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.     

Cytonuclear discordance and the species status of Myotis myotis and Myotis blythii (Chiroptera)

We used both highly variable mitochondrial and nuclear loci to investigate the large mouse‐eared bat species complex in the Western Palaearctic to clarify their systematic position. Although mitochondrial lineages show no species segregation and some haplotypes are shared between Myotis myotis and Myotis blythii sensu lato, Bayesian clustering methods based on multilocus genotypes indicate highly concordant nuclear and morphological species assignment. These multilocus, nuclear analyses detected only a single putative F₁ hybrid in the extensive areas of sympatry sampled, thus confirming the biological species status of M. myotis and M. blythii s.l. We propose that the strong cytonuclear discordance in these species complex results from a combination of prior spatial isolation of the two species in different glacial refugia, followed by a succession of mitochondrial introgression events that occurred during the eastward and westward expansions of M. myotis and of M. blythii, respectively. The nuclear markers further indicate the presence of a notable genetic discontinuity within M. myotis that broadly separates populations into an eastern and a western component with an overlap zone in the Balkans. This eastern and western discontinuity is also apparent in the mitochondrial lineages with the D haplogroup largely confined to samples found in Thrace and Asia Minor. None of these genetic discontinuities correspond to the distribution of the two commonly recognized M. myotis subspecies (myotis and macrocephalicus). We also show that distinct morphological subspecies within M. blythii (oxygnathus, omari, risorius and lesviacus) in Europe and the near‐East do not correlate with significant evolutionarily units, whether identified by mitochondrial or nuclear data and thus only represent local morphological variants with little taxonomic relevance.     

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 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.     

Phylogeography of the large Myotis bats (Chiroptera: Vespertilionidae) in Europe,Asia Minor,and Transcaucasia

The large Myotis complex in continental Europe, Asia Minor, and Transcaucasia comprises two sibling bat species, the greater mouse‐eared bat, Myotis myotis, and the lesser mouse‐eared bat, Myotis blythii, also referred to as Myotis oxygnathus. Here, we investigate the phylogeography of these bats using two mitochondrial markers: the second hypervariable domain of the control region (HVII) and a fragment of the cytochrome b gene (cyt b). The HVII haplotypes formed six distinct haplogroups associated with different geographical regions. Most of the European HVII haplotypes were exclusive to M. myotis, whereas the majority of HVII haplotypes found in Asia Minor were exclusive to M. blythii/M. oxygnathus. The phylogenetic reconstruction based on the concatenated cyt b and HVII fragments recovered two major lineages. The first lineage comprised samples from Europe (western lineage), and the second lineage included samples from Asia Minor, Transcaucasia, Crimea, Western Ukraine, Thrace, the Balkans, and Eastern Europe (eastern lineage). The mitochondrial lineage of M. blythii, reported from Kyrgyzstan, was not present in Asia Minor and Transcaucasia. Therefore, we consider the possibility that the M. blythii/M. oxygnathus found in Europe, Asia Minor, and Transcaucasia are not recent descendants of the Central Asian M. blythii. Instead, we suggest that M. blythii/M. oxygnathus and M. myotis diverged through allopatric speciation in Asia Minor and Europe, and that they are represented by the eastern and western mitochondrial lineages. We also examine an alternative hypothesis: that the large Myotis complex consists of more than two species that diverged independently in Asia Minor and Europe through ecological speciation. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, ??, ??–??.     

Is the Gibraltar Strait a barrier to gene flow for the bat Myotis myotis (Chiroptera: Vespertilionidae)?

Because of their role in limiting gene flow, geographical barriers like mountains or seas often coincide with intraspecific genetic discontinuities. Although the Strait of Gibraltar represents such a potential barrier for both plants and animals, few studies have been conducted on its impact on gene flow. Here we test this effect on a bat species (Myotis myotis) which is apparently distributed on both sides of the strait. Six colonies of 20 Myotis myotis each were sampled in southern Spain and northern Morocco along a linear transect of 1350 km. Results based on six nuclear microsatellite loci reveal no significant population structure within regions, but a complete isolation between bats sampled on each side of the strait. Variability at 600 bp of a mitochondrial gene (cytochrome b) confirms the existence of two genetically distinct and perfectly segregating clades, which diverged several million years ago. Despite the narrowness of the Gibraltar Strait (14 km), these molecular data suggest that neither males, nor females from either region have ever reproduced on the opposite side of the strait. Comparisons of molecular divergence with bats from a closely related species (M. blythii) suggest that the North African clade is possibly a distinct taxon warranting full species rank. We provisionally refer to it as Myotis cf punicus Felten 1977, but a definitive systematic understanding of the whole Mouse-eared bat species complex awaits further genetic sampling, especially in the Eastern Mediterranean areas.     

The mitochondrial and nuclear genetic structure of Myotis capaccinii (Chiroptera: Vespertilionidae) in the Eurasian transition, and its taxonomic implications

Allopatric isolation in glacial refugia has caused differentiation and speciation in many taxa globally. In this study, we investigated the nuclear and mitochondrial genetic differentiation of the long fingered bat, Myotis capaccinii during the ice ages in south-eastern Europe and Anatolia. The mitochondrial DNA (mtDNA) analyses indicated a suture zone similar to those recorded in other animal species, including bats, suggesting the association of more than one refugium with the region. Contrary to most of the other species where a suture zone was seen in Anatolia, for M. capaccinii the geographical location of the genetic break was in south-eastern Europe. This mitochondrial differentiation was not reflected in the nuclear microsatellites, however, suggesting that the lack of contact during the ice ages did not result in reproductive isolation. Hence taxonomically, the two mitochondrial clades cannot be treated as separate species.     

Male roosting habits and mating behaviour of Myotis myotis

Attic-dwelling male Myotis myotis use different roost sites, but prefer one (or a few) of them and visit others sporadically. Roost-site preference can change, especially during the mating season. The males use their roost sites over years. Mean occupancy of all roost sites shows a maximum during the mating season. Females of different nursery colonies meet at male roost sites as far as 12 km from their colonies. They stay, on average, four days with one male, may join several males during one mating season and may visit their mating sites over years. Males appear to show no special behaviour to attract females but extend the time they are present at the roost site during the mating season. Each male is visited by about seven adult mates, on average. Differences in reproductive success are indicated by the times males were joined by females and by the numbers of mates and copulations. Two typical mating postures, copulation and five different social calls are described. Since females of different colonies meet at the male roost sites, the mating system of Myotis myotis may favour genetic exchange between colonies.     

Paternity assessment and population subdivision in a natural population of the larger mouse-eared bat Myotis myotis

A hypervariable simple sequence locus and mitochondrial D-loop sequences were used to analyse genetically a natural population of the larger mouse-eared bat Myotis myotis in southern Bavaria. Tests for population subdivision and direct observations suggest that females return to their natal sites, while males disperse. The males present in female nursery colonies are not related to the females. Paternity assessment for 46 offspring from a particular nursery colony showed that there are no males that monopolize the reproduction, and that the resident males in the colony had only a small mating success. Instead, the results suggest that females actively seek matings outside their colony. Most interestingly, it appears that a group of males about 16 km away from the nursery colony had a relatively high mating success and that this group of males may be related to the females of the nursery colonies. If this finding can be confirmed in a larger study, it may have important consequences for future conservation strategies.     

同地共栖三种鼠耳蝠食性差异及其生态位分化

2005年9—11月在贵州省安龙县笃山乡暗河村,分析了共栖同一山洞3种鼠耳蝠的形态特征和食性。在体型上,华南水鼠耳蝠体重为(4.46±0.53)g,前臂长为(34.63±1.45)mm;毛腿鼠耳蝠体重为(5.15±1.76)g,前臂长为(35.20±1.07)mm;西南鼠耳蝠体重为(10.94±0.87)g,前臂长为(45.21±1.15)mm。3种鼠耳蝠的体重两两之间差异显著,西南鼠耳蝠与另外2种鼠耳蝠的前臂长相比两两之间差异显著。在食物组成上,华南水鼠耳蝠主要捕食近水面活动的双翅目及其幼虫,体积百分比和频次百分比分别为79.7%和100%;毛腿鼠耳蝠主要捕食双翅目和小型鞘翅目,体积百分比分别占59.6%和28.8%,频次百分比分别为91.3%和80.1%;西南鼠耳蝠的食物组成主要为近地面或在地表活动的鞘翅目步甲科和埋葬虫科昆虫,体积百分比和频次百分比分别为80.8%和100%;3种鼠耳蝠食物组成存在显著差异。结果表明,同地共栖3种鼠耳蝠除了形态结构上出现差异,食物组成也存在明显的差异。据此,推测3种鼠耳蝠可能采取不同的捕食生境和捕食策略,从而导致捕食生态位分离,避免出现激烈竞争,使得3种近缘鼠耳蝠能够同地共栖。     

Genetic structure and diversity of a rare woodland bat, <Emphasis Type="Italic">Myotis bechsteinii</Emphasis>: comparison of continental Europe and Britain

The Bechstein’s bat (Myotis bechsteinii) is a rare sedentary bat considered to be highly reliant on the presence of ancient woodland. Understanding the genetic connectivity and population structure of such elusive mammals is important for assessing their conservation status. In this study, we report the genetic diversity and structure of M. bechsteinii across Britain and Europe. Assessments were made using 14 microsatellite markers and a 747 bp region of the mitochondrial cytochrome b gene. Nuclear DNA (microsatellites) showed high levels of genetic diversity and little inbreeding across the species range, though genetic diversity was slightly lower in Britain than in mainland Europe. Bayesian and spatial PCA analysis showed a clear separation between the British and European sites. Within Europe, the Italian population south of the Alps was isolated from the other sites. In Britain, there was genetic structuring between the northern and southern part of the geographical range. Despite there being little genetic divergence in mitochondrial DNA (mtDNA) sequences throughout most of Europe, the mtDNA patterns in Britain confirmed this separation of northern and southern populations. Such genetic structuring within Britain—in the absence of any obvious physical barriers—suggests that other factors such as land-use may limit gene-flow.     

贵州省鼠耳蝠属一新纪录——狭耳鼠耳蝠

2010年10月在贵州省荔波县进行翼手目动物调查过程中,于荔波县洞塘乡捕获14只鼠耳蝠,经鉴定为狭耳鼠耳蝠Myotisblythii。主要鉴别特征:体型中等,前臂长64.60(60.78～67.72)mm(n=14,SD=1.71);耳狭长;耳屏直而细长,顶端尖锐;第三指最长、第五指最短;后足长约为胫长之半;无距缘膜;尾长67.99(60.56～73.82)mm(n=14,SD=3.7);雄性个体无阴茎骨,阴茎长12.58(8.42～15.72)mm(n=5,SD=2.86);颅全长24.09(23.48～24.72)mm(n=6,SD=0.55);头骨窄长;听泡发达;矢状脊较低;上颌内门齿有1个主尖和1个小附尖,外门齿小,与上犬齿分离;第2上前臼齿(P3)稍位于齿列内侧。此种鼠耳蝠为贵州省翼手目新纪录。     

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.     

Post-Pleistocene range expansion of the recently imperiled eastern little brown bat (Myotis lucifugus lucifugus) from a single southern refugium

Myotis lucifugus, once among the most widespread and common bats in North America, has been forecast to be extirpated east of the Rockies in as few as 16 years by the spread of white-nose syndrome. Recent genetic research has demonstrated that this species is paraphyletic and part of a broader species complex; however, only one lineage (Myotis lucifugus lucifugus [M. l. lucifugus]) is present in eastern North America. I used molecular tools and niche modeling to validate this and investigate the role that historical biogeography has played in the phylogenetic and population genetic structure of this species to determine if the eastern subspecies represents an evolutionarily distinct population.To establish the genetic structure within M. l. lucifugus, I densely sampled maternity colonies in Minnesota and sequenced 182 individuals for a portion of cytochrome b. Phylogenetic reconstruction and a haplotype network were used to infer the relationships among mitochondrial haplotypes. Population growth statistics were calculated to determine if there was evidence of significant expansion, and an environmental niche model (ENM) was constructed based on conditions during the last glacial maximum (LGM) to illustrate potential glacial refugia. All individuals derived from a single mitochondrial lineage. Genetic evidence points to population growth starting approximately 18 kya. ENM results show that there was likely a single large southern refugium extending across the southeastern United States and possibly several isolated refugia in western North America. Myotis lucifugus lucifugus likely maintained both a large range and a large population during the peaks of the glacial cycles, and its population appears to have expanded following the retreat of the Laurentide ice sheet. This imperiled lineage likely diverged in isolation from other members of the M. lucifugus/western long-eared Myotis during the Pleistocene.     

北京房山发现华南水鼠耳蝠

目前我国基础资源调查中有关翼手目物种多样性及其分布的项目正在开展,着手建立翼手目物种多样性和分布数据库,对翼手目物种在中国的分布情况亟需广泛调查和深入研究。我们于2021年9月在北京市开展翼手目物种调查时,在北京房山十渡镇四御洞使用竖琴网捕获8只蝙蝠,经过COI和Cyt b序列比对和系统发育关系重建以及外部形态数据鉴定为华南水鼠耳蝠（Myotis laniger）。此次发现华南水鼠耳蝠是北京市翼手目物种分布新记录种。     

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.     

A population genetic analysis of migration: the case of the noctule bat (Nyctalus noctula)

Although rarely assessed, the population genetics of hibernating colonies can help to understand some aspects of population structure, even when samples from nursery or mating colonies are not available, or in studies of migration when both types of samples are available and can be compared. Here we illustrate both points in a survey of mitochondrial DNA (mtDNA) control region sequences used to study the population genetics of hibernating colonies of a migrating species, the noctule bat (Nyctalus noctula). Lacking samples from Scandinavian nursery colonies, we use a North European hibernacula to suggest that Scandinavian populations are isolated from Central and East European colonies. Then, we compare genetic diversities of nursery and hibernating colonies. We find a significantly higher haplotype diversity in hibernacula, confirming that they consist of individuals from different nursery colonies. Finally, we show that pairwise comparisons of the haplotype frequencies of nursery and hibernating colonies contain some information on the migration direction of the noctule bat.     

Genetic structuring of northern myotis (Myotis septentrionalis) at multiple spatial scales

Although groups of bats may be genetically distinguishable at large spatial scales, the effects of forest disturbances, particularly permanent land use conversions on fine-scale population structure and gene flow of summer aggregations of philopatric bat species are less clear. We genotyped and analyzed variation at 10 nuclear DNA microsatellite markers in 182 individuals of the forest-dwelling northern myotis (Myotis septentrionalis) at multiple spatial scales, from within first-order watersheds scaling up to larger regional areas in West Virginia and New York. Our results indicate that groups of northern myotis were genetically indistinguishable at any spatial scale we considered, and the collective population maintained high genetic diversity. It is likely that the ability to migrate, exploit small forest patches, and use networks of mating sites located throughout the Appalachian Mountains, Interior Highlands, and elsewhere in the hibernation range have allowed northern myotis to maintain high genetic diversity and gene flow regardless of forest disturbances at local and regional spatial scales. A consequence of maintaining high gene flow might be the potential to minimize genetic founder effects following population declines caused currently by the enzootic White-nose Syndrome.     

Cryptic diversity in European bats

Different species of bat can be morphologically very similar. In order to estimate the amount of cryptic diversity among European bats we screened the intra- and interspecific genetic variation in 26 European vespertilionid bat species. We sequenced the DNA of subunit 1 of the mitochondrial protein NADH dehydrogenase (ND1) from several individuals of a species, which were sampled in a variety of geographical regions. A phylogeny based on the mitochondrial (mt) DNA data is in good agreement with the current classification in the family. Highly divergent mitochondrial lineages were found in two taxa, which differed in at least 11% of their ND1 sequence. The two mtDNA lineages in Plecotus austriacus correlated with the two subspecies Plecotus austriacus austriacus and Plecotus austriacus kolombatovici. The two mtDNA lineages in Myotis mystacinus were partitioned among two morphotypes. The evidence for two new bat species within Europe is discussed. Convergent adaptive evolution might have contributed to the morphological similarity among distantly related species if they occupy similar ecological niches. Closely related species may differ in their ecology but not necessarily in their morphology. On the other hand, two morphologically clearly different species (Eptesicus serotinus and Eptesicus nilssonii) were found to be genetically very similar. Neither morphological nor mitochondrial DNA sequence analysis alone can be guaranteed to identify species.     

Direction and extent of organelle DNA introgression between two spruce species in the Qinghai-Tibetan Plateau

A recent model has shown that, during range expansion of one species in a territory already occupied by a related species, introgression should take place preferentially from the resident species towards the invading species and genome components experiencing low rates of gene flow should introgress more readily than those experiencing high rates of gene flow. Here, we use molecular markers from two organelle genomes with contrasted rates of gene flow to test these predictions by examining genetic exchanges between two morphologically distinct spruce Picea species growing in the Qinghai-Tibetan Plateau. The haplotypes from both mitochondrial (mt) DNA and chloroplast (cp) DNA cluster into two distinct lineages that differentiate allopatric populations of the two species. By contrast, in sympatry, the species share the same haplotypes, suggesting interspecific genetic exchanges. As predicted by the neutral model, all sympatric populations of the expanding species had received their maternally inherited mtDNA from the resident species, whereas for paternally inherited cpDNA introgression is more limited and not strictly unidirectional. Our results underscore cryptic introgressions of organelle DNAs in plants and the importance of considering rates of gene flow and range shifts to predict direction and extent of interspecific genetic exchanges.