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The occurrence of emerging infectious diseases and their relevance to human health has increased the interest in bats as potential reservoir hosts and vectors of zoonotic pathogens. But while previous and ongoing research activities predominantly focused on viral agents, the prevalence of pathogenic bacteria in bats and their impact on bat mortality have largely neglected. Enteric pathogens found in bats are often considered to originate from the bats' diet and foraging habitats, despite the fact that little is known about the actual ecological context or even transmission cycles involving bats, humans and other animals like pets and livestock. For some bacterial pathogens common in human and animal diseases (e.g. Pasteurella, Salmonella, Escherichia and Yersinia spp.), the pathogenic potential has been confirmed for bats. Other bacterial pathogens (e.g. Bartonella, Borrelia and Leptospira spp.) provide evidence for novel species that seem to be specific for bat hosts but might also be of disease importance in humans and other animals. The purpose of this review is to summarize the current knowledge of bacterial pathogens identified in bats and to consider factors that might influence the exposure and susceptibility of bats to bacterial infection but could also affect bacterial transmission rates between bats, humans and other animals. [ABSTRACT FROM AUTHOR]

Abstract Background The emergence of important viral diseases and their potential threat to humans has increased the interest in bats as potential reservoir species. Whereas the majority of studies determined the occurrence of specific zoonotic agents in chiropteran species, little is known about actual bat pathogens and impacts of disease on bat mortality. Combined pathological and microbiological investigations in free-ranging bats are sparse and often limited by small sample sizes. In the present study about 500 deceased bats of 19 European species (family Vespertilionidae) were subjected to a post-mortem examination followed by histo-pathological and bacteriological investigations. The bat carcasses originated from different geographical regions in Germany and were collected by bat researchers and bat rehabilitation centers. Results Pathological examination revealed inflammatory lesions in more than half of the investigated bats. Lung was the predominantly affected organ (40%) irrespective of bat species, sex and age. To a lesser extent non-inflammatory organ tissue changes were observed. Comparative analysis of histo-pathology and bacteriology results identified 22 different bacterial species that were clearly associated with pathological lesions. Besides disease-related mortality, traumatic injuries represented an additional major cause of death. Here, attacks by domestic cats accounted for almost a half of these cases. Conclusions The present study shows that free-ranging bats not only serve as a reservoir of infectious agents, they are also vulnerable to various infectious diseases. Some of these microbial agents have zoonotic potential, but there is no evidence that European bats would pose a higher health hazard risk to humans in comparison to other wildlife.

Several bat-associated circoviruses and circular rep-encoding single-stranded DNA (CRESS DNA) viruses have been described, but the exact diversity and host species of these viruses are often unknown. Our goal was to describe the diversity of bat-associated circoviruses and cirliviruses, thus, 424 bat samples from more than 80 species were collected on four continents. The samples were screened for circoviruses using PCR and the resulting amino acid sequences were subjected to phylogenetic analysis. The majority of bat strains were classified in the genus Circovirus and some strains in the genus Cyclovirus and the clades CRESS1 and CRESS3. Some strains, however, could only be classified at the taxonomic level of the order and were not classified in any of the accepted or proposed clades. In the family Circoviridae, 71 new species have been predicted. This screening of bat samples revealed a great diversity of circoviruses and cirliviruses. These studies underline the importance of the discovery and description of new cirliviruses and the need to establish new species and families in the order Cirlivirales., (© 2023. The Author(s).)

Seven genotypically distinct strains assigned to the genus Erysipelothrix were isolated in different laboratories from several animal sources. Strain D17_0559-3-2-1 T and three further strains were isolated from samples of duck, pig and goose. The strains had >99 % 16S rRNA gene sequence similarity to each other and to strain VA92-K48 T and two further strains isolated from samples of medical leech and a turtle. The closest related type strains to the seven strains were those of Erysipelothrix inopinata (96.74 %) and Erysipelothrix rhusiopathiae (95.93 %). Average nucleotide identity, amino acid identity and in silico DNA-DNA hybridization results showed that the strains represented two separate novel species. One further phylogenetically distinct strain (165301687 T ) was isolated from fox urine. The strain had highest 16S rRNA gene sequence similarity to the type strains of Erysipelothrix tonsillarum (95.67 %), followed by Erysipelothrix piscisicarius (95.58 %) and Erysipelothrix larvae (94.22 %) and represented a further novel species. Chemotaxonomic and physiological data of the novel strains were assessed, but failed to unequivocally differentiate the novel species from existing members of the genus. MALDI-TOF MS data proved the discrimination of at least strain 165301687 T from all currently described species. Based on the presented phylogenomic and physiological data, we propose three novel species, Erysipelothrix anatis sp. nov. with strain D17_0559-3-2-1 T (=DSM 111258 T = CIP 111884 T =CCM 9044 T ) as type strain, Erysipelothrix aquatica sp. nov. with strain VA92-K48 T (=DSM 106012 T =LMG 30351 T =CIP 111492 T ) as type strain and Erysipelothrix urinaevulpis sp. nov. with strain 165301687 T (=DSM 106013 T = LMG 30352 T = CIP 111494 T ) as type strain.

The phylogenetic tree of the Staphylococcus aureus complex consists of several distinct clades and the majority of human and veterinary S. aureus isolates form one large clade. In addition, two divergent clades have recently been described as separate species. One was named Staphylococcus argenteus , due to the lack of the "golden" pigment staphyloxanthin. The second one is S. schweitzeri , found in humans and animals from Central and West Africa. In late 2021, two additional species, S. roterodami and S. singaporensis , have been described from clinical samples from Southeast Asia. In the present study, isolates and their genome sequences from wild Straw-coloured fruit bats ( Eidolon helvum ) and a Diamond firetail ( Stagonopleura guttata , an estrildid finch) kept in a German aviary are described. The isolates possessed staphyloxanthin genes and were closer related to S. argenteus and S. schweitzeri than to S. aureus . Phylogenetic analysis revealed that they were nearly identical to both, S. roterodami and S. singaporensis . We propose considering the study isolates, the recently described S. roterodami and S. singaporensis as well as some Chinese strains with MLST profiles stored in the PubMLST database as different clonal complexes within one new species. According to the principle of priority we propose it should be named S. roterodami . This species is more widespread than previously believed, being observed in West Africa, Southeast Asia and Southern China. It has a zoonotic connection to bats and has been shown to be capable of causing skin and soft tissue infections in humans. It is positive for staphyloxanthin, and it could be mis-identified as S. aureus (or S. argenteus ) using routine procedures. However, it can be identified based on distinct MLST alleles, and " S. aureus " sequence types ST2470, ST3135, ST3952, ST3960, ST3961, ST3963, ST3965, ST3980, ST4014, ST4075, ST4076, ST4185, ST4326, ST4569, ST6105, ST6106, ST6107, ST6108, ST6109, ST6999 and ST7342 belong to this species., Competing Interests: DG is employed by a company, Illumina, but he performed experiments for this study before commencing this employment. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Monecke, Schaumburg, Shittu, Schwarz, Mühldorfer, Brandt, Braun, Collatz, Diezel, Gawlik, Hanke, Hotzel, Müller, Reinicke, Feßler and Ehricht.)

Staphylococcus aureus can be a harmless coloniser, but it can also cause severe infections in humans, livestock and wildlife. Regarding the latter, only few studies have been performed and knowledge on virulence factors is insufficient. The aim of the present study was to study S. aureus isolates from deceased wild beavers (Castor fiber). Seventeen isolates from eleven beavers, found in Germany and Austria, were investigated. Antimicrobial and biocide susceptibility tests were performed. Isolates were characterised using S. aureus-specific DNA microarrays, spa typing and whole-genome sequencing. From two isolates, prophages were induced by mitomycin C and studied by transmission electron microscopy. Four isolates belonged to clonal complex (CC) 8, CC12, and CC398. Twelve isolates belonged to CC1956 and one isolate was CC49. The CC49 and CC1956 isolates carried distinct lukF/S genes related to the Panton-Valentine leukocidin (PVL) from human isolates of S. aureus. These genes were located on related, but not identical, Siphovirus prophages. The beavers, from which those isolates originated, suffered from abscesses, purulent organ lesions and necrotising pneumonia, i.e., clinical manifestations resembling symptoms of severe PVL-associated disease in humans. It might thus be assumed that the "Beaver Leukocidin (BVL, lukF/S-BV)"-positive strains are beaver-specific pathogens, and further studies on their clinical role as well as on a possible transmissibility to other species, including humans, are warranted., (© 2021. The Author(s).)

Bats are known to be reservoirs of several highly pathogenic viruses. Hence, the interest in bat virus discovery has been increasing rapidly over the last decade. So far, most studies have focused on a single type of virus detection method, either PCR, virus isolation or virome sequencing. Here we present a comprehensive approach in virus discovery, using all three discovery methods on samples from the same bats. By family-specific PCR screening we found sequences of paramyxoviruses, adenoviruses, herpesviruses and one coronavirus. By cell culture we isolated a novel bat adenovirus and bat orthoreovirus. Virome sequencing revealed viral sequences of ten different virus families and orders: three bat nairoviruses, three phenuiviruses, one orbivirus, one rotavirus, one orthoreovirus, one mononegavirus, five parvoviruses, seven picornaviruses, three retroviruses, one totivirus and two thymoviruses were discovered. Of all viruses identified by family-specific PCR in the original samples, none was found by metagenomic sequencing. Vice versa, none of the viruses found by the metagenomic virome approach was detected by family-specific PCRs targeting the same family. The discrepancy of detected viruses by different detection approaches suggests that a combined approach using different detection methods is necessary for virus discovery studies.

Background: Hypothermic preservation of boar semen is considered a potential method for omitting antibiotics from insemination doses, thereby contributing to the global antibiotic resistance defence strategy. The main challenges are chilling injury to spermatozoa and bacterial growth during semen storage leading to reduced fertility., Objectives: To examine chilling injury and the number and type of bacteria in boar semen stored at 5 °C in the absence of antibiotics, and to assess the applicability of hypothermic semen storage under field conditions., Material and Methods: Boar ejaculates were extended with AndroStar® Premium, stored at 17 °C with and at 5 °C without antibiotics and tested for functional sperm parameters by flow cytometry. Raw semen and extended samples were investigated bacteriologically. Fertility was evaluated after once-daily inseminations of 194 sows in a field study., Results: Lethal sperm damage assessed by motility and membrane integrity was low throughout storage in both experimental groups. Sublethal chilling effects based on the decrease of viable spermatozoa with low membrane fluidity were higher (P < 0.05) up until 72 h in sperm stored at 5 °C compared to 17 °C but did not differ after 144 h. After 72 h, incubation in capacitating medium for 60 min induced a similar decrease in viable sperm with high mitochondria membrane potential and low cytosolic calcium in both groups. In semen stored at 5 °C, bacteria counts were below 10 3  CFU/mL and the bacteria spectrum was similar to that of raw semen. In 88% of 34 boars, cooled semen fulfilled the requirements for insemination. Fertility was high and did not differ (P > 0.05) between sow groups inseminated with semen stored antibiotic-free at 5 °C and semen stored at 17 °C with antibiotics., Conclusion: Despite subtle chilling effects and low bacterial numbers, antibiotic-free hypothermic storage of boar semen offers the possibility to reduce the use of antibiotics in pig insemination. However, strict sanitary guidelines must be maintained and further evidence of efficiency under field conditions is considered desirable.

Bats have been gaining attention as potential reservoir hosts of numerous viruses pathogenic to animals and man. Issyk-Kul virus, a member of the family Nairoviridae, was first isolated in the 1970s from vespertilionid bats in Central Asia. Issyk-Kul virus has been described as human-pathogenic virus, causing febrile outbreaks in humans with headaches, myalgia and nausea. Here we describe the detection of a novel strain of Issyk-Kul virus from Eptesicus nilssonii in Germany. This finding indicates for the first time the prevalence of these zoonotic viruses in Europe.

Immunity and parasites have been linked to the success of invasive species. Especially lower parasite burden in invasive populations has been suggested to enable a general downregulation of immune investment (Enemy Release and Evolution of Increased Competitive Ability Hypotheses). Simultaneously, keeping high immune competence towards potentially newly acquired parasites in the invasive range is essential to allow population growth. To investigate the variation of immune effectors of invasive species, we compared the mean and variance of multiple immune effectors in the context of parasite prevalence in an invasive and a native Egyptian goose (Alopochen aegyptiacus) population. Three of ten immune effectors measured showed higher variance in the invasive population. Mean levels were higher in the invasive population for three effectors but lower for eosinophil granulocytes. Parasite prevalence depended on the parasite taxa investigated. We suggest that variation of specific immune effectors, which may be important for invasion success, may lead to higher variance and enable invasive species to reduce the overall physiological cost of immunity while maintaining the ability to efficiently defend against novel parasites encountered.

The avian pathogen Ornithobacterium rhinotracheale (ORT) has been implied in the etiology of poultry respiratory disease in recent years. To evaluate whether Whatman® Flinders Technology Associates (FTA®) cards can be used for hazard-free transport and storage of ORT samples for posterior DNA amplification, a controlled assay was performed. Three 10-fold dilutions of an ORT culture suspension were spotted on FTA cards and stored at room temperature (RT) for 6 mo. Sterile swabs were immersed in the same three 10-fold culture dilutions and stored at RT and 4 and -20 C without storage medium for the same time. DNA was extracted from both the FTA cards and swabs 1 day, 1 and 6 wk, and 6 mo following sample preparation and stored at -20 C. At the end of the experiment, real-time PCR amplification of the 16S ribosomal RNA gene was performed from DNA extracted throughout a 6-mo period from all ORT samples stored on both FTA cards and swabs. The obtained threshold cycle values for each ORT DNA extraction date were within the same range for all samples in a dilution-dependent fashion, regardless of storage temperature or used material. Pure ORT colonies could be reisolated 1 day after sample preparation from the swab dilutions stored at all temperatures but not from the FTA cards. We conclude that the efficiency of ORT DNA amplification from samples stored on FTA cards or in swabs is similar. However, FTA cards have the advantage of preventing microorganism growth, thus allowing safe transport and storage, for at least 6 mo, for bacterial dilutions down to at least 104-105 colony-forming units/ml.

Leptospirosis is a zoonotic disease of importance to public health and in livestock productions. It causes significant economic losses in pig breeding farms worldwide. However, actual transmission cycles and disease epidemiology in the pig population remain largely unknown. Despite the fact that the potential risk of venereal transmission of pathogenic Leptospira serovars in pigs has been a topic of discussion since the 1970s, reliable data are still lacking compared to other livestock species. Consequently, antibiotics are added to semen extenders to reduce bacterial contamination including pathogens like Leptospira. In view of the global threat of antimicrobial resistances, the routine use of antibiotics in porcine semen extenders is now under debate. Information about the prevalence of Leptospira infections in boar used for artificial insemination is needed for the development of novel antimicrobial concepts in pig insemination.This short report provides a summary of the state of knowledge, together with negative results from real-time PCR analyses for the detection of pathogenic Leptospira DNA in boar semen. Molecular analyses were performed on 96 raw and extended samples obtained from normospermic ejaculates of 58 boar housed in six different studs in Germany. In the absence of reliable data, it is important to raise the awareness for a subject that can represent a challenge for pig productions in keeping reproductive health and food safety at high levels. The present molecular results indicate that Leptospira might not be a common threat in boar semen. Conclusive evidence would require results from a systematic serological surveillance of boar, combined with seasonal molecular analyses of semen to identify potential carriers, and assess actual seroprevalences, associated Leptospira serovars and transmission events.

The arctic fox ( Vulpes lagopus ) is the only native terrestrial mammal in Iceland. While red foxes ( V. vulpes ) are known to be epidemiologically important carriers of several vector-borne pathogens in Europe, arctic foxes have never been evaluated in a similar context on this continent. This has become especially relevant in the last decade, considering the establishing populations of the tick species Ixodes ricinus in Iceland. In this study, liver DNA extracts of 60 arctic foxes, hunted between 2011-2012, were molecularly screened for vector-borne protozoan parasites ( Trypanosomatidae , Babesia , Theileria , Hepatozoon ) and bacteria ( Anaplasma , Ehrlichia , Rickettsia , Borrelia , hemotropic Mycoplasma ). One sample was real-time qPCR positive for Anaplasma phagocytophilum , though this positivity could not be confirmed with sequencing. Samples were negative for all other tested vector-borne pathogens. Results of this study indicate that, except for A. phagocytophilum , Icelandic arctic foxes were apparently "not yet infected" with vector-borne pathogens in 2011-2012, or their infections were "below the detection limit" of applied methods. Taking into account the broad range of target microorganisms analyzed here, as well as the warming climate and increasing presence of the vector I. ricinus in Iceland, our results will be very useful as baseline data for comparison in future monitoring of the emergence of ticks and tick-borne diseases in this country.

Novel catalase-negative, Gram-stain-positive, beta-haemolytic, coccus-shaped organisms were isolated from Chacoan peccaries that died from respiratory disease. The initial API 20 Strep profiles suggested Streptococcus agalactiae with acceptable identification scores, but the 16S rRNA gene similarity (1548 bp) to available sequences of streptococci was below 98 %. Next taxa of the genus Streptococcus , displaying highest similarities to the strains from this study, were S. bovimastitidis NZ1587 T (97.5 %), S. iniae ATCC 29178 T (97.5 %), S. hongkongensis HKU30 T (97.4 %), S. parauberis DSM 6631 T (97.1 %), S. penaeicida CAIM 1838 T (97.1 %), S. pseudoporcinus DSM 18513 T (97.0 %), S. didelphis DSM 15616 T (96.6 %), S. ictaluri 707-05 T (96.6 %), S. uberis JCM 5709 T (96.5 %) and S. porcinus NCTC 10999 T (96.4 %). All other Streptococcus species had sequence similarities of below 96.4 %. A sodA gene as well as whole genome-based core genome phylogeny of three representative strains and 145 available Streptococcus genomes confirmed the unique taxonomic position. Interstrain average nucleotide identity (ANI) and amino acid identity (AAI) values were high (ANI >96 %; AAI 100%), but for other streptococci clearly below the proposed species boundary of 95-96 % (ANI <75 %; AAI <83 %). Results were confirmed by genome-to-genome distance calculations. Pairwise digital DNA-DNA hybridization estimates were high (>90 %) between the novel strains, but well below the species boundary of 70 % for closely related Streptococcus type strains (23.5-19.7 %). Phenotypic properties as obtained from extended biochemical profiles and MALDI-TOF mass spectrometry supported the outstanding rank. Based on the presented molecular and physiological data of the six strains, we propose a novel taxon for which we suggest the name Streptococcus catagoni sp. nov. with the type strain 99-1/2017 T (=DSM 110457 T =CCUG 74072 T ) and five reference strains.

Background: Bats are hosts for a variety of microorganisms, however, little is known about the presence of Chlamydiales and hemotropic mycoplasmas. This study investigated 475 captive and free-living bats from Switzerland, Germany, and Costa Rica for Chlamydiales and hemotropic mycoplasmas by PCR to determine the prevalence and phylogeny of these organisms., Results: Screening for Chlamydiales resulted in a total prevalence of 31.4%. Positive samples originated from captive and free-living bats from all three countries. Sequencing of 15 samples allowed the detection of two phylogenetically distinct groups. These groups share sequence identities to Chlamydiaceae, and to Chlamydia-like organisms including Rhabdochlamydiaceae and unclassified Chlamydiales from environmental samples, respectively. PCR analysis for the presence of hemotropic mycoplasmas resulted in a total prevalence of 0.7%, comprising free-living bats from Germany and Costa Rica. Phylogenetic analysis revealed three sequences related to other unidentified mycoplasmas found in vampire bats and Chilean bats., Conclusions: Bats can harbor Chlamydiales and hemotropic mycoplasmas and the newly described sequences in this study indicate that the diversity of these bacteria in bats is much larger than previously thought. Both, Chlamydiales and hemotropic mycoplasmas are not restricted to certain bat species or countries and captive and free-living bats can be colonized. In conclusion, bats represent another potential host or vector for novel, previously unidentified, Chlamydiales and hemotropic mycoplasmas.

Hypothermic storage of boar semen provides the possibility to omit antibiotics from semen extenders so long as sperm quality is maintained and bacterial growth prevented. The objective of this study was to determine an optimal cooling-rate frame for boar semen preserved at 5°C in an antibiotic-free extender. Semen from eight boars extended in AndroStar® Premium was cooled from 30°C to 5°C using seven different cooling rates, ranging initially from 0.01 to 0.36°C min-1 and reaching 5°C between 2 h and 24 h after dilution. Sperm motility, membrane integrity, membrane fluidity, mitochondrial membrane potential and the response to the capacitation stimulus bicarbonate remained at a high level for 144 h at 5°C when the semen was initially cooled in a cooling-rate frame ranging from 0.01 to 0.09°C min‑1 in the temperature zone from 30 to 25°C, followed by 0.02 to 0.06°C min-1 to 10°C and 0.01 to 0.02°C min‑1 to the final storage temperature. A cooling rate of 0.07°C min-1 in the temperature zone from 30 to 10°C led to a reduced response to bicarbonate (P < 0.01) and fast cooling to 5°C within 1 h with a cooling rate of 0.31°C min-1 resulted in lower values (P > 0.05) of all sperm parameters. In a further experiment, slow cooling with a holding time of 6 h at 22°C induced after 6 h storage a temporary increase in Escherichia coli of 0.5 × 103 to 2.4 × 103 CFU mL-1 in the sperm-free inoculated extender. Overall, the load of mesophilic bacteria in the stored semen was below 6 × 103 CFU mL-1, a level that is not regarded as critical for sperm quality. In conclusion, appropriate cooling protocols were established for the antibiotic-free storage of boar semen at 5°C, allowing the application of hypothermic preservation in research and in artificial insemination., Competing Interests: The authors have declared that no competing interests exist.

From a phlegmon in a dog an aerobic and facultatively anaerobic, indole-, oxidase- and catalase-negative, non-motile bacterium was isolated in 2019 in Germany that stained Gram-negative and showed a pleomorphic, rod-shaped, non-spore-forming appearance. Based on the results of 16S rRNA gene sequence analyses, strain IHIT1603-19 T was assigned to the genus Streptobacillus with sequence similarities of 98.6, 98.0, 97.9, 97.1 and 94.4 % to the type strains of Streptobacillus felis , Streptobacillus notomytis , Streptobacillus ratti , Streptobacillus moniliformis and Streptobacillus hongkongensis , respectively. Strain IHIT1603-19 T could also clearly be differentiated from other Streptobacillus species by rpoB , groEL and recA gene, nucleotide and amino acid sequence analyses as well as by core genome phylogeny. Regarding DNA-DNA relatedness, strain IHIT1603-19 T demonstrated an average nucleotide identity of 83.00 and 82.28 % compared to S. felis 131000547 T and S. moniliformis DSM 12112 T , respectively. Chemotaxonomic and physiological data of strain IHIT1603-19 T were in congruence with other closely related members of the family Leptotrichiaceae , represented by highly similar enzyme profiles and fatty acid patterns. MALDI-TOF MS analysis also proved suitable in unequivocally discriminating strain IHIT1603-19 T from all currently described taxa of the genus Streptobacillus . On the basis of these data, we propose the novel species Streptobacillus canis sp. nov. with the type strain IHIT1603-19 T (=DSM 110501 T =CCUG 74118 T =CIP 111795 T ). The G+C content of the DNA of the type strain is 26.6 mol%, genome size is 1.60 Mbp.

Bats are well known reservoir hosts for RNA and DNA viruses. The use of captive bats in research has intensified over the past decade as researchers aim to examine the virus-reservoir host interface. In this study, we investigated the effects of captivity on the fecal bacterial microbiome of an insectivorous microbat, Mops condylurus, a species that roosts in close proximity to humans and has likely transmitted viral infections to humans. Using amplicon 16S rRNA gene sequencing, we characterized changes in fecal bacterial community composition for individual bats directly at the time of capture and again after six weeks in captivity. We found that microbial community richness by measure of the number of observed operational taxonomic units (OTUs) in bat feces increases in captivity. Importantly, we found the similarity of microbial community structures of fecal microbiomes between different bats to converge during captivity. We propose a six week-acclimatization period prior to carrying out infection studies or other research influenced by the microbiome composition, which may be advantageous to reduce variation in microbiome composition and minimize biological variation inherent to in vivo experimental studies., Competing Interests: The authors have declared that no competing interests exist.

Bats are reservoir hosts for several emerging and re-emerging viral pathogens causing morbidity and mortality in wildlife, animal stocks and humans. Various viruses within the family Phenuiviridae have been detected in bats, including the highly pathogenic Rift Valley fever virus and Malsoor virus, a novel Banyangvirus with close genetic relation to Huaiyangshan banyangvirus (BHAV)(former known as Severe fever with thrombocytopenia syndrome virus, SFTSV) and Heartland virus (HRTV), both of which have caused severe disease with fatal casualties in humans. In this study we present the whole genome of a novel Banyangvirus, named Zwiesel bat banyangvirus, revealed through deep sequencing of the Eptesicus nilssonii bat virome. The detection of the novel bat banyangvirus, which is in close phylogenetic relationship with the pathogenic HRTV and BHAV, underlines the possible impact of emerging phenuiviruses on public health.

Streptococcus castoreus is a rarely encountered beta-haemolytic group A Streptococcus with high tropism for the beaver as host. Based on 27 field isolates under study, evidence strongly suggests that S. castoreus behaves as an opportunistic pathogen in beavers. Although it belongs to the resident mucosal microbiota, this Streptococcus species is associated with purulent lesions in diseased animals. With few exceptions, isolates proved to be highly similar in a panel of phenotypic (including biochemistry, resistance pattern, MALDI-TOF mass spectrometry and Fourier transform-infrared spectroscopy) and classic molecular (16S rRNA and sodA gene) analyses, and thus did not show any specific pattern according to host species or spatio-temporal origin. Conversely, S. castoreus isolates were differentiated into a multitude of pulsed-field gel electrophoresis 'pulsotypes' that did not seem to reflect true epidemiologic lineages. In contrast, single reactions of genomic fingerprinting using BOX-, (GTG) 5 - and RAPD-PCRs revealed at least subclusters with respect to host species, geographic origin or year, and confirmed the co-colonization of individuals with more than one isolate. In addition to isolates from free-ranging Eurasian beavers (Castor fiber), this study includes S. castoreus from captive North American beavers (Castor canadensis) for the first time.

Virulence-associated traits have frequently been studied in enterococci and are considered to contribute towards the pathogenicity of infections. In the present study, Enterococcus isolates were collected during diagnostic investigations from meat turkeys in Germany. Twenty-eight isolates of three different Enterococcus species were analyzed for five selected putative virulence traits to understand their potential role in the pathogenicity using the chicken embryo lethality assay. Ten E. faecalis , ten E. faecium , and eight E. gallinarum isolates were examined for the presence of common virulence genes and their phenotypic expression, namely, the cytolysin operon, five individual cyl genes ( cylL L , cylL S , cylM , cylB , and cylA ), gelatinase ( gelE ), hyaluronidase ( hyl Efm ), aggregation substance ( asa1 ), and enterococcal surface protein ( esp ). The Enterococcus isolates showed significant species-dependent differences in the presence of genotypic traits ( p < 0.001 by Fisher's exact test; Cramer's V  = 0.68). At least one gene and up to three virulence traits were found in E. faecalis , while six E. faecium isolates and one E. gallinarum isolate did not display any virulence-associated pheno- or genotype. More than half of the Enterococcus isolates ( n  = 15) harbored the gelE gene, but only E. faecalis ( n  = 10) expressed the gelatinase activity in vitro . The hyl Efm gene was found in five E. gallinarum isolates only, while seven isolates showed the hyaluronidase activity in the phenotypic assay. In Cramer's V statistic, a moderate association was indicated for species ( V  ≤ 0.35) or genotype ( V  < 0.43) and the results from the embryo lethality assay, but the differences were not significant. All E. gallinarum isolates were less virulent with mortality rates ranging between 0 and 30%. Two E. faecalis isolates were highly virulent, harboring the whole cyl -operon as well as gelE and asa1 genes. Likewise, one E. faecium isolate caused high embryo mortality but did not harbor any of the investigated virulence genes. For the first time, Enterococcus isolates of three different species collected from diseased turkeys were investigated for their virulence properties in comparison. The results differed markedly between the Enterococcus species, with E. faecalis harboring the majority of investigated genes and virulence traits. However, the genotype did not entirely correlate with the phenotype or the isolates' virulence potential and pathogenicity for chicken embryos., Competing Interests: The authors declare that there are no conflicts of interest related to this article., (Copyright © 2019 Julia Maasjost et al.)

Wastewater treatment is crucial to environmental hygiene in urban environments. However, wastewater treatment plants (WWTPs) collect chemicals, organic matter, and microorganisms including pathogens and multi-resistant bacteria from various sources which may be potentially released into the environment via WWTP effluent. To better understand microbial dynamics in WWTPs, we characterized and compared the bacterial community of the inflow and effluent of a WWTP in Berlin, Germany using full-length 16S rRNA gene sequences, which allowed for species level determination in many cases and generally resolved bacterial taxa. Significantly distinct bacterial communities were identified in the wastewater inflow and effluent samples. Dominant operational taxonomic units (OTUs) varied both temporally and spatially. Disease associated bacterial groups were efficiently reduced in their relative abundance from the effluent by the WWTP treatment process, except for Legionella and Leptospira species which demonstrated an increase in relative proportion from inflow to effluent. This indicates that WWTPs, while effective against enteric bacteria, may enrich and release other potentially pathogenic bacteria into the environment. The taxonomic resolution of full-length 16S rRNA genes allows for improved characterization of potential pathogenic taxa and other harmful bacteria which is required to reliably assess health risk.

Members of the genus Sneathia are fastidious bacteria that predominantly colonise the female genital tract and are significantly associated with reproductive disorders and genital and neonatal disease. From a taxonomical perspective, the genus only comprises the species Sneathia sanguinegens . Numerous reports on a second species, ' Sneathia amnii ', have been published, but the name has never been validated. The same is the case for ' Leptotrichia amnionii ', which was previously shown to belong to the same species as ' Sneathia amnii '. We studied strains DSM 16631 T and DSM 16630, which have been identified and deposited as ' Leptotrichia amnionii ' previously. At the time of isolation, these strains were found to be most closely related to, but clearly different from, Sneathia sanguinegens based on 16S rRNA gene sequence similarities. Both strains proved to be almost indistinguishable from ' Sneathia amnii ' based on molecular, morphological and physiological traits. The 16S rRNA gene sequence analysis revealed that strain DSM 16631 T was assigned to the genus Sneathia with a sequence similarity of 95.47 % to Sneathia sanguinegens CCUG 41628 T , followed by type strains of Caviibacter abscessus (93.03 %), Oceanivirga salmonicida (92.68 %) and Oceanivirga miroungae (91.97 %) as the next closely related members of the Leptotrichiaceae . The novel species was also clearly differentiated from other related taxa by core genome phylogeny, average nucleotide and amino acid identities, in silico DNA-DNA hybridization and MALDI-TOF MS. With respect to chemotaxonomic and physiological patterns, strains DSM 16631 T and DSM 16630 were again highly similar to Sneathia sanguinegens . On the basis of these data, we propose the novel species Sneathia vaginalis sp. nov. with the type strain DSM 16631 T (=CCUG 52977 T =CCUG 52889A T ) and a second strain DSM 16630 (=CCUG 52976=CCUG 52888) that were both isolated from bloodstream infections in women with puerperal fever in France. The G+C content of the DNA of the type strain is 28.4 mol% and the genome size is 1.28 Mbp. Based on the observed extremely high similarities of genotypic and phenotypic traits of the novel proposed species to those reported for ' Sneathia amnii ', we recommend using this new name in all further publications on this taxon.

Background: Ornithobacterium (O.) rhinotracheale is an emerging bacterial pathogen in poultry and not fully understood to date. Because of its importance particularly for the global turkey meat industry, reliable diagnostic and characterization methods are needed for early treatment and in future for better vaccine production. The host range of birds infected by O. rhinotracheale or carrying the bacterium in their respiratory tract has constantly increased raising important epidemiological and taxonomic questions for a better understanding of its diversity, ecology and transmission cycles. The purpose of this study was to introduce partial rpoB gene sequencing for O. rhinotracheale into routine diagnostics to differentiate strains isolated from poultry and more diverse avian hosts (i.e., birds of prey, corvids and pigeons) and to compare phylogenetic relationships with results from 16S rRNA gene analysis and multilocus sequence typing (MLST)., Results: Partial 16S rRNA gene analysis revealed a high level of homogeneity among the 65 investigated O. rhinotracheale sequences with similarity values ranging from 98.6 to 100% between sequences from non-galliform and poultry species. The corresponding rpoB gene sequences were heterogeneous and ranged in their similarity values from 85.1 to 100%. The structure of the rpoB tree was in strong correlation with previous MLST results revealing three main clusters A (poultry and birds of prey), B (poultry, birds of prey and corvids) and C (pigeons), which were clearly separated from each other., Conclusions: By using partial sequences from a single gene, the rpoB gene analysis is in good agreement with MLST results with a slight decrease in resolution to distinguish more similar strains. The present results provide strong evidence that traditional phenotypic and genetic methods may not properly represent the heterogeneous group of bacteria classified as O. rhinotracheale. From housekeeping gene analyses, it is very likely that the genus Ornithobacterium includes additional species and partial rpoB gene sequencing can be recommended as fast, cost-effective and readily available method to identify strains and differentiate between O. rhinotracheale and Ornithobacterium-like bacteria.

Toxigenic Corynebacterium ulcerans may cause both respiratory and cutaneous diphtheria in humans. As a zoonotic emerging pathogen it has been isolated from a wide variety of animals living in captivity, such as livestock, pet, zoo and research animals and additionally in a large number of different wild animals. Here we report the isolation of tox-positive C. ulcerans in four hedgehogs with cutaneous diphtheria and pneumonia, respectively.

Antimicrobial resistance of Staphylococcus aureus is a major problem in human and veterinary medicine. The aim of this study was to characterise S. aureus isolates from wild and zoo animals mainly associated with bacterial infections. In total, 23 S. aureus isolates, including nine from wild animals and 14 from zoo animals, were obtained during routine diagnostics. All isolates were subjected to multilocus sequence typing (MLST), spa typing, macrorestriction analysis with subsequent SmaI pulsed-field gelelectrophoresis (PFGE), antimicrobial susceptibility testing and S. aureus-specific DNA-microarray analysis. Resistant isolates were also tested for their respective resistance genes by PCR. Isolates from zoo animals and wildlife showed a high diversity of MLST types, spa types and PFGE patterns. Nineteen different spa types were identified, including three novel types and 16 main macrorestriction patterns. Only few isolates were resistant to members of four classes of antimicrobial agents and harboured the respective resistance genes (β-lactams [blaZ, mecA, mecC], tetracyclines [tet(K), tet(L)] and chloramphenicol [cat pC221 ]) or mutations (fluoroquinolones). The DNA microarray analysis identified one isolate from a zoo animal harbouring the toxic shock syndrome toxin gene tst1. Moreover, several enterotoxin genes were detected in five S. aureus isolates. All isolates were negative for Panton-Valentine leukocidin (PVL) genes, but the animal-associated leukocidin genes lukM/lukF-P83 were found in three isolates from two animals., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The genus Brucella comprises various species of both veterinary and human medical importance. All species are genetically highly related to each other, sharing intra-species average nucleotide identities (ANI) of > 99%. Infections occur among various warm-blooded animal species, marine mammals, and humans. Until recently, amphibians had not been recognized as a host for Brucella. In this study, however, we show that novel Brucella species are distributed among exotic frogs worldwide. Comparative recA gene analysis of 36 frog isolates from various continents and different frog species revealed an unexpected high genetic diversity, not observed among classical Brucella species. In phylogenetic reconstructions the isolates consequently formed various clusters and grouped together with atypical more distantly related brucellae, like B. inopinata, strain BO2, and Australian isolates from rodents, some of which were isolated as human pathogens. Of one frog isolate (10RB9215) the genome sequence was determined. Comparative genome analysis of this isolate and the classical Brucella species revealed additional genetic material, absent from classical Brucella species but present in Ochrobactrum, the closest genetic neighbor of Brucella, and in other soil associated genera of the Alphaproteobacteria. The presence of gene clusters encoding for additional metabolic functions, flanked by tRNAs and mobile genetic elements, as well as by bacteriophages is suggestive for a different ecology compared to classical Brucella species. Furthermore it suggests that amphibian isolates may represent a link between free living soil saprophytes and the pathogenic Brucella with a preferred intracellular habitat. We therefore assume that brucellae from frogs have a reservoir in soil and, in contrast to classical brucellae, undergo extensive horizontal gene transfer., Competing Interests: The authors have declared that no competing interests exist.

The sudden emergence of Ornithobacterium rhinotracheale (ORT) in commercially raised poultry species and its presence in non-galliform birds raise important epidemiological issues about the role of interspecies transmission. In the present study, 21 ORT strains isolated from pigeons and from birds of prey were analyzed using the recently established multilocus sequence typing (MLST) scheme. Results were compared to MLST sequence data available from ORT strains isolated mainly from turkeys and chickens, but also single strains from pheasant, guineafowl and rook. The pigeon-derived ORT strains (n=11) were closely related amongst themselves representing their own cluster distant from ORT strains of non-columbiform avian hosts. ORT strains isolated from birds of prey (n=10) revealed a higher genetic heterogeneity that corresponded well to their host family relationships but grouped within the two mainly poultry-based clusters. None of these strains had a sequence type identical to strains investigated previously. However, three strains isolated from common kestrels and a single strain from a turkey vulture shared one or two out of seven gene loci, respectively, with strains of turkey and chicken origin. The MLST results of ORT isolated from pigeons and birds of prey likely reflect evolutionary bacterial host adaptations but might also indicate a potential for interspecies transmission. Definite conclusions should be drawn carefully as so far a few strains from non-galliform birds were analyzed by MLST. By extending the number of ORT isolates and the range of potential avian hosts, the MLST database can provide a valuable resource in understanding transmission dynamics., (© 2016 The Authors.)

Endogenous bornavirus-like L (EBLL) elements are inheritable sequences derived from ancient bornavirus L genes that encode a viral RNA-dependent RNA polymerase (RdRp) in many eukaryotic genomes. Here, we demonstrate that bats of the genus Eptesicus have preserved for more than 11.8 million years an EBLL element named eEBLL-1, which has an intact open reading frame of 1,718 codons. The eEBLL-1 coding sequence revealed that functional motifs essential for mononegaviral RdRp activity are well conserved in the EBLL-1 genes. Genetic analyses showed that natural selection operated on eEBLL-1 during the evolution of Eptesicus. Notably, we detected efficient transcription of eEBLL-1 in tissues from Eptesicus bats. To the best of our knowledge, this study is the first report showing that the eukaryotic genome has gained a riboviral polymerase gene from an ancient virus that has the potential to encode a functional RdRp.

Ornithobacterium rhinotracheale (ORT) is an economically important bacterial pathogen of turkeys and chickens worldwide. Since its first detection, a variety of typing methods have been used to gain basic knowledge about the bacterial population structure, an issue that still needs to be addressed. Serological characterization revealed at least 18 different serotypes (A-R) with ORT of serotype A to be predominate among poultry. This study aimed to establish a multilocus sequence typing (MLST) scheme for ORT that could easily be used by other laboratories and allows for worldwide comparison of sequence data. For this purpose, 87 ORT strains from different poultry hosts, geographical origins, years of isolation and serotypes were included in the analysis to identify correlations. Fourteen different sequence types (ST) were found. The most common ST1 was identified in 40 ORT strains from turkeys and chickens on 4 continents and in 3 different European countries. Together with ST9, both STs represented over three quarters (77%) of ORT strains used in the MLST analysis and included strains of frequently cross-reacting ORT serotypes A, E and I. Nine STs were only represented by one ORT strain and might indicate possible avian host, disease or serotype-specific relationships. In contrast, discrepancies between serotype and phylogenetic relatedness were clearly demonstrated by ORT strains that belonged to identical serotypes but differed in their ST. The overall identified low genetic diversity among strains isolated from turkeys and chickens independent of host and geographical origins suggests that ORT has only recently been introduced into domestic poultry and dispersed worldwide.

Five bacterial strains isolated from bats of the family Vespertilionidae were characterized by phenotypic tests and multilocus sequence analysis (MLSA) using the 16S rRNA gene and four housekeeping genes (rpoA, rpoB, infB, recN). Phylogenetic analyses of individual and combined datasets indicated that the five strains represent a monophyletic cluster within the family Pasteurellaceae. Comparison of 16S rRNA gene sequences demonstrated a high degree of similarity (98.3-99.9%) among the group of bat-derived strains, while searches in nucleotide databases indicated less than 96% sequence similarity to known members of the Pasteurellaceae. The housekeeping genes rpoA, rpoB, infB and recN provided higher resolution compared with the 16S rRNA gene and subdivided the group according to the bat species from which the strains were isolated. Three strains derived from noctule bats shared 98.6-100% sequence similarity in all four genes investigated, whereas, based on rpoB, infB and recN gene sequences, 91.8-96% similarity was observed with and between the remaining two strains isolated from a serotine bat and a pipistrelle bat, respectively. Genome relatedness as deduced from recN gene sequences correlated well with the results of MLSA and indicated that the five strains represent a new genus. Based on these results, it is proposed to classify the five strains derived from bats within Vespertiliibacter pulmonis gen. nov., sp. nov. (the type species), Vespertiliibacter genomospecies 1 and Vespertiliibacter genomospecies 2. The genus can be distinguished phenotypically from recognized genera of the Pasteurellaceae by at least three characteristics. All strains are nutritionally fastidious and require a chemically defined supplement with NAD for growth. The DNA G+C content of strain E127/08(T) is 38.2 mol%. The type strain of Vespertiliibacter pulmonis gen. nov., sp. nov. is E127/08(T) ( = CCUG 64585(T) = DSM 27238(T)). The reference strains of Vespertiliibacter genomospecies 1 and 2 are E145/08 and E157/08, respectively., (© 2014 IUMS.)

In Germany, rabies in bats is a notifiable zoonotic disease, which is caused by European bat lyssaviruses type 1 and 2 (EBLV-1 and 2), and the recently discovered new lyssavirus species Bokeloh bat lyssavirus (BBLV). As the understanding of bat rabies in insectivorous bat species is limited, in addition to routine bat rabies diagnosis, an enhanced passive surveillance study, i.e. the retrospective investigation of dead bats that had not been tested for rabies, was initiated in 1998 to study the distribution, abundance and epidemiology of lyssavirus infections in bats from Germany. A total number of 5478 individuals representing 21 bat species within two families were included in this study. The Noctule bat (Nyctalus noctula) and the Common pipistrelle (Pipistrellus pipistrellus) represented the most specimens submitted. Of all investigated bats, 1.17% tested positive for lyssaviruses using the fluorescent antibody test (FAT). The vast majority of positive cases was identified as EBLV-1, predominately associated with the Serotine bat (Eptesicus serotinus). However, rabies cases in other species, i.e. Nathusius' pipistrelle bat (Pipistrellus nathusii), P. pipistrellus and Brown long-eared bat (Plecotus auritus) were also characterized as EBLV-1. In contrast, EBLV-2 was isolated from three Daubenton's bats (Myotis daubentonii). These three cases contribute significantly to the understanding of EBLV-2 infections in Germany as only one case had been reported prior to this study. This enhanced passive surveillance indicated that besides known reservoir species, further bat species are affected by lyssavirus infections. Given the increasing diversity of lyssaviruses and bats as reservoir host species worldwide, lyssavirus positive specimens, i.e. both bat and virus need to be confirmed by molecular techniques.

White-nose syndrome (WNS) has claimed the lives of millions of hibernating insectivorous bats in North America. Its etiologic agent, the psychrophilic fungus Geomyces destructans, causes skin lesions that are the hallmark of the disease. The fungal infection is characterized by a white powdery growth on muzzle, ears and wing membranes. While WNS may threaten some species of North American bats with regional extinction, infection in hibernating bats in Europe seems not to be associated with significant mortality. We performed histopathological investigations on biopsy samples of 11 hibernating European bats, originating from 4 different countries, colonized by G. destructans. One additional bat was euthanized to allow thorough examination of multiple strips of its wing membranes. Molecular analyses of touch imprints, swabs and skin samples confirmed that fungal structures were G. destructans. Additionally, archived field notes on hibernacula monitoring data in the Harz Mountains, Germany, over an 11-year period (2000-2011) revealed multiple capture-recapture events of 8 banded bats repeatedly displaying characteristic fungal colonization. Skin lesions of G. destructans-affected hibernating European bats are intriguingly similar to the epidermal lesions described in North American bats. Nevertheless, deep invasion of fungal hyphae into the dermal connective tissue with resulting ulceration like in North American bats was not observed in the biopsy samples of European bats; all lesions found were restricted to the layers of the epidermis and its adnexae. Two bats had mild epidermal cupping erosions as described for North American bats. The possible mechanisms for any difference in outcomes of G. destructans infection in European and North American bats still need to be elucidated.

The genome of bat adenovirus 2 was sequenced and analyzed. It is similar in size (31,616 bp) to the genomes of bat adenovirus 3 and canine adenoviruses 1 and 2. These four viruses are monophyletic and share an identical genome organization, with one E3 gene and four E4 genes unique to this group among the mastadenoviruses. These findings suggest that canine adenoviruses may have originated by interspecies transfer of a vespertilionid bat adenovirus.

In recent years novel human respiratory disease agents have been described in South East Asia and Australia. The causative pathogens were classified as pteropine orthoreoviruses with strong phylogenetic relationship to orthoreoviruses of flying foxes inhabiting these regions. Subsequently, a zoonotic bat-to-human transmission has been assumed. We report the isolation of three novel mammalian orthoreoviruses (MRVs) from European bats, comprising bat-borne orthoreovirus outside of South East Asia and Australia and moreover detected in insectivorous bats (Microchiroptera). MRVs are well known to infect a broad range of mammals including man. Although they are associated with rather mild and clinically unapparent infections in their hosts, there is growing evidence of their ability to also induce more severe illness in dogs and man. In this study, eight out of 120 vespertilionid bats proved to be infected with one out of three novel MRV isolates, with a distinct organ tropism for the intestine. One isolate was analyzed by 454 genome sequencing. The obtained strain T3/Bat/Germany/342/08 had closest phylogenetic relationship to MRV strain T3D/04, isolated from a dog. These novel reoviruses provide a rare chance of gaining insight into possible transmission events and of tracing the evolution of bat viruses.

The zoonotic potential of paramyxoviruses is particularly demonstrated by their broad host range like the highly pathogenic Hendra and Nipah viruses originating from bats. But while so far all bat-borne paramyxoviruses have been identified in fruit bats across Africa, Australia, South America, and Asia, we describe the detection and characterization of the first paramyxoviruses in free-ranging European bats. Moreover, we examined the possible impact of paramyxovirus infection on individual animals by comparing histo-pathological findings and virological results. Organs from deceased insectivorous bats of various species were sampled in Germany and tested for paramyxovirus RNA in parallel to a histo-pathological examination. Nucleic acids of three novel paramyxoviruses were detected, two viruses in phylogenetic relationship to the recently proposed genus Jeilongvirus and one closely related to the genus Rubulavirus. Two infected animals revealed subclinical pathological changes within their kidneys, suggestive of a similar pathogenesis as the one described in fruit bats experimentally infected with Hendra virus.Our findings indicate the presence of bat-born paramyxoviruses in geographic areas free of fruit bat species and therefore emphasize a possible virus-host co-evolution in European bats. Since these novel viruses are related to the very distinct genera Rubulavirus and Jeilongvirus, a similarly broad genetic diversity among paramyxoviruses in other Microchiroptera compared to Megachiroptera can be assumed. Given that the infected bats were either found in close proximity to heavily populated human habitation or areas of intensive agricultural use, a potential risk of the emergence of zoonotic paramyxoviruses in Europe needs to be considered.