Molecular identification of Baylisascaris melis (Gedoelst, 1920) from the Eurasian badger (Meles meles) and ascarids from other wild carnivores in Kazakhstan.

Introduction: The presence of gastrointestinal nematodes, including zoonotic ascarids, in wild canids, felids and mustelids as definitive hosts in Central Asian countries has been documented in many studies based on traditional morphological methods. In contrast, relevant data for the badger are scarce. The aim of this study was the molecular identification of ascarid nematodes from five wild carnivore species in different regions of Kazakhstan. Methods: A total of 211 adult ascarids were collected from gray wolves (Canis lupus, 8 of 83 infected with 2-6 Toxascaris leonina), red foxes (Vulpes vulpes, 26 of 53, with 2-8 Toxascaris leonina), corsac foxes (Vulpes corsac, 6 of 11, 3-6 Toxascaris leonina), lynx (Lynx lynx, 2 of 3, with 2-5 Toxocara cati) and badgers (Meles meles, 2 of 4, with 2-7 Baylisascaris melis). Genomic DNA was extracted from the worms and ribosomal DNA, including the first and second internal transcribed spacer genes, was amplified by polymerase chain reaction using specific oligonucleotide primers and then sequenced. Results: Toxascaris leonina, but not Toxocara canis, was molecularly identified in the wild canids, Toxocara cati in the lynx and Baylisascaris melis in the badger. The maximum likelihood phylogenetic tree showed three distinct clades: the canid Toxascaris leonina was placed in one clade, Toxocara cati in another and Baylisascaris melis in a third. Discussion: The study provides the world's first molecular data and phylogenetic analysis of Baylisascaris melis, identified for the second time since its description over 100 years ago. This species was shown to be genetically distinct from other Baylisascaris spp. (B. columnaris, B. procyonis, B. transfuga, B. devosi). The possible zoonotic significance of ascarids from wild carnivores is discussed in the light of conditions in Central Asia.

Topography and morphology of the Eira barbara diaphragm.

BACKGROUND: The diaphragm, the main muscle involved in respiration and one of those responsible for maintaining life, is still little explored in terms of its morphology in wild animals. There are few studies on the anatomy of Eira barbara, a carnivorous mustelid that is a victim of the urbanization process. In order to contribute to the conservation of the species, we described the topography and morphology of the diaphragm, which may be involved in injuries caused by the impacts of human activities. RESULTS: We studied five specimens of Eira barbara, whose diaphragmatic muscle had a dorsal insertion on the 14th thoracic vertebra, laterally between the 8th and 13th intercostal space (EIC) and ventrally on the 8th EIC, with attachment to the xiphoid process. Consisting of three muscle regions (lumbar, costal and sternal), the diaphragm in Eira barbara showed radially arranged bundles, with the right costal muscle being slimmer than the left; the left pillar wider than the right and between them were the aortic and esophageal hiatuses. The Y-shaped tendinous center housed the foramen of the vena cava bordering the right costal region. In the most dorsal portion of the diaphragm, between the costal regions and the diaphragmatic pillar, we found two triangular-shaped regions devoid of muscle. CONCLUSIONS: Our findings, when compared with the current literature, indicate that the location and positioning of the diaphragm are independent of the physical conformation of the species, and that the right costal region, as well as the triangular areas devoid of musculature, may be fragile points for herniation in cases of Eira barbara being run over.

Sonographic features of abdominal organs in healthy captive Neotropical otters (Lontra longicaudis).

This study aimed to assess the sonographic features of abdominal organs in healthy captive Neotropical otters (Lontra longicaudis). The sonograph showed that the urinary bladder was located in the caudal abdomen and its content was uniformly anechoic. The bladder wall had three layers: outer hyperechoic serosa, hypoechoic muscular layer and hyperechoic mucosa. The mean total wall thickness was 0.10 cm. The splenic parenchyma had a homogeneous echotexture with greater echogenicity than that of the left renal cortex and liver. The borders were regular and tapered. Rugal folds were observed in the stomach when undistended. The mean gastric wall thickness was 0.28 cm and five distinct layers were visible. The liver had smooth contours, and division of the hepatic lobes was not precisely visualized. The hepatic parenchyma had a homogeneous echotexture with greater echogenicity as compared to the right kidney and lesser echogenicity than that of the spleen. The gallbladder appeared as an oval or rounded structure in the transverse plane, filled with homogeneous anechogenic content; the wall was thin, regular and hyperechoic, with a mean thickness of 0.09 cm. The kidneys had a lobulated appearance with renicular subunits. The mean total length of the kidneys in longitudinal plane was 6.18 cm for the left and 6.27 cm for the right. Each reniculus was covered by an echogenic capsule, and the medullary region was more hypoechogenic than the cortical region. In conclusion, sonographic features in most of the abdominal organs in Neotropical otters are similar to the ultrasound patterns observed in healthy dogs and cats, except for kidneys with reniculi.

Diversity of Crenosoma species in mustelids with the first molecular characterization of C. melesi and C. petrowi.

Species of genus Crenosoma have a wide distribution and are reported in Europe, the Americas, and Asia. Currently, the genus includes 14 nominal species, out of which 9 are parasitic in mustelids. Two species are mostly reported in mustelids from Europe, namely C. melesi and C. petrowi. Up to now, no genetic sequences are deposited in GenBank for any of the two. The aims of this study were to investigate the distribution, prevalence, and diversity of Crenosoma spp. infecting mustelids in Romania and to genetically characterize the species. Mustelids (n = 247) were collected over a period of 7 years from different locations in Romania and the respiratory tract was removed and examined for nematodes. Detected nematodes were morphologically identified and fragments of two genes were sequenced. Sampled mustelids included Eurasian badger, Meles meles (n = 102), Eurasian otter, Lutra lutra (n = 20), beech marten, Martes foina (n = 36), European pine marten, Martes martes (n = 5), steppe polecat, Mustela eversmanii (n = 1), European mink, Mustela lutreola (n = 1), least weasel, Mustela nivalis (n = 2), European polecat, Mustela putorius (n = 78), and marbled polecat, Vormela peregusna (n = 1). Nematodes from Eurasian badgers were morphologically identified as C. melesi (n = 13, 12.74%) and C. petrowi (n = 3, 2.94%). Nematodes from the beech martens were identified as C. petrowi (n = 6, 16.66%), C. vulpis (n = 1, 2.78%) and Crenosoma spp. (n = 3, 8.33%). Co-infections with two Crenosoma species were detected in one beech marten (C. petrowi + C. vulpis, n = 1, 2.77%) and in one European pine marten [C. petrowi + C. vulpis (n = 1, 20%)]. Two genes of Crenosoma melesi and C. petrowi were partly sequenced for the first time. We report new host-parasite associations for M. martes and C. vulpis. However, further studies are needed in order to determine the host-parasite associations and to improve the understanding of the epidemiology of Crenosoma nematodes.

Discovery of a coronavirus in the Eurasian badger (Meles meles) belonging to a putative new genus.

In the aftermath of COVID-19, coronaviruses gained renewed attention by the scientific community. The study reports the identification and genetic characterization of a novel coronavirus in the European badger (Meles meles) obtained in the framework of passive surveillance implemented in Italian wildlife in response to the pandemic. Positive samples were characterized using next generation sequencing as well as genetic and phylogenetic analyses, aiming for taxonomic placement under ICTV guidelines of the viruses contained in each sample. Results obtained for six conserved domains within the polyprotein showed that the virus clustered as outgroup and shared <46% amino acid identity with other coronaviruses, supporting the assumption that it belongs to a new putative genus Epsiloncoronavirus. This finding highlights that mammals still hide diverse coronaviruses whose zoonotic and epizootic potential remains unknown.

Intestinal T-cell lymphoma in an Asian small-clawed otter: case report and literature review of lymphoma in the subfamily Lutrinae.

The Asian small-clawed otter (Aonyx cinereus) is an endangered species that is common in zoologic collections. A 17-y-old female Asian small-clawed otter under human care, with a clinical history of chronic renal disease, was euthanized because of deteriorating health. Histologically, the jejunal wall was infiltrated by a monomorphic population of small neoplastic lymphocytes that expanded the lamina propria of the villi and crypts, and on rare occasions invaded the submucosa. The tumor was composed of T cells (CD3+) with a proliferation index of 16%. Based on the World Health Organization (WHO) Classification of Hematopoietic Neoplasms in Domestic Animals, this lymphoma was classified as an enteropathy-associated T-cell lymphoma (EATL) type 2. We also present here a review of the literature on intestinal lymphoma in the subfamily Lutrinae (otters).

Detection and sequence analysis of Canine morbillivirus in multiple species of the Mustelidae family.

BACKGROUND: Canine morbillivirus (canine distemper virus, CDV) is a member of the Paramyxoviridae family. Canine distemper is a serious viral disease that affects many mammalian species, including members of the Mustelidae family. These animals have an elusive nature, which makes related virological studies extremely challenging. There is a significant knowledge gap about the evolution of their viruses and about the possible effects of these viruses to the population dynamics of the host animals. Spleen and lung tissue samples of 170 road-killed mustelids belonging to six species were collected between 1997 and 2022 throughout Hungary and tested for CDV with real-time RT-PCR. RESULTS: Three species were positive for viral RNA, 2 out of 64 Steppe polecats (Mustela eversmanii), 1 out of 36 European polecats (Mustela putorius) and 2 out of 36 stone martens (Martes foina); all 18 pine martens (Martes martes), 10 least weasels (Mustela nivalis) and 6 stoats (Mustela erminea) tested negative. The complete CDV genome was sequenced in five samples using pan-genotype CDV-specific, amplicon-based Nanopore sequencing. Based on the phylogenetic analysis, all five viral sequences were grouped to the Europe/South America 1 lineage and the distribution of one sequence among trees indicated recombination of the Hemagglutinin gene. We verified the recombination with SimPlot analysis. CONCLUSIONS: This paper provides the first CDV genome sequences from Steppe polecats and additional complete genomes from European polecats and stone martens. The infected specimens of various species originated from distinct parts of the country over a long time, indicating a wide circulation of CDV among mustelids throughout Hungary. Considering the high virulence of CDV and the presence of the virus in these animals, we highlight the importance of conservation efforts for wild mustelids. In addition, we emphasize the importance of full genomic data acquisition and analysis to better understand the evolution of the virus. Since CDV is prone to recombination, specific genomic segment analyses may provide less representative evolutionary traits than using complete genome sequences.

Immunogenicity, Safety, and Anti-Viral Efficacy of a Subunit SARS-CoV-2 Vaccine Candidate in Captive Black-Footed Ferrets (Mustela nigripes) and Their Susceptibility to Viral Challenge.

A preliminary vaccination trial against the emergent pathogen, SARS-CoV-2, was completed in captive black-footed ferrets (Mustela nigripes; BFF) to assess safety, immunogenicity, and anti-viral efficacy. Vaccination and boosting of 15 BFF with purified SARS-CoV-2 S1 subunit protein produced a nearly 150-fold increase in mean antibody titers compared to pre-vaccination titers. Serum antibody responses were highest in young animals, but in all vaccinees, antibody response declined rapidly. Anti-viral activity from vaccinated and unvaccinated BFF was determined in vitro, as well as in vivo with a passive serum transfer study in mice. Transgenic mice that received BFF serum transfers and were subsequently challenged with SARS-CoV-2 had lung viral loads that negatively correlated (p < 0.05) with the BFF serum titer received. Lastly, an experimental challenge study in a small group of BFF was completed to test susceptibility to SARS-CoV-2. Despite viral replication and shedding in the upper respiratory tract for up to 7 days post-challenge, no clinical disease was observed in either vaccinated or naive animals. The lack of morbidity or mortality observed indicates SARS-CoV-2 is unlikely to affect wild BFF populations, but infected captive animals pose a potential risk, albeit low, for humans and other animals.

Risks of SARS-CoV-2 transmission between free-ranging animals and captive mink in the Netherlands.

In the Netherlands, 69 of the 126 (55%) mink farms in total became infected with SARS-CoV-2 in 2020. Despite strict biosecurity measures and extensive epidemiological investigations, the main transmission route remained unclear. A better understanding of SARS-CoV-2 transmission between mink farms is of relevance for countries where mink farming is still common practice and can be used as a case study to improve future emerging disease preparedness. We assessed whether SARS-CoV-2 spilled over from mink to free-ranging animals, and whether free-ranging animals may have played a role in farm-to-farm transmission in the Netherlands. The study encompassed farm visits, farm questionnaires, expert workshops and SARS-CoV-2 RNA and antibody testing of samples from target animal species (bats, birds and free-ranging carnivores). In this study, we show that the open housing system of mink allowed access to birds, bats and most free-ranging carnivores, and that direct and indirect contact with mink was likely after entry, especially for free-ranging carnivores and birds. This allowed SARS-CoV-2 exposure to animals entering the mink farm, and subsequent infection or mechanical carriage by the target animal species. Moreover, mink can escape farms in some cases, and two SARS-CoV-2-positive mink were found outside farm premises. No other SARS-CoV-2-RNA-positive free-ranging animals were detected, suggesting there was no abundant circulation in the species tested during the study period. To investigate previous SARS-CoV-2 infections, SARS-CoV-2 antibody detection using lung extracts of carcasses was set up and validated. One tested beech marten did have SARS-CoV-2 antibodies, but the closest SARS-CoV-2-infected mink farm was outside of its home range, making infection at a mink farm unlikely. Knowing that virus exchange between different species and the formation of animal reservoirs affects SARS-CoV-2 evolution, continued vigilance and monitoring of mink farms and surrounding wildlife remains vital.

Extensive genome introgression between domestic ferret and European polecat during population recovery in Great Britain.

The European polecat (Mustela putorius) is a mammalian predator which occurs across much of Europe east to the Ural Mountains. In Great Britain, following years of persecution the range of the European polecat contracted and by the early 1900s was restricted to unmanaged forests of central Wales. The European polecat has recently undergone a population increase due to legal protection and its range now overlaps that of feral domestic ferrets (Mustela putorius furo). During this range expansion, European polecats hybridized with feral domestic ferrets producing viable offspring. Here, we carry out population-level whole-genome sequencing on 8 domestic ferrets, 19 British European polecats, and 15 European polecats from the European mainland. We used a range of population genomics methods to examine the data, including phylogenetics, phylogenetic graphs, model-based clustering, phylogenetic invariants, ABBA-BABA tests, topology weighting, and Fst. We found high degrees of genome introgression in British polecats outside their previous stronghold, even in those individuals phenotyped as "pure" polecats. These polecats ranged from presumed F1 hybrids (gamma = 0.53) to individuals that were much less introgressed (gamma = 0.2). We quantify this introgression and find introgressed genes containing Fst outliers associated with cognitive function and sight.

Evidence of antibodies against SARS-CoV-2 in wild mustelids from Brittany (France).

In the French region of Brittany, mainly in the department of the Côtes d'Armor, during the first half of 2021, seropositivity for SARS-CoV-2 was detected in five wild mustelids out of 33 animals tested (15.6%). Anti-SARS-CoV-2 IgG was detected against at least four out of five recombinant viral proteins (S1 receptor binding domain, nucleocapsid, S1 subunit, S2 subunit and spike) in three pine martens (Martes martes) and in two badgers (Meles meles) using the automated western blot technique. An ELISA test also identified seropositive cases, although these did not align with western blot results. Although the 171 qPCRs carried out on samples from the 33 mustelids were all negative, these preliminary results from this observational study nevertheless bear witness to infections of unknown origin. The epidemiological surveillance of Covid-19 in wildlife must continue, in particular with effective serology tools.

Multiple types of genomic variation contribute to adaptive traits in the mustelid subfamily Guloninae.

Species of the mustelid subfamily Guloninae inhabit diverse habitats on multiple continents, and occupy a variety of ecological niches. They differ in feeding ecologies, reproductive strategies and morphological adaptations. To identify candidate loci associated with adaptations to their respective environments, we generated a de novo assembly of the tayra (Eira barbara), the earliest diverging species in the subfamily, and compared this with the genomes available for the wolverine (Gulo gulo) and the sable (Martes zibellina). Our comparative genomic analyses included searching for signs of positive selection, examining changes in gene family sizes and searching for species-specific structural variants. Among candidate loci associated with phenotypic traits, we observed many related to diet, body condition and reproduction. For example, for the tayra, which has an atypical gulonine reproductive strategy of aseasonal breeding, we observed species-specific changes in many pregnancy-related genes. For the wolverine, a circumpolar hypercarnivore that must cope with seasonal food scarcity, we observed many changes in genes associated with diet and body condition. All types of genomic variation examined (single nucleotide polymorphisms, gene family expansions, structural variants) contributed substantially to the identification of candidate loci. This argues strongly for consideration of variation other than single nucleotide polymorphisms in comparative genomics studies aiming to identify loci of adaptive significance.

Differential susceptibility of SARS-CoV-2 in animals: Evidence of ACE2 host receptor distribution in companion animals, livestock and wildlife by immunohistochemical characterisation.

Angiotensin converting enzyme 2 (ACE2) is a host cell membrane protein (receptor) that mediates the binding of coronavirus, most notably SARS coronaviruses in the respiratory and gastrointestinal tracts. Although SARS-CoV-2 infection is mainly confined to humans, there have been numerous incidents of spillback (reverse zoonoses) to domestic and captive animals. An absence of information on the spatial distribution of ACE2 in animal tissues limits our understanding of host species susceptibility. Here, we describe the distribution of ACE2 using immunohistochemistry (IHC) on histological sections derived from carnivores, ungulates, primates and chiroptera. Comparison of mink (Neovison vison) and ferret (Mustela putorius furo) respiratory tracts showed substantial differences, demonstrating that ACE2 is present in the lower respiratory tract of mink but not ferrets. The presence of ACE2 in the respiratory tract in some species was much more restricted as indicated by limited immunolabelling in the nasal turbinate, trachea and lungs of cats (Felis catus) and only the nasal turbinate in the golden Syrian hamster (Mesocricetus auratus). In the lungs of other species, ACE2 could be detected on the bronchiolar epithelium of the sheep (Ovis aries), cattle (Bos taurus), European badger (Meles meles), cheetah (Acinonyx jubatus), tiger and lion (Panthera spp.). In addition, ACE2 was present in the nasal mucosa epithelium of the serotine bat (Eptesicus serotinus) but not in pig (Sus scrofa domestica), cattle or sheep. In the intestine, ACE2 immunolabelling was seen on the microvillus of enterocytes (surface of intestine) across various taxa. These results provide anatomical evidence of ACE2 expression in a number of species which will enable further understanding of host susceptibility and tissue tropism of ACE2 receptor-mediated viral infection.

SARS-CoV-2 in animals: From potential hosts to animal models.

Within only one year after the first detection of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), nearly 100 million infections were reported in the human population globally, with more than two million fatal cases. While SARS-CoV-2 most likely originated from a natural wildlife reservoir, neither the immediate viral precursor nor the reservoir or intermediate hosts have been identified conclusively. Due to its zoonotic origin, SARS-CoV-2 may also be relevant to animals. Thus, to evaluate the host range of the virus and to assess the risk to act as potential animal reservoir, a large number of different animal species were experimentally infected with SARS-CoV-2 or monitored in the field in the last months. In this review, we provide an update on studies describing permissive and resistant animal species. Using a scoring system based on viral genome detection subsequent to SARS-CoV-2 inoculation, seroconversion, the development of clinical signs and transmission to conspecifics or humans, the susceptibility of diverse animal species was classified on a semi-quantitative scale. While major livestock species such as pigs, cattle and poultry are mostly resistant, companion animals appear moderately susceptible, while several model animal species used in research, including several Cricetidae species and non-human primates, are highly susceptible to SARS-CoV-2 infection. By natural infections, it became obvious that American minks (Neovison vison) in fur farms, e.g., in the Netherlands and Denmark are highly susceptible resulting in local epidemics in these animals.

Pathology of Urinary Bladder in Pearsonema spp. Infected Wildlife from Central Italy.

The genus Pearsonema, in the nematode family Capillariidae, includes several species that parasitize the urinary bladders of wild and domestic carnivores. The infection has been reported worldwide from several wildlife species, including canids, mustelids, and felids, but the pathological aspects have seldom been investigated. In order to assess the presence and severity of the lesions in Pearsonema-infected wildlife, we performed a parasitological and pathological examination of urinary bladders from 72 animals, belonging to the families Canidae (red fox Vulpes vulpes, n = 28, and wolf Canis lupus, n = 29) and Mustelidae (beech marten Martes foina, n = 3; pine marten Martes martes, n = 2; and European badger Meles meles, n = 10). A greater prevalence of infection for canids (64.91%; 95% confidence interval (95% CI), 52.52-77.30%) than for mustelids (13.33%) (p < 0.001) was recorded. The prevalence of infection in red foxes was 75.0% (95% CI, 58.96-91.04%), in accordance with other reports from European countries, supporting the role of this species as a reservoir for infection. Eosinophilic cystitis was observed in 34 out of the 72 examined animals (47.22%). The influence of Pearsonema sp. infection on the occurrence of eosinophilic cystitis was statistically significant in wolves (p < 0.01), which were also affected by more severe histological lesions compared to foxes.

Coronaviruses Associated with the Superfamily Musteloidea.

Among the animal superfamily Musteloidea, which includes those commonly known as mustelids, naturally occurring and species-specific alphacoronavirus infections have been observed in both mink (Mustela vison/Neovison vison) and domestic ferrets (Mustela putorius furo). Ferret systemic coronavirus (FRSCV), in particular, has been associated with a rare but fatal systemic disease. In recent months, it has become apparent that both minks and ferrets are susceptible to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a betacoronavirus and the cause of the coronavirus disease 2019 (COVID-19) pandemic. Several mink farms have experienced SARS-CoV-2 outbreaks, and experimental models have demonstrated susceptibility of ferrets to SARS-CoV-2. The potential for pet ferrets to become infected with SARS-CoV-2, however, remains elusive. During the 2002-2003 SARS epidemic, it was also apparent that ferrets were susceptible to SARS-CoV and could be utilized in vaccine development. From a comparative standpoint, understanding the relationships between different infections and disease pathogenesis in the animal superfamily Musteloidea may help elucidate viral infection and transmission mechanisms, as well as treatment and prevention strategies for coronaviruses.

Cryopreservation of ferret (Mustela putorius furo) sperm collected by rectal massage and electroejaculation: Comparison of a decelerating and an accelerating freezing rate protocol.

The domestic ferret (Mustela putorius furo) provides a good model for developing new reproductive technologies for use with threatened related species. Such technologies could also be used in the reproductive management of this pet species. The present work reports an improved freezing protocol for ferret sperm. Semen was collected by electroejaculation plus rectal massage (in an attempt to reduce the electrical stimulation necessary) from five adult male ferrets, and then subjected to one of two freezing protocols: (a) from 5 to -35°C at 40°C/min, then from -35 to -65°C at 17°C/min, and finally from -65 to -85°C at 3°C/min-a decelerating freezing rate; and (b) from 5 to - 10°C at 5°C/min, and then from -10 to -130°C at 60°C/min-an accelerating freezing rate. After thawing, the viability and acrosomal integrity of the sperm frozen via the two-step accelerating method were better than those frozen via the three-step decelerating method (43.3 ± 3.5% and 71.2 ± 3.4% compared with 29.7 ± 3.7% and 58.8 ± 3.4% respectively; p < .05). No differences were seen between the methods with respect to sperm motility variables; most sperm (>90%) remained static with both freezing methods. In conclusion, although the method with accelerating freezing rate was associated with better post-thaw sperm viability and acrosome integrity values, neither of the two freezing methods tested provided adequate motility results after thawing. Combining rectal massage with electrical stimuli seemed to reduce the number of the latter required for successful sperm collection.

Molecular survey of Bartonella spp. and haemoplasmas in American minks (Neovison vison).

The aim of this study was to perform a molecular survey and characterize Bartonella spp. and haemotropic Mycoplasma (haemoplasmas) in invasive American minks (Neovison vison) from Southern Chile. Additionally, we addressed risk factors for positivity in both groups of agents. Blood and/or tissue samples from 246 minks were analysed by qPCR targeting the nuoG gene for Bartonella spp. and conventional (c)PCR for 16S rRNA for haemotropic Mycoplasma spp. nuoG qPCR-positive Bartonella spp. samples were submitted to cPCR assays (ITS, ribC, gltA, rpoB, pap-31 and ftsZ genes) to perform phylogenetic inferences. Haemotropic Mycoplasma spp. 16S-positive samples were further amplified by cPCR targeting RNaseP gene (160-210 bp) and by two overlapping 16S rRNA cPCR assays to amplify a larger portion of the gene (1,200bp) for phylogenetics. Bartonella DNA was detected in 8.9% of minks (22/246). Out of 22 nuoG qPCR-positive samples, one and two showed positive results in cPCR assays based on ITS and ribC, respectively. Consistent sequencing results were obtained for only one ITS sample (464 bp sequence), which shared 99.6% identity with B. clarridgeiae. Two per cent of minks (5/246) were positive for 16S rRNA haemotropic Mycoplasma-cPCR assay. Two concatenated sequences of 16S rRNA (1,176 and 1,230 bp) were obtained: one sample shared 97.87% identity with haemotropic Mycoplasma sp. from a wild rodent, and the other 96.49% identity with 'Candidatus Mycoplasma haematoparvum' from a dog. All BLAST results were supported by phylogenetic analysis. One haemoplasma RNase P sequence shared 94.86% identity with Mycoplasma haemofelis from a cat. No risk factors for PCR positivity were identified. In a nutshell, Bartonella clarridgeiae and a potentially novel haemoplasma closely related to haemoplasmas previously reported in rodents, dogs, domestic and wild cats were described for the first time in American minks.

Multi-species ELISA for the detection of antibodies against SARS-CoV-2 in animals.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a pandemic with millions of infected humans and hundreds of thousands of fatalities. As the novel disease - referred to as COVID-19 - unfolded, occasional anthropozoonotic infections of animals by owners or caretakers were reported in dogs, felid species and farmed mink. Further species were shown to be susceptible under experimental conditions. The extent of natural infections of animals, however, is still largely unknown. Serological methods will be useful tools for tracing SARS-CoV-2 infections in animals once test systems are evaluated for use in different species. Here, we developed an indirect multi-species ELISA based on the receptor-binding domain (RBD) of SARS-CoV-2. The newly established ELISA was evaluated using 59 sera of infected or vaccinated animals, including ferrets, raccoon dogs, hamsters, rabbits, chickens, cattle and a cat, and a total of 220 antibody-negative sera of the same animal species. Overall, a diagnostic specificity of 100.0% and sensitivity of 98.31% were achieved, and the functionality with every species included in this study could be demonstrated. Hence, a versatile and reliable ELISA protocol was established that enables high-throughput antibody detection in a broad range of animal species, which may be used for outbreak investigations, to assess the seroprevalence in susceptible species or to screen for reservoir or intermediate hosts.

Distribution and habitat use intensity of the Neotropical Otter (Lontra longicaudis) in a Colombian hydroelectric dam / Distribución e intensidad de uso del hábitat de la nutria neotropical (Lontra longicaudis Olfers, 1818) en una represa hidroeléctrica colombiana

Introduction: The Neotropical Otter, Lontra longicaudis, is a semi-aquatic mammal that ranges from Mexico to Argentina inhabiting near-pristine watercourses, but also human-dominated aquatic and riparian landscapes. Objective: We assessed the distribution and habitat use frequency of L. longicaudis in La Miel I hydroelectric power dam and its influence area in the Department of Caldas, Colombia. Methods: We carried out diurnal surveys across standardized transects between 2014 and 2018 looking for records (sightings, faeces, tracks, and dens) that indicate the presence of the species. Each yearly survey was done during 12 consecutive days over three seasonal sampling periods assessing the upstream, reservoir, and downstream waterscape areas. Results: We sampled a total of 875 km in a five-year period across the three main waterscape areas (upstream -103 km, reservoir -582 km, and downstream -190 km) registering a total of 1 496 records. Faeces were the most common record (~ 95 %) across the whole study area followed by sightings, dens (1.7 % each), and tracks (1.3 %). Spatial distribution analyses suggest that L. longicaudis prefers dwelling upstream watercourses (hot spots areas; Gi Z-score = 4.46, p < 0.001) and in a lesser extent, areas around the water reservoir (cold spot areas; Gi Z-score = -2.69, p = 0.007). Signs of otters were also recorded at downstream area, but these records were non-significant within the analysis (Gi Z-score = -0.11, p = 0.48), suggesting L. longicaudis uses this area opportunistically. Cluster and outlier analysis showed that even though L. longicaudis was commonly found upstream and in the reservoir area, only some specific sectors (Moro, La Miel and Tasajos rivers) had high (LMI Z-score = 5.63, p = 0.001) and low (LMI Z-score = 2.12, p = 0.001) clusters. Conclusions: The upstream waterscape area is key for the survival of L. longicaudis in this regulated system, likely providing enough shelter and food for the species to carry out living activities and have resident populations. In contrast, downstream areas require specific attention to understand in a better way the effects of caused by the dam on the species dynamics, also defining management strategies that avoid population fragmentation and movement reduction.

Introducción : La nutria neotropical Lontra longicaudis, es un mamífero semiacuático que se distribuye desde México hasta Argentina; habita en paisajes acuáticos y ribereños prístinos, pero también hábitats dominados por el hombre. Objetivo : Se evaluó la distribución e intensidad de uso del hábitat de L. longicaudis en la hidroeléctrica La Miel I y su área de influencia en el Departamento de Caldas, Colombia. Métodos: Se realizaron transectos diurnos estandarizados entre 2014 y 2018 en busca de registros (avistamientos, heces, huellas y madrigueras) que indicaban la presencia de la especie. Cada muestreo anual se realizó durante 12 días consecutivos en tres períodos de muestreo estacional (36 días), evaluando las áreas aguas arriba, el embalse y aguas abajo de la presa. Resultados : Se muestreó un total de 875 km a través de las tres áreas principales con un total de 1 496 rastros. Las heces fueron el registro más común (~ 95 %) en toda el área de estudio, seguido de avistamientos, madrigueras (1.7 % cada una), y huellas (1.3 %). Los análisis de distribución espacial sugieren que L. longicaudis prefiere los afluentes aguas arriba (áreas de puntos calientes; Gi Z-score = 4.46, p < 0.001) y, en menor medida, las zonas alrededor del embalse (áreas de puntos fríos; Gi Z-score = -2.69, p = 0.007). El área aguas abajo también mostró presencia de nutrias; sin embargo, esos registros no fueron significativos dentro del análisis (Gi Z-score = -0.11, p = 0.48), sugiriendo que L. longicaudis usa esta área esporádicamente. El análisis de conglomerados y datos atípicos mostró que, aunque L. longicaudis se encuentra comúnmente aguas arriba y en el área del embalse, solo algunos sectores específicos (ríos Moro, La Miel y Tasajos) tienen valores altos (LMI Z-score = 5.63, p= 0.001) y bajos (LMI Z-score = 2.12, p = 0.001) de conglomerados. Conclusiones : El paisaje y las características hidrológicas aguas arriba son claves para la supervivencia de la nutria neotropical en este sistema regulado, probablemente aportando suficiente refugio y alimento para que la especie lleve a cabo sus actividades vitales, permitiéndole tener poblaciones residentes. Se debe enfocar la atención en la zona aguas abajo para comprender el efecto de la presa en la dinámica de la especie, definiendo estrategias de manejo que eviten la fragmentación de la población y la reducción de los movimientos.