New insights into the taxonomy and phylogeny of social voles inferred from mitochondrial cytochrome b sequences

We sequenced the entire cytochrome b gene in Microtus paradoxus from Turkmenistan and Microtus socialis from Crimea and Kalmykia. Phylogenetic relationships among social voles were reconstructed by the inclusion into analyses of a further 23 published haplotypes belonging to six species. The two probabilistic methods which were used in phylogenetic analyses, the Bayesian inference and Maximum Likelihood, yielded very similar results. Both trees showed two highly divergent lineages which were further subdivided into seven species. The socialis lineage encompassed four species (M. socialis, M. irani, M. anatolicus, and M. paradoxus), and the remaining three species clustered into the guentheri lineage (M. guentheri, M. hartingi, M. dogramacii). The ranges for nucleotide divergences between seven species of social voles (4.95–9.28% and 4.18–8.81% for mean and net divergences, respectively) mainly exceeded 4.3%, which is frequently regarded as the conservative cut-off between sibling species in the specious genus Microtus.     

Social but lonely: Species delimitation of social voles and the evolutionary history of the only Microtus species living in Africa

The social vole of Cyrenaica, Libya, is the only extant representative of the Cricetidae family found in Africa. Its taxonomic status has been under debate, partly due to the problematic systematics of the entire group of social voles and partly due to the lack of morphological and molecular data from Cyrenaican specimens. In this study, we applied ancient DNA protocols to produce three cytochrome b (cytb) sequences of Cyrenaican voles and built a phylogenetic reconstruction (195 sequences in total) incorporating all available cytb sequences of the remaining social voles, other representatives of the genus Microtus and closely related cricetids. We used this phylogeny to test the performance of the model-based, single-locus, species delimitation approach implemented in mPTP and delimited nine species of social voles. Among them are the Cyrenaican vole, Microtus mustersi and its sister species M. guentheri, distributed along the Mediterranean coasts of southwestern Asia. Biogeographical reconstruction of ancestral area and molecular clock estimations of the time since the divergence of the two sister lineages suggest that their common ancestor dispersed into Africa through a coastal route and was isolated in Cyrenaica as a result of population fragmentation associated with Middle Pleistocenic pluvial/interpluvial cycles. Geographic isolation triggered the speciation process, but species distribution modeling gave evidence of subsequent niche divergence; M. guentheri has adapted to the xeric conditions of its distributional area, while M. mustersi benefited from the milder Cyrenaican climate. The Cyrenaican vole is a relict species more than 200,000 years old, has a small and isolated distribution and probably merits conservation.     

Chromosomal variation in social voles: a Robertsonian fusion in Günther’s vole

The study reports on chromosomes in several populations of social voles from south-eastern Europe and the Middle East. The standard karyotypes of individuals of Microtus hartingi and Microtus guentheri originating from both south-eastern Europe and Asia Minor comprised 54 mostly acrocentric chromosomes. However, variation between populations was found in the amount and distribution of C-heterochromatin in certain autosomes and the sex chromosomes. Furthermore, a specific pattern of argyrophilic nucleolar organizer region distribution was recorded in different geographic populations. In a population from Asia Minor, a heterozygous centric fusion of two autosomes was found. The G-banded karyotypes of M. guentheri and Microtus socialis were compared, and tandem fusions of autosomes were suggested as possible mechanism of the divergence. The karyotypes of the nine currently recognized species of social voles are reviewed, and implications of chromosomal data for systematics are evaluated.     

Heterochromatin study and geographical distribution of Microtus species (Rodentia,Arvicolinae) from Greece

In a sample of 108 underground voles from 23 Greek localities, the species Microtus felteni, M. guentheri, M. rossiaemeridionalis and M. subterraneus were identified, based on external body morphology and karyotype. Moreover, the implemented C-banding staining technique revealed the heterochromatin distribution in the chromosomes of the above species. All M. guentheri (2n=54, FN=54) and M. rossiaemeridionalis (2n=54, FN=56) specimens displayed the typical karyotypes of these species, respectively. The M. subterraneus specimens belonged to the chromosomal race with 2n=52, FN=60 of this species, apart from a single individual that demonstrated a medium-sized, subtelocentric autosome in heterozygous condition (2n=52, FN=61). Furthermore, M. felteni individuals, trapped again after many years, were karyologically studied (2n=54, FN=56) and the C-banding pattern for this species is hereby presented for the first time. Finally, the study of meiotic preparations in M. guentheri and M. rossiaemeridionalis males verified the asynaptic behaviour of their sex chromosomes. The karyotype of the four studied Microtus species does not seem to have diversified much from the putative ancestral arvicoline karyotype (2n=56, FN=56). On the other hand, the heterochromatin accumulation in the sex chromosomes, particularly prominent in M. felteni and M. rossiaemeridionalis, represents this common tendency in several Microtus species.     

Giardia microti in pet Microtus guentheri: Evidence of a parasite never detected in Italy

The genus Giardia includes several species distinguished by morphological, biological and molecular features. Currently, eight species within the genus are retained as valid. In Italy no identification of Giardia species other than Giardia duodenalis has been so far reported. Fecal samples were collected from two Günther's Voles (Microtus guentheri) positive to Giardia cysts by microscopic investigation and immunofluorescence. The voles were born in Milan (Northern Italy) from two gravid females imported from the Netherlands and kept for sale in a pet shop in Varese (Northern Italy). Positive feces were subjected to a nested PCR to amplify a 18S rRNA fragment for molecular characterization. A phylogenetic analysis was conducted to compare the obtained sequence with those of all other Giardia species available in GenBank for the 18S locus, using the Maximum Likelihood (ML) method by R software. Sequence analyses unambiguously identified the isolates as belonging to G. microti, showing 99% of identity with those of its isolates available in GenBank. A well-defined cluster, supported by significant bootstrap values and corresponding to the G. microti cluster, including sequences obtained from M. guentheri, was evidenced in the ML tree, confirming species assignment. The present finding represents the first report of G. microti from pet animals in Italy.     

Morphological variation of the five vole species of the genus Microtus (Mammalia,Rodentia, Arvicolinae) occurring in Greece

Morphometric data for the five vole species of the genus Microtus living in Greece are old, sparse, poor and insufficiently analysed. This work aims to give the first comprehensive morphometric analysis of body and skull inter‐ and intraspecific variation for M. (M.) guentheri, M. (M.) rossiaemeridionalis, M. (Terricola) subterraneus, M. (T.) felteni and M. (T.) thomasi, applying multivariate statistics to 28 linear morphometric variables. It was based on ample material (202 adult individuals) using samples from localities that adequately cover the entire distributional range of each species in Greece. The five species and the two subgenera (Microtus and Terricola) were morphometrically clearly distinguished and discriminating variables were revealed. However, morphometrics did not provide robust criteria to infer phylogenetic relations among species. Furthermore, three species, M. (M.) guentheri, M. (M.) rossiaemeridionalis and M. (T.) thomasi, exhibited considerable intraspecific size or shape variation, which was mostly random and not associated with geographical proximity. Comparisons with data in the literature, mainly concerning populations of these species from adjacent areas, indicate that the Greek M. (M.) guentheri, M. (M.) rossiaemeridionalis and M. (T.) thomasi tend to be smaller than their conspecifics, while M. (T.) subterraneus and M. (T.) felteni are about equal in size to their Balkan relatives.     

A complex scenario of glacial survival in Mediterranean and continental refugia of a temperate continental vole species (Microtus arvalis) in Europe

The role of glacial refugia in shaping contemporary species distribution is a long-standing question in phylogeography and evolutionary ecology. Recent studies are questioning previous paradigms on glacial refugia and postglacial recolonization pathways in Europe, and more flexible phylogeographic scenarios have been proposed. We used the widespread common vole Microtus arvalis as a model to investigate the origin, locations of glacial refugia, and dispersal pathways, in the group of “Continental” species in Europe. We used a Bayesian spatiotemporal diffusion analysis (relaxed random walk model) of cytochrome b sequences across the species range, including newly collected individuals from 10 Iberian localities and published sequences from 68 localities across 22 European countries. Our data suggest that the species originated in Central Europe, and we revealed the location of multiple refugia (in both southern peninsulas and continental regions) for this continental model species. Our results confirm the monophyly of Iberian voles and the pre-LGM divergence between Iberian and European voles. We found evidence of restricted postglacial dispersal from refugia in Mediterranean peninsulas. We inferred a complex evolutionary and demographic history of M. arvalis in Europe over the last 50,000 years that does not adequately fit previous glacial refugial scenarios. The phylogeography of M. arvalis provides a paradigm of ice-age survival of a temperate continental species in western and eastern Mediterranean peninsulas (sources of endemism) and multiple continental regions (sources of postglacial spread). Our findings also provide support for a major role of large European river systems in shaping geographic boundaries of M. arvalis in Europe.     

Multiple captures provide evidence of small mammal social behavior

Social behavior of small mammals living under natural conditions often is inferred from live-trapping data, particularly from incidents in which two or more individuals are captured together in a trap. We examined whether multiple-capture data from a long-term study of prairie voles (Microtus ochrogaster) and meadow voles (Microtus pennsylvanicus) were consistent with well-known species differences in social behavior (whereas prairie voles are highly social and display monogamy, meadow voles are less social and promiscuous). When possible, we also examined multiple captures of two nontarget species, northern short-tailed shrews (Blarina brevicauda) and western harvest mice (Reithrodontomys megalotis). Percent of total captures that were multiple captures and percent of total adult captures that were male–female captures were highest for prairie voles and lowest for meadow voles; values for harvest mice and shrews were in between those of the vole species, but more similar to values for meadow voles. Repeat captures of the same male–female pair occurred most commonly in prairie voles, and multiple captures of this species typically involved individuals from the same social group. Multiple captures of adults and juveniles were more common in prairie voles than meadow voles, except for captures of at least one adult male and at least one juvenile, which did not differ between the two vole species. Multiple capture data for prairie voles and meadow voles were largely consistent with established species differences in social behavior, suggesting that such data can provide an accurate indication of social and mating systems of small mammals.     

Integrative taxonomy of the Italian pine voles,Microtus savii group (Cricetidae,Arvicolinae)

The Italian endemic voles belonging to the Microtus savii group (de Sélys‐Longchamps, 1838) include five controversial taxa, namely M. savii savii, M. s. niethammericus Contoli 2003 ; M. s. tolfetanus Contoli 2003 , M. s. nebrodensis (Minà‐Palumbo, 1868) and M. brachycercus (Lehamann, 1961). A previous mtDNA phylogenetic analysis revealed three main mtDNA lineages that do not correspond to the recognized taxa. Here, we perform a comprehensive univariate and multivariate analysis of craniodental measurements in 151 specimens from 54 geographic samples belonging to the three main phylogroups. Qualitative data on the skull of 126 specimens were also obtained from 54 localities. Moreover, we sequenced the nuclear DNA β‐fibrinogen intron 7 (β‐fibint7) of 23 specimens from 12 localities and Cyt b sequences of 11 new specimens from seven localities. The pattern of morphological and cytogenomic divergence does not corroborate the current taxonomic framework and indicates that M. nebrodensis is a Sicilian endemic species.     

Sound communication in social voles (subgenus Sumeriomys)

ABSTRACT

The acoustic communication of three species of social voles from the subgenus Sumeriomys – Microtus socialis (two subspecies: M. s. socialis and M. s. goriensis), M. paradoxus and M. hartingi – are described. Vole sound communication includes two main signals: squeaks and singing. The sounds made by M. hartingi have significantly higher frequency parameters than those of other species. Voles of all species squeak in situations of distress, and the males sing during courtship of the females. However, singing in social voles is not a necessary pattern for sexual behaviour: less than half of M. s. socialis and M. paradoxus males sang, M. hartingi sang even more rarely and M. s. goriensis did not demonstrate this behaviour at all. Despite the great similarity of the squeaks, its parameters differ significantly between species and differ from those of the common voles. This introduces one more argument that M. paradoxus and M. socialis are independent species, as are the subgenera Sumeriomys and Microtus.     

A Newly Emerged Cutaneous Leishmaniasis Focus in Northern Israel and Two New Reservoir Hosts of Leishmania major

In 2006/7, 18 cases of cutaneous leishmaniasis (CL) were reported for the first time from Sde Eliyahu (pop. 650), a village in the Beit She''an valley of Israel. Between 2007–2011, a further 88 CL cases were diagnosed bringing the total to 106 (16.3% of the population of Sde Eliyahu). The majority of cases resided in the south-western part of the village along the perimeter fence. The causative parasite was identified as Leishmania major Yakimoff & Schokhor, 1914 (Kinetoplastida: Trypanosomatidae). Phlebotomus papatasi (Scopoli), 1786 (Diptera: Psychodidae) was found to be the most abundant phlebotomine species comprising 97% of the sand flies trapped inside the village, and an average of 7.9% of the females were positive for Leishmania ITS1 DNA. Parasite isolates from CL cases and a sand fly were characterized using several methods and shown to be L. major. During a comprehensive survey of rodents 164 Levant voles Microtus guentheri Danford & Alston, 1880 (Rodentia: Cricetidae) were captured in alfalfa fields bordering the village. Of these 27 (16.5%) tested positive for Leishmania ITS1 DNA and shown to be L. major by reverse line blotting. A very high percentage (58.3% - 21/36) of Tristram''s jirds Meriones tristrami Thomas, 1892 (Rodentia: Muridae), found further away from the village also tested positive for ITS1 by PCR. Isolates of L. major were successfully cultured from the ear of a wild jird found positive by ITS1 PCR. Although none of the wild PCR-positive voles exhibited external pathology, laboratory-reared voles that were infected by intradermal L. major inoculation, developed patent lesions and sand flies became infected by feeding on the ears of these laboratory-infected voles. This is the first report implicating M. guentheri and M. tristrami as reservoirs of Leishmania. The widespread co-distribution of M. guentheri and P. papatasi, suggests a significant threat from the spread of CL caused by L. major in the Middle East, central Asia and southern Europe.     

Geomorphometric differences among four species of Microtus in Turkey (Mammalia: Rodentia)

In order to determine the phenotypic associations of four morphologically similar species of Microtus occurring in Turkey, we applied landmark-based shape analysis. The skulls of Microtus anatolicus, M. dogramacii, M. guentheri and M. levis (= M. rossiaemeridionalis) were found to differ significantly in terms of both size and shape. M. guentheri had the biggest skull, while M. levis had the smallest. Sexual dimorphism was found in the shape of the skull in M. dogramacii and M. levis. The tympanic bulla area is enlarged in M. anatolicus compared to the other species. Mahalanobis distances (the distance between a point and the group mean, taking into account the within-group covariance-variance matrix) confirm the distinction of the arvalis group (M. levis) and socialis group (Microtus anatolicus, M. dogramacii, M. guentheri).     

Genetic Divergence Detected by ISSR Markers and Characterization of Microsatellite Regions in Mytilus Mussels

The wide distribution of microsatellites in mussels of the Mytilus edulis complex (Mytilidae) enables the analysis of inter-simple-sequence repeat (ISSR) markers. The aim of this investigation was to assess genetic differentiation in six sampling localities distributed along the European Atlantic coast to expose the potential of these markers in genetic studies requiring the detection of low polymorphism and as a source of sequences for developing microsatellite markers. We detected low genetic structuring within each member of the Mytilus edulis complex. Nei and Li distances and AMOVA clustered the individuals studied into two groups. On the basis of these results two sampling localities coming from the M. edulis × M. galloprovincialis mosaic hybrid zone in Western Europe were assigned to one species. On the other hand, mussels of a sampling locality in the Baltic Sea were not significantly different from a pure M. edulis locality supporting an extensive introgression of M. edulis in these individuals. Finally, 148 microsatellites were found in the sequences of 51 ISSR markers, and two polymorphic microsatellite markers were developed.     

Genetic structure, habitat fragmentation and bottlenecks in Danish bank voles (Clethrionomys glareolus)

Tissue-samples from 161 bank voles (Clethrionomys glareolus) collected in three forests (five sampling localities) situated in eastern Jutland (Denmark) were analysed by nine microsatellite loci. The genetic diversity found within the populations was high (H_e=0.753–0.806).Bank voles have specific habitat requirements favouring woodlots, hedgerows and deciduous forests as their prime living area. Hence, a natural or human-induced fragmentation of the forest may cause a sub-structuring of the populations and thereby a restriction of dispersal among populations. The sub-structuring indicated by the observed significant genetic differentiation among the five geographically distinct localities (F_st =0.033, P<0.05) could either result from habitat fragmentation or a combination of home range behaviour and different tree composition in the forests. A road situated between two adjacent forests was not found to exert any barrier effect to the gene flow of bank voles. In one out of five localities investigated, genetic evidence for a recent bottleneck-like situation was found. Bank voles are known to exhibit sometimes huge density fluctuations not only from year to year but also from season to season. The bottleneck-like situation found could therefore be due to the low number of individuals during the low-density phase.     

From widespread to microendemic: molecular and acoustic analyses show that Ischnocnema guentheri (Amphibia: Brachycephalidae) is endemic to Rio de Janeiro, Brazil

Many species of tropical amphibians are restricted to very small ranges, and this microendemism coupled with ongoing habitat loss and susceptibility to emerging pathogens imperils the long-term persistence of these species. Incomplete taxonomic and distributional knowledge may obscure conservation assessment, particularly in putatively widespread species that are typically considered to be of Least Concern in Red List assessments, but that in fact may constitute complexes of partly microendemic species. Such is the case in the Steindachner’s Robber Frog, Ischnocnema guentheri which, together with the recently recognized Ischnocnema henselii, is thought to occupy most of the Brazilian Atlantic Forest. To test whether these taxa may constitute a species complex of range-restricted and thus potentially threatened species, we analyzed 160 samples of I. guentheri and/or I. henselii for two molecular markers, 16S rRNA (16S) and recombination activation gene 1 (RAG1). To verify the monophyly of the complex, closely related species were also included in the 16S analysis. Congruent evidence from the molecular data and from analyses of advertisement calls support the existence of six distinct species within the complex: I. guentheri and I. henselii as well as four candidate new species. The lineages are distributed as a mosaic in the Atlantic Forest and are sympatric at some localities without indication of admixture. Their phylogeographical pattern partially agrees with paleo-models for the Atlantic Forest, but also suggests the existence of micro-refugia in less stable areas. I. guentheri, previously considered to be widespread, was found only in its type locality, a reserve within the urban area of Rio de Janeiro city. Although none of the species studied appears highly threatened with extinction, we recommend their IUCN threat status to be re-evaluated carefully for the next comprehensive update of the Red List of Brazil’s amphibians.     

Predation on two vole species by a shared predator: antipredatory response and prey preference

In prey communities with shared predators, variation in prey vulnerability is a key factor in shaping community dynamics. Conversely, the hunting efficiency of a predator depends on the prey community structure, preferences of the predator and antipredatory behavioural traits of the prey. We studied experimentally, under seminatural field conditions, the preferences of a predator and the antipredatory responses of prey in a system consisting of two Myodes species of voles, the grey-sided vole (M. rufocanus Sund.) and the bank vole (M. glareolus Schreb.), and their specialist predator, the least weasel (Mustela nivalis nivalis L.). To quantify the preference of the weasels, we developed a new modelling framework that can be used for unbalanced data. The two vole species were hypothesised to have different habitat-dependent vulnerabilities. We created two habitats, open and forest, to provide different escape possibilities for the voles. We found a weak general preference of the weasels for the grey-sided voles over the bank voles, and a somewhat stronger preference specifically in open habitats. The weasels clearly preferred male grey-sided voles over females, whereas in bank voles, there was no difference. The activity of voles changed over time, so that voles increased their movements immediately after weasel introduction, but later adjusted their movements to times of lowered predation risk. Females that were more active had an elevated mortality risk, whereas in the case of males, the result was the opposite. We conclude that, in vulnerability to predation, the species- or habitat-specific characteristics of these prey species are playing a minor role compared to sex-specific characteristics.     

Habitat use by singing voles and tundra voles in the southern Yukon

Summary We investigated how far competitive interactions influence the use of habitats and relative abundance of two species of Microtus in the southwestern Yukon. We worked in the ecotone between alpine tundra and subalpine shrub tundra where populations of singing voles (Microtus miurus) and tundra voles (M. oeconomus) overlap little.We removed tundra voles from shrub tundra on one live-trapping area to look at the effect on the contiguous population of singing voles in alpine tundra. The removal of tundra voles did not affect the distribution or relative abundance of singing voles. The spatial distribution of these species and their movements within habitats suggest that they have a strong habitat preference.Populations of small mammals in the area are extremely dynamic and the relative importance of competitive interactions may change as density varies. At present we have no evidence that competition affects habitat use in M. miurus.     

Influence of edge effects on common vole population abundance in an agricultural landscape of eastern France

Vole demographics are often modified close to wooded environments. Population monitoring in such settings reveals substantial edge effects, although these have seldom been quantified. This study compares the abundance of common voleMicrotus arvalis (Pallas, 1778) populations at various distances from three types of wooded environment (hedgerow, copse and forest) and in two types of habitat (temporary and permanent grassland). In temporary grassland, substantial edge effects were detected along the boundaries of forests and hedgerows. At 100 m from the forest edge, voles were twice more abundant than at 50 m, four times more than at 25 m and eight times more than at 5 m. In permanent grassland, the edge effect was such that voles were practically absent up to 100 m from the forest edge. For both types of grassland, the edge effect was more pronounced for forest habitat than for hedgerows, that is, for elements with a large surface area of wooded environment. Our results suggest that landscape context may greatly affect the local abundance of voles. Given the considerable damage done byM. arvalis, the presence of wooded patches in grassland areas leads to lower vole densities in adjacent areas and this might be an alternative method to controlM. arvalis as a pest species.     

Microhabitat use by wild-ranging Cabrera voles <Emphasis Type="Italic">Microtus cabrerae</Emphasis> as revealed by live trapping

The conservation problems affecting the endangered rodent Microtus cabrerae are mainly related to habitat loss and fragmentation. In the present study, we examined the presence/absence of the species in a group of potentially suitable habitat patches, and then evaluated the microhabitat attributes of locations used by wild-ranging individuals within occupied patches. Our results revealed that shallow flooding and abundant grass cover appear to be important factors in determining the presence among localities, but high rush cover strongly determined the presence and habitat use by voles both among and within localities. The preservation of these microhabitat characteristics may be required to effectively ensure the occurrence and stability of M. cabrerae colonies.     

Giant sex chromosomes retained within the Portuguese lineage of the field vole (<Emphasis Type="Italic">Microtus agrestis</Emphasis>)

The field vole (Microtus agrestis) is characterised by extremely large blocks of heterochromatin on both the X and Y chromosome. Some other Microtus also have blocks of heterochromatin on their sex chromosomes but not as extensive and always of independent origin from the heterochromatic expansion found in M. agrestis. Coupled with evidence of geographic variation in large heterochromatic blocks within other species (e.g. in the western hedgehog Erinaceus europaeus), it might be expected that field voles would show substantial variation in size and disposition of the sex chromosome heterochromatin. In fact, only minor variation has been described up to now. Those studies conducted previously were largely on field voles from central and northern Europe. Here, we describe the karyotype of field voles from Portugal, of interest because recent molecular studies have shown field voles from western Iberia to be a separate evolutionary unit that might be considered a cryptic species, distinct from populations further to the east. The two Portuguese field voles (one female, one male) that we examined also had essentially the same karyotype as seen in other field voles, including the giant sex chromosomes, but with small differences in the structure of the Y chromosome from that described previously. The finding that field voles throughout Europe show relatively little variation in their giant sex chromosomes is consistent with molecular data which suggest a recent origin for this complex of species/near-species.