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Since 2001, high-mortality outbreaks of border disease (BD) have negatively affected populations of Pyrenean chamois (Rupicapra pyrenaica pyrenaica). Studies in the affected areas determined that sympatric wild ruminants did not seem to have an epidemiologic role in the circulation of border disease virus (BDV). However, the recent increase in European mouflon (Ovis aries musimon) densities might enhance the risk of pathogen transmission among chamois and mouflons. We conducted a serologic and virologic investigation of BDV in European mouflon from the Spanish Pyrenees, with the aim of determining potential changes in the role of this species in BDV epidemiology. From 2018 to 2022, we detected antibodies against BDV in 31/185 (16.7%) animals but did not detect BDV RNA in any spleen sample (0/65). These results indicate that BDV infection is occurring in these mouflon populations to a greater extent than previously described, which could shift the current understanding of BD epidemiology in the Pyrenees and cause an unpredictable effect on both chamois and mouflon populations. Further studies on the molecular identification of BDV in mouflon and chamois are required to better understand the contribution of mouflon in the epidemiology of BD., (© Wildlife Disease Association 2023.)

Since 2001, Pyrenean chamois (Rupicapra pyrenaica pyrenaica) populations have been affected by border disease virus (BDV) causing mortalities of more than 80% in some areas. Field studies carried out in France, Andorra, and Spain have shown different epidemiological scenarios in chamois populations. This study was designed to confirm the presence of BDV strains of a high and low virulence in free-ranging chamois populations from Pyrenees and to understand the implications of these findings to the diverse epidemiological scenarios. An experimental infection of Pyrenean chamois with a high-virulence (Cadí-6) and low-virulence (Freser-5) BDV strains was performed. Pregnant and non-pregnant animals with and without antibodies against BDV were included in each group. Cadí-6 BDV strain was confirmed to be of high virulence for seronegative adults and their foetuses. The antibody negative chamois infected with Freser-5 BDV strain did not show symptoms, presented less viral distribution and RNA load in tissues than Cadí-6 group, and cleared the virus from the serum. However, foetuses died before the end of the experiment and RNA virus was detected in sera and tissues although with lower RNA load than the Cadí-6 group. Chamois from both groups presented lesions in brain but the ones infected with the low-virulence Freser-5 BDV strain were mild and most likely transient. In both groups, seropositive pregnant females and all but one of their foetuses did not present viraemia or viral RNA in tissues. The existence of a low-virulence strain has been confirmed experimentally and related to chamois population infection dynamics in the area where it was isolated. Such strain may persist in the chamois population through PI animals and may induce cross-protection in chamois against high-virulence strains. This study demonstrates that viral strain diversity is a significant factor in the heterogeneity of epidemiological scenarios in Pyrenean chamois populations., (© 2019 Blackwell Verlag GmbH.)

Border Disease Virus (BDV) causes health and economic impact on livestock and is also of importance in wildlife conservation as it causes high mortality outbreaks in Pyrenean chamois (Rupicapra pyrenaica pyrenaica). Pastoral practices are known as a main interspecies pathogen transmission. Hence, the presence of pestivirus in transhumant sheep flocks and sympatric chamois was assessed in areas with different epidemiological scenarios of chamois BDV infections. Moreover, the present study had also the goal to identify if inter-specific infections occurred and when they happened. Five sheep flocks grazing in two alpine areas in the Pyrenees with two different BDV epidemiological scenarios in chamois populations were studied during two transhumant seasons. Sheep were sampled before and after transhumance. Pyrenean chamois sera and spleen samples from both areas where also studied during the same period. Antibodies against BDV were assessed by means of ELISA and VNT. A qRT-PCR was used in order to detect the virus. Seroprevalence in sheep ranged between 0 and 91.1% at the flock level. Chamois were found to have high seroprevalences (52.9-77.7%) in both areas, and four new BDV isolates were sequenced. One sheep farm presented persistent BDV circulation and three showed low BDV circulation. The after-transhumance period was identified as the moment when viral transmission occured in the first farm, associated to BDV strains of domestic origin, according to VNT results. However, the BDV isolate was genetical closely related to previous BDV strains from chamois origin. In another farm, antibodies in two of the three positive sera were associated to infection with a chamois-like BDV strain. Altogether indicates that occasional viral transmission from chamois to sheep may occur., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Border disease virus (BDV) affects a wide range of ruminants worldwide, mainly domestic sheep and goat. Since 2001 several outbreaks of disease associated to BDV infection have been described in Pyrenean chamois (Rupicapra pyrenaica pyrenaica) in Spain, France and Andorra. In order to reconstruct the most probable places of origin and pathways of dispersion of BDV among Pyrenean chamois, a phylogenetic analysis of 95 BDV 5'untranslated sequences has been performed on chamois and domestic ungulates, including novel sequences and retrieved from public databases, using a Bayesian Markov Chain Monte Carlo method. Discrete and continuous space phylogeography have been applied on chamois sequences dataset, using centroid positions and latitude and longitude coordinates of the animals, respectively. The estimated mean evolutionary rate of BDV sequences was 2.9×10-3 subs/site/year (95% HPD: 1.5-4.6×10-3). All the Pyrenean chamois isolates clustered in a unique highly significant clade, that originated from BDV-4a ovine clade. The introduction from sheep (dated back to the early 90s) generated a founder effect on the chamois population and the most probable place of origin of Pyrenean chamois BDV was estimated at coordinates 42.42 N and 1.9 E. The pathways of virus dispersion showed two main routes: the first started on the early 90s of the past century with a westward direction and the second arise in Central Pyrenees. The virus spread westward for more than 125 km and southward for about 50km and the estimated epidemic diffusion rate was about 13.1 km/year (95% HPD 5.2-21.4 km/year). The strong spatial structure, with strains from a single locality segregating together in homogeneous groups, and the significant pathways of viral dispersion among the areas, allowed to reconstruct both events of infection in a single area and of migrations, occurring between neighboring areas., Competing Interests: The authors have declared that no competing interests exist.

In 2001, border disease virus (BDV) was identified as the cause of a previously unreported disease in Pyrenean chamois (Rupicapra pyrenaica) in Spain. Since then, the disease has caused a dramatic decrease, and in some cases collapse, of chamois populations and has expanded to nearly the entire distribution area in the Pyrenees. Chamois BDV was characterized as BDV-4 genotype and experimental studies confirmed that it was the primary agent of the disease. The infection has become endemic in the Central and Eastern Pyrenees. However, while most Pyrenean chamois populations have been severely affected by the disease, others have not, despite the circulation of BDV in apparently healthy individuals, suggesting the existence of different viral strategies for persisting in the host population. Changes in the interplay of pathogen, host and environmental factors may lead to the formation of different disease patterns. A key factor influencing disease emergence may be pathogen invasiveness through viral mutation. Host factors, such as behavior, immunity at the population level and genetic variability, may also have driven different epidemiological scenarios. Climatic and other ecological factors may have favored secondary infections, such as pneumonia, that under particular circumstances have been major contributing factors in the high mortality observed in some areas.

Border disease virus (BDV) causes high mortality in Pyrenean chamois (Rupicapra pyrenaica) on the French and Spanish sides of the Pyrenees Mountains. We investigated the pathology induced by BDV in pregnant chamois via experimental infection. Three females were inoculated during the second third of pregnancy with a BDV-4 subgroup strain isolated from a wild Pyrenean chamois during an acute epizootic. A fourth pregnant chamois and one nonpregnant ewe were kept as negative controls. Animals were monitored to assess clinical signs, hematology, viremia, and serology. Postmortem examinations included necropsy, histopathology, and quantification of viral RNA in organs. Pregnancy was unsuccessful in all inoculated animals. One died 24 days postinoculation (dpi) without showing any precursory clinical signs. The second animal had profuse diarrhea from 13 dpi to its death at 51 dpi. The third aborted at 46 dpi and was euthanized at 51 dpi. All animals were viremic from 4 dpi until death. Neutralizing antibodies against BDV-4 were detected from 12 dpi. Necropsies showed generalized lymphadenomegaly, associated in one case with disseminated petechial hemorrhages in the digestive tract. Seventy-eight of 79 organs from inoculated adults and their fetuses had detectable viral RNA. The main histologic lesions in adults were mild lymphohistiocytic encephalitis associated with moderate or moderately severe lymphoid depletion. Control animals remained negative for virus (in blood and organs), antibody, and lesions upon postmortem examination. BDV infection during pregnancy in Pyrenean chamois causes severe disease leading to abortion, then death. Despite 100% fetal death following inoculation, viral RNA was recovered from all organs of infected fetuses, suggesting that persistently infected offspring could be born. Our results may help explain the reported decrease in chamois populations in several areas and suggest that great care must be taken when interpreting infection status for wildlife.

The Principality of Andorra is surrounded by areas in which Pyrenean chamois (Rupicapra pyrenaica pyrenaica) populations were severely affected by infection with border disease virus (BDV) which caused disease outbreaks between 2001 and 2009. Nevertheless, the Andorran chamois populations were not affected during this period. In light of the severe impact of BDV on several of the neighboring Pyrenean chamois populations, we monitored local Andorran populations in an effort to detect pestivirus antibodies and BDV in wild ungulates. In addition, an episode of mortality between 2009 and 2010 in chamois was investigated. We analyzed samples (spleen or serum) from 175 Pyrenean chamois, 284 European mouflon (Ovis orientalis musimon), 13 roe deer (Capreolus capreolus capreolus), and five wild boars (Sus scrofa castilianus). With the exception of three dead chamois found between 2009 and 2010, all samples came from healthy animals hunted during the hunting season. A commercial blocking enzyme-linked immunosorbent assay (ELISA) was used to test sera for antibodies against pestivirus. Positive sera were tested with a comparative virus neutralization test (VNT) using three BDV strains and a bovine viral diarrhea virus strain. Reverse-transcription-polymerase chain reaction (RT-PCR) was performed on all sera and spleen homogenates. Antibodies against pestivirus were detected by ELISA in four of the 69 chamois (5%; 95% CI= 1.29-13.11). The VNT confirmed three of these chamois were infected with a BDV. Viral RNA was detected by RT-PCR in three chamois-one apparently healthy animal hunted in 2009 and two dead animals. Viral sequences showed that the three chamois were infected with a BDV-4, the same genotype that was involved in previous episodes of mortality in the Pyrenees. Although Pyrenean chamois from Andorra had had little contact with the pestiviruses until 2009, in this year BDV was associated with a severe disease outbreak.

Since 2001 several outbreaks of a new disease associated with Border disease virus (BDV) infection have caused important declines in Pyrenean chamois (Rupicapra pyrenaica) populations in the Pyrenees. The goal of this study was to analyze the post-outbreak BDV epidemiology in the first two areas affected by disease with the aim to establish if the infection has become endemic. We also investigated if BDV infected wild and domestic ruminants sharing habitat with chamois. Unexpectedly, we found different epidemiological scenarios in each population. Since the disease outbreaks, some chamois populations recuperated quickly, while others did not recover as expected. In chamois from the first areas, prevalence was high (73.47%) and constant throughout the whole study period and did not differ between chamois born before and after the BDV outbreak; in all, BDV was detected by RT-PCR in six chamois. In the other areas, prevalence was lower (52.79%) and decreased during the study period; as well, prevalence was significantly lower in chamois born after the disease outbreak. No BDV were detected in this population. A comparative virus neutralisation test performed with four BDV strains and one Bovine viral diarrhoea virus (BVDV) strain showed that all the chamois had BDV-specific antibodies. Pestivirus antibodies were detected in all the rest of analyzed species, with low prevalence values in wild ruminants and moderate values in domestic ruminants. No viruses were detected in these species. These results confirm the hypothesis that outbreaks of BDV infection only affect the Pyrenean chamois, although other wild ruminants can occasionally be infected. In conclusion, two different scenarios have appeared since the first border disease outbreaks in Pyrenean chamois: on the one hand frequent BDV circulation with possible negative impact on population dynamics in some areas and on the other, lack of virus circulation and quick recovery of the chamois population.

Since 2001, severe outbreaks of disease associated with border disease virus (BDV) infection have been reported in Pyrenean chamois. The disease is characterized by variable degrees of cachexia, alopecia and neurological manifestations prior to death. The aim of this study was to investigate this disease under experimental conditions. To assess viral virulence, humoral immune response, dissemination and probable routes of transmission, seven chamois (five seronegative and two seropositive for BDV) were inoculated with a BDV isolated from a naturally infected chamois. A group of three chamois were maintained as uninfected controls. The five seronegative chamois became viraemic from day 2 post-inoculation (p.i.) until their death (three animals) or the end of the experiment (on day 34 p.i.) and developed neutralizing antibodies from day 18 p.i. until the end of the study. Continuous shedding of the virus was detected by RT-PCR in oral, nasal and rectal swabs in viraemic chamois from day 5 p.i. Despite none of the viraemic chamois showing obvious neurological signs, all of them had a non-suppurative meningoencephalitis as seen in naturally infected chamois. The two inoculated BDV-seropositive chamois did not become viraemic. This study confirms that BDV is the primary agent of the disease that has been affecting chamois populations in recent years in the Pyrenees and that previously acquired humoral immunity is protective.

In 2005 and 2006 an outbreak of disease associated with border disease virus (BDV) infection caused high mortality in the Pyrenean chamois (Rupicapra pyrenaica) in the Catalan Pyrenees (NE Spain). The aim of this study was to determine values for different haematological and serum biochemical analytes in 32 free-ranging Pyrenean chamois affected by the disease and to compare them with those obtained from healthy chamois. In the affected chamois red blood cell counts, haemoglobin concentrations, packed cell volumes, mean corpuscular volumes and lymphocyte counts were all lower, while the neutrophil and platelet counts were higher. Glucose, lactate, triglycerides, creatinine, total protein concentrations and alkaline phosphatase activity were also lower, in contrast to the concentrations of total bilirubin, urea and aspartate aminotransferase activity, which were higher. Most of the observed changes could be associated with cachexia and inflammation in the affected chamois. Lymphopenia could be directly related to the BDV, which would lead to immunosuppression and explain the high rate of secondary infection observed in these animals., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Pyrenean chamois (Rupicapra pyrenaica) populations of the central and eastern Pyrenees have been affected by severe outbreaks associated with Border disease virus (BDV) since 2001. Eight Pyrenean chamois (7 males and 1 female) from 1 to 8 years of age with clinical signs consistent with BDV infection were studied. At necropsy, whole blood, tissue samples (skin, brain, prescapular lymph node, thyroid gland, lung, liver, spleen, kidney, small intestine, bone marrow, and testicle), urine, and nasal, oral, and rectal swabs were obtained. The fetus from a pregnant female was also studied. Reverse transcription polymerase chain reaction (RT-PCR) was used to detect the virus in all samples, and virus isolation was performed. Sera and tissue samples were positive to RT-PCR, and the virus was isolated from all chamois. The nasal, oral, and rectal swabs and urine samples were RT-PCR positive in 100%, 85.71%, 71.43%, and 100% of chamois, respectively, confirming the excretion of the virus via these 4 routes. In addition, sera were tested for BDV antibodies using enzyme-linked immunosorbent assay and seroneutralization techniques, with negative results. Sequence analysis of the 5' untranslated region in 7 of the chamois confirmed that the virus is grouped into the BDV-4 genotype, the same BDV previously described in Pyrenean chamois. To the authors' knowledge, this is the first study of naturally infected Pyrenean chamois, providing evidence that infected animals shed BDV through nasal, oral, fecal, and urinary excretion routes.

Between 2001 and 2007, several outbreaks of disease associated with Border disease virus (BDV) infection were reported in the central Pyrenees (northeast Spain) and were associated with a major reduction in chamois (Rupicapra pyrenaica) populations. At the same time, wild boars (Sus scrofa) from the same area were found to be seropositive to this pestivirus, without showing clinical signs. The present study examines the susceptibility of domestic swine and the course of the infection with a BDV strain isolated from naturally infected chamois. Twenty pigs were inoculated with 1 x 10(7) TCID(50) (50% tissue culture infective dose) by oronasal route, and 16 control pigs received Eagles sterile Minimal Essential Medium. Serologic (enzyme-linked immunosorbent assay and virus neutralization test) and reverse transcription polymerase chain reaction assays were performed on serum samples obtained at 0, 3, 7, 10, 14, 21, and 31 days postinoculation (dpi). All infected pigs were viremic from 3 to 14 dpi. After 14 dpi, all infected animals developed an antibody response against the homologous virus. Clinical signs or histologic lesions were not observed in inoculated pigs. The present work demonstrates the susceptibility of domestic swine to a BDV strain of chamois origin.

Approximately 3,000 Pyrenean chamois (Rupicapra pyrenaica pyrenaica) died in northeastern Spain during 2005-2007. Border disease virus infection was identified by reverse transcription-PCR and sequencing analysis. These results implicate this virus as the primary cause of death, similar to findings in the previous epizootic in 2001.

In 2001 and 2002, an outbreak of a previously unreported disease, associated with a border disease virus (BDV), caused high mortality in the Southern chamois (Rupicapra pyrenaica) population in the Alt Pallars-Aran National Hunting Reserve in the Catalan Pyrenees (NE Spain). Between 2002 and 2006, sera and/or tissue samples taken from 116 healthy chamois shot during the hunting season, plus 42 from chamois affected by different diseases, were studied. A blocking enzyme-immunosorbent assay (ELISA) was used to study pestivirus seroprevalence in 114 healthy hunted and 31 diseased chamois, yielding positive results in 73.7 and 22.6% of the chamois, respectively. Comparative virus neutralization tests (VNT) performed on 42 seropositive samples with 6 pestivirus strains yielded statistically higher titres to BDV Spain 97, followed by BDV chamois, BDV 137/4, BDV Moredun, Bovine Diarrhoea virus-1 (BVDV-1) NADL and BVDV-2 atypical. Virological investigations for pestivirus detection were performed using an antigen ELISA test in 82 healthy and 18 diseased chamois, RT-PCR in 16 healthy and in all diseased chamois, and virus isolation in 14 diseased chamois. No viral antigen was detected in any of the healthy animals. A pestivirus, characterized as BDV by monoclonal antibodies, was detected in the 10 chamois showing clinical signs consistent with BDV infection. Sequence analysis in the 5' untranslated region (5'-UTR) revealed that they were grouped into the BDV-4 genotype. In the remaining chamois, infectious keratoconjunctivitis, pneumonia, trauma and contagious ecthyma were diagnosed. The cause of death was unknown in five chamois. The results suggest that the infection has become endemic in the population and that it could have a significant impact on chamois population dynamics.

An outbreak of a previously unreported disease affecting southern chamois (Rupicapra pyrenaica) in the central Pyrenees (NE Spain) was recorded in 2001 and 2002. There was a marked temporal distribution, most animals being found between February and June. After the outbreak, the population was found to have decreased by about 42%, most probably due to the disease. We examined 20 affected chamois. Clinical manifestations included depression, weakness and movement difficulties in all cases. Three chamois presented abnormal behaviour, with absence of flight reaction, and 16 showed different degrees of alopecia with skin hyperpigmentation. At necropsy cachexia was observed in all animals, four chamois had abscesses in different parts of the body, four had pneumonia, one had an extensive subcutaneous infection on the head and neck and one had severe orchitis. Microscopic lesions were found in the brain, mainly edema, gliosis, espongiosis, cariorrexis and neuronal multifocal necrosis. A perivascular mononuclear inflammatory infiltrate was present in three of them. Skin lesions included marked follicular atrophy, mild to moderate epidermal hyperplasia with orthokeratotic hyperkeratosis and follicular hyperkeratosis, and hypermelanosis. In 13 chamois there were haemosiderin deposits in the spleen, and in three individuals kidney "cloissone" was observed. Intraeritrocitic parasites were detected either by direct observation or PCR in 8 of 17 chamois. A pestivirus was isolated and detected by RT-PCR from 12 of 13 affected chamois and antigenic characterized as border disease virus by monoclonal antibodies. This is the first time a border disease virus has been associated with an outbreak of a high-mortality disease in a wild species.