Chloroplast microsatellites in Prunus,Rosaceae

Ten chloroplast microsatellite markers were developed from Japanese plum (Prunus salicina) based on nucleotide sequences of c. 4300 bp from six chloroplast regions. Out of 10 microsatellites, seven markers contained mononucleotide repeats. Almost all microsatellites displayed discrete amplified fragments for 17 species in Prunus. The microsatellites generated 46 different fragment types and differentiated all used species. Polymorphism was also observed within species for all microsatellites.     

Differentiation of golden-ringed dragonfly Anotogaster sieboldii (Selys, 1854) (Cordulegastridae: Odonata) in the insular East Asia revealed by the mitochondrial gene genealogy with taxonomic implications

Molecular phylogeographical analyses of Anotogaster sieboldii (Selys, 1854) were conducted to reveal the differentiation process of insular populations. The gene genealogy based on 845 bp of the mitochondrial genes (cytochrome oxidase subunit I and subunit II) indicated that A. sieboldii includes two deeply separated lineages. These two major lineages seem to have differentiated in Miocene before the formation of the insular East Asia. One lineage includes three inner clades that correspond to the populations of northern area (the Japanese main islands, Korean Peninsula, Yakushima), Amamioshima and Okinawajima. Populations of Central Ryukyu, including Amamioshima and Okinawajima, might have been divided from the northern populations in early Pleistocene. The other major lineage includes populations of the Yaeyama Group, Taiwan and East China. The former two populations were reconstructed as a reciprocal monophyletic group. Populations of Taiwan and Yaeyama Groups would have been separated from the continental ones in Pleistocene. These two highly divergent lineages should be recognized as distinct species. Furthermore, the mitochondrial lineages revealed six genetically distinct and geographically isolated assemblages: (1) northern populations, (2) Amamioshima, (3) Okinawajima, (4) Yaeyama Group, (5) Taiwan and (6) East China.     

Polymorphic microsatellites in Buff-throated partridge developed by cross-species amplification

Microsatellite loci for the buff-throated partridge (Tetraophasis szechenyii), an endemic pheasant species of China, are here described for the first time. Twenty-five microsatellite markers from chicken and Japanese quail were tested on buff-throated partridge DNA by means of cross-amplification. Twenty (80%) primers yielded specific products and polymorphisms were tested in a wild population of buff-throated partridge. Twelve (48%) proved to be polymorphic with an average of two alleles per locus. Current results of buff-throated partridge microsatellites loci could be employed in population genetic studies and on other endangered pheasant species.     

Isolation and characterization of microsatellite markers for Ligularia hodgsonii Hook. (Asteraceae)

Ligularia hodgsonii is one of widely distributed Ligularia species in south China, central China, Far East area of Russia and Japan. In this study, we described the development of 14 microsatellite markers from the genome of L. hodgsonii using the FIASCO protocol. Polymorphism of each locus was assessed in 30 adult individuals of the Ligularia. The average allele number of the microsatellites was 3.0 per locus, ranging from 2 to 5. The observed (H _O) and expected (H _E) heterozygosities varied from 0.2000 to 0.7333 and from 0.3910 to 0.7598. The marker transferability of the 14 primer pairs was tested on other two congeneric species that also occur in China.     

Isolation and cross‐species amplification of microsatellite loci in the freshwater minnow Zacco pachycephalus (Teleostei: Cyprinidae) for diversity and conservation genetic analysis

Asian minnows of the genus Zacco are dominant fish in various freshwater ecosystems. Two species, Zacco pachycephalus and Z. platypus, occur in Taiwan and are favourite targets for local sport anglers. The introduction of Zacco fish into nonindigenous habitats in Taiwan has become a conservation issue. We developed eight polymorphic microsatellites for Z. pachycephalus (average H_E = 0.779) and these microsatellites were applicable to Z. platypus, which showed a comparable polymorphic level (average H_E = 0.784). These loci can be used as genetic markers for identifying conservation units and studying population differentiation for both Zacco species.     

Isolation and development of microsatellite markers for the Japanese dormouse, Glirulus japonicus

Eight microsatellite markers were developed for the Japanese dormouse (Glirulus japonicus), a natural monument and near‐threatened species in Japan. The markers amplify in individuals from all of the mitochondrial lineages detected in a previous study. Numerous polymorphisms were detected in specimens from a local population in central Honshu (11–21 alleles per locus; n = 31) and from the entire distribution range of the species (19–41 alleles per locus; n = 152). These microsatellites will be useful in conservation genetic studies of G. japonicus.     

Isolation and characterization of 12 microsatellite loci from <Emphasis Type="Italic">Suzukia shikikunensis</Emphasis> (Lamiaceae), a genus endemic to Taiwan and Ryukyus

Suzukia shikikunensis Kudo is an endemic plant in Taiwan and suffers habitat destruction caused by human overexploitation. In this study, we developed 12 microsatellite primer pairs for genetic study. These markers were screened for 24 samples collected from wild populations distributed in Taiwan, and for a sister species S. luchuensis collected from Yonaguni and Lutao Islands. The number of alleles ranged from 5 to 14. The expected (H _E) and observed (H _O) heterozygosities were 0.65–0.922 and 0–0.625, respectively. All loci were significantly deviated from Hardy–Weinberg equilibrium due to the heterozygote deficiency. These primers amplifying microsatellites in the two species may provide a useful tool for population genetics to establish conservation strategy.     

A migration analysis of the rice planthopper Nilaparvata lugens from the Philippines to East Asia with three-dimensional computer simulations

Migrations of the rice planthopper Nilaparvata lugens (Stål) (Delphacidae) from the Philippines to Taiwan, southern China, and southern Japan were analyzed using three-dimensional migration simulations. The results strongly suggested that the Southeast Asian population of N. lugens mixes with the East Asian population. This highlighted the possibility that planthoppers from the Southeast Asian population, which have properties different from those in the East Asian population such as feeding of resistant rice varieties and wing polymorphism, could migrate to Japan via southern China and Taiwan. This study, therefore, emphasizes the special care that should be taken to monitor the properties of immigrants to Japan.     

Isolation and characterization of 22 polymorphic microsatellite DNA markers of Japanese sea bass (Laterolabrax japonicus)

Japanese sea bass (Laterolabrax japonicus) inhabits the coast of East Asia and is cage‐cultured currently in China as well. Twenty‐two microsatellite DNA markers were developed and used to type 35 individuals collected along the Chinese coast. The number of alleles per locus ranged from three to 23. The expected heterozygosity ranged from 0.111 to 0.938, while the observed heterozygosity ranged from 0.114 to 0.914. All 22 loci are neutral and independent of each other; two deviated significantly from Hardy–Weinberg equilibrium. These microsatellite DNA markers are moderately informative, which will certainly facilitate the management and exploitation of the genetic resource of L. japonicus.     

Isolation and characterization of microsatellite loci in the endangered freshwater fish <Emphasis Type="Italic">Varicorhinus alticorpus</Emphasis> (Cyprinidae)

Varicorhinus alticorpus (Cyprinidae) is an endangered freshwater fish with a scattered distribution in the southern and eastern regions of Taiwan. We described the development of nine polymorphic microsatellite loci in V. alticorpus for genetic studies. These new markers were tested in 20 individuals of the rare species. The number of alleles ranged from 4 to 13. Expected (H _E) and observed (H _O) heterozygosities ranged from 0.692 to 0.892 (averaged at 0.821) and from 0.000 to 0.350 (averaged at 0.088), respectively. All loci are significantly deviated from Hardy–Weinberg expectations due to the heterozygote deficiency. This suite of highly polymorphic microsatellites provides the first chance to undertake a conservation program for this species in Taiwan. It is essential to evaluate its genetic diversity and the fine-scale population structure. Tzen-Yuh Chiang, Teh-Wang Lee, and Feng-Jiau Lin contributed equally to this work.     

Refugia and phylogeography of Taiwania in East Asia

Aim The East Asia endemic Taiwania cryptomerioides Hayata is an iconic and relictual monotypic conifer whose main extant populations are now restricted to the Yunnan–Myanmar border, northern Vietnam and Taiwan. It has also been reported from several localities in Guizhou, Hubei and Fujian Provinces, China. Its fossil record indicates that, while it was more widely distributed in the Northern Hemisphere and grew under a range of different ecological conditions, it has remained almost unchanged in its morphology for over 100 Myr. We investigate whether these remaining extant, disjunct populations have diverged genetically; when such a divergence may have occurred; and which, if any, of the extant populations exhibit refugial characteristics. Location East Asia. Methods Sequences of five chloroplast DNA markers (petG–trnP, trnH–psbA, trnV–trnM, trnC–ycf6 and trnL–trnF) from all extant populations of T. cryptomerioides were analysed to reveal their phylogeography. Molecular clock models with fossil calibrations were used to estimate divergence times between extant populations. Results Extremely low nucleotide diversity was found in the overall population (π = 0.00077) with only nine haplotypes distinguished. The mainland Asia populations share one major ancestral haplotype. The insular populations in Taiwan all possess a unique haplotype with at least an eight‐mutational‐step difference to the mainland Asia haplotype. Molecular clock estimations demonstrated that the mean divergence time between the predominant insular population haplotype and the mainland Asia haplotype occurred at c. 3.23–3.41 Ma, followed by a split into Vietnamese and Yunnan–Myanmar populations (c. 1.0–1.39 Ma). Main conclusions Strong genetic differentiation exists between insular (Taiwan) and mainland Asia populations. The split between insular and mainland haplotypes can be dated back to the end of the Pliocene. The Yunnan–Myanmar border area, northern Vietnam and Taiwan are identified here as potential refugia for T. cryptomerioides. Other populations in mainland China are unlikely to be the result of historical fragmentation and their origins require further investigation.     

Identification and characterization of nuclear microsatellites in Mediterranean cedars (Cedrus sp.)

We developed primers for the amplification of six polymorphic nuclear microsatellites in Mediterranean Cedrus taxa. Microsatellites originated from two Cedrus atlantica genomic libraries enriched for TC (four markers) and TG (two markers) motifs. No distortion from Mendelian segregation was observed. There were up to eight alleles per locus and mean expected heterozygosity was 0.75. Four microsatellites also amplified in the Himalayan cedar. These markers should be helpful for species identification, diversity studies, parentage analysis and genome mapping and to monitor biodiversity in endangered Mediterranean Cedrus forests.     

Isolation and characterization of polymorphic microsatellite loci in <Emphasis Type="Italic">Ardisia crenata</Emphasis> (Myrsinaceae)

Ardisia crenata, an evergreen shrub native to East Asia, has been a serious invasive plant to the southeastern USA. Here 13 polymorphic microsatellite loci were isolated and characterized from an enrichment genomic library of A. crenata. The average allele number of these microsatellites was four per locus, ranging from two to seven. The ranges of observed and expected heterozygosity were 0.000–1.000 and 0.239–0.789, respectively. These microsatellite markers will be useful for investigating population genetics and reproductive ecology of A. crenata.     

Characterization of Microsatellites in the IGF-2 and GH Genes of Asian Seabass (Lates calcarifer)

Four microsatellites were identified by screening the DNA sequences of Asian seabass (Lates calcarifer) deposited to GenBank. Two markers each are located in the growth hormone gene (GH) and in the insulin-like growth factor II gene (IGF-2), respectively. The markers were characterized by genotyping 34 Asian seabass individuals. All 4 microsatellites showed polymorphism: the number of alleles per locus ranged from 2 to 11 (average, 5.0), while the expected heterozygosity ranged from 0.51 to 0.85 (average, 0.63) at the 4 loci. Cross-priming with all 4 primer pairs was tested in species belonging to 5 different genera, but no bands were amplified. These microsatellites are the first genomic DNA markers characterized in L. calcarifer; thus they may be valuable for research and aquaculture production of this species. Received April 10, 2000; accepted July 13, 2000.     

Microsatellite markers for species of the genus Dionda (Cyprinidae) from the American southwest

Thirty-eight microsatellite markers were developed from an enriched genomic DNA library of the cyprinid fish (minnow) Dionda episcopa. The microsatellites include 31 perfect-repeat motifs (29 dinucleotide, 1 trinucleotide, and 1 tetranucleotide) and 7 imperfect-repeat dinucleotide motifs. The microsatellite primers were used to amplify microsatellites from five related congeners: D. argentosa, D. diaboli, D. episcopa, D. nigrotaeniata, and D. serena. One species (D. diaboli) is listed as threatened and critically imperiled and two species (D. argentosa and D. serena) are listed as imperiled; the conservation status of D. nigrotaeniata is unknown. The number of experimentally tractable microsatellite markers varied from 28 for D. diaboli to 34 for D. episcopa. The number of polymorphic microsatellites conforming to Hardy–Weinberg expectations (following Bonferroni correction) ranged from 19 (D. diaboli) to 27 (D. argentosa). One pairwise comparison of microsatellites (in D. nigrotaeniata) deviated significantly from expectations of genotypic equilibrium. The microsatellite markers will be useful for conservation and population-genetic studies of these and other species in genus Dionda.     

Genetic introgression between an introduced babbler, the Chinese hwamei Leucodioptron c. canorum, and the endemic Taiwan hwamei L. taewanus: a multiple marker systems analysis

Identification of interspecific hybrids is often a subject of primary concern in the development of conservation strategies. Here we performed a genetic analysis combining mitochondrial DNA (mtDNA), microsatellites and single nucleotide polymorphic sites (SNPs) to assay the level of hybridization and introgression between an introduced babbler, Chinese hwamei Leucodioptron canorus, and its close relative, the endemic Taiwan hwamei L. taewanus in Taiwan. Fifty‐five Chinese hwameis from the Asian mainland and 69 Taiwan hwameis, including nine morphological hybrids, were sampled and analyzed. Evidences of mitochondrial introgression were found in three hybrids and one Taiwan hwamei. Five unlinked interspecific SNPs were identified at nine anonymous nuclear loci with interspecific differentiation (total Fst=0.77) that was much higher than that at seven highly polymorphic microsatellite loci combined (total Fst=0.1). Bayesian cluster analysis based on five interspecific SNP loci and two highly differentiated microsatellite loci (Fst>0.08) suggested that twelve individuals sampled in Taiwan were likely F2 or backcross hybrids, among which eight were morphological intermediates. A total of 20.3% (14/69) individuals sampled in Taiwan were suggested to be hybrids, suggesting that fitness reduction in hybrids might be negligible. These results imply that without an effective management strategy, the entire Taiwan hwamei population could easily become an admixed with Chinese hwamei and loose its evolutionary integrity. To reduce introgressive hybridization, illegal trade of Chinese hwamei should be strictly regulated and only the expensive male Chinese hwameis should be legally imported to minimize the chance for Chinese hwameis being released into the field. In our study we also found interspecific SNP markers to outperform microsatellite loci in detecting hybridization and introgression between two closely related species, which may be ascribed to the lower level of homoplasy of SNP loci.     

Phytoplankton community diversity is influenced by environmental factors in the coastal East China Sea

Surface seawater was collected in four different seasons in the coastal East China Sea adjacent to the Yangtze River Estuary and phytoplankton community diversity was analysed using rbcL genetic markers. Phytoplankton diversity (Shannon Index) was found to be highest in autumn and lowest in summer, which was mainly controlled by seawater temperature, river runoff, the Taiwan Warm Current and possibly other environmental factors. For taxa characterized by Form IAB rbcL, the abundance of Chlorophyta was much greater than those of Proteobacteria and Cyanobacteria throughout the year with the most dominant taxa being Bathycoccus prasinos (Chlorophyta) in spring and Micromonas sp. (Chlorophyta) in other seasons. For taxa identified by Form ID rbcL, Coscinodiscophyceae (diatoms) constituted the largest group (most clones) in the phylogenetic tree. Dinophysis fortii (a dinoflagellate) was found to be the most abundant species in winter and spring and Skeletonema spp. (a diatom) dominated the phytoplankton community in summer and autumn. The seasonal dominance of Dinophysis fortii agreed well with the recently increasing proportion of dinoflagellates in the phytoplankton community in the coastal East China Sea. The abundance of Dinophysis fortii was negatively correlated with seawater temperature, suggesting that harmful algal blooms caused by this species may primarily occur in spring.     

Molecular phylogeny of the genus Pieris (Ericaceae) with special reference to phylogenetic relationships of insular plants on the Ryukyu Islands

Phylogenetic relationships within the genus Pieris (Ericaceae) were investigated based on the rbcL and matK genes along with five spacer sequences of chloroplast DNA to address questions regarding the phylogeography of the genus in association with insular plants on the Ryukyu Islands. The most parsimonious trees indicated that P. floribunda from eastern North America is a sister taxon to the remaining taxa examined, and suggested that the East Asian taxa examined are monophyletic. A morphologically cohesive group, section Pieris, was revealed to be paraphyletic. Within the East Asian clade, insular endemics from the Ryukyu Islands, Taiwan, and mainland Japan formed a sister group to P. formosa from the Himalayas and southern China. Our data suggest that the insular endemics of the Ryukyu Islands and Taiwan arose via allopatric divergence as a result of a paleogeographical land configuration of a landbridge during the early–middle Pleistocene in the Quaternary Period.     

Zooplankton assemblages as indicators of seasonal changes in water masses in the boundary waters between the East China Sea and the Taiwan Strait

Hydrology and trophic relationships are frequently reported for inducing changes in mesozooplankton communities. This study investigated the distribution and abundance of mesozooplankton in the boundary waters between the East China Sea and the Taiwan Strait. Samples were collected using a NOR-PAC zooplankton net towed horizontally at a depth of 2 m, at eight stations along a transect, in March (spring) and October (autumn) 2005. The abundance of mesozooplankton was significantly higher during autumn than spring. Densities of many groups (e.g., Noctiluca scintillans, pteropods, copepods, mysids, euphausiids, and other larva) increased in October. During both seasons, copepods represented more than 50% of the total zooplankton abundance. Noctiluca scintillans, appendicularians, and Calanus sinicus were dominant in spring, indicating their association with the cold waters of the East China Coastal Current. Appendicularians and N. scintillans were mainly associated with the coastal waters of Taiwan; whereas C. sinicus was concentrated in the offshore waters along the coast of Mainland China. Chaetognaths, Temora turbinata, Acrocalanus spp., and radiolarians were dominant in autumn, showing their association with the warm waters of the Kuroshio Branch Current. Oncaea venusta was relatively abundant during both seasons. Our study shows that, in addition to the influence of seasonal changes in the water masses, the distribution and composition of mesozooplankton are highly influenced by trophic interactions between zooplankton taxa, in the boundary waters of the Taiwan Strait and the East China Sea.     

Characterization of seven genomic and one dbEST-derived microsatellite loci in the river mangrove<Emphasis Type="Italic"> Aegiceras corniculatum</Emphasis> (Myrsinaceae)

We developed seven microsatellite markers from the genome of Aegiceras corniculatum using the FIASCO protocol and one dbEST-derived microsatellite marker. Polymorphism of each locus was assessed in 45 individuals from China, Thailand and Australia. The average allele number of these microsatellites was 4.4 per locus, ranging from 2 to 8. The observed (H _O ) and expected (H _E ) heterozygosity were from 0.178 to 1.000 and from 0.164 to 0.828, respectively. These microsatellite markers would provide a useful tool for genetic and conservation studies of A. corniculatum.