Drivers of longitudinal telomere dynamics in a long‐lived bat species,Myotis myotis

Age‐related telomere shortening is considered a hallmark of the ageing process. However, a recent cross‐sectional ageing study of relative telomere length (rTL) in bats failed to detect a relationship between rTL and age in the long‐lived genus Myotis (M. myotis and M. bechsteinii), suggesting some other factors are responsible for driving telomere dynamics in these species. Here, we test if longitudinal rTL data show signatures of age‐associated telomere attrition in M. myotis and differentiate which intrinsic or extrinsic factors are likely to drive telomere length dynamics. Using quantitative polymerase chain reaction, rTL was measured in 504 samples from a marked population, from Brittany, France, captured between 2013 and 2016. These represent 174 individuals with an age range of 0 to 7+ years. We find no significant relationship between rTL and age (p = .762), but demonstrate that within‐individual rTL is highly variable from year to year. To investigate the heritability of rTL, a population pedigree (n = 1744) was constructed from genotype data generated from a 16‐microsatellite multiplex, designed from an initial, low‐coverage, Illumina genome for M. myotis. Heritability was estimated in a Bayesian, mixed model framework, and showed that little of the observed variance in rTL is heritable (h² = 0.01–0.06). Rather, correlations of first differences, correlating yearly changes in telomere length and weather variables, demonstrate that, during the spring transition, average temperature, minimum temperature, rainfall and windspeed correlate with changes in longitudinal telomere dynamics. As such, rTL may represent a useful biomarker to quantify the physiological impact of various environmental stressors in bats.     

Spectral call features provide information about the aggression level of greater mouse-eared bats (Myotis myotis) during agonistic interactions

Bats rely heavily on acoustic signals in order to communicate with each other in a variety of social contexts. Among those, agonistic interactions and accompanying vocalizations have received comparatively little study. Here, we studied the communicational behaviour between male greater mouse-eared bats (Myotis myotis) during agonistic encounters. Two randomly paired adult males were placed in a box that allowed us to record video and sound synchronously. We describe their vocal repertoire and compare the acoustic structure of vocalizations between two aggression levels, which we quantified via the bats’ behaviour. By inspecting thirty, one-minute long encounters, we identified a rich variety of social calls that can be described as two basic call types: echolocation-like, low-frequency sweeps and long, broadband squawks. Squawks, the most common vocalization, were often noisy, i.e. exhibited a chaotic spectral structure. We further provide evidence for individual signatures and the presence of nonlinear phenomena in this species’ vocal repertoire. As the usage and acoustic structure of vocalizations is known to encode the internal state of the caller, we had predicted that the spectral structure of squawks would be affected by the caller’s aggression level. Confirming our hypothesis, we found that increased aggression positively correlated with an increase in call frequency and tonality. We hypothesize that the extreme spectral variability between and within squawks can be explained by small fluctuations in vocal control parameters (e.g. subglottal pressure) that are caused by the elevated arousal, which is in turn influenced by the aggression level.     

A new perspective on the zoogeography of the sibling mouse-eared bat species Myotis myotis and Myotis blythii: morphological, genetical and ecological evidence

The actual geographic distribution of the two sibling mouse-eared bat species Myotis myotis and Myotis blythii , which occur widely sympatrically in the western Palaearctic region, remains largely controversial. This concerns particularly the specific attribution of marginal populations from the Mediterranean islands and from adjacent areas of North Africa and Asia, which are morphologically intermediate between continental M. myotis and M. blythii from Europe. This study attempts to clarify this question by using four different approaches: cranial morphology, external morphology, genetics and trophic ecology. The three latter methods show unambiguously that North Africa, Malta, Sardinia and Corsica are presently inhabited by monospecific populations of M. myotis . In contrast, cranial morphometrics do not yield conclusive results. These results contradict all recent studies, which attribute North African and Maltese mouse-eared bats to M. blythii and consider that Sardinia and Corsica harbour sympatric populations of the two species. As concerns south-eastem populations, doubts are also expressed about the attribution of the subspecific taxon omari which may actually refer to M. myotis instead of M. blythii. Protein electrophoresis is presently the only absolute method available for determining M. myotis and M. blythii throughout their distribution ranges. However, species identification may be approached by relying on less sophisticated morphometrical methods as presented in this study. Species-specific habitat specializations are probably responsible for the differences observed between the geographic distributions of M. myotis and M. blythii , as they provide a logical groundwork for a coherent model of speciation for these two bat species.     

Population density of the greater mouse-eared bat (Myotis myotis), local diet composition and availability of foraging habitats

We studied the regional variation in population density of Myotis myotis (Borkhausen 1797) in south-eastern Bavaria, Germany, and its relations to diet composition and the availability of potential foraging habitats. We monitored colony size and juvenile mortality from 1991 to 2003, conducted faecal analyses in 1993 and determined land-use patterns around colonies. The numbers of individuals counted in the nursery colonies showed only small fluctuations over the years. However, data on colony size demonstrated a pronounced regional variation. Epigeic arthropods, mainly Carabidae, were the most important prey. The diet included prey taxa of forest as well as grassland habitats. The percentage of those prey taxa in the diet that originated in grassland managed with different intensity varied according to the availability of these potential foraging habitats around the nursery roosts. The calculated population density of the bats was positively correlated to forest area and especially to the area of mixed forest around the nursery roosts. Our results indicate that the availability of foraging habitats is a limiting factor for local population densities in the greater mouse-eared bat.     

Italy as a major Ice Age refuge area for the bat Myotis myotis (Chiroptera: Vespertilionidae) in Europe

The distribution of biota from the temperate regions changed considerably during the climatic fluctuations of the Quaternary. This is especially true for many bat species that depend on warm roosts to install their nursery colonies. Surveys of genetic variation among European bats have shown that the southern peninsulas (Iberia and the Balkans) harbour endemic diversity, but to date, no such surveys have been conducted in the third potential glacial refuge area, the Apennine peninsula. We report here the phylogeographical analysis of 115 greater mouse-eared bats ( Myotis myotis ) sampled throughout Italy, and show that 15 of the 18 different haplotypes found in the mitochondrial control region of these bats were unique to the Apennine peninsula. Colonies within this region also showed substantial genetic structure at both mitochondrial ( Φ_ST  = 0.47, P  < 0.001) and nuclear markers ( F _ST = 0.039, P  < 0.001). Based on a comprehensive survey of 575 bats from Europe, these genetic markers further indicate that central Italian populations of M. myotis are more closely related to Greek samples from across the Adriatic Sea, than to other Italian bats. Mouse-eared bat populations from the Apennine peninsula thus represent a complex mixture of several endemic lineages, which evolved in situ , with others that colonized this region more recently along an Adriatic route. Our broad survey also confirms that the Alps represent a relatively impermeable barrier to gene flow for Apennine lineages, even for vagile animals such as bats. These results underline the conservation value of bats from this region and the need to include the Apennine peninsula in phylogeographical surveys in order to provide a more accurate view of the evolution of bats in Europe.     

Genetic variation and population structure in the endangered greater horseshoe bat Rhinolophus ferrumequinum

Following a dramatic decline last century, the British population of the endangered greater horseshoe bat Rhinolophus ferrumequinum is highly fragmented. To examine the consequences of fragmentation and limited dispersal on patterns of genetic structure and variation, we used microsatellite markers to screen bats from around 50% of the known maternity colonies in Britain, and two areas from continental Europe. Analyses revealed that Welsh and English colonies were genetically isolated. This, and lower variability in Britain than north France, may result from either genetic drift, or the species' colonization history. Gene flow among most neighbouring colonies was not generally restricted, with one exception. These findings have important implications for the ongoing conservation management of this species.     

Temporal dynamics of European bat Lyssavirus type 1 and survival of Myotis myotis bats in natural colonies

Many emerging RNA viruses of public health concern have recently been detected in bats. However, the dynamics of these viruses in natural bat colonies is presently unknown. Consequently, prediction of the spread of these viruses and the establishment of appropriate control measures are hindered by a lack of information. To this aim, we collected epidemiological, virological and ecological data during a twelve-year longitudinal study in two colonies of insectivorous bats (Myotis myotis) located in Spain and infected by the most common bat lyssavirus found in Europe, the European bat lyssavirus subtype 1 (EBLV-1). This active survey demonstrates that cyclic lyssavirus infections occurred with periodic oscillations in the number of susceptible, immune and infected bats. Persistence of immunity for more than one year was detected in some individuals. These data were further used to feed models to analyze the temporal dynamics of EBLV-1 and the survival rate of bats. According to these models, the infection is characterized by a predicted low basic reproductive rate (R(0) = 1.706) and a short infectious period (D = 5.1 days). In contrast to observations in most non-flying animals infected with rabies, the survival model shows no variation in mortality after EBLV-1 infection of M. myotis. These findings have considerable public health implications in terms of management of colonies where lyssavirus-positive bats have been recorded and confirm the potential risk of rabies transmission to humans. A greater understanding of the dynamics of lyssavirus in bat colonies also provides a model to study how bats contribute to the maintenance and transmission of other viruses of public health concern.     

Paternity assessment in wild groups of toque macaques Macaca sinica at Polonnaruwa, Sri Lanka using molecular markers

Genetic variation at four microsatellite loci in conjunction with that at a highly variable allozyme locus was used to analyse paternity over a 12-year period in 13 social groups of toque macaques Macaca sinica inhabiting a natural forest in Polonnaruwa, Sri Lanka. Paternity exclusion analysis revealed that the set of offspring produced by a female usually consists of half-siblings because few males father more than one offspring with a particular female. No evidence of offspring produced by matings between first degree relatives was found. The social unit in toque macaques was not identical to the reproductive unit and the possibility of paternity by males outside the social group should be considered when estimating male reproductive output. Although it was common for multiple males to father offspring in a social group each year, reproduction within a group during a breeding season tended to be limited to a few males. The mean number of males reproducing per group per year was independent of the number of males in a group. The paternity data suggests that many males may father relatively few offspring during their entire lives and that the effective population size for toque macaques may be much smaller than indicated by demographic data.     

Telomere shortening correlates with harsh weather conditions in the bat species Myotis myotis

The relationship of telomere shortening and cellular ageing in cultured cells such as fibroblasts is straightforward: telomeres shorten with an increasing number of cell divisions until they trigger replicative senescence which prevents further mitotic cycles. But studies investigating the relationship between telomere shortening and ageing in whole organisms show contrasting results: while there is a clear decline in telomere length (TL) with chronological age in some species such as humans, no such decline is observed in others. In this issue of Molecular Ecology, Foley et al. (2020) show that experiencing harsh weather conditions correlates with longitudinal telomere shortening in the bat species Myotis myotis, whereas chronological age does not (Foley et al., 2020). Further, the authors investigated whether genetics influence TL and find a low heritability (h² = 0.01–0.06) again suggesting that environmental effects are the dominant drivers of variation in TL in this species. These are important findings as there is disagreement in the literature about the relative magnitude of genetic and environmental effects contributing to TL variation in different species. This paper investigating the impact of environmental effects makes a novel and important contribution to the literature on TL in free‐living mammals.     

Paternity assessment in rhesus macaques (Macaca mulatta): Multilocus DNA fingerprinting and PCR marker typing

Establishing kinship relations in primates using modern molecular genetic techniques has enhanced the ability to scrutinize a number of fundamental biological issues. We screened 51 human short tandem repeats (STRs) for cross-species PCR amplification in rhesus macaques (Macaca mulatta) and identified 11 polymorphic loci with heterozygosity rates of at least 0.6. These markers were used for paternity testing in three social groups (M, R, and S) of rhesus macaques from Cayo Santiago, Puerto Rico. Several consecutive birth cohorts were analyzed in which approximately 200 males were tested for paternity against more than 100 mother/infant pairs. Despite a combined exclusion rate of more than 99.9% in all three groups, some cases could not be solved unequivocally with the STR markers and additional testing of the MHC-associated DQB1 polymorphism. A final decision became possible through multilocus DNA fingerprinting with one or more of the oligonucleotide probes (GATA)₄, (CA)₈, and (CAC)₅. Paternity assessment by multilocus DNA analysis with probe (CAC)₅ alone was found to have limitations in rhesus macaques as regards the number of potential sires which might be involved in a given case. Multilocus DNA fingerprinting requires large amounts of DNA, and the ensuing autoradiographic patterns present difficulties in comparisons across gels and even within the same gel across remote lanes. Computer-assisted image analysis was incapable of eliminating this problem. Therefore, a dual approach to DNA typing has been adopted, using STR markers to reduce the number of potential sires to a level where all remaining candidates can be tested by multilocus DNA fingerprinting on a single gel, preferably in lanes adjacent to the mother/infant pair. Am. J. Primatol. 44:1–18, 1998. © 1998 Wiley-Liss, Inc.     

Contrasted patterns of mitochondrial and nuclear structure among nursery colonies of the bat Myotis myotis

Thirteen nursery colonies of Myotis myotis were sampled in central Europe to investigate the dispersal behaviour of this bat species. Mitochondrial DNA sequences of 260 bats reveal the occurrence of three evolutionary lineages that have probably originated in distinct glacial refugia and meet in a contact zone near the Alps. Moreover, the strong haplotypic segregation (Φ_ST=0.540) suggests that breeding females are philopatric. Contrastingly, the low population structure at 15 microsatellite loci (F_ST=0.022), suggests the homogenizing effect of nuclear gene flow. The different perspectives given by these two markers are consistent with strong male‐biased dispersal. As a result of female philopatry, the local haplotypic variability seems to be largely influenced by historical processes of colonization. Conversely, the homogeneity of nuclear variability within roosts that are located north of the Alps seems to mainly reflect contemporary gene flow. Finally, despite the fact that females are faithful to their natal colony, movements of both males and females occur outside the breeding period. Mitochondrial survey of individuals sampled exclusively in nurseries may thus poorly reflect the metapopulation dynamics of this species.     

Patterns of population subdivision and gene flow in the ant Nothomyrmecia macrops reflected in microsatellite and mitochondrial DNA markers

The Australian endemic ant Nothomyrmecia macrops is renowned for having retained a large proportion of 'primitive' morphological and behavioural characters. Another less studied peculiarity of this species is the production of short-winged (brachypterous) female sexuals, which presumably are poor dispersers. The males, in contrast, bear a full set of normally developed wings and thus may disperse widely. We investigated patterns of genetic differentiation within and among three distantly separated populations in South Australia using nine polymorphic microsatellite loci and four regions of mitochondrial DNA (COI, COII, Cytb, lrRNA). We sampled eight subpopulations, one in the Lake Gilles CP, two near Penong and five around Poochera where distances ranged from 360 km to sites separated by 2-10 km. Only little differentiation was found at the local scale (within the assumed dispersal distance of males) using nuclear markers, whereas the three distant locations were moderately differentiated (FST = 0.06). Mitochondrial DNA genetic structure was much more pronounced on all scales (phiST = 0.98), with regular differences in both haplotype composition and frequency even occurring among closely located sites. This lack of congruence between nuclear and mitochondrial markers strongly suggests limited female dispersal and male-biased gene flow among populations. As to the conservation status of the species there is no evidence for severe population reductions in the recent past, which would have left populations genetically depauperate.     

Fine-scale genetic structuring in a natural population of European wild rabbits (Oryctolagus cuniculus)

The genetic structure of a free-living tagged population of European wild rabbits (Oryctolagus cuniculus) was investigated for two consecutive years (1990 and 1991) using 10 polymorphic microsatellite loci. A specific social behaviour, the formation of stable breeding groups, influenced the genetic structure of the population. These breeding groups were shown to constitute genetically differentiated units with low levels of gene flow between them. The average relatedness among members of a social group was higher than within the population as a whole. As a result of female philopatry coupled with male-biased natal dispersal, the relatedness of females was higher than that of males, both within social groups and in the whole population. Furthermore, the average relatedness of females within groups was twice the relatedness of females between groups. This study reveals marked fine-scale, intrapopulation genetic structure, which is attributable to the social behaviour of the European wild rabbit.     

Genetic consequences of the ice ages on nurseries of the bat Myotis myotis: a mitochondrial and nuclear survey

Analyses of mitochondrial DNA (mtDNA) control region polymorphism and of variation at 10 nuclear microsatellite loci were used to investigate the mechanisms and genetic consequences of postglacial expansion of Myotis myotis in Europe. Initial sampling consisted of 480 bats genotyped in 24 nursery colonies arranged along a transect of approximately 3000 km. The phylogeographical survey based on mtDNA sequences revealed the existence of major genetic subdivisions across this area, with several suture zones between haplogroups. Such zones of secondary contact were found in the Alps and Rhodopes, whereas other potential barriers to gene flow, like the Pyrenees, did not coincide with genetic discontinuities. Areas of population admixture increased locally the genetic diversity of colonies, which confounded the northward decrease in nucleotide diversity predicted using classical models of postglacial range expansion. However, when analyses were restricted to a subset of 15 nurseries originating from a single presumed glacial refugium, mtDNA polymorphism did indeed support a northwards decrease in diversity. Populations were also highly structured (PhiST = 0.384). Conversely, the same subset of colonies showed no significant latitudinal decrease in microsatellite diversity and much less population structure (FST = 0.010), but pairwise genetic differentiation at these nuclear markers was strongly correlated with increasing geographical distance. Together, this evidence suggests that alleles carried via male bats have maintained enough nuclear gene flow to counteract the effects of recurrent bottlenecks generally associated with recolonization processes. As females are highly philopatric, we argue that the maternally transmitted mtDNA marker better reflects the situation of past, historical gene flow, whereas current levels of gene flow are better reflected by microsatellite markers.