Wild Animals in Captivity: An Analysis of Parasite Biodiversity and Transmission among Animals at Two Zoological Institutions with Different Typologies.

We have conducted a 10-year-long coprological study of the animals housed in two zoological institutions (ZooAquarium and Faunia, Madrid, Spain) to assess the parasite biodiversity, prevalence, and their relation with host class, diet, and enclosure type (soil type and level of isolation from wild fauna). A total of 4476 faecal samples from 132 mammal species and 951 samples from 86 avian species were examined. The results indicated that only 12.8% of avian species had parasites at least once during the study period, whereas 62.1% of mammal species tested positive. Predominantly, protists (Entamoeba, flagellates, and ciliates) and nematodes (mainly Trichuris) were identified in the findings. Carnivorous species were primarily infected by nematodes, while herbivorous and omnivorous species were mainly infected by protists. The number of infected herbivorous and omnivorous species was significantly greater than carnivorous species. Differences were observed based on soil type (artificial, natural, mixed) and isolation level (isolated/accessible), but these differences were not statistically significant. Several parasites (Entamoeba spp., Giardia spp., Balantidoides coli, Trichuris spp.) could potentially be transmitted between humans and some mammals and birds. Regular animal analyses and a personnel health program in the institutions would minimise transmission risks between zoo animals, wildlife, and humans.

Widespread circulation and transmission risk of Mycobacterium avium subsp. paratuberculosis at the livestock-wildlife-environment interface in a Mediterranean agro-forestry farmstead.

Mycobacterium avium subsp. paratuberculosis (MAP) is the etiological agent of paratuberculosis, a chronic infection affecting ruminants and other species worldwide. Information on the ecological factors that increase infection risk at the livestock-wildlife-environment interface remains scarce. Thus, this work aimed at determining which factors modulate the exposure of a mammal community within a Mediterranean agro-forestry farmstead to MAP. Through field, molecular and ecological modeling approaches, MAP prevalence, distribution and spatial risk at the livestock-wildlife-environment was estimated in the study area by screening 436 samples (cattle, n = 150; wildlife, n = 206; soil, n = 80). Using molecular detection of IS900 as proxy, MAP was identified in ten wild mammal species. Being a central prey of mesocarnivores in Portugal, the high prevalence of MAP in the wild rabbit (19%) may be related with red fox's (22%). MAP was also detected in cattle managed in the farmstead (animal and herd prevalence, 54% and 100%) and in soil (44%), which may perpetuate intraspecies and interspecies transmission. Wildlife diversity showed a positive influence on MAP presence in wild mammals, while wildlife abundance showed a negative effect. Land use variables exerted distinct degrees of impact upon MAP detection in specific groups of mammals: mixed forest cover showed positive influence on carnivores, and shrubland showed positive effect on wild rabbits. The prevalence of MAP in cattle showed a negative influence on the detection of MAP in lagomorph, which may stem from wild rabbit lower density and avoidance of cattle areas. Based on explanatory variables, the spatial prediction of MAP occurrence in wildlife indicated two hotspots with increased exposure risk but future studies are needed to confirm this projection. This work represents the most comprehensive molecular survey of MAP occurrence and determinants in Mediterranean agroecosystems leveraging the principles and tools of community ecology, debating potential biological and ecological effects underlying MAP transmission.

Vector-Borne Disease in Wild Mammals Impacted by Urban Expansion and Climate Change.

Ecologies of zoonotic vector-borne diseases may shift with climate and land use change. As many urban-adapted mammals can host ectoparasites and pathogens of human and animal health concern, our goal was to compare patterns of arthropod-borne disease among medium-sized mammals across gradients of rural to urban landscapes in multiple regions of California. DNA of Anaplasma phagocytophilum was found in 1-5% of raccoons, coyotes, and San Joaquin kit foxes; Borrelia burgdorferi in one coyote, rickettsiae in two desert kit foxes, and Yersinia pestis in two coyotes. There was serological evidence of rickettsiae in 14-37% of coyotes, Virginia opossums, and foxes; and A. phagocytophilum in 6-40% of coyotes, raccoons, Virginia opossums, and foxes. Of six flea species, one Ctenocephalides felis from a raccoon was positive for Y. pestis, and Ct. felis and Pulex simulans fleas tested positive for Rickettsia felis and R. senegalensis. A Dermacentor similis tick off a San Joaquin kit fox was PCR-positive for A. phagocytophilum. There were three statistically significant risk factors: risk of A. phagocytophilum PCR-positivity was threefold greater in fall vs the other three seasons; hosts adjacent to urban areas had sevenfold increased A. phagocytophilum seropositivity compared with urban and rural areas; and there was a significant spatial cluster of rickettsiae within greater Los Angeles. Animals in areas where urban and rural habitats interconnect can serve as sentinels during times of change in disease risk.

Mammal Roadkills in Lithuanian Urban Areas: A 15-Year Study.

We investigated roadkills in urban areas in Lithuania from 2007 to 2022, including two periods with COVID-19 restrictions on people's movement. We analyzed the proportions of wild and domestic animals in roadkill, annual trends, the predominant species involved, and monthly changes during the restrictions. Urban roads were characterized by a low species diversity of roadkilled mammals, with roe deer (Capreolus capreolus) dominating. Total numbers increased exponentially during the study period. The proportion of domestic animals, 12.2%, significantly exceeded that on non-urban roads in the country. The proportion of domestic animals decreased from over 40% in 2007-2009 to 3.7-5.4% in 2020-2022, while the proportion of wild mammals increased from 36.1-39.6% to 89.9-90.6%, respectively. During the periods of COVID-19 restrictions, the number of roadkills in urban areas was significantly higher than expected based on long-term trends. Compared to 2019, the number of roadkilled roe deer in 2020-2021 almost doubled from 700 to 1281-1325 individuals. These anthropause effects were, however, temporary. The imbalance between the roadkill number and transport intensity might require new mitigation strategies to sustain mammal populations in urban areas, at least through improving driver awareness on the issue.

Occurrence of Leishmania infantum in Wild Mammals Admitted to Recovery Centers in Spain.

Zoonotic leishmaniasis caused by Leishmania infantum is distributed worldwide and affects humans and domestic and wild mammals. In Europe, specifically in the Mediterranean basin, leishmaniasis is endemic due to the concurrence of the phlebotomine vectors and reservoir mammals, including carnivorous wildlife species and other less studied wild species. In this article, spleen, skin, and eye or oral swabs taken from 134 wild mammals admitted to five wildlife recovery centers in Spain were used. PCR employing fragments of the Repeat region, ITS1, and SSUrRNA were used for detection, and positive samples were processed for sequencing. L. infantum was detected in three out of the nine species analyzed, including European hedgehog, European badger, and red squirrel, with percentages ranging from 11.53 to 35.71%, depending on the species. Most of the species showed higher percentages of positivity in spleen samples than in skin samples. A small number of animals from the remaining six species tested negative, including Algerian hedgehog, stone marten, least weasel, garden dormouse, western polecat, and Egyptian mongoose. Hedgehogs and badgers are good candidates for consideration as epidemiological sentinels and pose a higher risk as potential reservoirs of leishmaniasis based on their percentage of infection and wide distribution.

Prevalence of gastrointestinal parasitic infections in wild mammals of a safari park and a zoo in Bangladesh.

In safari parks and zoos, wild animals are kept mainly for recreational purposes. Animals in these enclosures are also crucial for the education, research, and conservation aspect. To ensure better management and good health of wild animals in captivity, it is essential to monitor the occurrence of gastrointestinal (GI) parasitic (helminths and protozoa) infections. The current investigation was undertaken to investigate the prevalence of GI parasitic infections in wild mammals at Bangabandhu Sheikh Mujib (BSM) safari park and Chattogram (CTG) zoo of Bangladesh. A total of 72 individual faecal samples were collected from 25 species of wild mammals. Routine qualitative (e.g. direct smear, sedimentation, and flotation) and quantitative (e.g. McMaster technique) tests were performed to identify the eggs or oocysts of helminths and protozoa. Results demonstrated that wild mammals of both BSM safari park and CTG zoo were infected with a total of 17 genera/species of helminths and protozoa. The overall prevalence of GI parasitic infections in wild mammals of both zoological parks was 65.3% (95% confidence interval [CI]: 53.14-76.12), whereas it was 72.4% (95% CI: 52.76-87.27) in the BSM safari park and 60.5% (95% CI: 44.41-75.02) in the CTG zoo. In both zoological parks, infection with nematodes was more frequent compared to other helminth into the wild mammals. The herbivores were more infected with GI parasites than carnivores and omnivores of both BSM safari park and CTG zoo. The mean eggs/oocysts per gram of faeces was the highest in the carnivores compared to herbivores and omnivores of both enclosures. The findings of the current study demonstrated that wild mammals of both BSM safari park and CTG zoo suffered from various GI parasitic infections. Regular monitoring along with proper therapeutic measures may reduce the severe consequences of GI parasitic infections in captive wild animals.

"Mi Casa, Tu Casa": the coati nest as a hub of Trypanosoma cruzi transmission in the southern Pantanal biome revealed by molecular blood meal source identification in triatomines.

BACKGROUND: The study of the ecology of Trypanosoma cruzi is challenging due to its extreme adaptive plasticity, resulting in the parasitism of hundreds of mammal species and dozens of triatomine species. The genetic analysis of blood meal sources (BMS) from the triatomine vector is an accurate and practical approach for gathering information on which wild mammal species participate in a local transmission network. South American coatis, Nasua nasua, act as important reservoir host species of T. cruzi in the Pantanal biome because of their high rate of infection and elevated parasitemia, with the main discrete typing unit (DTU) lineages (TcI and TcII). Moreover, the carnivore coati is the only mammal species to build high arboreal nests for breeding and resting that can be shared by various vertebrate and invertebrate species. Herein, we applied the sensitive and specific methodology of DNA barcoding and molecular cloning to study triatomines found in a coati nest to access the diversity of mammal species that explore this structure, and therefore, may be involved in the parasite transmission network. METHODS: Twenty-three Triatoma sordida were collected in one coati's nest in the subregion of Nhecolândia, Pantanal. The DNA isolated from the gut of insects was subjected to BMS detection by PCR using universal primers that flank variable regions of the cytochrome b (cytb) and 12S rDNA mitochondrial genes from vertebrates. The Trypanosoma spp. diagnosis and DTU genotyping were based on an 18S rDNA molecular marker and also using new cytb gene primers designed in this study. Phylogenetic analyses and chord diagrams were constructed to visualize BMS haplotypes, DTU lineages detected on vectors, and their interconnections. RESULTS: Twenty of 23 triatomines analyzed were PCR-positive (86.95%) showing lineages T. cruzi DTU TcI (n = 2), TcII (n = 6), and a predominance of TcI/TcII (n = 12) mixed infection. Intra-DTU diversity was observed mainly from different TcI haplotypes. Genetic analyses revealed that the southern anteater, Tamandua tetradactyla, was the unique species detected as the BMS of triatomines collected from the coati's nest. At least three different individuals of T. tetradactyla served as BMS of 21/23 bugs studied, as indicated by the cytb and 12S rDNA haplotypes identified. CONCLUSIONS: The identification of multiple BMS, and importantly, different individuals of the same species, was achieved by the methodology applied. The study demonstrated that the southern anteaters can occupy the South American coati's nest, serving as the BMS of T. sordida specimens. Since anteaters have an individualist nonsocial behavior, the three individuals detected as BMS stayed at the coati's nest at different times, which added a temporal character to BMS detection. The TcI and TcII infection, and significantly, a predominance of TcI/TcII mixed infection profile with different TcI and TcII haplotypes was observed, due to the discriminatory capacity of the methodology applied. Tamandua tetradactyla, a host which has been little studied, may have an important role in the T. cruzi transmission in that Pantanal subregion. The data from the present study indicate the sharing of coatis' nests by other mammal species, expanding the possibilities for T. cruzi transmission in the canopy strata. We propose that coatis' nests can act as the true hubs of the T. cruzi transmission web in Pantanal, instead of the coatis themselves, as previously suggested.

Advancements in veterinary medicine: the use of Flowgy for nasal airflow simulation and surgical predictions in big felids (a case study in lions).

Flowgy is a semi-automated tool designed to simulate airflow across the nasal passage and detect airflow alterations in humans. In this study, we tested the use and accuracy of Flowgy in non-human vertebrates, using large felids as the study group. Understanding the dynamics of nasal airflow in large felids such as lions (Panthera leo) is crucial for their health and conservation. Therefore, we simulated airflow during inspiration through the nasal passage in three lions (Panthera leo), two of which were siblings (specimens ZPB_PL_002 and ZPB_PL_003), without breathing obstructions. However, one of the specimens (ZPB_PL_001) exhibited a slight obstruction in the nasal vestibule, which precluded the specimen from breathing efficiently. Computed tomography (CT) scans of each specimen were obtained to create detailed three-dimensional models of the nasal passage. These models were then imported into Flowgy to simulate the airflow dynamics. Virtual surgery was performed on ZPB_PL_001 to remove the obstruction and re-simulate the airflow. In parallel, we simulated the respiration of the two sibling specimens and performed an obstructive operation followed by an operation to remove the obstruction at the same level and under the same conditions as the original specimen (ZPB_PL_001). Thus, we obtained a pattern of precision for the operation by having two comparable replicas with the obstructed and operated specimens. The simulations revealed consistent airflow patterns in the healthy specimens, demonstrating the accuracy of Flowgy. The originally obstructed specimen and two artificially obstructed specimens showed a significant reduction in airflow through the right nostril, which was restored after virtual surgery. Postoperative simulation indicated an improvement of >100% in respiratory function. Additionally, the temperature and humidity profiles within the nostrils showed marked improvements after surgery. These findings underscore the potential of Flowgy in simulating nasal airflow and predicting the outcomes of surgical interventions in large felids. This could aid in the early detection of respiratory diseases and inform clinical decision-making, contributing to improved veterinary care and conservation efforts. However, further research is needed to validate these findings in other species and explore the potential of integrating Flowgy with other diagnostic and treatment tools in veterinary medicine.

Worldwide Disseminated IncX4 Plasmid Carrying mcr-1 Arrives to Wild Mammal in Portugal.

The mcr-1 gene spread is worldwide recognized as a public health threat at multidrug-resistant infections therapy level. Here, we report for the first time, to the best of our knowledge, the detection of the globally distributed IncX4 plasmid carrying mcr-1 (mcr-1/IncX4) in Escherichia coli isolated from a wild mammal in Portugal and Europe. This plasmid was found in a ST533 E. coli isolate with a multidrug-resistant profile, virulence potential, and possibly phylogenetically related to human isolates. Our work contributes to highlight the importance of antimicrobial resistance (AMR) surveillance in wildlife, an important compartment of the whole ecosystem often overlooked in the fight against AMR. IMPORTANCE Colistin resistance mediated by plasmids is recognized worldwide as an emergency problem connected with the whole ecosystem, since is well described in the interface of the human-animal-environment. The plasmid IncX4 is reported as one of the most prevalent plasmids harboring the gene mcr-1. On an European scale the plasmid IncX4 carrying mcr-1 has been described in humans, the environment, and animals, including wildlife, but only in wild birds. This study shows the first report of the plasmid IncX4 harboring mcr-1 in a wild mammal in Portugal and Europe, identified in a ST533 E. coli commensal that is, curiously, more related to isolates from humans than from livestock. Our findings show that the plasmid IncX4 harboring mcr-1 is well established in a colistin resistance drive embracing the whole ecosystem.

Antimicrobial Resistance of Enterococcus Species Isolated from Wild Mammals in Aragón, Spain.

Introduction: Antimicrobial resistance is currently one of the major public health threats. In order to prevent its spread, the WHO, OIE and FAO have formed an alliance to promote the study of antibiotic resistance evolution in human, animal and environmental bacteria posing a public health threat; however, the studies performed in wild animals are scarce so far. The main objective of this study was to assess the antibiotic resistance of Enterococcus spp. isolated from wild mammals in Aragón, Spain. Material and Methods: Rectal samples were collected from 103 wild mammals - 70 hunt prey and 33 rescued animals. Isolates were identified by matrix-assisted laser desorption/ionisation-time of flight mass spectrometry and susceptibility tests to 10 antibiotics were also carried out. Statistical analysis was performed (P ≤ 0.05). Results: A total of 126 isolates of seven different Enterococcus species were recovered. Among them, E faecalis (37.60%), E. casseliflavus (20.63%) and E. faecium (17.46%) were the most prevalent. The antibiotics quinupristin-dalfopristin and ciprofloxacin most frequently lost efficacy against the isolates. Multi-drug resistance was more prevalent in enterococci isolated from the rescued mammals. Conclusion: This study found resistance widely distributed among enterococci isolated from the studied mammals. This points to the need for additional study of its genetic determinants and investigation of the sources and measures to avoid contributory environmental contamination.

Flea infestation on small wild mammals in Gharyan, Northwest Libya.

Background: Fleas play a major role as vectors for many pathogens that affect humans and livestock. Wild animals, especially wild rodents, are the most important hosts, acting as reservoir hosts for many flea species and pathogens. Aim: This study aimed to identify seasonality and hosts of fleas that parasitize small wild mammals in Gharyan, north-western Libya. Methods: Fleas were collected from seasonally infested hosts from summer 2017 to winter 2018. Results: This survey identified three flea species: Pulex irritans, Xenopsylla cheopis, and Leptopsylla segnis. Pulex irritans was collected from porcupines, X. cheopis from hedgehogs and jerboas, whereas L. segnis from gundis. The highest flea prevalence was in porcupines (35.00%) and the lowest was in gundis (11.11%). The highest intensity was in porcupines (10.43 ± 4.37), and the lowest was in jerboas (1.28 ± 0.24). The highest mean flea abundance was among porcupines of 3.65, whereas in hedgehogs, jerboas and gundis were less than 0.50 flea/host. Pulex irritans was collected during all seasons, while X. cheopis was collected during all seasons except winter, whereas L. segnis was collected only in spring. Conclusion: The finding in this study has increased our awareness that small wild mammals can act as hosts for medically important flea species. All collected flea species have a great specialty in choosing their hosts. Flea intensity is affected by seasonal variations.

Investigation of Leishmania (Viannia) braziliensis Infection in Wild Mammals in Brazil.

PURPOSE: Leishmaniasis are infectious and zoonotic diseases and present in cutaneous and visceral forms. Cutaneous leishmaniasis is endemic and widely distributed throughout the state of Espírito Santo, Brazil. Several cases of cutaneous leishmaniasis in humans and dogs associated with Leishmania (Viannia) braziliensis have been reported in the state. This study aimed to investigate the occurrence of natural infection by Leishmania(Viannia) braziliensis in wild mammals found dead (by trampling or natural death) in the Sooretama Biological Reserve, Espírito Santo State. METHODS: From January 2018 to December 2019, 60 animals were collected. Of these, 47 animals from 12 different species were analyzed. The results were demonstrated using descriptive analysis of the observations to calculate the absolute and relative frequencies of the data. RESULTS: In the PCR, using specific primers for the genus Leishmania (D1, D2, and D3) and the species Leishmania (Viannia) braziliensis (ISVB/ISVC), 4 positive animals (8.5%) were detected: 1 Cuniculus paca (paca) (25%) and 3 Callithrix geoffroyi (white-faced marmoset) (25%). In the histopathological analysis, the parasitic amastigote form was not observed. CONCLUSIONS: The natural infection, detected by PCR, by Leishmania (Viannia) braziliensis in Cuniculus paca (paca) and Callithrix geoffroyi (white-faced marmoset) constitutes the first report of infection of this rodent and primate species in the literature. Despite the confirmation of Leishmania (Viannia) braziliensis infection in rodents and primates, the role of these species in the transmission of this zoonosis still needs further observational studies to identify their seasonal variation, transmissibility, infection stability, and the effects of a given parasite on the population and/or individual.

Seasonal shift of the gut microbiome synchronizes host peripheral circadian rhythm for physiological adaptation to a low-fat diet in the giant panda.

Characteristics of the gut microbiome vary synchronously with changes in host diet. However, the underlying effects of these fluctuations remain unclear. Here, we performed fecal microbiota transplantation (FMT) of diet-specific feces from an endangered mammal (the giant panda) into a germ-free mouse model. We demonstrated that the butyrate-producing bacterium Clostridium butyricum was more abundant during shoot-eating season than during the leaf-eating season, congruent with the significant increase in host body mass. Following season-specific FMT, the microbiota of the mouse model resembled that of the donor, and mice transplanted with the microbiota from the shoot-eating season grew faster and stored more fat. Mechanistic investigations revealed that butyrate extended the upregulation of hepatic circadian gene Per2, subsequently increasing phospholipid biosynthesis. Validation experiments further confirmed this causal relationship. This study demonstrated that seasonal shifts in the gut microbiome affect growth performance, facilitating a deeper understanding of host-microbe interactions in wild mammals.

Wild Mammals in the Economy of Wroclaw (Poland) as an Example of a Medieval and Modern Era City in the Light of Interdisciplinary Research.

The purpose of this article was to determine the role of wild animals in the economy of a historical city on the basis of archaeological and cultural layers of medieval and early modern Wroclaw from the 11th to the 17th century. Archaeozoological analyses were applied, mainly encompassing the percentage share of particular animal species and the research of material culture, i.e., items manufactured from bones, antlers and hides of wild animals. The collected data were compared with written sources. As a result of the following analysis, a low but stable frequency of bone remains in urban layers and is evidence for occasional breaching of the medieval hunting laws by burghers, possibly driven by the opportunity to sell meat and other wild animal products on the markets. Moreover, the relatively low amounts of items made from bones, antlers and wild animal leather may indicate low availability or seasonality (shed antler) of the materials, which might have indirectly raised the product price. Additionally, the area around Wroclaw did not feature large forest complexes, which are habitats of wild game, thus explaining the low frequency of wild animal remains in the archaeozoological material.

'Candidatus Mycoplasma haematonasua' and tick-borne pathogens in ring-tailed coatis (Nasua nasua Linnaeus, 1976) from the Iguaçu National Park, Paraná State, southern Brazil.

The Iguaçu National Park (INP) is the largest remnant of Atlantic Forest in southern Brazil, representing an ecological continuum with Argentina. The INP harbours a diverse fauna, with ring-tailed coatis (Nasua nasua Linnaeus, 1976, Carnivora: Procyonidae) in close contact with tourists either begging and/or snatching food from visitors. A potentially novel haemotropic Mycoplasma sp. has been previously detected in the ring-tailed coatis from central-western and southern Brazil. Therefore, the aims of this study were to investigate the occurrence of haemotropic Mycoplasma sp. and tick-borne pathogens in wild ring-tailed coatis from the INP, Foz do Iguaçu municipality, Paraná State, southern Brazil. Blood samples were collected from 18 wild ring-tailed coatis and evaluated by conventional PCR (cPCR) assays for haemotropic Mycoplasma spp. (16S and 23S rRNA), Theileria/Babesia spp. (18S rRNA) and Ehrlichia/Anaplasma spp. (16S rRNA, sodB, dsb and groEL). Eight out of 18 (44.44%; 95% CI: 24.56%-66.28%) animals were positive for haemotropic Mycoplasma spp. All ring-tailed coatis tested negative for Theileria/Babesia spp. and only one out of 18 (5.56%; 95% CI: 0.99%-25.76%) animals tested positive for Ehrlichia/Anaplasma spp. by the 16S rRNA cPCR. Unfortunately, multiple attempts to sequence the 16S rRNA gene of the Ehrlichia/Anaplasma-positive sample have failed. Phylogenetic and network analysis of the hemoplasma 16S and 23S rRNA gene fragments confirmed that animals were infected by a potentially novel haemotropic Mycoplasma sp. previously reported in ring-tailed coatis from Brazil. The name 'Candidatus Mycoplasma haematonasua' is proposed for this novel organism.

The spleen bacteriome of wild rodents and shrews from Marigat, Baringo County, Kenya.

BACKGROUND: There is a global increase in reports of emerging diseases, some of which have emerged as spillover events from wild animals. The spleen is a major phagocytic organ and can therefore be probed for systemic microbiome. This study assessed bacterial diversity in the spleen of wild caught small mammals so as to evaluate their utility as surveillance tools for monitoring bacteria in an ecosystem shared with humans. METHODS: Fifty-four small mammals (rodents and shrews) were trapped from different sites in Marigat, Baringo County, Kenya. To characterize their bacteriome, DNA was extracted from their spleens and the V3-V4 regions of the 16S rRNA amplified and then sequenced on Illumina MiSeq. A non-target control sample was used to track laboratory contaminants. Sequence data was analyzed with Mothur v1.35, and taxomy determined using the SILVA database. The Shannon diversity index was used to estimate bacterial diversity in each animal and then aggregated to genus level before computing the means. Animal species within the rodents and shrews were identified by amplification of mitochondrial cytochrome b (cytb) gene followed by Sanger sequencing. CLC workbench was used to assemble the cytb gene sequences, after which their phylogenetic placements were determined by querying them against the GenBank nucleotide database. RESULTS: cytb gene sequences were generated for 49/54 mammalian samples: 38 rodents (Rodentia) and 11 shrews (Eulipotyphyla). Within the order Rodentia, 21 Acomys, eight Mastomys, six Arvicanthis and three Rattus were identified. In the order Eulipotyphyla, 11 Crucidura were identified. Bacteria characterization revealed 17 phyla that grouped into 182 genera. Of the phyla, Proteobacteria was the most abundant (67.9%). Other phyla included Actinobacteria (16.5%), Firmicutes (5.5%), Chlamydiae (3.8%), Chloroflexi (2.6%) and Bacteroidetes (1.3%) among others. Of the potentially pathogenic bacteria, Bartonella was the most abundant (45.6%), followed by Anaplasma (8.0%), Methylobacterium (3.5%), Delftia (3.8%), Coxiella (2.6%), Bradyrhizobium (1.6%) and Acinetobacter (1.1%). Other less abundant (<1%) and potentially pathogenic included Ehrlichia, Rickettsia, Leptospira, Borrelia, Brucella, Chlamydia and Streptococcus. By Shannon diversity index, Acomys spleens carried more diverse bacteria (mean Shannon diversity index of 2.86, p = 0.008) compared to 1.77 for Crocidura, 1.44 for Rattus, 1.40 for Arvicathis and 0.60 for Mastomys. CONCLUSION: This study examined systemic bacteria that are filtered by the spleen and the findings underscore the utility of 16S rRNA deep sequencing in characterizing complex microbiota that are potentially relevant to one health issues. An inherent problem with the V3-V4 region of 16S rRNA is the inability to classify bacteria reliably beyond the genera. Future studies should utilize the newer long read methods of 16S rRNA analysis that can delimit the species composition.

Examples of proper reporting for evaluation (Stage 2 validation) of diagnostic tests for diseases listed by the World Organisation for Animal Health.

Reporting and design standards are key indicators of the quality of diagnostic accuracy (validation) studies but, with the exception of aquatic animal diseases and paratuberculosis in ruminants, there is limited guidance for designing these studies in animals. There is, therefore, a need for generic guidelines that are based on disease characteristics, such as mode of transmission, latent period and pathogenesis. Comprehensive, clear and transparent reporting of primary test accuracy studies for diseases listed by the World Organisation for Animal Health (OIE) has value for the end users of diagnostic tests and, ultimately, for decision-makers, who require systematic reviews and meta-analysis of multiple tests for specified diseases and testing purposes. The recent publication of reporting standards for Bayesian latent class models, to analyse test-accuracy data from naturally occurring disease events, fills an important gap as these methods are being increasingly used for OIE-listed diseases. Adherence to design and reporting standards, as well as to guidelines, helps to ensure that research funding for test validation studies is used appropriately and that the strengths and limitations of single tests or test combinations are made clear to test users. The authors provide a review of key points that are often overlooked or misinterpreted in test validation studies, as well as two concrete examples of good practice for use as a reference point for future studies.

Les normes de notification et de conception sont des indicateurs essentiels de la qualité des études de validation des tests destinées à déterminer leur exactitude diagnostique ; or, en dehors des maladies des animaux aquatiques et de la paratuberculose chez les ruminants, il n'existe guère de lignes directrices pour concevoir ce type d'études pour les tests utilisés en santé animale. À la connaissance des auteurs, il n'existe pas non plus de normes de conception applicables aux études de validation en santé humaine. Par conséquent, il conviendrait de disposer de lignes directrices génériques fondées sur les caractéristiques des maladies telles que leurs modalités de transmission, leur période de latence et leur pathogénie. Une notification complète, claire et transparente des études d'exactitude des tests primaires pour les maladies listées par l'Organisation mondiale de la santé animale (OIE) serait une aide précieuse pour les utilisateurs finaux des tests de diagnostic, mais aussi pour les responsables de l'élaboration des politiques, dont les décisions reposent sur des examens et des méta-analyses systématiques couvrant un grand nombre de tests pour certaines maladies ou pour certains usages d'un test. La publication récente des normes de notification applicables aux modèles bayésiens à classe latente pour analyser les données de performance d'un test à partir de foyers naturels de maladie comble une lacune importante dans la mesure où ces méthodes sont de plus en plus utilisées pour les maladies listées par l'OIE. L'adhésion à des normes de conception et de notification ainsi qu'à des lignes directrices en la matière permettra de garantir que les fonds alloués aux études de validation des tests sont bien utilisés et que les atouts et les limitations de certains tests individuels ou associations de tests sont clairement perçus par les utilisateurs. Les auteurs passent en revue certains points essentiels qui sont souvent ignorés ou mal interprétés lors des études de validation des tests et proposent deux exemples concrets de bonnes pratiques qui pourront servir de références pour les études à venir.

Las normas de comunicación y diseño son indicadores básicos de la calidad de los estudios encaminados a determinar la exactitud de diagnóstico (validación) pero, con la excepción de las enfermedades de los animales acuáticos y la paratuberculosis en rumiantes, hay escasas directrices que se apliquen al diseño de esos estudios en animales. Además, hasta donde saben los autores, en el ámbito de la salud humana no hay normas de diseño. De ahí la necesidad de directrices genéricas que estén basadas en las características de las enfermedades, como modo de transmisión, período de latencia o patogénesis. La comunicación exhaustiva, clara y transparente de estudios primarios sobre la exactitud de pruebas de diagnóstico de enfermedades incluidas en las listas de la Organización Mundial de Sanidad Animal (OIE) reviste utilidad no solo para los usuarios finales de la prueba, sino también, en última instancia, para los órganos decisorios, que necesitan metaanálisis y estudios sistemáticos de múltiples pruebas que se apliquen a una u otra enfermedad y sirvan para una u otra finalidad. La reciente publicación de normas de comunicación de modelos bayesianos de clases latentes para analizar los datos de exactitud de pruebas a partir de episodios de enfermedad de origen natural viene a colmar una importante laguna, en la medida en que estos métodos se aplican cada vez más al diagnóstico de enfermedades incluidas en las listas de la OIE. El cumplimiento de las normas de diseño y comunicación, y también de las directrices, ayuda a garantizar que los fondos de investigación destinados a estudios de validación de pruebas sean utilizados debidamente y que el usuario final de una prueba reciba información clara sobre los puntos fuertes y las limitaciones de una prueba o combinación de pruebas. Los autores pasan revista a los principales aspectos que se suelen pasar por alto o malinterpretar en los estudios de validación de pruebas y ofrecen dos ejemplos concretos de buenas prácticas que se pueden utilizar como referencia en futuros estudios.

Tick-Borne Pathogens in Ticks Collected from Wild Ungulates in North-Eastern Poland.

This study was carried out in north-eastern Poland during two hunting seasons between 2018 and 2020. Ticks (Ixodes ricinus and Dermacentor reticulatus) were removed from wild cervids and boars and examined for the presence of Borrelia spirochetes and Rickettsiales members: Rickettsia spp. and Anaplasma phagocytophilum. The present study contributes to the knowledge of even-toed ungulates, which are an important reservoir of the above-mentioned pathogens and a potential source of infections for humans through ticks as vectors. Almost 40% of the collected ticks (191 out of 484) were infected with the following pathogens: 3.3% with Borrelia spp., 19.2% with A. phagocytophilum and 26.9% with Rickettsia spp. Only the ticks collected from cervids carried Borrelia. Typing of the species DNA confirmed the presence of B. afzelii, B. garinii, B. lusitaniae and B. miyamotoi. An analysis of Rickettsia spp. sequences using the GenBank data revealed the presence of R. helvetica, R. raoultii and R. monacensis. Monoinfections (79.1%) dominated over co-infections (20.9%). Among co-infections, the most frequent was A. phagocytophilum/Rickettsia spp. (70%), however co-infections, including B. afzelii/A. phagocytophilum, B. afzelii/Rickettsia spp., B. miyamotoi/A. phagocytophilum and B. afzelii/B. garinii/B. lusitaniae, were also noted. Significant differences were observed in the affinity of some pathogens to their vectors. Thus, Borrelia spp. and A. phagocytophilum were more frequently detected in I. ricinus (5.3% and 23.1%) than in D. reticulatus (1.2% and 15.3%). Infection frequency with Rickettsia spp. was similar (approximately 25-29%) in both tick species. The prevalence of A. phagocytophilum and Rickettsia spp. in ticks removed from cervids was 19.8% and 27.1%, and in ticks from wild boars it was 13.3% and 24.4%, respectively.

Relationships between vector-borne parasites and free-living mammals at the Brazilian Pantanal.

In nature, parasitic infections must be addressed as complex systems involving parasite-host relationships on a temporal and spatial scale. Since the parasites cover a great biological diversity, we can expect that wildlife are exposed simultaneously to different parasites. In this sense, the objective of this work was to determine the relationships between free-living mammals and their associated hemoparasites in the Brazilian Pantanal. We used the data published during 2017 and 2018 by de Sousa et al. regarding the detection of vector-borne pathogens (VBP), namely Anaplasma, Babesia, Bartonella, Cytauxzoon, Ehrlichia, Hepatozoon, Mycoplasma, and Theileria, in nine species of free-living mammals belonging to orders Carnivora, Rodentia, and Didelphimorphia. We assume as infected an individual positive on any of parasitological, molecular, and/or serological tests. We observed a strong association between the wild felid Leopardus pardalis with Cytauxzoon, the wild canid Cerdocyon thous with Hepatozoon, the small rodent Thrichomys fosteri with Bartonella, and the procyonid Nasua nasua with Mycoplasma and Theileria. Therefore, N. nasua, C. thous, T. fosteri, and the small rodent Oecomys mamorae can be considered key species for the maintenance of selected VBP in the Pantanal region, because they showed a high number of single and coinfections. Together, our results highlighted the importance of coinfection as a common phenomenon in nature.

Ovarian gene expression in collared peccary (Pecari tajacu Linnaeus, 1758) subjected to gonadotropin stimulation protocols.

Ovarian response of collared peccaries (Pecari tajacu), after hormonal stimulation with gonadotropin association (eCG/hCG), was accessed by both gene expression and follicular development. Thus, collared peccaries (n = 8) were treated with the dose used for sows (swine dose, SWD) or with dose adjusted for peccary's weight (allometric dose, ALD). The gene expression of receptors was evaluated for both gonadotropins (FSHR and LHCGR) and growth factors (proteins codified by TGFßR-1, BMPR1-A and BMPR2 genes) in antral follicles, cortex and corpora haemorrhagica (CH). Five days after gonadotropin injection, all females presented CH. The ovulation rate was similar (p > .05) between SWD (4.00 ± 1.17) and ALD (2.50 ± 0.43) group. The total number of follicles per animal and amounts of small (<3 mm), medium (3-5 mm) and large (>5 mm) follicles was similar among groups. However, SWD produced large follicles heavier than ALD group, as accessed by weight of follicular wall biopsies. Ovarian follicles expressed both gonadotropin and growth factor receptors at levels which are independent from gonadotropin dose. In conclusion, the two gonadotropin doses (SWD and ALD) can be used for ovarian stimulation of collared peccary. Additionally, FSH and growth factors (TGFßR-1, BMPR1-A and BMPR2) receptors are more expressed in the early follicle development, while LH receptor seems to be more important in the final of follicular growth.