Characterization of 14 polymorphic microsatellite loci for the lesser horseshoe bat,Rhinolophus hipposideros (Rhinolophidae,Chiroptera)

Fourteen polymorphic microsatellites isolated in the lesser horseshoe bat, Rhinolophus hipposideros, are described. A subset of eight loci successfully coamplified in a multiplex polymerase chain reaction (PCR), and allowed to confirm that expected heterozygosity was relatively high for the eight loci (0.56–0.83). This set of eight microsatellites indeed permits to build genetic tags that distinguish all individuals in colonies consisting of up to more than 1000 individuals.     

Isolation and characterization of 13 microsatellite loci in Rhinolophus pusillus (least horseshoe bat) with cross-amplification in five related species

We used the enriched genomic library method to isolate and characterize dinucleotide microsatellite loci in the least horseshoe bat, Rhinolophus pusillus. Seventeen loci were obtained and tested on 31 individuals sampled from Guangxi Province in southern China. Thirteen of these markers were polymorphic with expected heterozygosity ranging from 0.821 to 0.909. A total of 164 alleles were detected and the number of alleles per locus ranged from 9 to 16 (mean 12.6). These polymorphic markers will be used to assess population structure in R. pusillus. In addition, successful cross-amplification in five congeneric bat species suggests most of these markers will also be useful for studying related species.     

Microsatellite loci for the greater horseshoe bat,Rhinolophus ferrumequinum (Rhinolophidae,Chiroptera) and their cross‐utility in 17 other bat species

We have isolated 14 polymorphic greater horseshoe bat, Rhinolophus ferrumequinum microsatellite loci. The number of alleles varied from two to 12 in 58 individuals. These loci will be used to assign paternity in order to characterize patterns of breeding and reproductive success in a wild R. ferrumequinum population. Loci were also tested in 17 other bat species. Twelve loci cross‐amplified in other species and three loci were polymorphic in all eight Rhinolophus species tested.     

中国大陆菊头蝠科一新纪录——单角菊头蝠(Rhinolophus monoceros Andersen,1905)

During a survey of bat species diversity from July 2008 to February 2009 in Songtao county of Guizhou Province, one specimen of Rhinolophus was collected from Guizhou. It was identified as a Formosan lesser horseshoe bat, based on the following features: small body size, forearm length of 36. 3mm, the greatest length of skull was 15.16 mm, the cells of lancet were shorter and appeared triangular and showed an acute angle, and forward bending, the lancet was shorter and approximately triangular and had a narrow top, the anterior median swellings of the skull were higher and more vertical, the rear of the sagittal crest was thin, the upper molars were shorter and the length of C~1 - M~3 was 5. 36 mm, the width of M~3 M~3 was 5.24 mm. Rhinolophus monoceros, which is new record in Guizhou and the first found on the Chinese mainland, supports Simmons' prediction that Rhinolophus monoceros may occur in the south of China.     

Pleistocene climatic cycling drives intra‐specific diversification in the intermediate horseshoe bat (Rhinolophus affinis) in Southern China

The repeated formation and loss of land‐bridges during the Pleistocene have had lasting impacts on population genetic structure. In the tropics, where island populations persisted through multiple glacial cycles, alternating periods of isolation and contact are expected to have driven population and taxonomic divergence. Here, we combine mitochondrial and nuclear sequence data with microsatellites to dissect the impact of Pleistocene climate change on intra‐specific diversification in the horseshoe bat Rhinolophus affinis. This taxon shows considerable morphological and acoustic variation: two parapatric subspecies (himalayanus and macrurus) occur on mainland China and a third (hainanus) on Hainan Island. Our phylogeographic reconstruction and coalescent analyses suggest the island subspecies formed from an ancestral population of himalayanus via two colonization events c. 800 000 years before present. R. a. hainanus then recolonized the mainland, forming macrurus and thus a secondary contact zone with himalayanus. Finally, macrurus recolonized Hainan following the LGM. We found that all three biological events corresponded to known periods of land‐bridge formation. Evidence of introgression was detected between macrurus and both its sister taxa, with geographical proximity rather than length of separation appearing to be the biggest determinant of subsequent genetic exchange. Our study highlights the important role of climate‐mediated sea level changes have had in shaping current processes and patterns of population structure and taxonomic diversification.     

广东五种菊头蝠的核型分析

本用蝙蝠的新鲜肺组织和尾椎进行组织培养,然后在光学显微镜下计数30个染色体分散良好的中期分裂相细胞,并进行摄影、剪贴和测量。分析了广东地区5种菊头蝠的核型,即小菊头蝠的核型为2n=62,FN=60;角菊头蝠的核型为2n=62,FN=60;中菊头蝠的核型为2n=62,FN=60;大耳菊头蝠的核型为2n=62,FN=60;中华(栗黄)菊头蝠的核型为2n=36,FN=60。大耳菊头蝠的核型为首次报道。角菊头蝠和中菊头蝠的核型与前人报道的结果基本相同。中华(栗黄)菊头蝠的核型(2n=36)与张维道报道的鲁氏菊头蝠相同,而与印度和斯里兰卡产R．rouxii的核型2n=56迥异。最后对东亚地区菊头蝠多样性进行了探讨。     

Development of 10 polymorphic microsatellite loci primers for Codonopsis pilosula Nannf. (Campanulaceae)

Codonopsis pilosula Nannf., as an important traditional Chinese medicinal plant species, has been used to treat fatigue, thirst and loss of appetite. In this study, we developed 10 new microsatellite loci primers from the genome of this species using the combined biotin capture method. The polymorphism of each locus was further assessed through 27 individuals from four geographically distant populations. The number of alleles per locus ranged from 6 to 11. The observed and expected heterozygosity ranged from 0.15 to 0.24 and 0.27 to 0.40, respectively. We further performed cross-priming tests of these loci in the other three congeneric species. These microsatellite markers will be useful for investigating genetic diversity within and between populations of these species.     

A new species of horseshoe bat of the genus Rhinolophus from China (Chiroptera: Rhinolophidae)

A new species of the Rhinolophus philippinensis group (Chiroptera: Rhinolophidae) is described from Guangdong, Guangxi, and Jiangxi Provinces in China. Rhinolophus huananus n. sp. is characterized by the horseshoe, as well as by external and cranial characteristics that separate it at the species level from the other members of the philippinensis group. One of the small species of the philippinensis group, R. huananus is intermediate in size between smaller R. siamensis and larger R. macrotis.     

The complete mitochondrial genome of the greater horseshoe bat subspecies, Rhinolophus ferrumequinum korai (Chiroptera: Rhinolophidae)

The total length of the mitogenome of Rhinolophus ferrumequinum korai is 16,839?bp with a total base composition of 31.8% A, 25.4% T, 28.7% C, and 14.0% G. The mitogenome consists of 13 protein-coding genes, 2 rRNA (12S and 16S RNA) genes, 22 tRNA genes, and 1 control region.     

Characterization and multiplex genotyping of 16 polymorphic microsatellite loci in the endangered bumble-bee bat, Craseonycteris thonglongyai (Chiroptera: Craseonycteridae)

The bumble-bee bat (Craseonycteris thonglongyai) is an endangered species with a limited distribution range and a suspected declining population. Of the 30 microsatellites developed and tested in C. thonglongyai, 16 were amplified in two multiplex polymerase chain reactions (PCRs). Based on genotypes of 64 individuals from two populations, expected heterozygosities, averaged 0.50 in Myanmar and 0.61 in Thailand. Only one pair of loci showed linkage disequilibrium in one population and no deviation from Hardy–Weinberg equilibrium was detected. This set of 16 microsatellites will be useful to study population structure and gene flow in C. thonglongyai. Tin Nwe, retired.     

Ontogenesis of auditory fovea representation in the inferior colliculus of the Sri Lankan rufous horseshoe bat,Rhinolophus rouxi

Summary This report describes the ontogenesis of tonotopy in the inferior colliculus (IC) of the rufous horseshoe bat (Rhinolophus rouxi). Horseshoe bats are deaf at birth, but consistent tonotopy with a low-to-high frequency gradient from dorsolateral to ventromedial develops from the 2nd up to the 5th week. The representation of the auditory fovea is established in ventro-mediocaudal parts of the IC during the 3rd postnatal week (Fig. 3). Then, a narrow frequency band 5 kHz in width, comprising 16% of the bat's auditory range, captures 50–60 vol% of the IC (Fig. 3c). However, foveal tuning is 10–12 kHz (1/3 octave) lower than in adults; foveal tuning in females (65–68 kHz) is 2–3 kHz higher than in males (62–65 kHz). Thereafter, foveal tuning increases by 1–1.5 kHz per day up to the 5th postnatal week, when the adult hearing range is established (Figs. 4, 5). The increase of sensitivity and of tuning sharpness of single units also follows a low-to-high frequency gradient (Fig. 6).Throughout this development the foveal tuning matches the second harmonic of the echolocation pulses vocalised by these young bats. The results confirm the hypothesis of developmental shifts in the frequency-place code for the foveal high frequency representation in the IC.Abbreviations BF best frequency - CF constant frequency - FM frequency modulation - IC inferior colliculus - IHC inner hair cell; - OHC outer hair cell - RR Rhinolophus rouxi     

Eight polymorphic microsatellite loci for the Chinese medicinal plant Artemisia annua L. (Asteraceae)

Artemisia annua is an important medicinal plant from which Artemisinin was extracted to cure malaria effectively. We developed eight microsatellite markers from the genome of A. annua using the FIASCO protocol. Polymorphism of each locus was assessed in 54 individuals from two Chinese populations. The average allele number of these microsatellites was 3.1 per locus, ranging from 2 to 6. The observed (H _O) and expected (H _E) heterozygosity were from 0.019 to 0.907 and from 0.055 to 0.793, respectively. These microsatellite markers would provide a useful tool for genetic studies of A. annua. H.-R. Huang and G. Zhou have contributed equally to this work.     

Development and characterization of microsatellite loci for lotus (Nelumbo nucifera)

This paper reports the development of microsatellite primers for Nelumbo nucifera Gaerten. By screening genomic libraries enriched with 10 kinds of probes, Seventeen polymorphic loci were isolated and primers were designed. Polymorphism of these 17 loci was assessed in 24 individuals. All the 17 loci are polymorphic and the number of alleles ranged from two to seven. Observed heterozygosity and expected heterozygosity ranged from 0.0000 to 0.9176 and from 0.2837 to 0.7917 respectively. These microsatellite loci should be useful for studying the genetic diversity of N. nucifera.     

Historical introgression and the persistence of ghost alleles in the intermediate horseshoe bat (Rhinolophus affinis)

Phylogenetic conflicts between genetic markers can help to disentangle complex histories of phylogeography and introgression among taxa. We previously proposed that the Chinese mainland subspecies of the intermediate horseshoe bat Rhinolophus affinis himalayanus colonized Hainan Island to form the subspecies R. a. hainanus. Subsequent recolonization of the mainland formed a third taxon, R. a macrurus, and a secondary contact zone with the ancestral himalayanus. To test for historical and recurrent genetic exchange between these mainland subspecies, we sampled populations of each from two parapatric zones and undertook analyses using one mtDNA marker, three nuclear genes and 14 microsatellites. Nuclear DNA, echolocation call frequencies and morphological data all recovered two taxa; however, a mtDNA phylogeny revealed two himalayanus clades, of which one clustered with macrurus, as well as some shared or related mtDNA haplotypes in eastern populations. Isolation‐with‐migration (IM) models suggested some mtDNA gene flow from macrurus to himalayanus. However, strong population structure in himalayanus raises the possibility that macrurus captured mtDNA from a coastal population of himalayanus that has since become rare or extinct. To reconcile these two sets of results, we suggest that the IM estimates might reflect historical mtDNA gene flow among populations of himalayanus, before mtDNA was subsequently captured by macrurus. Finally, microsatellite‐based ABC analyses supported the island origin of macrurus; however, mtDNA‐based ABC analyses suggest this taxon might have evolved on the mainland. Our findings highlight the importance of understanding population history and structure for interpreting hybridization and introgression events.     

Isolation and characterization of eight polymorphic microsatellite loci for the critically endangered Arenaria nevadensis (Caryophyllaceae)

We report on the isolation and characterization of eight microsatellite markers from enriched libraries for Arenaria nevadensis, one of the most critically endangered plant species in the Iberian Peninsula. These are the first microsatellite loci reported for Arenaria species. The number of alleles ranged from two to eight, and the expected heterozygosity from 0.067 to 0.873. These markers will be useful for characterizing the genetic diversity in A. nevadensis and understanding its population structure, and will provide important genetic data for the conservation and recovery of this species.     

Development of twelve polymorphic microsatellite loci in polyploid endangered Omphalogramma vincaeflora Franch. (Primulaceae)

A total of twelve polymorphic microsatellite loci were developed from polyploid endangered species, Omphalogramma vincaeflora (Primulaceae). These loci were screened for variability among 45 individuals from three populations in China. The primers amplified loci with allele number ranging from 3 to 9, with an average of 4.25 per locus. Polymorphism information content ranged from 0.23 to 0.86. Nei’s genetic diversity ranged from 0.34 to 0.86. These primers provide an opportunity to use polymorphic DNA markers to study the population genetic structure and its breeding system in this species.     

Development of twenty-five polymorphic microsatellite markers for the endangered red-cockaded woodpecker (Picoides borealis)

Twenty-five polymorphic microsatellite markers were developed for the endangered red-cockaded woodpecker (Picoides borealis). The number of alleles ranged from two to five and observed heterozygosities ranged from 0.036 to 0.750. These loci should be useful tools for conducting research towards the management and conservation of this species.     

Development and characterization of novel microsatellite markers from the Chinese rufous horseshoe bat (Rhinolophus sinicus) with cross-species amplification in closely related taxa

We isolated nine polymorphic microsatellite markers from the Chinese rufous horseshoe bat (Rhinolophus sinicus) using an enriched library method. We assessed genetic polymorphism at these loci in 42 individuals from a single population. We recorded high genetic diversity with four to 17 alleles per locus, and estimated expected and observed heterozygosity values ranging from 0.492 to 0.910 and from 0.462 to 0.881, respectively. No locus departed from Hardy-Weinberg equilibrium following Bonferroni correction, and no linkage disequilibrium was detected. Most loci successfully cross-amplified congeneric species. These loci will be used to characterize phylogeographical history of Rhinolophus sinicus in China.