trinidad, but a serologic survey of cattle serum, collected before the importation of the buffalo revealed antibody to the virus in cattle on farms located in diverse areas of Florida.

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.

Culicoides spp. (Diptera: Ceratopogonidae) associated with cattle in St. Croix, Virgin Islands, and their relevance to bluetongue virus.

Potential biting midge vectors were collected at two sites on St. Croix as part of an ongoing study on the epidemiology of bluetongue viruses in the Caribbean region. Six species of Culicoides were trapped in a New Jersey light trap (mean = 173 biting midges/trap night) near cattle on a dairy farm. C. furens (Poey) and C. insignis Lutz were the predominant phototactic species, and C. pusillus Lutz, C. trilineatus Fox, C. jamaicensis Edwards, and C. phlebotomus (Williston) were collected less frequently. Four species of Culicoides were aspirated from bovine bait during the morning and evening crepuscular periods with a modified car vacuum. C. furens was aspirated primarily from the ventral portions of the bovine host, whereas C. insignis and C. pusillus were collected principally from the dorsum, and C. trilineatus was collected equally from all aspects. At least one individual of each of these aspirated species was blood engorged. Because C. furens, C. insignis, C. pusillus, and C. trilineatus were most abundant and feed on cattle, they deserve further consideration as vectors of bluetongue virus on St. Croix.

Vector competence parameters of Culicoides variipennis (Diptera: Ceratopogonidae) for bluetongue virus serotype 2.

Culicoides variipennis (Coquillett), the only proven vector of bluetongue virus (BLU) in the western hemisphere, was evaluated as a vector of bluetongue virus serotype 2 (BLU 2). This serotype was isolated from sentinel cattle in south Florida at a site devoid of C. variipennis. Culicoides variipennis readily fed on a mixture of defibrinated blood and BLU 2 through chicken skin membrane. An infection rate of 46.2% was obtained. A growth curve of virus titers recovered from orally infected flies showed a linear relationship between the virus titers and the period of extrinsic incubation. Culicoides variipennis also became infected when inoculated intrathoracically with BLU 2. Peak titers were higher and more rapidly attained in inoculated flies when compared with orally infected flies. Infected C. variipennis also transmitted BLU 2 to sheep via bite. These results demonstrate that C. variipennis is a potential biological vector of BLU 2 in the laboratory. The implication of this on the epidemiology of BLU 2 in the United States is that BLU 2 should have become more widespread in ruminants in the United States. The fact that this has not occurred during the past 10 yr is discussed.

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.

The search for viruses in bovine semen, a review.

Viruses reported in bovine semen include those of foot and mouth disease, bluetongue, bovine leukemia, infectious bovine rhinotracheitis, bovine viral diarrhea, ephemeral fever, and lumpyskin disease. Bovine enteroviruses, a parapoxvirus (paravaccinia), and several uncharacterized viruses have also been isolated. Their presence in semen has been recognized by a variety of animal inoculation and cell culture techniques. The prevalence of viruses in semen and the attendant impact on fertility is largely unknown. The widespread distribution of frozen semen, an ideal system for preserving viral infectivity, indicates this product could serve as an important vehicle for transmission of viruses to uninfected herds or areas. This concern has placed restrictions on international movement of semen. Improved methods for detecting viruses in semen are necessary to assess the importance of the problem and to develop realistic precautionary measures.

Preliminary observations on transplacental infection of bluetongue virus in sheep-a possible overwintering mechanism.

Sheep infected mid-gestation with bluetongue virus type 4 and type 16 produced clinically normal lambs that were viraemic at birth. Viraemia persisted for two months in some lambs even though they received colostrum. It is suggested that transplacental infection of bluetongue virus in sheep may be an overwintering mechanism for the virus in some areas of the world.

The isolation of adenoviruses from goats affected with peste des petits ruminants in Nigeria.

Mucosal scrapings from the large intestine of two goats that had died from peste des petits ruminants (PPR) in separate outbreaks in Nigeria were examined for viruses. A mixed viral infection of PPR virus (morbillivirus) and adenovirus was confirmed in both goats. The adenoviruses did not conform to any of the ovine and bovine serotypes recognised; the two isolates were considered different serotypes. It is concluded that, although the role of adenoviruses in the epizootiology of PPR in Nigeria is difficult to appraise, they are probably commensals. This is believed to be the first report of the isolation of adenoviruses from goats.