Determining prevalence of bluetongue and epizootic hemorrhagic disease viruses in mule deer in Arizona (USA) using whole blood dried on paper strips compared to serum analyses

We investigated the feasibility of using whole blood dried on paper strips as a means to collect antibody prevalence data for the epizootic hemorrhagic disease viruses (EHDV) and bluetongue viruses (BTV) from hunter-harvested male mule deer (Odocoileus hemionus) in October 2002 from Arizona, USA. We compared antibody prevalence estimates in mule deer from paired paper strip and serum samples. Prevalence data obtained from elution of dried blood on paper strips proved to be consistent with results from serum in 94% of the samples tested. The paper strip method allows easy collection of blood from dead animals, with a smaller amount of blood being needed for analyses. Also, samples do not need to be refrigerated before analyses. We also used serum samples to determine hemorrhagic disease (HD) serotype exposure status of mule deer harvested from 4 distinct areas in Arizona. Antibodies to BTV and EHDV were identified in 3 of the 4 areas, with positive results to EHDV-1, EHDV-2, BTV-10, and BTV-11 being most common. Many animals did not have antibodies against the BTV serotypes. Exposure varied geographically and potentially with elevation. Hemorrhagic disease viruses commonly infect Arizona mule deer, except on the Kaibab Plateau in northern Arizona.     

Cross-protection between epizootic hemorrhagic disease virus serotypes 1 and 2 in white-tailed deer

Viruses in the epizootic hemorrhagic disease (EHD) serogroup are the most frequent cause of hemorrhagic disease in the southeastern United States, but nothing is known about cross-protection between the two EHD serotypes (EHDV-1 and EHDV-2) present in this region. We experimentally tested whether deer surviving EHDV-2 infection would be protected against subsequent infection with EHDV-1, and used field data to examine the possibility of reciprocal cross-protection. Eleven white-tailed deer fawns (Odocoileus virginianus) were experimentally infected with EHDV-2 and later challenged with EHDV-1. Two EHDV-2-na?ve fawns also were infected with EHDV-1. Deer were monitored via physical examination, complete blood counts, clotting profiles, viral isolation, and serology, and each animal was assigned a quantitative clinical disease severity score based on presence of certain physical and clinical parameters. Infection of na?ve controls with EHDV-1 caused severe clinical disease and death of both fawns, whereas deer previously infected with EHDV-2 exhibited no or minimal signs of disease. Thus, infection with EHDV-2 conferred protection against disease caused by subsequent EHDV-1 infection. Although prior EHDV-2 exposure protected deer from severe clinical disease, it did not prevent infection nor viremia indicating they could still act as virus amplifying hosts. These experimental infections suggest that EHDV-1 and 2 may exist in a state of mutual permissiveness.     

Epizootiology of an epizootic hemorrhagic disease outbreak in West Virginia

An outbreak of epizootic hemorrhagic disease virus, serotype 2 (EHDV-2) was responsible for localized white-tailed deer (Odocoileus virginianus) mortality in Hardy and Hampshire counties, West Virginia (USA), in the summer and fall of 1993. Using available historical data on regional herd immunity, data opportunistically collected during the epizootic, and postepizootic sampling of hunter-harvested deer, we grossly estimate certain epidemiologic parameters and compare findings to a hypothesis about hemorrhagic disease outbreaks in the Appalachian Mountains. During the epizootic, 57.9 km(2) were actively searched and 228 dead deer were found. Epizootic hemorrhagic disease virus, serotype 2 was isolated from seven of nine deer sampled in Hardy and Hampshire counties. Preepizootic exposure of deer to EHD viruses was unknown, but available data suggest that it was negligible. The geographic distribution of the outbreak was defined by plotting the locations of dead deer found during the outbreak, as well as the locations of deer harvested by hunters after the outbreak that had antibodies to EHDV-2 on a map sectioned into 16.65 km(2) rectangular sections. Sections that included one or more dead deer or hunter-harvested deer with antibodies to EHDV-2 were included in the defined outbreak area. Postoutbreak sampling revealed monospecific EHDV-2 antibodies in 12% of deer harvested by hunters within the defined outbreak area. Based on the available data and accepting certain assumptions, gross calculations suggest that this outbreak appears to have been isolated and probably killed a high percentage of the deer that were infected. This is consistent with the hypothesis that sporadic hemorrhagic disease outbreaks in the Appalachian Mountains are usually localized and severe.     

Susceptibility of white-tailed deer (Odocoileus virginianus) to experimental infection with epizootic hemorrhagic disease virus serotype 7

During the fall of 2006, in Israel, epizootic hemorrhagic disease virus (EHDV) serotype 7 caused an intense and widespread epizootic in domestic cattle that resulted in significant economic losses for the dairy industry. The susceptibility of potential North American vector and ruminant hosts to infection with EHDV-7 is not known but is essential to understanding the potential for establishment of this exotic orbivirus in North America if it were introduced. Our primary objective was to determine whether white-tailed deer (WTD; Odocoileus virginianus) are susceptible to infection with EHDV-7. Six, 8-mo-old WTD were experimentally infected with EHDV-7, and all became infected and exhibited varying degrees of clinical disease. Clinical signs, clinicopathologic abnormalities, and postmortem findings were consistent with previous reports of orbiviral hemorrhagic disease (HD) in this species. Four of six animals died or were euthanized because of the severity of disease, one on postinoculation day (PID) 5 and the remaining WTD on PID 7. All deer had detectable viremia on PID 3, which peaked on PID 5 or 6 and persisted for as long as PID 46 in one animal. Deer surviving the acute phase of the disease seroconverted by PID 10. Based on the 67% mortality rate we observed, this strain of EHDV-7 is virulent in WTD, reaffirming their role as a sentinel species for the detection of endemic and nonendemic EHDV. Further, the observed disease was indistinguishable from previous reports of disease caused by North American EHDV and bluetongue virus serotypes, highlighting the importance of serotype-specific diagnostics during suspected HD outbreaks.     

Dynamics of maternal antibodies to hemorrhagic disease viruses (Reoviridae: Orbivirus) in white-tailed deer

Enzootic stability, potentially associated with acquired resistance and subsequent transfer of maternal antibodies, innate resistance, or both, has been hypothesized to explain the lack of reports of hemorrhagic disease (HD) in white-tailed deer (Odocoileus virginianus) from Texas. The objectives of this research were to determine the following: how long maternal antibodies to epizootic hemorrhagic disease (EHD) and bluetongue (BT) viruses persist; whether fawns from an enzootic site are naturally exposed to EHD and BT viruses while maternal antibodies are present; and whether field-challenged fawns develop clinical disease. Twelve of 52 fawns from Texas were moved to an indoor facility. All 12 (100%) were positive for maternal antibodies to EHD or BT viruses by agar gel immunodiffusion (AGID) and serum neutralization (SN) tests. Weekly monitoring demonstrated that precipitating antibodies disappeared by 23 wk of age and serum neutralizing antibodies disappeared by 17-18 wk of age. Fawns that remained outdoors in Texas were not observed with signs of HD. At 14-21 wk of age (October), 39 of 40 (98%) fawns that had remained outdoors were positive for EHD and/or BT virus antibodies by AGID and 32 (80%) had SN antibody titers to one or more of five viruses (EHDV-1, EHDV-2, BTV-10, BTV-11, BTV-17). Antibody titers to EHDV-1, EHDV-2, and BTV-11 all exceeded titers of same-age indoor fawns, suggesting recent exposure. Epizootic hemorrhagic disease viruses were isolated from seven (18%) of the outdoor fawns and all 40 remained clinically normal. Natural exposure of deer to EHD and BT viruses occurred at this site in the presence of maternal antibodies without causing disease. This may be due to acquired immunity and the subsequent transfer of maternal antibodies, but it does not exclude innate resistance as a possible factor in the enzootic stability of EHD and BT viruses at this location.     

Oral and fecal shedding of epizootic hemorrhagic disease virus, serotype 1 from experimentally infected white-tailed deer

Epizootic hemorrhagic disease (EHD), one of the most important infectious diseases of white-tailed deer (Odocoileus virginianus), is vectored by species of midges in the genus Culicoides. Although vector borne, fecal shedding of EHD virus, serotype 2 has been reported from infected deer in a previous study. To evaluate the potential for fecal and oral shedding, oral and rectal swabs were obtained on day 8 post-inoculation from white-tailed deer fawns experimentally infected with EHD virus, serotype 1 (EHDV-1). Eight deer were viremic for EHDV-1; virus was detected in oral swabs from three (38%) and in rectal swabs from four (50%). The ability to isolate EHDV-1 in oral secretions or feces was not dependent on being able to detect clinical disease. These results indicate that in a relatively large proportion of EHDV-1 infected deer, virus can be detected in feces and oral secretions. Although more work is necessary, such shedding may be important in experimental studies or pen situations where deer-to-deer contact is prevalent and intense.     

Molecular characterization of epizootic hemorrhagic disease virus serotype 1 associated with a 1999 epizootic in white-tailed deer in the eastern United States

During the autumn of 1999 (mid-August-late September), an outbreak of hemorrhagic disease in white-tailed deer (Odocoileus virginianus) caused by epizootic hemorrhagic disease virus serotype 1 (EHDV-1) occurred along the east coast of the United States from Georgia to New Jersey. An EHDV-1 epizootic of such magnitude had not been described in this region since 1975. To determine the genetic relatedness among the 1999 viruses, as well as among additional EHDV-1 isolates from the eastern and western United States, portions of the S10 and L2 gene segments were sequenced and compared utilizing phylogenetic analyses. Nearly all of the 1999 eastern isolates were identical in nucleotide sequence at one or both loci. Additionally, confirmed cases of EHDV-1 in white-tailed deer occurred in a south (Georgia)-to-north (New Jersey/Virginia) progression over a short period of approximately six weeks. Taken together, these results indicate that this outbreak resulted from the spread of a single viral strain. The phylograms derived from analysis of the entire sample set displayed eastern and western region-specific clusterings (topotypes), as well as an eastern versus western difference in branch lengths, which may reflect the influence of epizootic versus enzootic transmission patterns on viral genetic diversity.     

Hemorrhagic disease in bighorn sheep in Arizona

Two bighorn sheep from Arizona (USA) were submitted for necropsy. One was a Rocky Mountain bighorn (Ovis canadensis canadensis) and the other was a desert bighorn (Ovis canadensis mexicana). Both had lesions consistent with those of hemorrhagic disease (HD). Epizootic hemorrhagic disease virus (EHDV) type-2 and bluetongue virus (BTV) type-17, respectively, were isolated from the sheep tissues. To our knowledge, HD caused by either EHDV or BTV infection has not been documented previously in Arizona bighorn sheep.     

Serologic survey of selected pathogens in white-tailed and mule deer in western Nebraska

Exposure of free-ranging white-tailed deer (Odocoileus virginianus) and mule deer (Odocoileus hemionus) in western Nebraska to selected livestock pathogens was determined by serology and attempted virus isolation. Antibodies to bluetongue virus, epizootic hemorrhagic disease virus, and bovine respiratory syncytial virus were present in both species of deer. No serologic reactors to Brucella or Anaplasma were found. Attempts to isolate bluetongue virus were negative.     

Experimental adenovirus hemorrhagic disease in white-tailed deer fawns

Infection with a newly described endotheliotropic adenovirus was the cause of a 1993 epizootic reminiscent of hemorrhagic disease in California mule deer (Odocoileus hemionus columbianus and O. hemionus hemionus). Pulmonary edema and intestinal luminal hemorrhage, or necrotizing stomatitis associated with systemic or localized vasculitis, respectively, were common lesions seen in animals that died during the epizootic. In order to determine if white-tailed deer (Odocoileus virginianus) also are susceptible to infection and fatal disease with the deer adenovirus, eight white-tailed deer fawns (4- to 6-mo-old) were inoculated with purified deer adenovirus. Four were inoculated intravenously and four were inoculated through the mucous membranes. Seven days post-inoculation, one of the fawns inoculated intravenously died. Pulmonary edema and hemorrhagic enteropathy were associated with pulmonary and intestinal vasculitis with systemic multiorgan distribution of endotheliotropic adenovirus as demonstrated by transmission electron microscopy and immunohistochemistry. Adenovirus was reisolated from lung homogenates of the fawn that died of adenovirus hemorrhagic disease.     

Experimental bluetongue and epizootic hemorrhagic disease virus infection in California black-tailed deer.

Four adult black-tailed deer (Odocoileus hemioneus columbianus) and five fawns were inoculated with bluetongue virus (BTV) and one adult deer was inoculated with epizootic hemorrhagic disease (EHD) virus to produce clinical signs and lesions of hemorrhagic disease. Serologic response was monitored using the agar gel immunodiffusion (AGID) test and the competitive enzyme-linked immunosorbent assay (C-ELISA). Embryonating chicken eggs and vero cells were used to detect viremia. No animal exhibited clinical or pathologic signs of hemorrhagic disease. Bluetongue viremia was detected as early as 2 days post-inoculation (DPI-2) and in some animals, persisted until at least DPI-12. The earliest detection of BTV antibodies using the AGID was DPI-8. Two adult deer remained seropositive for BTV antibodies for > 9 mo and 1 yr, respectively, using both the AGID and C-ELISA tests. We observed cross reactions between BT and EHD antibodies using the AGID tests. Also, the AGID test did not consistently detect exposure to BTV. Viremia was not detected in the deer inoculated with EHD although this animal was AGID positive between DPI-6 and DPI-49.     

Hemorrhagic disease in Kansas: enzootic stability meets epizootic disease

Kansas (USA) could represent a transition area between contrasting epidemiologic patterns of hemorrhagic disease (HD) in the midwestern United States. In this study, we compare the distribution of reported clinical HD with serologic data to determine whether the risk of HD in white-tailed deer (Odocoileus virginianus) is associated with geographic location corresponding to the reported distribution of two white-tailed deer subspecies. On the basis of a high prevalence of antibodies (91-100%) to multiple serotypes of epizootic hemorrhagic disease virus (EHDV) and bluetongue virus (BTV), with correspondingly few reports of clinical HD, it appears that a state of enzootic stability exists in central and western Kansas. This area corresponds to the reported range of O. virginianus texanus. In contrast, in the eastern third of the state, which corresponds to the reported range of O. virginianus macronurus, antibody prevalence is significantly lower (45%), EHDV serotypes appear to predominate, and HD, as confirmed by virus isolation, has been consistently reported. These results suggest an abrupt demarcation between enzootic stability in central and western Kansas to a pattern of epizootic HD within the eastern part of this state. Understanding host, vector, and environmental variables responsible for these contrasting patterns could have application to understanding the risk of HD in the midwestern United States.     

An epizootic of hemorrhagic disease in white-tailed deer in Missouri

As part of a white-tailed deer (Odocoileus virginianus) survival study in Missouri (USA) we were actively monitoring 97 radio-collared deer when 8 (8%) died. This mortality, which occurred from 20 August to 23 September 1996, consisted of five adult females, two yearling females and one yearling male. Based on the seasonality of this mortality and the isolation of epizootic hemorrhagic disease virus (EHDV) serotype 2 from one of these animals, we believe that these losses resulted from an epizootic of hemorrhagic disease. The remains of five unmarked deer that may have died from HD also were found on the study area during this same period. During the fall following this mortality, we tested serum from 96 deer taken by hunters in the immediate area. Fifteen (16%) were positive for EHDV or bluetongue virus (BTV) antibodies as determined by agar gel immunodiffusion tests. Serum neutralization test results indicated that previous infections were caused by EHDV virus serotype 2. Based on these data, and assuming that there was no prior exposure to EHDV serotype 2 in this population, the exposure rate for this epizootic was 24% of which 8% died. We noted hoof interruptions in only two of the 96 deer sampled. During this mortality event, the Missouri Department of Conservation received no reports of dead deer, and without the radio-monitored animals the event would have been undetected.     

Landscape Seroprevalence of Three Hemorrhagic Disease-Causing Viruses in a Wild Cervid

Disease plays a major role in shaping wildlife populations worldwide, and changes in landscape conditions can significantly influence risk of pathogen exposure, a threat to vulnerable wild species. Three viruses that cause hemorrhagic disease affect cervid populations in the USA (Odocoileus hemionus adenovirus, bluetongue virus, and epizootic hemorrhagic disease virus), but little is known of their distribution and prevalence in wild populations. We explored the distribution and co-occurrence of seroprevalence of these three pathogens in southern mule deer (Odocoileus hemionus fuliginatus), a subspecies of conservation concern and a harvested species native to southern California, to evaluate the distribution of exposure to these pathogens relative to landscape attributes. We found that habitat type, level of development, and proximity to livestock may affect hemorrhagic disease seroprevalence in southern mule deer. Continued monitoring of hemorrhagic disease-causing viruses in areas where deer are in proximity to cattle and human development is needed to better understand the implications of future outbreaks in wild populations and to identify opportunities to mitigate disease impacts in southern mule deer and other cervid species.

     

An outbreak of a hemorrhagic disease in white-tailed deer in Kentucky.

In 1971, an outbreak of a hemorrhagic disease occurred in captive and free-ranging white-tailed deer (Odocoileus virginianus) in Mammoth Cave National Park, Kentucky, Clinical signs and gross pathological lesions were consistent with those of epizootic hemorrhagic disease and bluetongue, as were serological and histopathological findings for samples sent to other laboratories. The infection rate among the 104 captive deer was 88-92%, and that among the free-ranging Park deer appeared to be similar. Mortality was negligible in the Park deer, but 65 (62%) of the captive deer died. The deaths were bimodally distributed over a 36-day period, and the mortality rate decreased from 97-100% for deer clinically ill during the first 17 days of the outbreak to 58% for deer first exhibiting clinical signs on day 16 or later. Mortality was equal in males and females, but less in yearlings than among fawns or adults. Winter mortality among survivors of the initial outbreak was associated with low ambient temperatures and sometimes fungal and bacterial abscesses, possibly sequelae or complications of the hemorrhagic disease. The pregnancy and birth rates among surviving does appeared to be normal.     

Innate resistance to epizootic hemorrhagic disease in white-tailed deer

Differences in innate disease resistance at the sub-species level have major implications for wildlife management. Two subspecies of white-tailed deer, Odocoileus virginianus borealis and O. virginianus texanus were infected with epizootic hemorrhagic disease (EHD) viruses. These viruses are highly virulent pathogens of white-tailed deer and are endemic within the range of O. virginianus texanus but not within the range of O. virginianus borealis. Two experimental infections were performed. Five O. virginianus texanus fawns and five O. virginianus borealis fawns were infected with 10(7.1) median tissue culture infective doses (TCID50) of EHD virus, serotype 1 and five of each subspecies were infected with 10(7.1) TCID50 of EHD virus, serotype 2. Infections with both EHD virus serotypes caused severe clinical disease and mortality in O. virginianus borealis fawns, whereas disease was mild or nondetectable in O. virginianus texanus fawns. Virus titers and humoral immune response were similar in both subspecies suggesting that differences in innate disease resistance explain the differences seen in clinical disease severity. In white-tailed deer, innate disease resistance may vary at the subspecies level. Should this phenomenon occur in other species, these findings have major implications for managing wildlife populations, both endangered and non-endangered, using tools such as translocation and captive propagation.     

Culicoides, the vector of epizootic hemorrhagic disease in white-tailed deer in Kentucky in 1971.

The biting gnat, Culicoides variipennis (Coquillett), was shown to be a vector of epizootic hemorrhagic disease (EHD) in white-tailed deer, Odocoileus virginianus, in Kentucky because of virus isolations from parous females. Epidemiological evidence showed a close relationship of this vector to the animal host during an outbreak of EHD in penned deer. Larval breeding sites of C. variipennis were found and C. variipennis was the most abundant biting fly present during the outbreak. Females of C. variipennis were commonly observed biting deer, swine, cattle and, occasionally, man.     

Serosurvey for selected disease agents in white-tailed deer from Mexico

Serum samples from 350 white-tailed deer (Odocoileus virginianus texanus) collected in March 1994 from northeastern Mexico were tested for the prevalence of antibody activity against five infectious diseases of ruminants. The prevalence rate was 81% for bluetongue virus (BTV) of all serotypes, 72% for epizootic hemorrhagic disease virus (EHDV), 3% for Borrelia burgdorferi, 69% for Anaplasma marginale, and 0% for Brucella abortus, B. melitensis, and B. ovis. These are diseases that affect domestic ruminants, and deer may act as a reservoir of infection. In addition, if deer are translocated, they may introduce pathogens to formerly disease-free areas. The high seroprevalence of BTV and EHDV cannot be related to the presence of hemorrhagic disease in the deer in this region. This is the first report to indicate the presence of B. burgdorferi infection of deer in Mexico. Despite the high prevalence of A. marginale titers, it is uncertain that deer play a role in the epizootiology of cattle anaplasmosis in the region. Apparently, white-tailed deer are unimportant in the epizootiology of brucellosis of both cattle and goats in northeastern Mexico.     

Serological studies on sympatric Barbary sheep and mule deer from Palo Duro Canyon, Texas.

Sera were collected from 12 Barbary sheet (Ammotragus lervia) and 11 mule deer (Odocoileus hemionus) occupying sympatric ranges in Palo Duro Canyon, Texas. These were tested for leptospirosis, brucellosis, bovine virus diarrhea, anaplasmosis, vesicular stomatitis, bluetongue (BT), epizootic hemorrhagic disease (EHD), infectious bovine rhinotracheitis (IBR), and coccidioidomycosis. Serologic reactors were found to IBR in 3 Barbery sheep, BT in 6 Barbary sheep and 6 mule deer and EHD in 3 Barbary sheep and 4 mule deer. Possible ramifications of evidence for these diseases in wild herbivore populations in this area are discussed.