Cross-species amplification and characterization of microsatellite DNA loci from Gallus gallus in Bambusicola thoracica

The Chinese Bamboo Partridge(Bambusicola thoracica) is a gamebird endemic to China.Ten polymorphic microsatellite(simple sequence repeat) markers were obtained through cross-species amplification for this partridge from its relative species Gallus gallus.The number of alleles per locus varied from 4 to 13.The observed heterozygosity ranged from 0.1220 to 1.0000 and the expected heterozygosity from 0.1183 to 0.8898.Four microsatellite loci showed significant differences from the Hardy-Weinberg equilibrium.These polymorphic loci provide a valuable tool for the investigation of the phylogeography and conservation genetics of this partridge.

Isolation of microsatellite loci in the pollinating fig wasp of Ficus hispida, Ceratosolen solmsi

Microsatellite loci were isolated for Ceratosolen solmsi, pollinator of the dioecious Ficus hispida. We developed nine polymorphic microsatellite loci based on the method of polymerase chain reaction isolation of microsatellite arrays （PIMA）. Enrichment of genomic libraries was performed by random amplified polymorphic DNA （RAPD）. A subset of 38 positive clones was sequenced; 15 clones showed microsatellite loci. We tested 15 designed primer pairs and nine of them produced polymorphic amplification in 48 individual wasps collected from different fruits of the dioecious host fig Ficus hispida in China. Among the 48 individuals, 49 alleles were obtained at the nine loci. The observed heterozygosity ranged between 0.357 and 0.634.

Isolation and Characterization of 17 Microsatellite DNA Loci for Odorrana margaretae(Anura: Ranidae)

We isolated and characterized 17 microsatellite DNA loci for the odorous frog Odorrana margaretae from its transcriptome sequence data. These loci were screened with 24 individuals from Mt. Emei. All loci were polymorphic, with the number of alleles ranging from 2 to 8. The observed and expected heterozygosity, polymorphism information content, ranged from 0.04 to 1, 0.04 to 0.81, and 0.040 to 0.763, respectively. All loci were in linkage equilibrium and six loci were significantly deviated from Hardy-Weinberg equilibrium after sequential Bonferroni corrections. Cross- species amplification test was conducted for ten odorous frog species, and 12 loci were amplifiable in most species. With the high cross-species amplification rates, thesc markers will provide useful molecular tools for conservation genetic and phylogeographic studies on the genus Odorrana and Bamburana.

Development of novel microsatellite markers for Holothurian scabra (Holothuriidae), Apostichopusjaponicas (Stichopodidae) and cross-species testing in other sea cucumbers

Thirty-five new microsatellite loci from the sea cucumbers Holothurian scabra （Jaeger, 1833） and Apostichopus japonicas （Selenka, 1867） were screened and characterized using the method of magnetic bead enrichment. Of the twenty-four polymorphic loci tested, eighteen were consistent with Hardy-Weinberg equilibrium after a modified false discovery rate （B-Y FDR） correction, whereas six showed statistically significant deviations （CHS2 and CHSll： P〈0.014 790; FCS1, FCS6, FCS8 and FCS14：P〈0.015 377）. Furthermore, four species ofplesiomorphous and related sea cucumbers （Holothurian scabra, Holothuria leucospilota, Stichopus horrens and Apostichopus japonicas） were tested for mutual cross-amplification using a total of ninety microsatellite loci. Although transferability and universality of all loci were generally low, the results of the cross-species study showed that the markers can be applied to identify individuals to species according to the presence or absence of specific microsatellite alleles. The microsatellite markers reported here will contribute to the study of genetic diversity, assisted breeding, and population conservation in sea cucumbers, as well as allow for the identification of individuals to closely related species,

Development and Characterization of Fifteen Polymorphic M icosatellites for Extremely Endangered Salamander, Echinotriton chinhaiensis

Echinotriton chinhaiensis is a critically endangered salamander and its distribution is restricted to Ningbo, Zhejiang Province of China. In this study, we developed and characterized fifteen polymorphic microsatellite loci for E. ehinhaiensis from dinucleotide- and tetranucleotide- enriched library. The number of alleles ranges from 4 to 12 with an average of 7.27 alleles per locus. The observed and expected heterozygosities values were from 0.250 to 0.844 and 0.511 to 0.872 with an average value of 0.596 and 0.722; respectively. The polymorphic microsatellite loci described in this paper are useful in the further study on genetic diversity and gene flow, which would be helpful to formulate effective conservation strategies for the E. chinhaiensis.

Impact of anthropogenic pressure on the formation of population structure and genetic diversity of raccoon dog Nyctereutes procyonoides

The raccoon dog Nyctereutes procyonoides experienced an active introduction and acclimatization in the European part of Russia followed by its migration to and colonization in the neighboring countries. Eventually, it has spread rapidly into many European countries. N. procyonoides probably invaded Lithuania from the neighboring countries of Belarus and Latvia where the species was introduced. However, the data on genetic diversity and population structure of the raccoon dogs in the recently invaded territories are still scarce. The objectives of this study were to investigate genetic diversity of N. procyonoides in Lithuania after acclimatization, and to assess the impact of anthropogenic pressure on the formation of population structure. A total of 147 N. procyonoides individuals collected from different regions of Lithuania were genotyped using 17 microsatellite markers. The microsatellite analysis of raccoon dogs indicated high levels of genetic diversity within the population. The Bayesian clustering analysis in STRUCTURE identified 4 genetic clusters among sampled raccoon dogs that could not reveal a clear separation between subpopulations. The widespread distribution of raccoon dogs in Lithuania, high level of genetic variation observed within subpopulations, and low level of variation portioned among subpopulations suggest migration and gene flow among locations. The significant correlation between genetic and geographic distances indicated isolation that reflected the distance between locations. The fencing of highways and very intensive traffic could be barriers to gene flow between the western and eastern sampling areas of raccoon dogs.

Augmenting the conservation value of rehabilitated wildlife by integrating genetics and population modeling in the post-rehabilitation decision process

Insular populations are particularly vulnerable to the effects of stochastic events, epidemics, and loss of genetic diversity due to inbreeding and genetic drift. The development of successful man- agement options will require accurate baseline data, establishment of clear objectives, and finally monitoring and implementation of corrective measures, if and when required. This study assessed management options for the genetic rehabilitation of highly inbred woylies obtained from wildlife rehabilitation centers. The study generated genetic data for the woylie Bettongia penicillata from a conservation reserve and calculated measures of genetic diversity and individual relatedness. These data were fed into a population viability analysis （PVA） to test genetic outcomes in relation to different management actions. We demonstrated that a careful selection of the founder cohort produced a population with an expected heterozygosity of -70% for a window of approximately 10 years. A proposal to increase the size of the reserve available to the colony was shown to almost double the time at which the colony would retain heterozygosity levels of≥70%. Additionally, developing a regular program of supplementation of unrelated woylies would result in a further improvement in their genetic value. This study demonstrated how the application of molecular techniques in combination with PVA can be beneficial for the management of rehabilitated wildlife otherwise considered of little conservation value. This approach can be applied to the management of breeding programs, but also to small, closed populations such as those found on islands, fenced enclosures, insurance populations, and in zoological collections.

Patterns of geographic variation between mitochondrial and nuclear markers in Heaviside’s(Benguela)dolphins(Cephalorhynchus heavisidii)

The Heaviside’s(or Benguela)dolphin(Cephalorhynchus heavisidii)is endemic to the west coast of southern Africa.The present study investigated the population genetic structure across a large portion of the species dis­tribution using mitochondrial control region and nuclear(microsatellite)markers.A total of 395 biopsy skin samples were analyzed;they were collected from free-ranging Heaviside’s dolphins in 7 locations along 1650 km of coast between Table Bay,South Africa and Walvis Bay,Namibia.Both genetic markers rejected the hy­pothesis of 1 homogenous population but revealed contrasting results in the genetic structuring of putative pop­ulations.Mitochondrial DNA suggested either 2 populations or a fine-scale division with 6(sub)populations,while microsatellite markers were indicative of 2 widespread populations with measurable gene flow between them.Neutrality tests and mismatch distribution of the mitochondrial sequences indicated a departure from mu­tation-drift equilibrium due to a population expansion at the 2 extremes of the geographic range,but not to­wards the middle of the distribution.These results highlight the importance of evaluating multiple genetic mark­ers to gain reliable insights into population processes and structure.