Management of a caseous lymphadenitis outbreak in a new Iberian ibex (Capra pyrenaica) stock reservoir.

BACKGROUND: In 2010, an Iberian ibex (Capra pyrenaica hispanica) stock reservoir was established for conservation purposes in north-eastern Spain. Eighteen ibexes were captured in the wild and housed in a 17 hectare enclosure. Once in captivity, a caseous lymphadenitis (CLA) outbreak occurred and ibex handlings were carried out at six-month intervals between 2010 and 2013 to perform health examinations and sampling. Treatment with a bacterin-based autovaccine and penicillin G benzatine was added during the third and subsequent handlings, when infection by Corynebacterium pseudotuberculosis was confirmed. Changes in lesion score, serum anti-C. pseudotuberculosis antibodies and haematological parameters were analyzed to assess captivity effects, disease emergence and treatment efficacy. Serum acute phase proteins (APP) Haptoglobin (Hp), Amyloid A (SAA) and Acid Soluble Glycoprotein (ASG) concentrations were also determined to evaluate their usefulness as indicators of clinical status. Once in captivity, 12 out of 14 ibexes (85.7%) seroconverted, preceding the emergence of clinical signs; moreover, TP, WBC, eosinophil and platelet cell counts increased while monocyte and basophil cell counts decreased. After treatment, casualties and fistulas disappeared and both packed cell volume (PCV) and haemoglobin concentration significantly increased. Hp, SAA and ASG values were under the limit of detection or showed no significant differences. CONCLUSIONS: A role for captivity in contagion rate is suggested by the increase in antibody levels against C. pseudotuberculosis and the emergence of clinical signs. Although boosted by captivity, this is the first report of an outbreak of caseous lymphadenitis displaying high morbidity and mortality in wild ungulates. Treatment consisting of both vaccination and antibiotic therapy seemed to prevent mortality and alleviate disease severity, but was not reflected in the humoural response. Haematology and APP were not useful indicators in our study, perhaps due to the sampling frequency. Presumably endemic and irrelevant in the wild, this common disease of domestic small ruminants is complicating conservation efforts for the Iberian ibex in north-eastern Spain.

Using a top predator as a sentinel for environmental contamination with pathogenic bacteria: the Iberian wolf and leptospires.

The Iberian wolf (Canis lupus) is the top predator in the Iberian environments in which it lives, feeding on a wide range of species, thus encountering a wide range of disease agents. Therefore, the wolf can serve as sentinel of environmental contamination with pathogens. We investigated the exposure of free-living wolves to 14 serovars of Leptospira interrogans sensu lato. Kidney samples from 49 wolves collected from 2010-2013 in northwestern Spain were analysed by culture, direct immunofluorescence and polymerase chain reaction. Tissue fluids were analysed for antibodies by a microscopic agglutination test. Ten wolves (observed prevalence: 20%, 95% confidence interval = 11-33%) showed evidence of contact with leptospires, eight through direct detection and nine through serology (7 wolves were positive according to both techniques). Titres below the cut-off level were also detected in seven cases. Serovars confirmed were Canicola (n = 4), Icterohaemorrhagiae (n = 3) and Sejroë, Ballum and Grippotyphosa (n = 1 each), indicating that wolves were infected with serovars for which dogs, rodents and ungulates, are the natural hosts and supporting the utility of the wolf and other large predators as environmental sentinels for pathogens.

Using a top predator as a sentinel for environmental contamination with pathogenic bacteria: the Iberian wolf and leptospires

The Iberian wolf (Canis lupus) is the top predator in the Iberian environments in which it lives, feeding on a wide range of species, thus encountering a wide range of disease agents. Therefore, the wolf can serve as sentinel of environmental contamination with pathogens. We investigated the exposure of free-living wolves to 14 serovars of Leptospira interrogans sensu lato. Kidney samples from 49 wolves collected from 2010-2013 in northwestern Spain were analysed by culture, direct immunofluorescence and polymerase chain reaction. Tissue fluids were analysed for antibodies by a microscopic agglutination test. Ten wolves (observed prevalence: 20%, 95% confidence interval = 11-33%) showed evidence of contact with leptospires, eight through direct detection and nine through serology (7 wolves were positive according to both techniques). Titres below the cut-off level were also detected in seven cases. Serovars confirmed were Canicola (n = 4), Icterohaemorrhagiae (n = 3) and Sejroë, Ballum and Grippotyphosa (n = 1 each), indicating that wolves were infected with serovars for which dogs, rodents and ungulates, are the natural hosts and supporting the utility of the wolf and other large predators as environmental sentinels for pathogens.

Detection of antibodies in wild ruminants to evaluate exposure to liver trematodes.

Wild ruminants sharing pastures with domestic livestock are at risk of infection by liver trematodes. Detection of antibodies provides a very useful tool to gain more knowledge about the distribution of these parasites. Non-lethal methods are strongly encouraged for the analysis of the risk of infection among wild ruminants. A seroepidemiological survey was conducted to analyze exposure to hepatic trematodes ( Fasciola hepatica and Dicrocoelium dendriticum ) in wild ruminants from southern Spain. Blood samples were collected from 69 bovids (Mouflon + Spanish ibex) and 143 cervids (red deer + fallow deer) from Sierra de Cazorla, Segura and Las Villas Natural Park. The samples were analyzed using the excretory/secretory antigens of each trematode to determine the IgG response. All the animals were examined at necropsy for the presence of flukes, and the species, age, and gender of the animals were recorded. Fasciola hepatica were only observed in cervids (3%; 95% confidence interval [CI]  =  2-8), while D. dendriticum specimens were recorded in 1% (0-8) of bovids and 4% (CI  =  2-9) of the cervids. The IgG-seroprevalence against F. hepatica was significantly higher in the cervids. Statistical differences according to gender were observed. The bovids exhibited the greatest percentages of positive cases to D. dendriticum antigens, and the DdES-seroprevalence was related to age of the animals. When considering all the factors, the FhES-seroprevalence was initially distributed according to the type of ruminant (cervids), gender (male), and age (>2 yr).

Importance of canine distemper virus (CDV) infection in free-ranging Iberian lynxes (Lynx pardinus).

Canine distemper virus (CDV) is a morbillivirus that is the etiological agent of one of the most important viral diseases affecting canids and an expanding range of other carnivores. Using real-time RT-PCR, CDV RNA was detected in organs of an Iberian lynx (Lynx pardinus) found dead in the Doñana National Park, Southwestern Andalusia, Spain. This finding may be of great importance for the conservation of the species; at present the Iberian lynx is the most critically endangered wild felid. The aim of the present study was to elucidate the significance of CDV for the Iberian lynx population. High viral loads were evident in the dead lynx, suggesting an etiological involvement of CDV in its death. When carnivores from the same region were analyzed by CDV RT-PCR, a stone marten (Martes foina) was positive. Phylogenetic analyses demonstrated high identity of the two detected CDVs and a close relationship to the European dog lineage of CDV. Antibodies to CDV were detected in 14.8% of 88 tested free-ranging Iberian lynxes. The sample seroprevalence was significantly higher in lynxes from the Doñana Natural Space (22.9%) than Sierra Morena (5%). The stone marten and a red fox (Vulpes vulpes) also tested seropositive. In conclusion, CDV is present in the Iberian lynx population, especially in the Doñana region, with sporadic cases of disease. To reduce the infectious pressure of CDV on this endangered population, a mass dog vaccination should be considered.

Leptospirosis in wild and domestic carnivores in natural areas in Andalusia, Spain.

Leptospirosis is a zoonosis that affects humans, domestic animals, and wildlife. Carnivores are at the top of the feeding chain, thus being exposed to pathogens through their preys. From June 2004 to April 2007, we analyzed for evidences of contact with 14 serovars of Leptospira interrogans Sensu Lato serum (analyzed by indirect Microscopic Agglutination Test) and urine or kidney samples (analyzed by microscopic observation, immunostaining and culture) collected from 201 wild and domestic carnivores, including 26 free-living Iberian lynx (Lynx pardinus), 33 red foxes (Vulpes vulpes), 33 Egyptian mongooses (Herpestes ichneumon), 25 common genets (Genetta genetta), two Eurasian badgers (Meles meles) and one Eurasian otter (Lutra lutra), and 53 free-roaming cats and 28 rural dogs in protected areas in Andalusia (southern Spain). Twenty-three percent of the animals presented evidences of contact, being the prevalence similar among wild (23.5%) and domestic species (22.2%). Contact with Lesptospira was detected in all the species but the otter. Prevalence was: lynx (11% by bacteriological detection, 32% by serology), fox (0%, 47%), mongoose (5%, 20%), genet (0%, 12%), badger (0%, 50%), cat (20%, 14%), dog (only serology: 36%). Serovar Icterohemorragiae accounted for 2/3 of the cases. Serovar Canicola was detected in half of the positive dogs and one lynx. Other serovars detected were Ballum, Sejroë, and Australis. No macroscopic lesions were observed in necropsied animals that showed evidence of contact with the agent, although histopathologic lesions (chiefly chronic interstitial nephritis) were observed in 7 out of the 11 microscopically analyzed individuals. Thus, L. interrogans may cause previously unrecorded disease in wild carnivores in Spain. Wild and free-roaming carnivores may not act as reservoir of L. interrogans but as a dead-end hosts, though the dog may act as reservoir of serovar Canicola. Carnivores are apparently good sentinels for the epidemiological monitorization of leptospirosis.

Disease threats to the endangered Iberian lynx (Lynx pardinus).

The Iberian lynx, (Lynx pardinus), is the most endangered felid in the world. To determine whether sympatric carnivores are reservoirs of pathogens posing a disease risk for the lynx, evidence of exposure to 17 viral, bacterial and protozoan agents was investigated in 176 carnivores comprising 26 free-living lynx, 53 domestic cats, 28 dogs, 33 red foxes (Vulpes vulpes), 24 Egyptian mongooses (Herpestes ichneumon), 10 common genets (Genetta genetta) and 2 Eurasian badgers (Meles meles) in the areas inhabited by the last two populations of Iberian lynx, both in Andalusia (South-Western Spain). The results indicated that the lynx had low rates of contact with viral pathogens, with one seropositive finding each for feline leukemia virus, parvovirus and canine adenovirus-1, whereas contact with bacteria and protozoa appeared more frequent. Active infections with parvovirus, Ehrlichia spp., Mycobacterium bovis, Leptospira interrogans and Cytauxzoon spp. were confirmed. In contrast, 53% of the domestic cats were exposed to some infectious agent (prevalence range 4.5-11.4%). Antibodies to canine distemper virus and parvovirus were frequently found in dogs (32% and 42%, respectively) and foxes (30% and 12%). Past or present infections with parvovirus, Ehrlichia spp., Chlamydophila spp., M. bovis, Salmonella enterica, L. interrogans, Toxoplasma gondii, and Neospora caninum were also detected in these and other species surveyed. Questionnaires to owners revealed that 14% of the dogs but none of the cats had been vaccinated, and no cat had been neutered. Based on the apparent absence of acquired immunity of the lynx against infectious agents, the frequent detection of agents among sympatric carnivores, and the reported lack of immunocompetence of the Iberian lynx, a disease outbreak among the local abundant carnivores may pose a serious disease risk for lynx conservation.

Disseminated bovine tuberculosis in a wild red fox (Vulpes vulpes) in southern Spain.

A wild-caught, adult red fox (Vulpes vulpes) from Doñana National Park (southern Spain), in very poor condition, died during anesthesia. At necropsy, the submandibular, retropharyngeal, mediastinal, axillary, mesenteric, and popliteal lymph nodes were enlarged, and the right submandibular and mesenteric lymph nodes had hard, white-colored concretions (calcifications). Multiple white pinpoint foci were observed in the lungs, and abscesses were present in the left dorsal lung lobes. No lesions were seen in tonsils, liver, or spleen. On histopathology multiple tuberculous granulomas, with Ziehl-Neelsen-positive bacilli, were observed in the lung, and multifocal necrotic areas with calcification were present in the submandibular and mesenteric lymph nodes. Nucleic acid amplification from lymph node samples demonstrated the presence of mycobacteria belonging to the Mycobacterium tuberculosis complex. One strain was isolated by culture and identified as Mycobacterium bovis. The suspected route of infection was oral, probably after repeated scavenging of infected wild ungulate carcasses. This is the first report of generalized bovine tuberculosis (bTB) in a wild canid. This finding raises concerns about bTB as a disease risk for protected species, livestock, and humans in Mediterranean Spain.

Exposure to disease agents in the endangered Iberian lynx (Lynx pardinus).

The Iberian lynx (Lynx pardinus) is the most endangered felid species in the world. Lynx populations have decreased dramatically in size and distribution in the last four decades, thus becoming increasingly vulnerable to catastrophic events such as epizooties. From 1989 to 2000, serum samples were obtained from 48 free-ranging lynx captured in the Doñana National Park (DNP, n = 31) and mountains of Sierra Morena (SM, n = 17) in southern Spain. Samples were tested for antibodies against Toxoplasma gondii, feline herpesvirus 1 (FHV-1), feline calicivirus (FCV), feline/canine parvovirus (FPV/CPV), feline coronavirus, feline immunodeficiency virus (FIV), feline leukaemia virus and canine distemper virus (CDV) and for FeLV p27 antigen, to document baseline exposure levels. Antibodies against T. gondii were detected in 44% of lynx, with a significantly greater prevalence in DNP (61%) than in SM (12%). In DNP, prevalence was significantly higher in adult (81%) than in juvenile and sub-adult (41%) lynx, but no such difference was observed in SM. Low prevalences (≤11%) of minimally positive titres were found for FHV-1, FCV and FPV/CPV. This, combined with the lack of evidence for exposure to CDV, FIV and FeLV, suggests that these lynx populations are naïve and might be vulnerable to a disease outbreak in the future. Because of the reduced size of lynx populations, the documented low level of genetic variation (particularly in the DNP population) coupled with the recently documented state of immune depletion in a majority of necropsied lynx, it is important to better understand the threat and potential impact that disease agents might pose for the conservation of this endangered species. Future surveillance programs must include possible disease reservoir hosts such as domestic cats and dogs and other wild carnivores.

Potential pathogens carried by Spanish ibex (Capra pyrenaica hispanica) in southern Spain.

The Spanish ibex (Capra pyrenaica hispanica) population of southern Spain was surveyed for potential pathogens associated with the conjunctiva, external ear canal, as well as reproductive and upper respiratory tracts. We sampled 321 ibex (131 adult males, 100 adult females, and 90 yearlings); these included 271 apparently healthy animals and 50 that were naturally infected with Sarcoptes scabiei. A total of 688 bacterial isolates were identified (377 gram-negatives, 225 gram-positives, and 86 Mycoplasma spp.); sex, age, location, infection with S. scabiei, and disposition of the animal (free-ranging versus captive) were evaluated as risk factors for infection. Infections with Mycoplasma agalactiae and Mycoplasma arginini were associated with age, having a higher frequency of isolation in young animals. With Escherichia coli, Mannheimia haemolytica, Pasteurella multocida biotype A, and Staphylococcus aureus, significantly higher isolation rates were associated with adults. The isolation frequency for E. coli was higher in females, whereas Moraxella bovis isolations were mostly associated with males. The presence of mange increased the risk of infection with both Streptococcus equi subsp. zooepidemicus and M. haemolytica. The geographic origin of sampled animals was related to the isolation of Branhamella ovis, M. agalactiae, and all Pasteurella sp. Isolations of M. haemolytica, P. multocida biotype A, E. coli, and B. ovis were more prevalent in samples from free-ranging rather than captive animals. Of the gram-positive bacteria, S. aureus represented the predominant species isolated from nasal, vaginal, and ocular samples. Mycoplasma agalactiae and M. arginini were the predominant Mycoplasma spp., and both were associated most often with the external ear canal. The most frequently isolated gram-negative bacteria included E. coli, M. haemolytica, P. multocida biotype A, and B. ovis. Isolation rates of gram-negative species varied by source. In nasal samples, M. haemolytica and P. multocida biotype A were isolated most frequently, whereas in ocular and vaginal samples, B. ovis and E. coli, respectively, were most frequently isolated.

Population effects of sarcoptic mange in Barbary sheep (Ammotragus lervia) from Sierra Espuña Regional Park, Spain.

The nonindigenous Barbary sheep population (Ammotragus lervia) of the Sierra Espuna Regional Park (Murcia, Spain) suffered an outbreak of sarcoptic mange between 1991 and 1995, which contributed to a population decrease of 86%. This study presents the results of two population surveys conducted in 1994 and 1999 based on the fixed point and itineraries method (FPI) and the excrement count (EC) method, as well as data from demographic estimates and clinical observations conducted by the Regional Administration of Murcia. Results of surveillance for mange are given between 1992 and 1995, because no animals were observed with sarcoptic mange in 1999. Prevalence of mange peaked in 1994 and then declined. During the regression phase of the epidemic, there was a higher infection rate in males (21.9%) than in females (16.6%) or young animals (5.1%). Males over 5 yr old were the worst affected age group, followed by subadults of both sexes. Few animals had generalized lesions of mange (7%), and most individuals (72%) had lesions of moderate severity. The most common locations of lesions were the neck, head, and back. The density of Barbary sheep in the Sierra Espuna Regional Park increased from introduction in 1972 until it peaked at 13 animals/km(2) in 1991, the year when the first case of sarcoptic mange was detected. After 2 yr of the mange epidemic, the average estimated density was 1.7 animals/km(2) in 1994, which increased to 5.0 animals/km(2) in 1999. The average group size also increased from 7.9 to 19.2 animals/group between 1994 and 1999. The sex ratio, expressed as the proportion of females in the total population observed, decreased from 0.61 in 1994 to 0.49 in 1999. The reproduction rate (kids per females per year) was essentially stable (0.59 in 1994 to 0.65 in 1999). Between 1994 and 1999 the population aged, with the number of young animals (<18 mo of age) decreasing from 45.3% to 36.6% from 1994 to 1999. In the same period, the proportion of males increased 21.4% to 32.6%. We believe sarcoptic mange acted as one of the regulating factors of population density after 1991 and that currently, although no sarcoptic mange lesions were observed in the 1999 survey, there is a demographic imbalance in sex ratio, age structure, and density.

Antibodies to selected viral and bacterial pathogens in European wild boars from southcentral Spain.

Serum samples from 78 European wild boars (Sus scrofa) harvested during the 1999-2000 hunting season were tested for antibodies to Brucella spp., classical swine fever virus, Erysipelothrix rhusiopathiae, Haemophilus parasuis, Leptospira interrogans serovar pomona, Mycoplasma hyopneumoniae, pseudorabies virus (PRV), porcine parvovirus (PPV), porcine reproductive and respiratory syndrome virus, Salmonella serogroups B, C, and D, Streptococcus suis, and swine influenza virus (SIV) serotypes H1N1 and H3N2. Samples were collected from Sierra Morena and Montes de Toledo in southcentral Spain. Antibodies were detected to PRV (36%), L. interrogans serovar pomona (12%), PPV (10%), E. rhusiopathiae (5%), SIV serotype H1N1 (4%), Salmonella serogroup B (4%), and Salmonella serogroup C (3%). Our results suggest that more research is needed to describe the epidemiology of infectious diseases of Spanish wild boars.