Prevalence of Paslahepevirus balayani in commercial swine food products from Spain.

Paslahepevirus balayani (formerly known as hepatitis E virus) is an emerging cause of foodborne disease in Europe, transmitted mainly by the consumption of raw or undercooked pork. Since little is known about the presence of the virus in several pork products that are eaten uncooked, our aim was to evaluate the prevalence of Paslahepevirus balayani in groups of commercial pork products intended for human consumption subjected to different processing techniques. A total of 1265 samples of pork products from Spain were divided into four groups and tested for the presence of Paslahepevirus balayani RNA: unprocessed pig and wild boar meat frozen at -20 °C (n = 389), dry-cured pork products (n = 391), dry-cured and salted pork products (n = 219), and boiled products (n = 266) (none of these products contained pork liver). Five samples were positive for Paslahepevirus balayani RNA (overall prevalence: 0.4%; 95% CI: 0.17% - 0.92%). All positive samples were from unprocessed meat stored at -20 °C, with a prevalence in this group of 1.3% (95% CI: 0.42-3.44); two samples came from pigs (1.1%; 95% CI: 0.13-3.81) and three from wild boar (1.5%; 95% CI: 0.31-4.28). None of the pork samples in the other groups was positive. In conclusion, Paslahepevirus balayani was found in unprocessed swine products form Spain, but not in processed products intended to be consumed undercooked, demonstrating that transmission of this zoonotic virus by eating these pork products should be more seriously considered.

Epidemiological survey and risk factors associated with Sarcoptes scabiei in wild lagomorphs in Spanish Mediterranean ecosystems.

Although mortality caused by Sarcoptes scabiei has been reported in European wild rabbit (Oryctolagus cuniculus) and Iberian hare (Lepus granatensis), there is a lack of detailed information regarding the exposure of wild lagomorph species to this parasite. Here, we aimed to determine the seroprevalence and potential risk factors associated with S. scabiei exposure in European wild rabbits and Iberian hares in Mediterranean ecosystems of southern Spain. Between 2018/2019 and 2021/2022 hunting seasons, serum samples from 464 wild rabbits and 132 Iberian hares were collected from 100 hunting grounds in Andalusia (southern Spain). Sera were tested using an in-house indirect ELISA to detect specific anti-S. scabiei antibodies based on the immunodominant protein Ssλ20ΔB3. The overall apparent individual seroprevalence was 15.9% (95/596; 95%CI: 13.0-18.9). Antibodies against S. scabiei were detected in 11.6% (54/464; 95%CI: 8.7-14.5) of the European wild rabbits and 31.1% (41/132; 95%CI: 23.2-39.0) of the Iberian hares. Species (Iberian hare), age (adults) and geographical area (western Andalusia) were identified as risk factors potentially associated with S. scabiei exposure using generalized estimating equation analysis. By applying spatial analysis, two significant cluster of high seropositivity were detected in western and central Andalusia, respectively. The seroprevalence values obtained provide evidence of endemic, widespread and heterogeneous exposure to S. scabiei among wild lagomorph populations in Spanish Mediterranean ecosystems. Our findings underscore the importance of implementing integrated surveillance programs for sarcoptic mange in wild lagomorphs as well as in other sympatric species.

Movement-driven modelling reveals new patterns in disease transmission networks.

Interspecific interactions are highly relevant in the potential transmission of shared pathogens in multi-host systems. In recent decades, several technologies have been developed to study pathogen transmission, such as proximity loggers, GPS tracking devices and/or camera traps. Despite the diversity of methods aimed at detecting contacts, the analysis of transmission risk is often reduced to contact rates and the probability of transmission given the contact. However, the latter process is continuous over time and unique for each contact, and is influenced by the characteristics of the contact and the pathogen's relationship with both the host and the environment. Our objective was to assess whether a more comprehensive approach, using a movement-based model which assigns a unique transmission risk to each contact by decomposing transmission into contact formation, contact duration and host characteristics, could reveal disease transmission dynamics that are not detected with more traditional approaches. The model was built from GPS-collar data from two management systems in Spain where animal tuberculosis (TB) circulates: a national park with extensively reared endemic cattle, and an area with extensive free-range pigs and cattle farms. In addition, we evaluated the effect of the GPS device fix rate on the performance of the model. Different transmission dynamics were identified between both management systems. Considering the specific conditions under which each contact occurs (i.e. whether the contact is direct or indirect, its duration, the hosts characteristics, the environmental conditions, etc.) resulted in the identification of different transmission dynamics compared to using only contact rates. We found that fix intervals greater than 30 min in the GPS tracking data resulted in missed interactions, and intervals greater than 2 h may be insufficient for epidemiological purposes. Our study shows that neglecting the conditions under which each contact occurs may result in a misidentification of the real role of each species in disease transmission. This study describes a clear and repeatable framework to study pathogen transmission from GPS data and provides further insights to understand how TB is maintained in multi-host systems in Mediterranean environments.

Outbreak of Contagious Ecthyma in Free-Ranging Iberian Ibex (Capra pyrenaica) in the Montgrí Massif Natural Park, Catalonia, Northeastern Spain.

Contagious ecthyma is a common, worldwide, and highly transmissible viral zoonotic skin disease caused by the orf virus (ORFV). It mainly affects farmed small ruminants, but it has also been described in a broad range of wild and domestic mammals, with Caprinae species most susceptible. Between November 2019 and January 2020, adults, juveniles, yearlings, and kids from an Iberian ibex (Capra pyrenaica) population established in the Montgrí Massif Natural Park (Catalonia, northeastern Spain) were observed with clinical signs and lesions compatible with ORFV infection. The carcass of an adult male with severe disease enabled sample collection for histopathology and molecular studies, confirming ORFV DNA in the skin lesions. Sequence analyses indicated that the ORFV strain detected had high homology (>98%) with strains previously obtained from other European wild ruminant species. The outbreak peaked in December 2019, with an estimated prevalence of 68.97% (95% confidence interval, 53.35-84.59). From February 2020 forward, no individuals with lesions were observed. The yearly counts for population monitoring corroborated the apparently negligible ORFV impact on the ibex population of the Montgrí Natural Park. There are no previous reports of contagious ecthyma in a free-ranging Iberian ibex population.

Serosurvey of Blood Donors to Assess West Nile Virus Exposure, South-Central Spain.

We analyzed West Nile Virus (WNV) exposure from 1,222 blood donors during 2017-2018 from an area of south-central Spain. Results revealed WNV seroprevalence of 0.08% (95% CI 0.004%-0.4%) in this population. Our findings underscore the need for continued surveillance and research to manage WNV infection in this region.

Identification and characterization of a novel circovirus in Iberian lynx in Spain.

Circoviruses cause severe disease in pigs and birds. Canine circovirus has thus far only been associated with respiratory and gastrointestinal disorders and systemic disease in dogs. The Iberian lynx (Lynx pardinus) is one of the most endangered carnivores in Europe and the most endangered felid worldwide. Exploring the virome of these animals may be important in terms of virus discovery and assessing the interspecies-circulation of viruses from related carnivores. In this study, 162 spleen samples from Iberian lynx were screened for CRESS DNA viruses. Overall, 11 (6.8%) of 162 samples tested positive using a consensus PCR. Partial rep sequences were tightly related to each other (96.6-100%). Specific molecular protocols were designed on the partial rep sequences of the novel virus, Iberian lynx-associated circovirus-1 (ILCV-1). By screening a subset of 45 spleen samples, the infection rate of ILCV-1 in Iberian lynxes was 57.8% (26/45). ILCV-1 strains formed a separate cluster intermingled with bat, rodent, mongoose, and felid circoviruses. The genome of the novel virus displayed the highest nucleotide identity (64.3-65.3%) to mongoose circoviruses, thus representing a novel candidate circovirus species. The detection of these viruses in the spleen tissues could suggest systemic infection in the animal host. Overall, these findings suggest that this novel circovirus is common in the Iberian lynx. Further studies are warranted to assess the possible health implications of ILCV-1 in this endangered species.

Occurrence of Coxiella burnetii in wild lagomorphs and their ticks in Spanish Mediterranean ecosystems.

BACKGROUND: Coxiella burnetii, the causative agent of Q fever, is a zoonotic multi-host vector-borne pathogen of major public health importance. Although the European Food Safety Authority has recently made the monitoring of this bacterium in wildlife a priority, the role of wild lagomorphs in the transmission and maintenance of C. burnetii is poorly understood. AIMS: The aims of this study were to determine the prevalence and risk factors associated with C. burnetii circulation in European wild rabbits (Oryctolagus cuniculus) and Iberian hares (Lepus granatensis) and to assess the presence of this pathogen in ticks that feed on them in Mediterranean ecosystems in Spain, the country with the highest number of reported cases of Q fever in Europe. METHODS: A total of 574 spleen samples were collected from 453 wild rabbits and 121 Iberian hares, and 513 ticks (processed in 120 pools) between the 2017/2018 and 2021/2022 hunting seasons. RESULTS: C. burnetii DNA was detected in 103 (17.9%; 95% CI: 14.8-21.1) of the 574 wild lagomorphs tested. By species, prevalence was 16.3% (74/453; 95% CI: 12.9-19.7) in the European wild rabbit and 24.0% (29/121; 95% CI: 16.4-31.6) in the Iberian hare. At least one positive lagomorph was found on 47.9% of the 96 hunting estates sampled and in every hunting season since 2018/2019. Two risk factors associated with C. burnetii infection were as follows: outbreak of myxomatosis on the hunting estate in the month prior to sampling and high tick abundance observed by gamekeepers on the hunting estate. C. burnetii DNA was also found in 33 of the 120 (27.5%; 95% CI: 19.5-35.5) tick pools tested. The pathogen was detected in 66.7% (4/6), 29.2% (26/89) and 21.4% (3/14) of Haemaphysalis hispanica, Rhipicephalus pusillus and Hyalomma lusitanicum pools respectively. CONCLUSIONS: This study provides new epidemiological data on C. burnetii in European wild rabbits and is the first survey on this zoonotic pathogen performed in Iberian hares. Our results indicate widespread endemic circulation of C. burnetii and highlight the importance of both wild lagomorph species as natural reservoirs of this zoonotic bacterium in Mediterranean ecosystems in southern Spain, which may be of public and animal health concern. The high prevalence and wide diversity of positive tick species suggest the possible role of ticks in the epidemiological cycle of C. burnetii, with the potential risk of transmission to sympatric species, including humans.

Effect of Myxoma Virus Species Jump on Iberian Hare Populations.

The myxoma virus species jump from European rabbits (Oryctolagus cuniculus) to Iberian hares (Lepus granatensis) has raised concerns. We assess the decline suffered by Iberian hare populations on the Iberian Peninsula and discuss the association between the effect of myxomatosis and the average abundance index, which we estimated by using hunting bags.

SARS-CoV-2 in Captive Nonhuman Primates, Spain, 2020-2023.

We conducted a serologic and molecular study to assess exposure of captive nonhuman primates (NHPs) to SARS-CoV-2 in Spain during the 2020-2023 COVID-19 pandemic. We found limited exposure of NHPs to SARS-CoV-2. Biosafety measures must be strictly maintained to avoid SARS-CoV-2 reverse-zoonotic transmission in the human-NHP interface.

Epidemiological surveillance of Leishmania infantum in wild lagomorphs in Spanish Mediterranean ecosystems.

Wild lagomorphs play a key epidemiological role as reservoirs of Leishmania infantum, causative agent of the largest outbreak of human leishmaniosis in Europe to date. A large-scale survey study was conducted on wild rabbit (Oryctolagus cuniculus) and Iberian hare (Lepus granatensis) populations in Spanish Mediterranean ecosystems to evaluate the exposure of L. infantum and investigate potential risk factors associated with exposure to this zoonotic parasite. Between 2018 and 2021, a total of 631 wild lagomorphs (471 wild rabbits and 160 Iberian hares) were collected in Andalusia (southern Spain) and tested for antibodies against L. infantum using the indirect fluorescent antibody test (IFAT). Spleen samples from 563 of the wild lagomorphs sampled (441 wild rabbits and 122 Iberian hares) were also evaluated by real-time quantitative PCR (qPCR) for detection of Leishmania kDNA. Exposure to L. infantum (positive by IFAT and/or qPCR) was detected in 56.4 % (356/631; 95 %CI: 52.3-60.3) of the lagomorphs analyzed. Anti-Leishmania antibodies were found in 12.8 % (81/631; 95 %CI: 10.2-15.5) of the animals, and L. infantum kDNA was detected in 59.0 % (332/563; 95 %CI: 54.9-63.0) of the spleen samples tested. Phylogenetic analysis revealed high homology (99.9-100 %) between L. infantum sequences obtained and strains previously isolated from humans in Spain. While apparent seroprevalence was significantly higher in Iberian hares (19.4 %; 95 %CI: 13.3-25.5) compared to wild rabbits (10.6 %; 95 %CI: 7.9-13.4), no significant differences in prevalence were found between wild rabbits (61.0 %; 95 %CI: 56.5-65.6) and Iberian hares (51.6 %; 95 %CI: 42.8-60.5). At least one positive animal was found on 64.8 % (70/108) of the hunting grounds sampled, and a high-risk spatial cluster (P < 0.001) was also identified in central Andalusia. The multivariable analysis identified bioclimatic level (meso-Mediterranean climate) and the presence of goats on hunting grounds as risk factors potentially associated with L. infantum exposure in wild lagomorphs. This study shows high, widespread exposure, but heterogeneous distribution of L. infantum in wild lagomorph populations in Mediterranean ecosystems in southern Spain. The results point to the need to promote integrated surveillance programs for the detection of Leishmania spp. in wild lagomorphs in order to establish effective control measures against human leishmaniosis under a One Health approach.

Emergence of epizootic hemorrhagic disease in red deer (Cervus elaphus), Spain, 2022.

Epizootic hemorrhagic disease (EHD) virus serotype 8 (EHDV-8) emerged in Spain in autumn 2022. In this study, we aimed to (1) characterize the clinical and lesional presentation of EHDV infection in European red deer (Cervus elaphus), and (2) study the spatial spread of the virus in wild ruminants in Spain after its introduction, in 2022/2023. We confirmed EHDV infection in two clinically compatible sick red deer by PCR and detection of anti-EHDV specific antibodies. EHDV infection occurred in red deer with hyperacute to acute clinical signs and lesions associated to vascular changes leading to death of the animals. Partial sequences of variable segment 2 (VP2) and segment 5 (NS1) genes of the detected viruses had >99% nucleotide identity with EHDV-8 sequences from Tunisia and Italy. In a cross-sectional serological study of EHDV in 592 wild ruminants, mainly red deer (n=578), in southwestern Spain, we detected anti-EHDV antibodies in 37 of 592 samples (6.3%; 95% confidence interval: 4.3-8.2), all from red deer and from the localities where clinical cases of EHD were confirmed in red deer. We conclude that EHDV-8 infection causes severe EHD in European red deer. The serosurvey revealed a limited spread of EHDV-8 in Spanish wild ruminant populations in the first year of virus detection in Spain.

Epidemiologic Survey of Crimean-Congo Hemorrhagic Fever Virus in Suids, Spain.

We conducted a cross-sectional study in wild boar and extensively managed Iberian pig populations in a hotspot area of Crimean-Congo hemorrhagic fever virus (CCHFV) in Spain. We tested for antibodies against CCHFV by using 2 ELISAs in parallel. We assessed the presence of CCHFV RNA by means of reverse transcription quantitative PCR protocol, which detects all genotypes. A total of 113 (21.8%) of 518 suids sampled showed antibodies against CCHFV by ELISA. By species, 106 (39.7%) of 267 wild boars and 7 (2.8%) of 251 Iberian pigs analyzed were seropositive. Of the 231 Iberian pigs and 231 wild boars analyzed, none tested positive for CCHFV RNA. These findings indicate high CCHFV exposure in wild boar populations in endemic areas and confirm the susceptibility of extensively reared pigs to CCHFV, even though they may only play a limited role in the enzootic cycle.

Mycobacterium tuberculosis complex in domestic goats in Southern Spain.

Tuberculosis (TB) is a zoonotic infectious disease caused by bacteria belonging to the Mycobacterium tuberculosis complex (MTC), which can affect a wide variety of domestic and wild animal species. Although the role of goats as a reservoir of MTC bacteria has been evidenced, information about the circulation of MTC strains in this species is still very scarce. The aim of the present study was to determine the seroprevalence, spatial distribution, risk factors and MTC spoligotypes circulating in goats from Andalusia (Southern Spain), the Spanish region with the largest goat census and a hotspot area of TB in both cattle and wild ungulates. A total of 2155 serum samples from 80 goat flocks were analyzed by an in-house ELISA using the P22 protein complex as a coating antigen. Antibodies against MTC were detected in 473 goats (21.9%, 95% CI: 20.2-23.7) and the true seroprevalence was 22.3% (95% CI: 20.6-24.1). Seropositivity was found in 72 (90.0%) of the 80 flocks analyzed. The generalized estimating equation model showed that the management system (higher seroprevalence on intensive and semi-intensive farms), and the presence of hospital pens inside the regular stables, were risk factors potentially associated with MTC exposure in goats in Southern Spain. The spatial analysis identified a significant spatial cluster (p < 0.001) in Eastern Andalusia. A total of 16 different MTC spoligotypes, including five of M. caprae and eleven of M. bovis, were identified in goats between 2015 and 2022 in the study area, with SB0157 as the most frequently isolated. The results obtained indicate widespread and non-homogeneous spatial distribution of MTC in goat herds from Southern Spain. The high individual and herd-level seroprevalence values found suggest that goats could play a significant role in the maintenance and transmission of MTC in the study area. Our results highlight the importance of implementing control measures in this species.

Surveillance of Sarcoptic Mange in Iberian Ibexes (Capra pyrenaica) and Domestic Goats (Capra hircus) in Southern Spain.

Sarcoptic mange is a highly contagious skin disease caused by Sarcoptes scabiei. Sera were collected from 411 Iberian ibexes, comprising 157 individuals with sarcoptic mange skin lesions and 254 clinically healthy animals, in 13 population nuclei across Andalusia (southern Spain) between 2015 and 2021. Skin samples from 88 of the 157 animals with mange-compatible lesions were also obtained. Moreover, 392 serum samples from domestic goats (Capra hircus) were collected in the same region and study period. Antibodies against S. scabiei were tested using an in-house indirect ELISA, while the presence of mites of S. scabiei was evaluated in the skin samples by potassium hydroxide digestion. Seropositivity was found in eight (3.1%) of the clinically healthy ibexes and in 104 (66.2%) of the animals with mange-compatible lesions. The presence of S. scabiei was confirmed in 57 (64.8%) out of the 88 skin samples analysed and anti-S. scabiei antibodies were found in 49 (86.0%) of these 57 mite-positive individuals. Seropositive animals were detected in population nuclei with previous records of sarcoptic mange, where S. scabiei mites were detected by potassium hydroxide digestion in individuals with sarcoptic mange-compatible external lesions. However, seropositivity was not observed in population nuclei that were historically free of this disease. None of the 392 domestic goats had antibodies against S. scabiei, suggesting an independent epidemiological cycle of sarcoptic mange in Iberian ibex populations in the study area, and a limited or null role of domestic goats in the transmission of the parasite to this wild species. Overall, our findings underscore the importance of maintaining and/or implementing integrated surveillance programs and control strategies in wildlife and livestock, to limit the risk of S. scabiei circulation in Iberian ibex populations.

Animal Exposure Model for Mapping Crimean-Congo Hemorrhagic Fever Virus Emergence Risk.

To estimate the determinants of spatial variation in Crimean-Congo hemorrhagic fever virus (CCHFV) transmission and to create a risk map as a preventive public health tool, we designed a survey of small domestic ruminants in Andalusia, Spain. To assess CCHFV exposure spatial distribution, we analyzed serum from 2,440 sheep and goats by using a double-antigen ELISA and modeled exposure probability with environmental predictors by using generalized linear mixed models. CCHFV antibodies detected in 84 samples confirmed low CCHFV prevalence in small domestic ruminants in the region. The best-fitted statistical model indicated that the most significant predictors of virus exposure risk were cattle/horse density and the normalized difference vegetation index. Model validation showed 99.7% specificity and 10.2% sensitivity for identifying CCHFV circulation areas. To map CCHFV exposure risk, we projected the model at a 1 × 1-km spatial resolution. Our study provides insight into CCHFV ecology that is useful for preventing virus transmission.

Detection of Rat Hepatitis E Virus in Pigs, Spain, 2023.

We identified rat hepatitis E virus (HEV) RNA in farmed pigs from Spain. Our results indicate that pigs might be susceptible to rat HEV and could serve as viral intermediaries between rodents and humans. Europe should evaluate the prevalence of rat HEV in farmed pigs to assess the risk to public health.

Approaching the complexity of Crimean-Congo hemorrhagic fever virus serology: A study in swine.

Crimean-Congo hemorrhagic fever virus (CCHFV) is a tick-borne zoonotic orthonairovirus of public health concern and widespread geographic distribution. Several animal species are known to seroconvert after infection with CCHFV without showing clinical symptoms. The commercial availability of a multi-species ELISA has led to an increase in recent serosurveillance studies as well as in the range of species reported to be exposed to CCHFV in the field, including wild boar (Sus scrofa). However, development and validation of confirmatory serological tests for swine based on different CCHFV antigens or test principles are hampered by the lack of defined control sera from infected and non-infected animals. For the detection of anti-CCHFV antibodies in swine, we established a swine-specific in-house ELISA using a panel of swine sera from CCHFV-free regions and regions with reported CCHFV circulation. We initially screened more than 700 serum samples from wild boar and domestic pigs and observed a correlation of ≃67% between the commercial and the in-house test. From these sera, we selected a panel of 60 samples that were further analyzed in a newly established indirect immunofluorescence assay (iIFA) and virus neutralization test. ELISA-non-reactive samples tested negative. Interestingly, only a subset of samples reactive in both ELISA and iIFA displayed CCHFV-neutralizing antibodies. The observed partial discrepancy between the tests may be explained by different test sensitivities, antibody cross-reactivities or suggests that the immune response to CCHFV in swine is not necessarily associated with eliciting neutralizing antibodies. Overall, this study highlights that meaningful CCHFV serology in swine, and possibly other species, should involve the performance of multiple tests and careful interpretation of the results.

Exposure to Coxiella burnetii in Wild Lagomorphs in Spanish Mediterranean Ecosystems.

Coxiella burnetii is an important zoonotic pathogen of worldwide distribution that can infect a wide range of wild and domestic species. The European wild rabbit (Oryctolagus cuniculus) can play a role as a reservoir for this bacterium in certain epidemiological scenarios, but, to date, a very limited numbers of large-scale serosurveys have been conducted for this species worldwide. Although exposure in hare species has also been described, C. burnetii in Iberian hare (Lepus granatensis) has never been assessed. Here, we aimed to determine the seroprevalence and risk factors associated with C. burnetii exposure in wild lagomorphs in the Mediterranean ecosystems of southern Spain. Between the 2018/2019 and 2021/2022 hunting seasons, blood samples from 638 wild lagomorphs, including 471 wild rabbits and 167 Iberian hares, were collected from 112 hunting grounds distributed across all eight provinces of Andalusia (southern Spain). The overall apparent individual seroprevalence was 8.9% (57/638; 95% CI: 6.8-11.4). Antibodies against C. burnetii were found in 11.3% (53/471; 95% CI: 8.4-14.1) of the wild rabbits and 2.4% (4/167; 95% CI: 0.1-4.7) of the Iberian hares. Seropositive animals were detected for 16 (14.3%; 95% CI: 7.8-20.8) of the 112 hunting grounds tested and in all the hunting seasons sampled. A generalized estimating equations model showed that the geographical area (western Andalusia) and presence of sheep were risk factors potentially associated with C. burnetii exposure in wild lagomorphs. A statistically significant spatial cluster (p < 0.001) was identified in the south-west of Andalusia. Our results provide evidence of moderate, endemic and heterogeneous circulation of C. burnetii in wild lagomorph populations in Spanish Mediterranean ecosystems. Risk-based strategies for integrative surveillance programs should be implemented in these species to reduce the risk of transmission of the bacterium to sympatric species, including humans.

Detection and genotyping of zoonotic microsporidia in the endangered Iberian lynx (Lynx pardinus).

Microsporidia is a diverse group of obligate, intracellular, and spore-forming parasites that infect a wide range of animals. Among them, Enterocytozoon bieneusi and Encephalitozoon spp. are the most frequently reported species in humans. Limited information is available about the presence and molecular diversity of microsporidian species in the endangered Iberian lynx (Lynx pardinus). Presence of Enterocytozoon bieneusi and Encephalitozoon spp. was investigated by molecular methods in wild and captive Iberian lynxes from Spain. Overall, E. bieneusi was detected in 3.2% (8/251) of the animals examined. None of the samples tested were positive for Encephalitozoon spp. Four known (D, EbfelA, PigEBITS7, and Type IV) and a novel (named as LynxSpEb1) E. bieneusi genotypes were identified. All the genotypes found belonged to the zoonotic Group 1 of E. bieneusi. This study provides the first genotyping data of E. bieneusi in Iberian lynx in Spain. Our result indicate that the Iberian lynx does not seem to play a relevant role in the epidemiology of Encephalitozoon spp., and that this endangered felid is likely acting as spillover host rather than a true reservoir of E. bieneusi. Additional studies should be conducted to assess the impact of this parasite in the health status of the endangered Iberian lynx.

Fatal leptospirosis in endangered Barbary macaques (Macaca sylvanus) kept in captivity: Assessing the role of sympatric rodents.

Between December 2020 and January 2021, an outbreak of acute mortality in endangered Barbary macaques (Macaca sylvanus) kept in captivity was detected in a zoo in Spain. The main findings observed in the two fatally affected animals at post-mortem evaluation were jaundice, renal tubular necrosis and interstitial nephritis. Leptospira spp. infection was confirmed by real time PCR (qPCR) in different tissues in both individuals. Analyses of secY gene from a positive individual showed 100% homology with a previously published sequence corresponding to Leptospira interrogans serovar Copenhageni. Free-living sympatric brown rats (Rattus norvegicus) from the affected zoo were also analyzed, and showed a prevalence and seroprevalence of Leptospira spp. of 18.2% (4/22; 95% CI: 2.1-34.3) and 41.9% (26/62; 95% CI: 29.7-54.2), respectively. We detected seropositive sera to five different serovars of Leptospira spp. (Copenhageni, Grippotyphosa, Pomona, Canicola and Hardjo) in the rodent population, with L. Copenhageni being the predominant one. This study describes for first time an outbreak of fatal leptospirosis in captive non-human primates in Europe. Our results show that Barbary macaques, an endangered species, are highly susceptible to Leptospira spp. infection, with sympatric wild rodents being the most likely reservoir animals involved in transmission in this outbreak. Our results suggest that rodent control could be an effective measure for minimizing exposure to Leptospira spp. in zoological collections. Given the potential implications for conservation, animal and public health, non-human primates and rodents should be included in surveillance programs for Leptospira spp. in zoos.