Anaplasma phagocytophilum infection in a fallow deer (Dama dama) population in a preserve of central Italy

From autumn 2004 to spring 2005, 70 fallow deer (Dama dama), 27 female and 43 male, living in a natural preserve of central Italy were examined by indirect immunofluorescence assay (IFA) to detect specific antibodies to Anaplasma phagocytophilum. Thirty-one (44.28%) sera scored positive: in particular 10 fallow deer (8 male and 2 female) scored positive at 40 antibody titer, 21 deer (8 male and 13 female) at > or = 80 titer. EDTA anticoagulated blood samples collected from 29 of the 70 deer examined were tested by a nested-PCR assay to disclose a 546 bp fragment, specific of A. phagocytophilum 16S rRNA gene. Twenty (72.41%) blood samples (8 male and 13 female deer) resulted positive. Fifteen PCR-positive deer also resulted positive to IFA, whereas the remaining six did not show specific antibodies. Three serologically positive animals gave negative results at the nested PCR. Five deer scored negative both to serological tests and PCR.     

Epidemiology of leptospirosis: observations on serological data obtained by a "diagnostic laboratory for leptospirosis" from 1995 to 2001

Serological data on leptospira infection were reported and discussed. From 1995 to 2001, the blood serum samples of 9885 domestic and wild animals and humans, living in Northern and Central Italy, were examined by the macroagglutination test (MAT) employing bratislava, ballum, canicola, grippotyphosa, icterohaemorrhagiae, pomona, hardjo and tarassovi serovars as antigens. Considering sera with > or = 1:400 antibody titers as positive, 674 (6.81%) animals scored positive. Sheep, horses, pigs and dogs gave the highest number of positive responses, particularly against the serovar bratislava and, for dogs, against icterohaemorrhagiae. The percentages of seropositivity observed in the most important animal species were: 12.13% in ovine (132 positive among 1088 tested animals), 11.40% in horses (107 positive animals among 938), 9.46% in swine (123 positive animals among 1299), 6.36% in dogs (278 positive animals among 4369), 2.39% in wild boars (11 positive animals among 459), 1.39% in deer (2 positive animals among 143), 0.48% in cattle (3 positive animals among 626). Among 250 human sera examined, 14 (5.60%) scored positive.     

Serologic survey for antibodies against Mycobacterium a vium subsp. paratuberculosis in free-ranging cervids from Norway

Affinity between protein-G and immunoglobulins from red deer (Cervus elaphus), moose (Alces alces), roe deer (Capreolus capreolus), and reindeer (Rangifer tarandus tarandus) was tested in a competition binding assay. Sera from red deer, reindeer, and moose inhibited the assay less than sera from cattle (less affinity), whereas sera from roe deer showed a slightly higher affinity to protein-G than did sera from cattle. The conclusion was made that protein-G could be used instead of anti-species antibodies for these cervid species, where the aim of the screening was to look for exposure or lack of exposure to mycobacteria in the tested populations. Serologic screening of 1,373 free-ranging cervids for antibodies against Mycobacterium avium subsp. paratuberculosis was conducted. All sera were tested by a protein-G-based antigen-absorbed enzyme-linked immunosorbent assay (ELISA). Seropositive moose (10/537; 1.9%), red deer (14/371; 3.8%), roe deer (6/49; 12.2%), and semidomesticated reindeer (11/325; 3.4%) were found, whereas wild reindeer (n = 91) were seronegative. In addition, the red deer sera were tested with a commercial ELISA, by which two animals tested positive and nine were suspicious of having M. avium subsp. paratuberculosis antibodies. Tissue samples and feces from 10 moose originating from a population with a clustering of seropositive animals were investigated by histology and bacteriology with negative results. Paratuberculosis has never been diagnosed in free-ranging or farmed cervid species in Norway. Thus, further studies are indicated to prove that the present findings reflect an infection with M. avium subsp. paratuberculosis.     

Prevalence of Neospora caninum and Toxoplasma gondii antibodies in wild ruminants from the countryside or captivity in the Czech Republic

In the Czech Republic, sera from 720 wild ruminants were examined for antibodies to Neospora caninum by screening competitive-inhibition enzyme-linked immunosorbent assay and confirmed by indirect fluorescence antibody test (IFAT); the same sera were also examined for antibodies to Toxoplasma gondii by IFAT. Neospora caninum antibodies were found in 14% (11 positive/79 tested) roe deer (Capreolus capreolus), 14% (2/14) sika deer (Cervus nippon), 6% (24/ 377) red deer (Cervus elaphus), 1% (2/143) fallow deer (Dama dama), 3% (3/105) mouflon (Ovis musimon), and none of 2 reindeer (Rangifer tarandus). Toxoplasma gondii antibodies were found in 50% (7/14) sika deer, 45% (169/377) red deer, 24% (19/79) roe deer, 17% (24/143) fallow deer, 9% (9/105) mouflon, and 1 of 2 reindeer. In 42 samples of wild ruminants that tested positive for N. caninum antibodies, 28 (67% of the positive N. caninum samples) reacted solely to N. caninum. This is the first evidence of N. caninum infection in mouflon, the first N. caninum seroprevalence study in farmed red deer, and the first survey of N. caninum in wild ruminants from the Czech Republic.     

Molecular survey of Anaplasma phagocytophilum and Ehrlichia canis in red foxes (Vulpes vulpes) from central Italy

During the 2007-2008 hunting season, 150 spleen samples were collected from free-ranging red foxes (Vulpes vulpes) in central Italy. The specimens were tested by two nested PCR assays to detect DNA of Anaplasma phagocytophilum, etiologic agent of granulocytic ehrlichiosis of animals and humans, and DNA of Ehrlichia canis, which causes the monocytic ehrlichiosis in canids. None of the foxes were PCR-positive for E. canis; 25 (16.6%) were positive for A. phagocytophilum. No specific gross alterations were detected at necropsy, and no histopathologic lesions found on PCR-positive spleen samples.     

Bovine virus diarrhea and the vector-borne diseases Anaplasmosis and Bluetongue: a sero-surveillance in free-ranging red deer (<Emphasis Type="Italic">Cervus elaphus</Emphasis>) in selected areas of Switzerland

Due to climate changes, diseases emerging from southerly adjacent areas (Mediterranean countries) are likely to spread northward. Expanded migration of red deer harbors the risk of introducing new pathogens into a naive population of either wild or domestic animals. Little is known about the importance of red deer as a potential reservoir for diseases of domestic ruminants in Switzerland. Deer is susceptible for all three agents that were selected in this study: bovine virus diarrhea virus (BVDV), Anaplasma marginale (AM), and Bluetongue virus (BTV). The goal of this project was to establish the serological status of red deer in Switzerland concerning BVDV, AM, and BTV, and to assess the possible impact of disease dynamics with a focus on potential transmission of these diseases from red deer to cattle or vice versa. Sampling areas were selected according the following criteria: abundance of red deer, potential insect vector distribution due to climatic conditions, and traditional alpine pasture husbandry along with known migration routes of red deer. Blood samples were collected during the regular hunting season 2004 and 2005 by hunters and gamekeepers. There was no serological evidence for the presence of the vector-borne diseases AM and BT in red deer in Switzerland. Four out of 234 sera showed a positive result for BVD, corresponding to a sero-prevalence of 1.7% (95% CI 0.46–4.38). Facing the fact of the high sero-prevalence for BVD in Swiss cattle (60–80%) disease transmission from red deer to cattle in these areas under investigation is rather unlikely.     

A serologic survey of mule deer and elk in Utah.

Sera from mule deer (Odocoileus hemionus) and elk (Cervus canadensis) in central and northern Utah were tested for the prevalence of antibodies to 11 diseases communicable to man or domestic livestock. Antibodies to Francisella tularensis (at 1:20) were found in 47 of 88 (53.4%) elk and 1 of 89 (1.1%) deer. A screening slide agglutination test for titers to Brucella (at 1:20) showed two reactors in elk but none in deer sera. No positive antibody titers were obtained in tests for anaplasmosis, Colorado tick fever, Rocky Mountain spotted fever, Q-fever, psittacosis, Powassan, western equine encephalitis, St. Louis encephalitis and California encephalitis.     

Transmission of Neospora caninum between wild and domestic animals

To determine whether deer can transmit Neospora caninum, brains of naturally infected white-tailed deer (Odocoileus virginianus) were fed to 4 dogs; 2 of these dogs shed oocysts. Oocysts from 1 of the dogs were tested by polymerase chain reaction and found to be positive for N. caninum and negative for Hammondia heydorni. The internal transcribed spacer 1 sequence of the new strain (designated NC-deer1) was identical to N. caninum from domestic animals, indicating that N. caninum is transmitted between wild and domestic animals, often enough to prevent divergent evolution of isolated populations of the parasite. NC-deerl oocysts were administered to a calf that developed a high antibody titer, providing evidence that N. caninum from wildlife can infect cattle. In addition, N. caninum antibody seroprevalence was detected in 64/164 (39%) free-ranging gray wolves (Canis lupus), 12/113 (11%) coyotes (Canis latrans), 50/193 (26%) white-tailed deer, and 8/61 (13%) moose (Alces alces). These data are consistent with a sylvatic transmission cycle of N. caninum between cervids and canids. We speculate that hunting by humans favors the transmission of N. caninum from deer to canids, because deer carcasses are usually eviscerated in the field. Infection of canids in turn increases the risk of transmitting the parasite to domestic livestock.     

Prevalence of Leptospira and Brucella antibodies in wild boars (Sus scrofa) in Tuscany, Italy

Five hundred sixty-two blood samples were collected from wild boars (Sus scrofa) shot in six districts of Tuscany, central Italy, between 1997 and 2000. Sera were examined for antibodies specific for Leptospira interrogans by microagglutination test and Brucella spp. by the Rose Bengal test and indirect enzyme-linked immunosorbent assay. Thirty-four (6.0%) samples tested positive for anti-Leptospira antibodies, 29 (5.1%) sera were positive for anti-L. interrogans serovar bratislava antibodies (titres ranging from 1:100-1:400), and 5 (0.9%) sera were positive for anti-L. interrogans serovar icterohaemorrhagiae antibodies (titres 1:100). All the examined sera were negative for anti-Brucella antibodies.     

Survey of Pestivirus infection in wild and domestic ungulates from south-western Italian Alps

The transmission of pestiviruses between domestic and wild ruminants has not been documented in communal alpine pastures shared between wildlife and livestock. The aim of this study was to investigate the role of domestic and wild ungulates species from Varaita Valley (SW Italian Alps) in the epidemiology of Pestivirus infections. Sera from free-ranging alpine chamois (Rupicapra rupicapra) and roe deer (Capreolus capreolus) were collected from 1994 to 2009 and 2001 to 2009, respectively. Also, sera from cattle and sheep sampled in 2009 were studied. Sera were tested for the presence of antibodies against pestivirus with an ELISA assay. Sera from positive animals were subsequently tested with a comparative virus neutralisation test using the BVDV-NADL and BDV-137/4 strains. Sera were tested for the presence of pestiviral antigen and the presence of viral RNA with a commercial ELISA assay and RT-PCR. Antibodies against Pestivirus were detected in 132 out of 312 (42%) chamois, in 30 out of 175 (17%) cattle and 6 out of 24 (25%) sheep. No antibodies were found in roe deer. No Pestivirus antigen or RNA was detected in any of the samples. Results indicate circulation of pestiviruses among the studied chamois, cattle and sheep populations. However the role of wild ungulates in the dynamics of Pestivirus infection is still unknown and monitoring the presence of these viruses in wild ungulates would be of importance, especially in the chamois population, where pestiviruses seem to circulate extensively.     

Ethiological agents of rickettsiosis and anaplasmosis in ticks collected in Emilia-Romagna region (Italy) during 2008 and 2009

Ticks are the main vectors of rickettsiae of the spotted fever group, as well as of a variety of other Rickettsiales, including bacteria of the genus Anaplasma, that might cause diseases in humans and animals. Here we present the result of a survey for ticks and for tick-associated Rickettsiales in the Emilia Romagna region (Northern Italy). The study was focused on ticks collected from wild-hunted animals. Out of 392 ticks collected from these animals, 282 (72%) were identified as Ixodes ricinus, 110 (28%) as Dermacentor marginatus. The former was found on four vertebrate species, whereas the latter appeared more specific for wild boar. The presence of rickettsiae was demonstrated in 22.5% of I. ricinus (57/253) and in 29% of D. marginatus (32/110). Five ticks of the species I. ricinus were also positive for Anaplasma phagocytophilum (2%). In addition, we collected ticks by dragging in a natural park of the same region. All of the ticks captured by dragging were identified as I. ricinus. Thirty-six out of 200 analyzed ticks proved positive for Rickettsia monacensis and R. helvetica (16.5 and 1.5%, respectively). Our results highlight that that ticks present in wild areas, widely exploited for recreation and hunting in Emilia-Romagna, represent a risk for the transmission of spotted fevers and anaplasmosis to humans.     

Factors driving the abundance of ixodes ricinus ticks and the prevalence of zoonotic I. ricinus-borne pathogens in natural foci

Environmental factors may drive tick ecology and therefore tick-borne pathogen (TBP) epidemiology, which determines the risk to animals and humans of becoming infected by TBPs. For this reason, the aim of this study was to analyze the influence of environmental factors on the abundance of immature-stage Ixodes ricinus ticks and on the prevalence of two zoonotic I. ricinus-borne pathogens in natural foci of endemicity. I. ricinus abundance was measured at nine sites in the northern Iberian Peninsula by dragging the vegetation with a cotton flannelette, and ungulate abundance was measured by means of dung counts. In addition to ungulate abundance, data on variables related to spatial location, climate, and soil were gathered from the study sites. I. ricinus adults, nymphs, and larvae were collected from the vegetation, and a representative subsample of I. ricinus nymphs from each study site was analyzed by PCR for the detection of Borrelia burgdorferi sensu lato and Anaplasma phagocytophilum DNA. Mean prevalences of these pathogens were 4.0% ± 1.8% and 20.5% ± 3.7%, respectively. Statistical analyses confirmed the influence of spatial factors, climate, and ungulate abundance on I. ricinus larva abundance, while nymph abundance was related only to climate. Interestingly, cattle abundance rather than deer abundance was the main driver of B. burgdorferi sensu lato and A. phagocytophilum prevalence in I. ricinus nymphs in the study sites, where both domestic and wild ungulates coexist. The increasing abundance of cattle seems to increase the risk of other hosts becoming infected by A. phagocytophilum, while reducing the risk of being infected by B. burgdorferi sensu lato. Controlling ticks in cattle in areas where they coexist with wild ungulates would be more effective for TBP control than reducing ungulate abundance.     

Use of recombinant antigens of Borrelia burgdorferi and Anaplasma phagocytophilum in enzyme-linked immunosorbent assays to detect antibodies in white-tailed deer

Serum samples obtained from white-tailed deer (Odocoileus virginianus) in Connecticut (n=218) and South Carolina (n=20) (USA) during the period 1992-2002 were analyzed for antibodies to whole-cell or recombinant antigens (i.e., fusion proteins) of Borrelia burgdorferi sensu stricto and Anaplasma phagocytophilum, etiologic agents of Lyme borreliosis and granulocytic ehrlichiosis, respectively. In enzyme-linked immunosorbent assays (ELISAs) with whole-cell B. burgdorferi, the overall seropositivity rate for Connecticut (53%) exceeded that for South Carolina (30%). In separate tests of seven recombinant antigens of B. burgdorferi by an ELISA, seroprevalence for the VlsE antigen was highest (48%) in Connecticut followed by outer surface protein (OspF) (21%), whereas serum reactivities to the protein (p) 41-G antigen (55%) and VlsE (25%) were most frequent for South Carolina sera. In analyses for antibodies to the recombinant protein (p) 44 antigen of A. phagocytophilum, seroprevalences of 52% and 25% were recorded for Connecticut and South Carolina samples, respectively. These findings paralleled those determined by indirect fluorescent antibody staining methods with whole cells (43% and 30%). Moreover, there was good agreement (74%) in results of Western blot analyses and an ELISA when a subset of 39 sera was screened with whole-cell or recombinant p44 antigens of A. phagocytophilum. An ELISA with highly specific recombinant VlsE or p44 antigens can be used in conjunction with other antibody tests to determine whether deer living in different regions of eastern United States were exposed to B. burgdorferi or A. phagocytophilum.     

A novel Giardia duodenalis assemblage A subtype in fallow deer

The molecular identification of species and genotypes of Giardia spp. infecting wild mammals represents the most reliable tool to understand the role played by these animals as reservoirs of cysts infectious for human and other animals. Of 139 fecal samples collected from fallow deer (Dama dama L.) hunted in a Natural Reserve of northern Italy, the prevalence of Giardia sp. was 11.5% (16 of 139 animals), and it was higher in fawns than in older animals. Fragments of the betagiardin and triose phosphate isomerase (tpi) genes were successfully polymerase chain reaction amplified and sequenced from 8 isolates. No sequence variation was observed between isolates at the 2 genetic loci. Sequence and phylogenetic analyses identified a Giardia duodenalis subtype that clusters with assemblage A isolates and that shows homologies of 98 and 97% at the beta-giardin and tpi loci, respectively, compared with the A1 subtype. Because the G. duodenalis subtype found in fecal samples of fallow deer has never been detected previously, its role as a pathogen for humans and domestic animals is unknown, but, considering its genetic distinctiveness, it is likely to be low.     

Cilia-associated respiratory (CAR) bacillus infection in adult red deer, chamois, and roe deer

Cilia-associated respiratory (CAR) bacillus is an unclassified bacterium that colonizes the ciliated epithelium of airways in laboratory rats, laboratory mice, and laboratory and conventionally reared rabbits, cattle, goats, and pigs. Data on the prevalence of CAR bacillus infection in wild animals are lacking. The present study demonstrated the occurrence of the organism in wild red deer (Cervus elaphus hippelaphus), chamois (Rupicapra rupicapra), and roe deer (Capreolus capreolus) from the Val Fontana in northern Italy. Prevalence ranged from 26% for red deer to 56% for chamois, with a statistically significant negative correlation between CAR bacilli infection and the presence of lymphoid follicles.     

Seroepidemiology of chlamydial infections of wild ruminants in Spain

Chlamydial infections were determined serologically among wild ruminants in the Nature Park of the Sierras de Cazorla, Segura y Las Villas (CNP; Spain). Sampling was done during the period from 1990-95. There were 1,244 blood samples collected, consisting of 490 from fallow deer (Dama dama), 343 from mouflon (Ovis mussimon), 283 from red deer (Cervus elaphus) and 128 from Spanish ibex (Capra pyrenaica). Specific complement-fixing antibodies of Chlamydia spp. were detected by means of microtechnique, using lipopolysaccharide antigen. The relationship of biological (species, sex, age), temporal (year) and territorial (central and peripheral areas) factors to seropositive prevalence was examined, and preliminary data were collected on whether or not sheep and goat herds grazing in the peripheral areas of the park also were infected with Chlamydia spp. Chlamydiosis was common in the four species of wild ruminants in the CNP in all the years studied. The prevalence of Chlamydia sp. in mouflon (37%) was significantly greater than in fallow deer (30%), and both had a significantly higher prevalence rate than Spanish ibex and red deer (both 24%). The four species of wild ruminants were similar in that they act as reservoirs of Chlamydia spp., although their receptivity may be different, and the infection can certainly be maintained among these animals by intra-group transmission. The differences in prevalences and geometric mean titers (GMT), both between the sexes (male versus female) and between different ages (adult versus juvenile), were insignificant in all four species. For all species of wild ruminants both prevalence rates and GMTs were greater in populations occupying the peripheral areas of the park than in those inhabiting the central area. Herds of sheep and goats had a high prevalence of chlamydiosis. Intertransmission of Chlamydia sp. between wild and domestic ruminants occurred through grazing on the same pastures. The highest mean prevalence (44%) of patent infections (CFT titers of > or =1:80) was detected in red deer, although this frequency was not significantly different from those observed in mouflon (39%), Spanish ibex (38%), and fallow deer (37%). The proportion of patent infection was higher in females than in males, and none of the juveniles (<2-yr-old) showed patent infections. The prevalence of predicted patent chlamydial infections was always higher in the peripheral areas of the park, although only among mouflon and fallow deer were the differences statistically significant.     

Abomasal nematodes of the red deer Cervus elaphus in north-eastern Italy

The composition of the abomasal helminth fauna and parasite diversity were studied in 298 red deer collected during 1997-2000 from three different sectors (Bolzano, Trento and Sondrio provinces) of the Stelvio National Park, one of the main protected areas of north-eastern Italy. The association between parasite burdens and geographical areas of the hosts was assessed using the negative binomial regression. A variety of abomasal helminths, both host specific and generalist, was found in all sectors. The most commonly observed parasites were the Spiculopteragia spiculoptera morph spiculoptera and Ostertagia leptospicularis morph leptospicularis, with prevalences of 79.5% and 40.9%, respectively. The minor morphs S. spiculoptera morph mathevossiani (prevalence 31.9%) and O. leptospicularis morph kolchida (18.8%) occurred at lower prevalences. Teladorsagia circumcincta morph circumcincta, Marshallagia marshalli morph marshalli and Haemonchus contortus were rarer, at prevalences of 1, 1.3 and 1.3%, respectively. Deviance analysis of the negative binomial regression model shows that the geographical area is significantly related to parasite burdens (P = 0.001). Prevalences of hosts with parasites were greater in the Sondrio (odds ratio = 1.31; 95% confidence interval: 0.16-10.85) and smaller in the Trento (odds ratio = 0.62; 95% confidence interval: 0.20-1.96) sectors with respect to Bolzano, but these differences were not statistically significant. Possible cross-infections by more generalist parasites between wild and domestic animals were also suggested, as deer and domestic ruminants (Bovinae, Caprinae and Ovinae) used the same feeding areas of the park sectors.     

Neutralizing antirabies antibodies in urban terrestrial wildlife in Brazil

The prevalence of rabies neutralizing antibodies (NA) in sera of wild animals from S?o Paulo City (Brazil) was investigated using the Rapid Fluorescent Focus Inhibition Test between 1994 and 1997. Sera from 547 specimens were examined. Marsupials represented 45% of the sample and primates 37%; carnivores, rodents, deer and edentates represented 6, 6, 3 and 2%, respectively. The overall prevalence of NA was 14%. The prevalence of NA was 18% in primates; whereas in marsupials, carnivores, edentates and rodents it was 13, 9, 8 and 6%, respectively. The stratification according to sex, age, and site of capture of the marsupials and primates showed a small predominance in males versus females and a large predominance of adults versus juveniles. The same relationship was seen in specimens captured near human habitations versus specimens captured in their own habitat. It is evident that there is circulation of rabies virus in wild animals, which are not recommended as pets since they represent a potential risk of exposure to rabies virus for both humans and domestic animals.     

Porcine reproductive and respiratory syndrome (PRRS) virus in wild boar and Iberian pigs in south-central Spain

Porcine reproductive and respiratory syndrome (PRRS) is a swine infectious disease causing major economic problems on the intensive pig industry. This virus has been reported worldwide in domestic pigs and there is evidence of PRRS virus (PRRSV) infection in wild boar (Sus scrofa). Nonetheless, the epidemiological role of wild boar and extensively kept domestic pigs remains unclear. The aim of this study was to determine the occurrence of PRRS in wild boar and Iberian pigs in the dehesa ecosystem of the Castile-La Mancha region of Spain, which boasts one of the most important free-roaming porcine livestock and hunting industries in the country. Using geo-spatial analysis of literature data, we first explored the relationship between domestic pig density and PRRS occurrence in wild boar in Europe. Results revealed that PRRS occurrence in wild boar may be influenced, albeit not significantly, by domestic pig density. Next, we analyzed sera from 294 wild boar and 80 Iberian pigs by indirect enzyme-linked immunosorbent assay for PRRSV antibodies. The sera and 27 wild boar tissue samples were analyzed by two real-time RT-PCR assays, targeting the most conserved genes of the PRRSV genome, ORF1 and ORF7. Seven wild boar (2.4 %) and one Iberian pig (1.3 %) were seropositive, while none of the animals tested positive for PRRSV by RT-PCR. Our results confirm the limited spread of PRRSV in free-roaming Iberian pigs and wild boar living in mutual contact. Further studies would be necessary to address whether this low seroprevalence found in these animals reflects transmission from intensively kept pigs or the independent circulation of specific strains in free-roaming pigs.     

Epizootic vesicular stomatitis in Colorado, 1982: some observations on the possible role of wildlife populations in an enzootic maintenance cycle

Sera obtained from wild ungulates, carnivores, and rodents in Colorado were tested for neutralizing (N) antibody against vesicular stomatitis, New Jersey serotype (VSNJ), virus to determine their involvement in the 1982 Colorado VSNJ epizootic in domestic animals. Viremic and N antibody responses of two local rodent species to a 1982 Colorado isolate of VSNJ were determined in the laboratory. The rodents produced only weak viremias, but all developed N antibody. N antibody prevalences for VSNJ in sera from wild ungulates was sufficiently high to indicate their involvement during the epizootic. In addition, the demonstration of N antibody in elk (Cervus elaphus) and mule deer (Odocoileus hemionus) prior to the epizootic in cattle and horses suggests that an enzootic cycle may exist in Colorado.