Redescription of Microphysogobiotungtingensis (Nichols, 1926) with the description of a new species of the genus (Cypriniformes, Gobionidae) from southern China.

Although Microphysogobiotungtingensis (Nichols, 1926) has been treated valid since it was described, its morphology remains vague, especially when comparing it with another similar species, M.elongatus (Yao & Yang, 1977). In this study, the types of both species were examined and also compared with several lots of specimens from a wide geographical range: there is no significant difference in morphology between them. Additionally, molecular evidence supported by mitochondrial gene sequence also showed low genetic distance in between. Thus, it is suggested that M.elongatus is a junior synonym of M.tungtingensis. While revising these two species, a new species, Microphysogobiopunctatus sp. nov., was discovered that has a similar distribution with them both. However, it can be distinguished from its congeners by having a globular or oval shaped posterior air-bladder chamber which length 58.6%-82.8% of eye diameter; a narrow upper jaw cutting edge which less than half mouth width; a slender caudal peduncle with depth 34.6%-48.5% of length; and a six-branched-ray anal fin. This new species also has numerous small black spots on all fins which is also unique. The new species is morphologically and molecularly close to M.bicolor (Nichols, 1930).

Genetic diversity and structure of the Chinese lake gudgeon (Sarcocheilichthys sinensis).

Mitochondrial DNA cytochrome b and d-loop sequences (2,137 bp) in 65 specimens of Sarcocheilichthys sinensis from five populations were identified as two lineages (I and II). The pairwise genetic distance between lineages I and II was 1.94%. SAMOVA analyses suggested that the best grouping occurred at three groups, Yangtze, Qiantang and Minjiang Rivers. High haplotype diversity (0.949) and low nucleotide diversity (θπ = 1.067%) were detected. The results of the neutrality tests, mismatch distribution and approximate Bayesian computation (ABC) did not support demographic expansions. The results of phylogenetic analysis, statistical dispersal-vicariance analysis (S-DIVA), ABC, MIGRATE-N and the time to the most recent common ancestor (TMRCA) indicated two colonization routes. First, before the Wuyi Mountains lifted, S. sinensis dispersed from the Yangtze River to the Minjiang River. Second, during glaciation, the continental shelf was exposed, which contributed to the dispersion of populations from the Yangtze River.

New insights on geographical/ecological populations within Coilia nasus (Clupeiformes: Engraulidae) based on mitochondrial DNA and microsatellites.

The Coilia nasus is an important fish species, which is of commercial values in China. In order to manage the fisheries resources and the development of sustainable fishing strategies to protect this species, 11 microsatellite loci polymorphisms and mtDNA COI sequences were used to examine the genetic diversity of C. nasus in Japan and China. In total, the 40 COI haplotypes and 93 microsatellite alleles were detected. The mtDNA phylogeny did not support population grouping, but the distribution patterns of mtDNA haplotypes and the results of STRUCTURE analysis based on microsatellite indicated a degree of genetic isolation in this species. Our study suggested that the lack of a population genetic structure might result in its amphidromous life cycle, and the geographical distance and habitat fragments might cause isolated populations. Thus, the sampling populations of C. nasus in Japan and China could be divided as four geographical/ecological populations.

Spatial genetic structure of the cyprinid fish Onychostoma lepturum on Hainan Island.

Population genetic structure of Onychostoma lepturum on Hainan Island was investigated based on mitochondrial CR + cyt b region in 63 specimens collected from four populations. Population analyses indicated significant genetic structure (FST = 0.749) and displayed a significant relationship between phylogeny and geography (NST = 0.750 and GST = 0.140). Thirty-one mtDNA haplotypes were classified into four lineages, and these lineages had an almost allopatric distribution. The results of a statistical dispersal-vicariance analysis suggest that the ancestral populations were distributed widely on Hainan Island, and the rising of the central mountainous area of Hainan Island, the Wuzhi and Yinggeling Mountain Range, separated these four drainages into independent lineages. According to a spatial analysis of molecular variance analysis, we divided these populations into three units: ND, CH and WQ + LS, running into Qiongzhou Strait, the Gulf of Tokin and the South China Sea, respectively. According to our study, the exposure of straits and shelf under water retreat gave chances for population dispersion during the glaciations.

The population history of Garra orientalis (Teleostei: Cyprinidae) using mitochondrial DNA and microsatellite data with approximate Bayesian computation.

BACKGROUND: The South China landmass has been characterized by a complex geological history, including mountain lifting, climate changes, and river capture/reversal events. To determine how this complexity has influenced the landmass's phylogeography, our study examined the phylogeography of Garra orientalis, a cyprinid widely distributed in South China, using sequences from the mitochondrial DNA control region and cytochrome b gene (1887 bp) and polymorphisms of thirteen microsatellite loci. RESULTS: In total, 157 specimens were collected from eight populations. All 88 mtDNA haplotypes were identified as belonging to three major lineages, and these lineages were almost allopatric in their distributions. The results of a statistical dispersal-vicariance analysis suggested that the ancestral populations of G. orientalis were distributed south of the Yunkai Mountains, including on Hainan Island. The mtDNA data revealed a strong relationship between phylogeny and geography. In the microsatellite analysis, a total of 339 alleles with an average of 26 alleles per locus were observed across thirteen microsatellite loci. A clustering algorithm for microsatellite data revealed an admixture-like genetic structure. Although the mtDNA and microsatellite data sets displayed a discordant population structure, the results of an approximate Bayesian computation approach showed that these two markers revealed congruent historical signals. The population history of G. orientalis reflects vicariance events and dispersal related to the complex geological history of South China. CONCLUSION: Our results (i) found that the discordances between mtDNA and microsatellite markers were accounted for by admixtures; (ii) showed that the Wuzhishan and Yinggeling mountain ranges and Qiongzhou Strait were important barriers limiting gene exchange between populations on both sides; (iii) indicated that during glaciation and inter-glacial periods, the strait and continental shelves were exposed and sank, which contributed with the dispersion and differentiation of populations; and (iv) displayed that the admixtures between lineages took place in coastal populations and then colonized the tributaries of the Pearl River.

The complete mitogenome of the oriental sucking barb, Garra orientalis (Cypriniformes, Cyprinidae): repetitive sequences in the control region.

The oriental sucking barb, Garra orientalis, is a small to moderate-sized freshwater fish. In this study, the complete mitochondrial genome of G. oriental was successfully sequenced for the first time with total length of 17,288 bp. The genome structure consists of 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA (tRNA) genes and a control region. Moreover, tandem repeat unit ranged from 259 to 260 bp (repeated 3-4 times) was identified in the control region among G. orientalis individuals.

[Review on taxonomical studies of Chinese inland-water fish].

We reviewed the taxonomy and systematics research history of freshwater fish in China based on 1 236 taxonomic literature records on Chinese freshwater fish. The research was divided into five research periods according to specific historical events: (1) period by foreign scholars, (2) period with Chinese scholars, (3) period during World War II and Civil War, (4) recovery period and (5) period of rapid development. There were representative studies and innovations in all periods. We also discuss here the characteristics of each period on the basis of literature analysis.

The complete mitochondrial genome sequence of Sarcocheilichthys nigripinnis (Cypriniformes, Cyprinidae).

We determined the complete mitochondrial genome (mitogenome) sequence of Sarcocheilichthys nigripinnis by using long polymerase chain reaction method. The total length of S. nigripinnis mitogenome is 16,680 bp, including 13 protein-coding genes, 22 transfer RNAs, 2 ribosomal RNAs, and a noncoding control region. The overall base composition of S. nigripinnis is 26.36% for T, 26.67% for C, 30.07% for A, and 16.90% for G, with a slight AT bias of 56.43%.

Complete mitochondrial genome of Osteochilus salsburyi (Cypriniformes, Cyprinidae).

Osteochilus salsburyi (Cypriniformes, Cyprinidae) is a small-sized fish of significant economic value. In this paper, the complete mitogenome sequence of O. salsburyi was first determined. It is 16,599 bp in length and consists of a typical vertebrate genome structure including 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA (tRNA) genes, and a control region. Except for eight tRNA and ND6 genes, all other mitochondrial genes are encoded on the heavy strand. The overall base composition of the heavy strand in descending order is A (33.0%), T (26.5%), C (25.4%), and G (15.0%), with a slight AT bias of 59.5%. This information of O. salsburyi mitogenome could contribute not only to the development of efficient conservation strategy on vulnerable genetic diversity but also to the identification of useful genetic markers for distinction across species boundary.

Complete mitochondrial genome of the endangered roughskin sculpin Trachidermus fasciatus (Scorpaeniformes, Cottidae).

In this study, the complete mitochondrial genome of the endangered roughskin sculpin, Trachidermus fasciatus, was first determined. The mitogenome (16,536 bp) consisted of 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes, and 1 control region. Except for the eight tRNA and ND6 genes, all other mitochondrial genes were encoded on the heavy strand. Mitochondrial DNA information can assist in species identification and conservation of the species' natural resources.

Comparative evolution of S7 Intron 1 and ribosomal internal transcribed spacer in Coilia nasus (Clupeiformes: Engraulidae).

Coilia nasus is widely distributed in the Yangtze River, the coastal waters of China, Korea and the Ariake Sound of Japan. Several ecotypes exist and this provides a useful model for the study of comparative diversity between molecular markers. Here we analyze and compare the nucleotide sequences between single-copy ribosomal protein S7 gene intron 1 (rpS7) and multiple-copy ribosomal internal transcribed spacer 1 (ITS1) in this species to compare the phylogenetic signal of the two nuclear genes. Nucleotide substitutions among the two gene sequences and partial sequence of mitochondrial cytochrome c oxidase subunit I (COI) gene were also analyzed. A total of 115 clones for rpS7 and 122 clones for ITS1 were obtained from 37 specimens. The nucleotide sequence length is 741 to 743 bp for rpS7 and 334 to 348 bp for ITS1. Intra- and inter-specimen variation in rpS7 results from nucleotide substitution, while such variation in ITS1 is mainly due to different numbers of short base repeats. The content of G + C is lower in rpS7 (43.5%) than in ITS1 (68.2%). Our results indicate that the proportion of the sequence variable sites is higher in rpS7 (61) than in ITS1 (23); the informative parsimony of rpS7 is evidently higher than that of ITS1 (26 vs. 2); the overall ratio between transitions and transversions in ITS1 is slightly lower than in rpS7, but remarkably lower than in COI. These results suggest that rpS7 is more suitable than ITS1 as a marker for genetic divergence of this group. Furthermore, gene flow is observed between the different geographic populations of C. nasus from the phylogeny of this species based on rpS7, showing that rpS7 has more evolutionary characteristics for understanding the processes of genomic evolution at the intraspecific level.

[Analysis of genetic diversity in wild populations of Trachidermus fasciatus by RAPD and the transformation of two SCAR markers].

Firstly, RAPD was conducted to analyze genetic diversity of Trachidermus fasciatus in the Fuchun River population (FR), Yellow River population (YR), Luan River population (LR), and Yalu River population (YL), with 32 polymorphic 10-bp random primers selected from 294 ones. Thirty wild individuals were detected in each population. The results indicated that the genetic diversity of T . fasciatus was relatively rich. The major results were as the following: 1) Altogether, 591 bands were detected and 515 of them were polymorphic, accounted for 87.14%. The range of proportion of polymorphic loci (P) was: FR(89.17%)>YR(87.99%)>YL(86.63%)>LR(83.25%). 2) The Shannon's information index(I(T)) and Nei's genetic diversity(H(T)) among populations were 0.3393-0.3566 and 0.2157-0.2279, respectively. Compare to other three populations, LR population had relative lower values. If took the populations as a whole, the total Nei's genetic diversity(H(T)) and Shannon's information index(I(T)) was 0.2336±0.1643 and 0.3710±0.2153, respectively. 3) The value of gene flow (N(m) ) (5.76103-19.84497) were high, indicating certain gene exchange existed among the four populations. But the AMOVA results exhibited significantly differentiation (P<0.05 or P<0.01) among the populations. 4) In the UPGMA tree constructed according to genetic distance, YL and YR populations clustered firstly, then with FR population, and finally they joined to LR population. Obviously, the YL, YR and FR populations had relatively close relationship according to their geographic distance, whereas LR population showed clear divergence to the other three populations. Secondly, out of the five special RAPD bands (S(1225)(525 bp), S(1225)(605 bp), S(1225)(841 bp), S(1345)(695 bp) and S(1345)(825 bp)), SCAR maker SCAR01(560 bp) and SCAR02(443 bp) were successfully transformed from S(1255)(605 bp) and S(1255)(841 bp), respectively. After large samples examination of the two markers, we found the highest frequency (96.67% and 93.33%) in the YL population, higher frequency (83.33% and 90%) in the FR population, high frequency (56.67% and 66.67%) in the YR population, and the lowest frequency (13.33% and 20 %) in the LR population. Therefore, SCAR01(560 bp) and SCAR02(443 bp) can be used as special molecular markers for the population identification between LR and other three populations.

Phylogeographical analysis on Squalidus argentatus recapitulates historical landscapes and drainage evolution on the island of Taiwan and mainland China.

Phylogeographical analyses on Squalidus argentatus samples from thirteen localities within mainland China and Taiwan were conducted for biogeographic studies, as their dispersal strictly depends on geological evolution of the landmasses. A total of 95 haplotypes were genotyped for mtDNA cyt b gene in 160 specimens from nine river systems. Relatively high levels of haplotype diversity (h = 0.984) and low levels of nucleotide diversity (π = 0.020) were detected in S. argentatus. Two major phylogenetic haplotype groups, A and B, were revealed via phylogenetic analysis. The degree of intergroup divergence (3.96%) indicates that these groups diverged about 4.55 myr (million years) ago. Haplotype network and population analyses indicated significant genetic structure (F(ST) = 0.775), largely concordant with the geographical location of the populations. According to SAMOVA analysis, we divided these populations into four units: Yangtze-Pearl, Qiantang-Minjiang, Jiulong-Beijiang and Taiwan groups. Mismatch distribution analysis, neutrality tests and Bayesian skyline plots indicated a significant population expansion for lineage A and B, approximately dated 0.35 and 0.04 myr ago, respectively. We found strong geographical organization of the haplotype clades across different geographic scales that can be explained by episodes of dispersal and population expansion followed by population fragmentation and restricted gene flow.

Complete mitochondrial genome of the mudskipper Boleophthalmus pectinirostris (Perciformes, Gobiidae): repetitive sequences in the control region.

The mudskipper, Boleophthalmus pectinirostris (Perciformes, Gobiidae), is an amphibious gobioid fish. In this paper, the complete mitochondrial genome of B. pectinirostris was firstly determined. The mitogenome (17,111 bp) comprises 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes and 1 putative control region. 130-bp tandem repeat was identified in the control region, which was almost identical among the 10 individuals examined, and three different frequencies of the repeat unit (five, six or seven) were found among these individuals.

Polymorphic microsatellite loci isolated from the Squalidus argentatus using PCR-Based Isolation of Microsatellite Arrays (PIMA).

Squalidus argentatus (Sauvage and Dabry de Thiersant 1874) is a small-sized freshwater fish which is distributed in Mainland China, Hainan Island and Taiwan. The populations of S. argentatus have dropped sharply probably due to overharvesting and water pollution recently. Eleven polymorphic microsatellite markers were developed for the cyprinid fish S. argentatus. These new markers were tested on 43 individuals collected from Yangtze River and Qiantang River. The number of alleles, observed and expected heterozygosity per locus, in two populations ranged from 3 to 14, from 0.333 to 0.954 and from 0.480 to 0.928, respectively. Only two loci are significantly deviated from Hardy-Weinberg expectations due to the heterozygote deficiency. No significant linkage disequilibrium was detected between the pairwise comparisons of these loci. These polymorphic microsatellite loci will enable us to study the genetic variation, population structure, and conservation genetics of this species in the future.

Behavioral migration diversity of the Yangtze River Japanese Eel, Anguilla japonica, based on otolith Sr/Ca ratios.

The Yangtze River estuary is the main production area of Anguilla japonica in China, as well as the only existing fishery area for adult eels. Japanese eels are distributed in the main rivers and many tributaries from the Yangtze River estuary to the upper Jinsha River, which extend to nearly 3 000 km. However, their migration behaviors remain relatively unknown. We analyzed the biological characteristics of 153 specimens of silver eels collected from the Jingjiang section of Yangtze River (31(o)30'N, 120(o)42'E) between September and November, 2008, and tested the sagittal Sr/Ca ratios of 27 specimens. Among the 153 specimens examined, 85 were female and 68 were male, which translated to a female-male ratio of 1 : 0.8. The ages of the female specimens ranged from 3 to 7 a (average 5.52) with an average total length (TL) of (669 ± 80) mm, average body weight (BW) of (555 ± 229)g, average condition factor of 1.77 ± 0.22, and average gonad somatic index (GSI) of 1.32 ± 0.31. The ages of the males ranged from 3 to 5 a (average:4.38) with an average TL of (518 ± 51) mm, average BW of (234 ± 76) g, average condition factor of 1.62 ± 0.18, and average GSI of 0.21 ± 0.11. All biological parameters of females were significantly larger than those of the male specimens (P<0.05). According to the average Sr/Ca ratio (7.99 ± 1.05) × 10(-3) of the elver mark of sagitta, 17 individuals (62.96%) were river eels and 10 individuals (37.04%) were estuarine eels. Of 16 females, 13 individuals (81.25%) were river eels and 3 were estuarine eels, while of 11 males, 36.36% were river eels and 63.64% were estuarine eels. The analysis on Sr/Ca ratios for every growth layer group (GLG) indicated there were no significant differences between second-age males and females. However, significant differences were observed between the third-age, fourth-age, and migration-age male and female specimens. This was likely related to the fact that second-age eels of both sexes stay in the same inhabitation waters; however, as they grow older, they move to different areas.

[A new record species, Heniochus diphreutes, Chaetodontidae from China].

A new record species of Chaetodontidae, Heniochus diphreutes, was found by comparative analysis of the morphological characteristics in a taxonomic revision of the family Chaetodontidae in China. It was distinguished from its relative species H. acuminatus by a combination of the following: Dorsal fin usually with 12 spines (vs. 11); 2-3 rows of teeth on both jaws (vs. 5-7); ventral profile of head convex (vs. almost straight); anal fin more angular, and black area on posterior part of anal fin usually extending anteriorly to longest soft ray (vs. more round, and usually not extending anteriorly to longest soft ray); and snout shorter than eye (vs. longer).

Morphological changes of silver and bighead carp in the Yangtze River over the past 50 years.

Multivariate analysis was adopted to analyze 30 morphometrical characteristics of 121 one-year-old juvenile silver carp (Hypophthalmichthys molitrix) and bighead carp (Aristichthys nobilis) bred during the 1950s ("the former population") and 2008 ("the current population") and collected from the middle reach of the Yangtze River. The average discriminant accuracies of the former and current silver and bighead carp population were 94.2% and 98.0%, respectively. Discriminant analysis also revealed that significant differences in morphology occurred between the former and current populations of both carp in overall characteristics. One-way analysis of variance indicated that between former and current populations, silver carp showed highly significant differences (P<0.01) in twelve of their characteristics and significant differences (P<0.05) in eight of their characteristics, while bighead carp showed highly significant differences (P<0.01) in eight of their characteristics and significant differences (P<0.05) in eight of their characteristics. Six head morphology variables of the current silver and bighead carp were significantly or highly significantly larger than the former populations; fourteen characteristics of silver carp and ten characteristics of bighead carp of the current populations, mainly reflecting truck and tail morphology, were significantly or very significantly smaller than the former populations. Our results indicate that silver and bighead carp have developed a larger head and smaller truck and tail during the last 50 years. Due to such morphological changes, it seems apparent that the heads of these fish species need to be considered in regards to human diets, particularly in relation to economic and nutritious value.

Ontogenetic changes in meristic measurements of silver carp and bighead carp.

The study analyzed 30 exterior meristic characters for 255 silver and bighead carp samples of 1- to 5-year-old collected from the National Primary Breeding Farm at Laojianghe Lake at the Middle Reach of the Yangtze River. Multivariate analysis was performed. In silver carp, the Euclidean distance was the greatest between the 1-year-old group and other age groups. Silver carp individuals were correctly classified at 98.0% accuracy with a discriminant function established by discriminant analysis based on meristic measurements. Similarly, bighead carp had the greatest distance between 1- to 2-year-old group and other age groups. Individuals of bighead carp were correctly classified at 90.7% accuracy by the discrimination function. The data showed that morphological transformation occurred during the life history of silver and bighead carp development. Eighteen meristic measurements showed highly significant differences, while four showed a significant difference between the two silver carp groups. Ten parameters decreased, while twelve measurements increased during development. In bighead carp, fourteen parameters were significantly different, while three parameters were significantly different between the two groups. Twelve parameters were significantly decreased and another five were increased during development. The results suggest allometric growth should be taken into account when identifying species, analyzing population differences and establishing germplasm standards based on morphology.