Serologic and virologic investigations into pestivirus infection in wild and domestic ruminants in the Pyrenees (NE Spain)

An outbreak of disease associated to a border disease virus was described in the Southern chamois (Rupicapra pyrenaica) in Spain in 2002. Sera and/or spleen samples from 57 mouflon, 15 red deer, 21 roe deer, 3 fallow deer, 55 sheep, 32 cattle, and 68 goats sharing the chamois habitat were studied. An antibody ELISA test yielded an inconclusive result in 2 mouflon and positive results in 5 goat sera. Comparative virus neutralization tests were performed on the 2 inconclusive mouflons, 3 of the 5 seropositive goats, 55 sheep and 32 cattle, using 6 pestivirus strains. Positive results were obtained in 1 mouflon, 2 goats, 69% of sheep and 78% of cattle. Virological investigations performed with an antigen ELISA test yielded negative results in 21 goats and 39 mouflons, the result in 1 mouflon being inconclusive. PCR performed on 12 goats and the inconclusive mouflon gave negative results. These results suggested that it is unlikely that chamois BDV is infecting wild and domestic ruminants.     

Retrospective study of pestivirus infection in Pyrenean chamois (Rupicapra pyrenaica) and other ungulates in the Pyrenees (NE Spain)

In 2001 a new Pestivirus (Family Flaviviridae) was associated with an outbreak of a previously unreported disease in Pyrenean chamois (Rupicapra pyrenaica) in the Pyrenees (NE Spain). Molecular characterization assigned this virus to the Border Disease Virus (BDV) cluster, BDV-4 genotype. A retrospective study was performed in archived sera and spleen of 74 Pyrenean chamois and in archived sera of 28 mouflon (Ovis ammon), 56 red deer (Cervus elaphus), 43 roe deer (Capreolus capreolus) and 29 fallow deer (Dama dama) from the Pyrenees between the years 1990 and 2000. Thirty six of 74 (48.6%) sera of Pyrenean chamois, one of mouflon and one of red deer were positive by an ELISA antibody test. Comparative virus neutralization tests were performed on 26 seropositive chamois, one mouflon and one red deer, using five pestivirus strains. An ELISA antigen test was performed on 37 seronegative chamois and yielded positive results in one chamois and inconclusive result in two. RT-PCR and virus isolation performed on spleen samples from these three animals gave positive results in the positive and one inconclusive animal. Sequence analysis in the 5' unstranslated region revealed that they were grouped into the BDV-4 genotype. Virological and serological data of the present study indicate that BDV infection has been present in the chamois population since at least 1990, 11 years before the first outbreak of disease. Therefore, the emergence of the disease in 2001 is apparently due to other factors rather than the introduction of a new virus in the chamois population.     

Paratuberculosis in farmed and free-living wild ruminants in the Czech Republic (1999-2001)

Due to the occurrence of the infection of Mycobacterium avium subspecies paratuberculosis among domestic ruminants and the rapid development of farmed deer industry and the market of cloven-hoofed game we have carried surveys of paratuberculosis, beginning in 1997, in the most common four species of wild ruminants in the Czech Republic [Pavlik et al., Vet. Microbiol. 77 (2000) 231-251]. From 1999 the prevalence of paratuberculosis has been slightly reduced in all three types of husbandry of wild ruminants. Nevertheless paratuberculosis has been diagnosed in wild ruminants in three districts, in four game parks and in five farms. M. a. paratuberculosis was isolated from 128 (5.3%) out of 2,403 wild ruminants of four animal species: 106 red deer, 2 roe deer, 4 fallow deer and 16 mouflons. In red deer farms, the highest number of clinical paratuberculosis cases was in yearling deer. RFLP type B-C1 of M. a. paratuberculosis predominated during the second period (1999-2001) in all types of husbandry with no relationship to wild ruminant species. New "cattle" RFLP types B-C5 and B-C16 of M. a. paratuberculosis were described in infected farmed red deer and one "intermediate" RFLP type R-I4 in fallow deer from one game park. The survival of M. a. paratuberculosis was found to be 4 months during winter in the pasture after destocking of all cattle infected with paratuberculosis. We found that non-vertebrates, wild ruminants or non-ruminant wildlife can be vectors and potentially become a risk factor in the spread of M. a. paratuberculosis infection.     

Immunoglobulin G class identification from wild ungulates by cross-reactivity with antisera to domestic animals

Seven species of Spanish ungulates were tested for the presence of homologous immunoglobulin G (IgG) with a gel-diffusion test using bovine, ovine, caprine and porcine IgG antisera. Homologous ovine and caprine IgG were detected in sera from chamois (Rupicapra pyrenaica), Spanish ibex (Capra pyrenaica hispanica), mouflon (Ovis orientalis musimon), red deer (Cervus elaphus), fallow deer (Dama dama) and roe deer (Capreolus capreolus). Homologous porcine IgG was detected in wild boar (Sus scrofa) serum. Immunoelectrophoretic assays were performed to compare the electrophoretic mobility of IgG from domestic and wild species.     

Seroprevalence of pestivirus in four species of alpine wild ungulates in the High Valley of Susa, Italy

Wildlife, once infected, can serve as a reservoir of infectious diseases that form a constant threat to domestic livestock. To make control and eradication programs successful in the long-term, presence of pestivirus in wildlife populations should be monitored. The goal of this study was to investigate seroprevalence of pestivirus in four alpine wild ungulates in the High Valley of Susa, north-west Italy. Species studied were: red deer (Cervus elaphus), roe deer (Capreolus capreolus), wild boar (Sus scrofa) and chamois (Rupicapra rupicapra). A further goal was using virus neutralisation tests (VNT) for four strains of pestivirus in chamois and wild boar. Three hundred and seventy-five serum samples collected during the hunting season of 1999 were tested for pestivirus specific antibodies. Positive sera of chamois and wild boar were subsequently tested in a VNT with four major subtypes of pestivirus, and virus isolation was performed. No antibodies were found in the 73 samples of roe deer, while 7 (12.5%), 8 (5.9%) and 28 (25.5%) of 56, 136 and 110 samples of wild boar, red deer and chamois were ELISA-positive, respectively. Different ranges of titers were found in the VNT and no pestivirus was isolated in the ELISA-positive wild boar and chamois samples. Several possibilities, which might explain the high seroprevalence in chamois are discussed. Pestivirus antibodies were found in three out of four large alpine ungulates in the High Valley of Susa. Seroprevalence was particularly high in chamois. Further investigation is needed to characterise the pestiviruses that circulate in these animals.     

Seroepidemiologic studies of the distribution of respiratory syncytial virus in wild ruminants

Sera from wild ruminants (302 roe, 39 red deer, 17 fallow deer, 1 mouflon) were tested for antibodies against respiratory syncytial virus (RSV). 22 sera (6.1%) were positive: 20 roe, 1 red deer, 1 mouflon. The findings are discussed from the epizootiological point of view.     

Blood-feeding patterns of horse flies in the French Pyrenees

Horse flies can mechanically transmit Besnoitia besnoiti, the agent of bovine besnoitiosis. Although previously limited to enzootic areas, especially the French Pyrenees Mountains, bovine besnoitiosis is now considered a re-emerging disease in western Europe. To improve understanding of the role of horse flies as mechanical vectors, this study investigated their blood-feeding ecology in the eastern French Pyrenees, in two high-altitude summer pastures whose main domestic ungulates were cattle, and in a wildlife park with native fauna. Species-specific PCR assays were conducted to identify the sources of blood meals: wild boar, horse, cattle (or bison), sheep (or mouflon), goat, red deer, roe deer and izard (or Pyrenean chamois). In La Mouline pasture, tabanids (N = 20) fed on red deer (70%) and cattle (30%). In Mantet pasture, tabanids (N = 24) fed on cattle (52%), red deer (20%), wild boar (16%), horse (8%) and sheep (4%). In the wildlife park, Tabanus bromius (N = 32), the most abundant species collected, fed on red deer (85%), bison (9%) and wild boar (6%). Despite relatively high densities in both the pastures and in the wildlife park, small wild ungulates (izard, mouflon and roe deer) were not detected as a source of blood meals. Only two mixed blood meals were identified in two specimens of T. bromius: cattle/horse for the specimen collected in the pastures, and bison/wild boar for the specimen collected in the wildlife park. Our findings showed that tabanids display a level of opportunistic feeding behaviour, in addition to a preference for red deer, the latter being particularly true for Philipomyia aprica, the most abundant species collected in the pastures.     

The emergence of Mycobacterium paratuberculosis in farmed deer in New Zealand - a review of 619 cases

AIM: To review cases in which Mycobacterium paratuberculosis was identified in farmed deer in New Zealand. METHODS: Case histories were reviewed where M. paratuberculosis was identified in deer by either culture or a polymerase chain reaction (PCR) test using primers from IS900. RESULTS: Between 1986 and 2000, M. paratuberculosis was identified by bacterial culture and/or PCR in 619 farmed deer from 299 herds, representing approximately 6% of deer herds in New Zealand. Over 85% of cases were identified during the last 6 years. In 60% of the infected herds, only one infected animal was identified. The maximum number of cases identified in a single deer herd was 47, and these were identified over a period of 8 years. Only 36 (5.8%) cases came from clinically affected animals identified on farms by veterinarians. The majority (89.7%) of the 619 cases were identified from lesions in mesenteric lymph nodes, including the ileocaecal lymph nodes, identified at meat inspection as being macroscopically either typical or equivocal of bovine tuberculosis (M. bovis). While the overwhelming majority of lesions were identified in mesenteric lymph nodes, M. paratuberculosis was also identified in 27 lesions in lymph nodes of the head, especially the retropharyngeal lymph node. CONCLUSIONS: The figures presented underestimate the true prevalence of infection with M. paratuberculosis, especially since not all suspect cases were submitted for culture or PCR. However, they do show that M. paratuberculosis appears to be spreading in farmed deer in New Zealand and highlight the possibility that Johne's disease is emerging as a potential major problem affecting this species. Identification of the organism by bacterial culture or PCR is required in many cases to distinguish lesions in mesenteric lymph nodes and lymph nodes of the head caused by M. paratuberculosis from those caused by M. bovis and M. avium.     

Liver microsomal biotransformation of albendazole in deer, cattle, sheep and pig and some related wild breeds

Albendazole (ABZ) biotransformation was studied in vitro in liver microsomes of adult noncastrated male farm animals (ram, buck, bull and boar), castrated adult males (wether, billy and hog), and free living males (fallow buck, red deer stag, mouflon ram, roe buck and wild boar). Liver microsomal fractions were incubated with either ABZ or racemic albendazole sulphoxide (ABZSO). ABZ was extensively metabolized to the (+) and (-) enantiomers of ABZSO, whereas ABZSO underwent a slow oxidation to albendazole sulphone (ABZSO2) in all species. In all species both ABZSO enantiomers were detected. The chiral ratio, (+)-ABZSO/(-)-ABZSO, was greater than one in farm animals, mouflon and wild boar, and less than one in three species of deer. For total ABZ sulphoxidation, deer like species had lower values compared to the other species. Mouflon ram and ram had lower total sulphoxidation rates compared to wethers, as well as ABZ suphoxidation towards (+)-ABZSO. No significant difference occurred comparing ABZSO formation in mouflon ram and ram, but ABZSO2 formation rate in mouflon ram was higher than in rams and wethers. Roe deer stag, fallow buck and red deer stag did not differ in both total-ABZSO and (-)-ABZSO synthesis rates and roe deer stag and fallow buck did not differ in synthesis rates of (+)-ABZSO and ABZSO2. The bull differed from other species in all metabolites studied, except for red deer stag and boar in (-)-ABZSO synthesis rate. The extent of ABZSO sulphonation to ABZSO2 in bull microsomes was more than twice that of other species.     

Shedding of foodborne pathogens and microbial carcass contamination of hunted wild ruminants

To assess the shedding of selected bacterial foodborne pathogens, fecal samples from 239 hunted wild red deer, roe deer, chamois, and ibex were examined. All samples tested negative for Salmonella spp. and L. monocytogenes, but other Listeria species were occasionally found. Of the 239 fecal samples, 32.6% tested positive for stx (Shiga toxins), 6.7% for eae (intimin) and 13.8% for both stx and eae genes. Among the 56 isolated Shiga toxin-producing Escherichia coli (STEC) strains, 44.6% harbored genes for the Stx2 group, 30.4% for the Stx1 group, and 21.4% for both Stx1 and Stx2. Only two of these strains harbored eae. Hence, wild ruminants constitute a reservoir for STEC, but further characterization data of the isolated strains are required to assess their actual human pathogenicity. In addition, 328 carcasses from hunted wild red deer, roe deer, and chamois were examined for total viable counts (TVC) and Enterobacteriaceae by swabbing. For the examined animal species, average TVC (4.0-4.2 log CFU cm(-2)) and average Enterobacteriaceae counts/detection rates (2.3-2.6 log CFU cm(-2); 87.5-90%) were at comparable levels. On the other hand, the microbial status of carcasses differed between certain abattoirs by several orders of magnitude. Strict compliance with good hunting and hygiene practices during any step from shooting, through evisceration in the field, to dehiding, cooling, and processing is therefore of central importance to avoid contaminations and to prevent foodborne pathogens carried by the animals from entering the food chain.     

Epidemiology of paratuberculosis in wild ruminants studied by restriction fragment length polymorphism in the Czech Republic during the period 1995-1998

In two studies carried out during the period 1995-1998, paratuberculosis was diagnosed in domestic and wild ruminants in the Czech Republic. The isolated Mycobacterium avium subspecies paratuberculosis strains were analysed by standardised restriction fragment length polymorphism (RFLP) [Pavlik, I., Horvathova, A., Dvorska, L., Bartl, J., Svastova, P., du Maine, R., Rychlik, I., 1999. J. Microbiol. Methods 38, 155-167]. In December 1992, 19 late pregnant Charolais heifers were imported to the Czech Republic from Hungary (original import from France to Hungary). One 11-month-old heifer roamed in the wild in a range of approximately 15-20km for 7 months from November 1993 to May 1994. Upon capture, the animal showed clinical signs of paratuberculosis (emaciation and diarrhoea). Seven other animals from the same herd were infected with the identical RFLP type B-C1 of M. paratuberculosis. During the period 1995-1996, samples were taken and examined from the small intestine and corresponding lymph nodes of 84 wild ruminants: 19 red deers (Cervus elaphus) and 65 roe-deers (Capreolus capreolus). These wild ruminants originated from 44 different locations within the same district from as the infected escaped heifer. Five M. paratuberculosis strains were isolated: one strain of RFLP type B-C1 from a stag and three strains of RFLP type B-C1 and one strain of RFLP type B-C9 from roe-deer. The three wild ruminants (one stag and two roe-deer) infected with the same RFLP type B-C1 were detected in the same area as the heifer, suggesting that this was the likely infection source. However, the infection source of the roe-deer infected with strain of RFLP type B-C9 was obviously different, and the stags that escaped from the farm were purchased from an area infected with this RFLP type. In the second study carried out during 1997-1998 in the whole Czech Republic (divided into 76 districts), 718 wild ruminants were examined from 90% of the districts. M. paratuberculosis was isolated from 25 (3.5%) animals from the wild, from farms and from game parks: 7.1% of 132 red deers, 1.5% of 336 roe-deers, 3.9% of 178 fallow deers (Dama dama), and 4.2% of 48 moufflons (Ovis musimon). This study discovered three RFLP types (B-C1, D-C12 and M-C16). A surprising finding was that of M. paratuberculosis (RFLP type B-C1) infection in roe-deer and a fallow deer in their natural habitat. The infection source was determined to have originated from two imported Holstein and Limousine cattle herds infected with the same strain. In the case of a mother and daughter roe-deer infected with RFLP type M-C16 and a fallow deer infected with RFLP type D-C12, all roaming in their natural habitat, the infection source was not discovered. The highest incidence of clinically ill wild ruminants was found in farmed red deer, and no relationship was found between the RFLP type or ruminant species and clinical status of animal.     

Mycobacterial diseases of deer

The most significant mycobacterial diseases of free-living, captive and farmed deer are bovine tuberculosis, caused by Mycobacterium bovis, Johne's disease (paratuberculosis), caused by Mycobacterium avium subsp paratuberculosis (basonym M. paratuberculosis), and avian tuberculosis, caused principally by M. avium subsp avium. The first case of M. bovis infection in farmed deer was identified in New Zealand in 1978. In 1983, a voluntary scheme was introduced in New Zealand to control tuberculosis in farmed deer, followed by a compulsory tuberculosis control scheme in 1990. The primary control measure is the slaughter of infected animals, detected by skin testing and blood testing, together with movement control and vector control. The number of infected deer herds peaked in the mid 1990s at over 160 herds, but by 30 June 2002 this had been reduced to 79 (1.45%), and to 67 (1.23%) by June 2003. Deer-to-deer transmission occurs, but the majority of herd breakdowns are believed to be from infected vectors. Factors likely to affect the susceptibility of deer include age, environment, population density, exposure and genetics. Avian tuberculosis occasionally causes clinical disease in wild, captive and farmed deer in New Zealand and overseas. Mycobacterium intracellulare, and subspecies of M. avium other than M. paratuberculosis, are widespread throughout New Zealand and are thought to be largely responsible for the high level of sensitisation to avian purified protein derivative (PPD), which is used for comparison purposes in tuberculosis skin testing of deer in this country. Infections with these organisms are usually subclinical in farmed deer, although M. avium subsp avium commonly causes lesions in retropharyngeal, mesenteric and ileocaecal lymph nodes. These lesions cause problems because of their gross and microscopic similarity to those due to M. bovis infection. Birds and domestic animals are most likely to become infected via environmental contamination of food, water, bedding litter or soil, while carnivores or scavengers may also become infected by ingesting infected carcasses. Johne's disease has been reported in deer in the wild and in zoos, especially in North America, the United Kingdom (UK) and Europe. Since first being confirmed in farmed deer in New Zealand in 1979, the incidence of Johne's disease has increased steadily. To date, M. paratuberculosis has been identified in >600 farmed deer on 300 properties. The majority of cases have been identified from suspected tuberculous lesions submitted from deer slaughter plants. Clinically, Johne's disease in deer is similar to the disease in sheep and cattle, with typical signs of loss of weight and condition, and diarrhoea. However, outbreaks of Johne's disease frequently occur in young red deer, 8-15 months of age, whereas the clinical disease in sheep and cattle is sporadic and usually affects adults 3-5 years of age. The disease is characterised by a chronic granulomatous enteritis and lymphadenitis, especially affecting the jejunum and ileum and the mesenteric lymph nodes. Deer affected subclinically may have lesions in these lymph nodes at slaughter, which are grossly indistinguishable from those due to bovine tuberculosis. Because of the antigenic similarity between M. intracellulare and all the subspecies of M. avium, including M. paratuberculosis, the diagnostic tests for Johne's disease lack sensitivity and specificity, making control difficult.     

Seroprevalence of Neospora caninum in non-carnivorous wildlife from Spain

Serum samples from 1034 non-carnivorous wildlife from Spain were tested for antibodies to Neospora caninum by competitive screening enzyme linked immunosorbent assay (ELISA) and confirmed by an indirect fluorescent antibody test (IFAT). High agreement was observed between results in both techniques (kappa value higher than 0.9). Prevalences of N. caninum antibodies positive by both techniques were 11.8% of 237 red deer (Cervus elaphus), 7.7% of 13 barbary sheep (Ammotragus lervia), 6.1% of 33 roe deer (Capreolus capreolus) and 0.3% of 298 wild boar (Sus scrofa). In one of 53 hares (Lepus granatensis), antibodies were found in the ELISA but could not be confirmed by IFAT due to lack of sample. Antibodies to N. caninum were not found in any of 251 wild rabbits (Oryctolagus cuniculus), 79 fallow deer (Dama dama), 27 mouflon (Ovis ammon), 40 chamois (Rupicapra pyrenaica) and three Spanish ibex (Capra pyrenaica). Statistically significant differences were observed between N. caninum seroprevalence in red deer and management of hunting estates (open versus fenced) with higher prevalence in fenced estates, and among sampling sites. Seroprevalence was particularly high in some areas (MO estate in South-Central Spain or some estates of Catalonia, North-East Spain), while no contact with N. caninum was observed in others. Results indicate that in certain areas of Spain, N. caninum is present in wildlife, especially in red deer. These results have important implications in both sylvatic cycles and may influence the prevalence of infection in cattle farms in those areas. To our knowledge, this is the first report of antibodies to N. caninum in wildlife from Spain and the first report of N. caninum antibodies in barbary sheep and wild boar.     

Factors affecting the herd level of antibodies against Mycobacterium avium subspecies paratuberculosis in dairy cattle

A case-control study was made of Norwegian dairy herds with high and low herd levels of antibodies against Mycobacterium avium subspecies paratuberculosis. A high proportion of the herds had a considerable number of seropositive cows, and environmental and management factors were examined for possible associations with the high serological levels of antibodies. The most important appeared to be: geographical location, red deer (Cervus elaphus) gaining access to the pastures for cattle, the observation of wild birds in the feed storage, and herds sharing common pasture with other herds of cattle. However, diagnostic tests showed that none of the animals in the case herds was infected with M a paratuberculosis.     

Immunoglobulin G Class Identification from Wild Ungulates by Cross‐reactivity with Antisera to Domestic Animals

Seven species of Spanish ungulates were tested for the presence of homologous immunoglobulin G (IgG) with a gel‐diffusion test using bovine, ovine, caprine and porcine IgG antisera. Homologous ovine and caprine IgG were detected in sera from chamois (Rupicapra pyrenaica), Spanish ibex (Capra pyrenaica hispanica), mouflon (Ovis orientalis musimon), red deer (Cervus elaphus), fallow deer (Dama dama) and roe deer (Capreolus capreolus). Homologous porcine IgG was detected in wild boar (Sus scrofa) serum. Immunoelectrophoretic assays were performed to compare the electrophoretic mobility of IgG from domestic and wild species.     

Prevalence of Toxoplasma gondii antibodies in red deer (Cervus elaphus) and other wild ruminants from Spain

Serum samples from 441 red deer (Cervus elaphus) and 161 other wild ruminant species, collected between 1993 and 2005 from six regions of Spain were tested for antibodies against Toxoplasma gondii by the modified agglutination test (MAT). Antibodies to T. gondii (MAT 1:25 or higher) were detected in 15.6% of red deer. Statistically significant differences were observed among sampling sites with seroprevalence in red deer from Catalonia (42.2%) being significantly higher compared with other Spanish regions (8.7%) (P < 0.05). Statistically significant differences were not observed between T. gondii seroprevalence and sex, age or management of hunting estates (open versus fenced). Seroprevalence of T.gondii infection in other ruminants species was 24% of 79 fallow deer (Dama dama), 21.8% of 33 in roe deer (Capreolus capreolus), 33.3% of three Spanish ibex (Capra pyrenaica), 20% of 10 chamois (Rupicapra pyrenaica), 10% of 10 barbary sheep (Ammotragus lervia), and 14.8% of 27 mouflon (Ovis ammon) in areas not including Catalonia, where no samples from these species were available. Serological results indicated a widespread exposure to T. gondii among wildlife in Spain and suggest that consumption of raw or inadequately cooked meat, as well as handling carcasses of wild game, should be taken into account as a source of infection for humans.     

Prevalence of Coxiella burnetti infection in wild and farmed ungulates

The aim of this study was to evaluate by serology and PCR analyses the prevalence of Coxiella burnetti infection in ungulates in Spain. Sera were collected from red deer (Cervus elaphus; n=116), roe deer (Capreolus capreolus; n=39), fallow deer (Dama dama; n=13) and cattle (n=79). Sera were tested for anti-C. burnetii antibody detection by means of an immunofluorescence antibody assay (IFA) and C. burnetii DNA was amplified by PCR in samples from ungulates that had antibodies to phase II antigens. Twenty-nine, 15 and 39 percent of the red deer, roe deer and cattle had antibodies against C. burnetii, respectively. None of the fallow deer sera tested positive. Seroprevalence was statistically higher in farmed than in wild red deer and higher in northern than in southern populations, whereas an inverse pattern was observed for the roe deer. Most of the seropositive animals had only anti-C. burnetii phase II antibodies, thus showing the acute nature of infections in the sampled ungulates. These results show that C. burnetii circulates in wild ungulates in Spain and suggest that they can act as pathogen reservoirs for both domestic animals and humans.     

Use of a polymerase chain reaction to subtype Mycobacterium avium subspecies paratuberculosis, an increasingly important pathogen from farmed deer in New Zealand

AIMS: To review the number of microbiologically-confirmed cases of Johne's disease in farmed deer since 2000, and determine the prevalence of the bovine and ovine subtypes of Mycobacterium avium subsp paratuberculosis (M. paratuberculosis), using a highly specific polymerase chain reaction (PCR) test on samples from infected herds. METHODS: The number of cases of M. paratuberculosis in farmed deer identified by culture or IS900 PCR was documented. A highly specific PCR test was applied to subtype M. paratuberculosis from BACTEC 12B cultures selected on the basis of one culture per deer herd, to give a wide coverage of herds in New Zealand. RESULTS: From January 2001 to October 2005, M. paratuberculosis was isolated from 1,141 farmed deer, and has now been identified by microbiological testing in over 600 deer herds in New Zealand. The bovine subtype of M. paratuberculosis was shown by a highly specific PCR test to be present in 91/95 herds examined; the ovine subtype was found in the remaining four herds. CONCLUSIONS: Since 2000, there has been a substantial increase in both the number of microbiologically-confirmed cases of Johne's disease in farmed deer and the number of infected herds. Johne's disease is now widespread and common in deer herds throughout New Zealand. Whilst the bovine subtype of M. paratuberculosis predominates in deer herds in New Zealand in which Johne's disease has been confirmed, the occasional finding of the ovine subtype highlights the need to consider both sheep and cattle as potential sources of infection for farmed deer.     

The occurrence of transmissible spongiform encephalopathies in roe deer (Capreolus capreolus), red deer (Cervus elaphus) and chamois (Rupicapra rupicapra) in Bavaria

Brain samples of 849 free-ranging ruminants (654 roe deer, 189 red deer and 6 chamois) from Bavaria were examined for TSE. In this respect, detections of PrPres were carried out by ELISA technique (BioRad) and immunohistochemistry using the monoclonal antibody (mAb) L42. Validation experiments of the ELISA technique for the detection of prionprotein from free-ranging ruminants showed that the examination of brains of wild ruminants lead to lower extinctions than that of bovine brain samples. Due to the fact that brain samples of 2 known CWD positive wapitis showed definite positive results, all samples were analysed by this test. There were no indications on the occurrence of TSE in any of the sample. In addition, immunohistochemical investigations of 10% of all samples did not show PrPres. Due to the large number of samples, the prevalence of TSE in Bavaria can be quoted under 0.5% for roe deer and under 1.5% for red deer with a certainty of 95%.     

Role of wild ruminants in the epidemiology of bluetongue virus serotypes 1, 4 and 8 in Spain

ABSTRACT: Although the importance of wild ruminants as potential reservoirs of bluetongue virus (BTV) has been suggested, the role played by these species in the epidemiology of BT in Europe is still unclear. We carried out a serologic and virologic survey to assess the role of wild ruminants in the transmission and maintenance of BTV in Andalusia (southern Spain) between 2006 and 2010.A total of 473 out of 1339 (35.3%) wild ruminants analyzed showed antibodies against BTV by both ELISA and serum neutralization test (SNT). The presence of neutralizing antibodies to BTV-1 and BTV-4 were detected in the four species analyzed (red deer, roe deer, fallow deer and mouflon), while seropositivity against BTV-8 was found in red deer, fallow deer and mouflon but not in roe deer. Statistically significant differences were found among species, ages and sampling regions. BTV RNA was detected in twenty-one out of 1013 wild ruminants (2.1%) tested. BTV-1 and BTV-4 RNA were confirmed in red deer and mouflon by specific rRT-PCR.BTV-1 and BTV-4 seropositive and RNA positive wild ruminants, including juveniles and sub-adults, were detected years after the last outbreak was reported in livestock. In addition, between the 2008/2009 and the 2010/2011 hunting seasons, the seroprevalence against BTV-1, BTV-4 and BTV-8 increased in the majority of provinces, and these serotypes were detected in many areas where BTV outbreaks were not reported in domestic ruminants. The results indicate that wild ruminants seem to be implicated in the dissemination and persistence of BTV in Spain.