Assessing bat droppings and predatory bird pellets for vector-borne bacteria: molecular evidence of bat-associated Neorickettsia sp. in Europe.

In Europe, several species of bats, owls and kestrels exemplify highly urbanised, flying vertebrates, which may get close to humans or domestic animals. Bat droppings and bird pellets may have epidemiological, as well as diagnostic significance from the point of view of pathogens. In this work 221 bat faecal and 118 bird pellet samples were screened for a broad range of vector-borne bacteria using PCR-based methods. Rickettsia DNA was detected in 13 bat faecal DNA extracts, including the sequence of a rickettsial insect endosymbiont, a novel Rickettsia genotype and Rickettsia helvetica. Faecal samples of the pond bat (Myotis dasycneme) were positive for a Neorickettsia sp. and for haemoplasmas of the haemofelis group. In addition, two bird pellets (collected from a Long-eared Owl, Asio otus, and from a Common Kestrel, Falco tinnunculus) contained the DNA of a Rickettsia sp. and Anaplasma phagocytophilum, respectively. In both of these bird pellets the bones of Microtus arvalis were identified. All samples were negative for Borrelia burgdorferi s.l., Francisella tularensis, Coxiella burnetii and Chlamydiales. In conclusion, bats were shown to pass rickettsia and haemoplasma DNA in their faeces. Molecular evidence is provided for the presence of Neorickettsia sp. in bat faeces in Europe. In the evaluated regions bat faeces and owl/kestrel pellets do not appear to pose epidemiological risk from the point of view of F. tularensis, C. burnetii and Chlamydiales. Testing of bird pellets may provide an alternative approach to trapping for assessing the local occurrence of vector-borne bacteria in small mammals.

Female pond bats hunt in other areas than males and consume lighter prey when pregnant.

Animals with large energy requirements are forced to optimize their hunting strategy, which may result in differentiation of the diet between sexes and across seasons. Here, we examined spatiotemporal variation in the diet of both sexes of the Pond Bat Myotis dasycneme, a species known to have spatial segregation of sexes when the young are born and lactating. Fecal pellets were collected from live animals for a period of 15 years at various locations in the Netherlands. A total of 535 pellets were successfully analyzed by microscopy and an additional 160 pellets by DNA metabarcoding. Morphological and molecular analyses showed that the diet of pregnant and lactating pond bats differed significantly from the diet of females with no reproductive investment. Further analyses of the data showed that pregnant female pond bats are highly dependent on small prey and pupae, mainly nonbiting midges and mosquitoes (Diptera: Chironomidae and Culicidae). These insects can be found in large quantities in peatlands intersected with shallow waterways, the habitat type in which female pond bats were observed more often than males. Our results suggest that during pregnancy the spatial segregation of sexes coincides with sex-specific diets, which might reflect habitat selection based on energy requirements, in addition to lowered intraspecific competition.

Male long-distance migrant turned sedentary; The West European pond bat (Myotis dasycneme) alters their migration and hibernation behaviour.

During autumn in the temperate zone, insectivorous male bats face a profound energetic challenge, as in the same period they have to make energy choices related to hibernation, mating and migration. To investigate these energetic trade-offs, we compared the body mass of male and female pond bats (Myotis dasycneme) through the summer season, characterized the known hibernacula in terms of male or female bias, and subsequently compared their population trend during two study periods, between 1930-1980 and 1980-2015. Towards the end of summer, males began losing weight whilst females were simultaneously accumulating fat, suggesting that males were pre-occupied with mating. We also found evidence for a recent adaptation to this energetic trade-off, males have colonised winter roosts in formerly unoccupied areas, which has consequently led to a change in the migration patterns for the male population of this species. As male bats do not assist in raising offspring, males have ample time to restore their energy balance after hibernation. Our results suggest that choosing a hibernacula closer to the summer range not only decreases energy cost needed for migration, it also lengthens the mating season of the individual male. Our findings have important conservation implications, as male and female biased hibernation assemblages may differ critically in terms of microclimate preferences.

Bats Swarm Where They Hibernate: Compositional Similarity between Autumn Swarming and Winter Hibernation Assemblages at Five Underground Sites.

During autumn in the temperate zone of both the new and old world, bats of many species assemble at underground sites in a behaviour known as swarming. Autumn swarming behaviour is thought to primarily serve as a promiscuous mating system, but may also be related to the localization and assessment of hibernacula. Bats subsequently make use of the same underground sites during winter hibernation, however it is currently unknown if the assemblages that make use of a site are comparable across swarming and hibernation seasons. Our purpose was to characterize the bat assemblages found at five underground sites during both the swarming and the hibernation season and compare the assemblages found during the two seasons both across sites and within species. We found that the relative abundance of individual species per site, as well as the relative proportion of a species that makes use of each site, were both significantly correlated between the swarming and hibernation seasons. These results suggest that swarming may indeed play a role in the localization of suitable hibernation sites. Additionally, these findings have important conservation implications, as this correlation can be used to improve monitoring of underground sites and predict the importance of certain sites for rare and cryptic bat species.

Screening of bat faeces for arthropod-borne apicomplexan protozoa: Babesia canis and Besnoitia besnoiti-like sequences from Chiroptera.

BACKGROUND: Bats are among the most eco-epidemiologically important mammals, owing to their presence in human settlements and animal keeping facilities. Roosting of bats in buildings may bring pathogens of veterinary-medical importance into the environment of domestic animals and humans. In this context bats have long been studied as carriers of various pathogen groups. However, despite their close association with arthropods (both in their food and as their ectoparasites), only a few molecular surveys have been published on their role as carriers of vector-borne protozoa. The aim of the present study was to compensate for this scarcity of information. FINDINGS: Altogether 221 (mostly individual) bat faecal samples were collected in Hungary and the Netherlands. The DNA was extracted, and analysed with PCR and sequencing for the presence of arthropod-borne apicomplexan protozoa. Babesia canis canis (with 99-100% homology) was identified in five samples, all from Hungary. Because it was excluded with an Ixodidae-specific PCR that the relevant bats consumed ticks, these sequences derive either from insect carriers of Ba. canis, or from the infection of bats. In one bat faecal sample from the Netherlands a sequence having the highest (99%) homology to Besnoitia besnoiti was amplified. CONCLUSIONS: These findings suggest that some aspects of the epidemiology of canine babesiosis are underestimated or unknown, i.e. the potential role of insect-borne mechanical transmission and/or the susceptibility of bats to Ba. canis. In addition, bats need to be added to future studies in the quest for the final host of Be. besnoiti.

Pan-European distribution of white-nose syndrome fungus (Geomyces destructans) not associated with mass mortality.

BACKGROUND: The dramatic mass mortalities amongst hibernating bats in Northeastern America caused by "white nose-syndrome" (WNS) continue to threaten populations of different bat species. The cold-loving fungus, Geomyces destructans, is the most likely causative agent leading to extensive destruction of the skin, particularly the wing membranes. Recent investigations in Europe confirmed the presence of the fungus G. destructans without associated mass mortality in hibernating bats in six countries but its distribution remains poorly known. METHODOLOGY/PRINCIPAL FINDINGS: We collected data on the presence of bats with white fungal growth in 12 countries in Europe between 2003 and 2010 and conducted morphological and genetic analysis to confirm the identity of the fungus as Geomyces destructans. Our results demonstrate the presence of the fungus in eight countries spanning over 2000 km from West to East and provide compelling photographic evidence for its presence in another four countries including Romania, and Turkey. Furthermore, matching prevalence data of a hibernaculum monitored over two consecutive years with data from across Europe show that the temporal occurrence of the fungus, which first becomes visible around February, peaks in March but can still be seen in some torpid bats in May or June, is strikingly similar throughout Europe. Finally, we isolated and cultured G. destructans from a cave wall adjacent to a bat with fungal growth. CONCLUSIONS/SIGNIFICANCE: G. destructans is widely found over large areas of the European continent without associated mass mortalities in bats, suggesting that the fungus is native to Europe. The characterisation of the temporal variation in G. destructans growth on bats provides reference data for studying the spatio-temporal dynamic of the fungus. Finally, the presence of G. destructans spores on cave walls suggests that hibernacula could act as passive vectors and/or reservoirs for G. destructans and therefore, might play an important role in the transmission process.