Phylogeography Of Rhodeus Ocellatus From South Of Yangtze River And Taiwan

Understanding how historical processes have either similarly, or differentially shaped the evolution of lineages or biotic assemblages is important for a broad of fields. Freshwater fishes in Southeast mainland China and Taiwan have been accomplished mainly by vicariant events associated with the complex geolocial history of this region. Rhodeus ocellatus, belonging to Rhodeus, Acheilognathinae, Cyprinidae, Cypriniformes, which is widely distributed in east Asia. In the paper, we collected 213 fish of 13 populations of the R. ocellatus from South of the Yangtze River and Taiwan, they were Qiantang river(KH), Feiyun river(WC), Minjiang river(WYS), Ganjiang river(YC), Mulan river(XY), Jiulong river(ZP), Hanjiang(SH), Beijiang of Pearl river(RY), Dongjiang of Pearl river(HY), Rongjiang river(JX), Moyang river(CW), Chang Hua river(LD), Er-ren river(TN). In this study, based on mitochondrial molecular markers(Cyt b gene and control area) and nuclear genetic markers(S7), we analyzed population genetic structure and phylogeography of R. ocellatus in this area. Through the genetic structure analysis for R. ocellatus of the area, rebuild the phylogenetic relationship of R. ocellatus among different drainage, then to speculate the correlation between genetic differentiation and geographic distribution. These results will provide the methods and measures to protect and manage the fish biodiversity in our country.Spotted barbell(Hemibarbus maculates)(Cypriniformes, Cyprinidae), a common and economically important species, is widely distributed throughout China’s rivers and lakes. The objective of this study was to investigate four hatchery populations(Shanghai, Hangzhou, Kaihua and Xianju) and one wild population(Yingshan) of spotted barbell(Hemibarbus maculates) to assess their genetic diversity level and investigate their population genetic structure.The main results were shown as follows: 1. Population genetic structure and phylogeography of Rhodeus ocellatus onnucleotide sequences of the mt DNA sequences.The mt DNA Cyt b gene complete sequences and control region part of the sequence were sequenced and analyzed the genetic diversity for population samples of R. ocellatus which collected from the south of the Yangtze River and Taiwan, then studied the population genetic variation and phylogeographic patterns. We analyzed 1947 base pairs of the mt DNA sequences(1141bp Cyt b and 806 bp D-loop region gene sequences) with 213 R.ocellatus individuals. The results were as follows:(1) Genetic diversity. There were 548 variable sites(28.15% of the total sequences) among 1947 base pairs of the mt DNA sequences, including 418 parsimoniously informative sites(21.47%) and 130 singleton sites(6.68%). A total of 149 haplotype is classified for these 213 sequences. The average contents of A, T, C and G were 27.7%, 31.4%, 25.5% and 15.4%, respectively. The content of A+T(59.1%) is bigger than G+C(40.9%). Genetic distances between the populations of R. ocellatus were calculated using the Kimura-2-parameter model. The result showed that the genetic distances ranged from 0.002 to 0.107.(2) Population genetic diversity. In each population, the haplotype diversity(h) and nucleotide diversity(π), ranged from 0.000 to 1.000 and from 0.00000 to 0.05209, average were 0.993 and 0.04993, respectively in 13 populations of R. ocellatus. In general, the genetic diversity was higher for all sampled populations.(3) Phylogenetic Analysis. ML/NJ tree of haplotypes for R. ocellatus revealed that the 13 populations can be divided into four major lineages. However, the samples of each lineage showed scattered distribution between populations of different geographic, and there was no significant correlation between genetic and geographical distance. In summary, R. ocellatus didn’t present a significant phylogeographic structure. Taiwan Population is a group in lineage C, and its closest living relative is the mainland drainage in Zhejiang feiyun river population and the qiantang river population. The recent ancestors directed to Zhejiang, and showed the fish is migrating from north to south.(4) Genetic differentiation and genetic structure among populations. The pairwise FST between populations of R. ocellatus was from-0.67208 to 1.00000, and FST of all populations was 0.65215, which showed the populations have a high genetic differentiation. AMOVA and SAMOVA revealed that the genetic variation mainly came from between individuals of within population. The results indicated that, genetic differentiation between populations of R. ocellatus was not completely, and gene exchange was frequent and R. ocellatus was approximated as a random mating group.(5) Demographic history. Neutrality test and nucleotide mismatch distribution analysis among all geographical populations showed that R. ocellatus had not experienced the population expansion. During the population evolution, R. ocellatus experienced a balanced selection function and maintained a stable state and population size. 2. Population genetic structure and phylogeography of Rhodeus ocellatus onnucleotide sequences of the S7 gene sequences.We invested the genetic structure and demographic history of 157 individuals of R. ocellatus using S7 gene sequences as the genetic marker.(1) Genetic diversity. There were 575 variable sites(66.17% of the total sequences) among 869 base pairs of the S7 gene sequences, including 450 parsimoniously informative sites and 125 singleton sites, and there were 205 insertion/deletion. A total of 111 haplotype is classified for these 157 sequences. The average contents of A, T, C and G were 31.7%, 31.1%, 17.1% and 20.1%, respectively. The content of A+T(62.8%) is bigger than G+C(37.2%). The genetic distance analysis based on S7 showed that the K-2-P distances between Changhua river(LD) population and Beijiang of Pearl river(RY) population was minimum(0.0104); and the K-2-P distances between Feiyun river(WC) population and Ganjiang river(YC) population was maximum(0.2120).(2) Population genetic diversity. In terms of nucleotide diversity analysis, we used π and θ to evaluate. The average π and θ of all populations were 0.08527 and 0.00569, beside the π value ranged from 0.00000 to 0.15309 and the θ value ranged from 0.00000 to 0.04058. In the analysis of the haplotype diversity(h), the average h of all populations was 0.966, and ranged from 0.000 to 1.000. These results showed that R.ocellatus populations had a very high genetic diversity.(3) Phylogenetic Analysis. We used MEGA 6.0 to rebuild relationship among individuals based on Neighbor-joining(NJ), the results showed all individuals present lineage A and lineage B on S7 loci. The populations of two lineages had no significant differentiation in geography. The samples showed scattered distribution and could not cluster according to the different geographical regions or drainages, then showed the two lineages of R.ocellatus populations were both in the lineage sorting. Among them, the population of Taiwan was closed to Zhejiang Feiyun river and the qiantang river population.(4) Genetic differentiation and genetic structure among populations. The pairwise FST between populations of R.ocellatus was from-0.20603 to 1.00000, and FST of all populations was 0.22107, which showed the populations have a high genetic differentiation. The index of genetic differentiation was 0.19588 between lineage A and lineage B, and reached significant level, then the results showed two lineages had a great difference. AMOVA and SAMOVA analysis results showed that most of the genetic variation came from between individuals of within population. The results indicated that there was not apparent geographical differentiation based on S7, namely the genetic differentiation was not affected by the geographic isolation.(5) History demographic analysis. We analysis neutrality test and mismatch distribution using DNASP, and found R.ocellatus had not experienced the population expansion. During the population evolution, R.ocellatus experienced a balanced selection function and maintained a stable state and population size.Mitochondrial and nuclear genes are adopted in our study, found that compared with the nuclear gene S7, mitochondrial genes(Cyt b + D-loop) have more polymorphic information, Tajima D test was significant only in nuclear gene S7. The variations of nuclear gene S7 distributed among the current geological districts, but the variations of mitochondrial gene distributed among the past geological districts. These discordant results between nuclear and mitochondrial structures could be the result of the different biological factors. Thus, these two genetic markers showed different phylogenetic patterns was not unexpected. In addition to this research, we found Taiwan’s population was shown closed to Zhejiang feiyun river population and the qiantang river population, and the tree bansed on the mitochondria DNA was more clear on sorting lineage. Thus, our researches combined with nuclear and mitochondrial genes two genetic markers to understand the evolutionary history of R. ocellatus fully. In general, considered that comparing mitochondrial and nuclear gene trees provides a way to identify instances of demographic history, and no moclecular system such as mitochondrial DNA is likely to be perfect for phylogenetic analysis. 3、 Population Genetic Structure in Wild and Hatchery Populations ofHemibarbus maculates Inferred from Microsatellites MarkersGenetic analysis of hatchery and wild populations of Hemibarbus maculates using the 12 microsatellite primers. A total of 135 alleles were detected for the 12 polymorphic markers in 112 individuals, with some unique alleles in each population. The mean number of alleles per locus in the wild and hatchery populations was 5.83 and 7.44. The mean expected heterozygosity per population ranged from 0.54(YS) to 0.66(KH)(average = 0.61), and the observed heterozygosity per population ranged from 0.48(HZ) to 0.68(SH). The average PIC value was highest in the SH and KH populations(0.62), followed by the XJ(0.58), HZ(0.56), and YS(0.51) populations. AMOVA analysis revealed that 77.72% of the total variation was within individuals, 3.28% was among individuals within populations, and 12.81% was among groups(FCT= 0.128). Results of AMOVA analysis according to wild and hatchery populations showed that most variances occurring within individuals. The dendrogram and STRUCTURE revealed that populations XJ, KH, and HZ jointly formed one cluster, to which populations SH and YS were sequentially adhered. The genetic diversity of hatchery populations was maintained better than for the wild population. This may be because the good seedling condition decreased the effect of selection on hatchery populations, which environmental pressure groups under the condition of natural, and made should be washed out of the individuals to stay, then maintained a high degree of genetic diversity in group.