The two sibling bat species Myotis myotis and Myotis blythii occur in sympatry over wide areas of Southern and Central Europe. Morphological, ecological and previous genetic evidence supported the view that the two species constitute two well-differentiated groups, but recent phylogenetic analyses have shown that the two species share some mtDNA haplotypes when they occur in sympatry. In order to see whether some genetic exchange has occurred between the two species, we sequenced a highly variable segment of the mitochondrial control region in both species living in sympatry and in allopatry. We also analysed the nuclear diversity of 160 individuals of both species found in two mixed nursery colonies located north and south of the Alps. MtDNA analysis confirmed that European M. blythii share multiple, identical or very similar haplotypes with M. myotis. Since allopatric Asian M. blythii presents mtDNA sequences that are very divergent from those of the two species found in Europe, we postulate that the mitochondrial genome of the European M. blythii has been replaced by that of M. myotis. The analysis of nuclear diversity shows a strikingly different pattern, as both species are well differentiated within mixed nursery colonies (F(ST) = 0.18). However, a Bayesian analysis of admixture reveals that the hybrids can be frequently observed, as about 25% of sampled M. blythii show introgressed genes of M. myotis origin. In contrast, less than 4% of the M. myotis analysed were classified as non-parental genotypes, revealing an asymmetry in the pattern of hybridization between the two species. These results show that the two species can interbreed and that the hybridization is still ongoing in the areas of sympatry. The persistence of well-differentiated nuclear gene pools, in spite of an apparent replacement of mitochondrial genome in European M. blythii by that of M. myotis, is best explained by a series of introgression events having occurred repeatedly during the recent colonization of Europe by M. blythii from Asia. The sharp contrast obtained from the analysis of mitochondrial and nuclear markers further points to the need to cautiously interpret results based on a single class of genetic markers.

In taxonomic accounts, three species of Myotis have been traditionally reported to occur on the island of Taiwan: Watase’s bat (M. formosus watasei Kishida), the Formosan broad-muzzled bat (M. muricola latirostris Kishida) and the Formosan mouse-eared bat (M. adversus taiwanensis Linde). The discovery in 1997 of an unknown taxon not fitting to the description of any of these species encouraged us to re-examine more thoroughly the systematics and phylogeny of Myotis bats inhabiting Taiwan. We used a combination of morphologic and molecular methods to aid the identification of the different taxa from this island and reconstruct their phylogenetic relationships. Multivariate analyses based on 17 craniodental characters of 105 specimens caught across Taiwan and further external characters allowed us to discriminate eight taxa of Myotinae co-occurring on this island. A subset of 80 specimens were further sequenced for the cytochrome b gene (1140 bp) and subjected to phylogenetic reconstructions including representative species from adjacent China and from all main lineages of the worldwide Myotis radiation. These molecular reconstructions showed that the Myotinae from Taiwan are phylogenetically diverse and are issued from several independent clades. The genetic results were completely congruent with the phenetic groupings based on craniodental and external morphology, as each of the eight Taiwanese taxa proved to be reciprocally monophyletic. Two unnamed taxa that did not fit into any of the known species were described as species new to science. Furthermore the taxon latirostris usually associated to the Asian M. muricola, was phylogenetically and morphologically distant from any other known Myotis and was assigned here to the fossil (Miocene) genus Submyotodon. Submyotodon latirostris, M. secundus sp. n. and M. soror sp. n. are endemic species from Taiwan, whereas the other five Myotis are more widespread and also found in the mainland. An identification key is provided to ease the discrimination of these Myotinae species in Taiwan and adjacent China.

Metabarcoding of feces has revolutionized the knowledge of animal diets by providing unprecedented resolution of consumed resources. However, it is still unclear how different methodological approaches influence the ecological conclusions that can be drawn from such data. Here, we propose a critical evaluation of several data treatments on the inferred diet of the bat Plecotus auritus using guano regularly collected from various colonies throughout the entire active season. First and unlike previous claims, our data indicates that DNA extracted from large amounts of fecal material issued from guano accumulates yield broader taxonomic diversity of prey than smaller numbers of pellets would do, provided that extraction buffer volumes are adapted to such increased amounts of material. Second, trophic niche analyses based on prey occurrence data uncover strong seasonality in the bat’s diet and major differences among neighboring maternity colonies. Third, while the removal of rare prey items is not always warranted as it introduces biases affecting particularly samples with greater prey species richness. Fourth, examination of distinct taxonomic depths in diet analyses highlights different aspects of food consumption providing a better understanding of the consumer’s diet. Finally, the biologically meaningful patterns recovered with presence-absence approaches are virtually lost when attempting to quantify prey consumed using relative read abundances. Even in an ideal situation where reference barcodes are available for most potential prey species, inferring realistic patterns of prey consumption remains relatively challenging. Although best practice in metabarcoding analyses will depend on the aims of the study, several previous methodological recommendations seem unwarranted for studying such diverse diets as that of brown long-eared bats.