Research In Ecology And Conservation Genetics Of Chinese Giant Salamander (Andrias Davidianus) In Guizhou

Objective:?1?To understand what water quality parameters are relevant to the survival of Andrias davidianus,whether human activities are the key factors to cause endangerment of A.davidianus,and whether A.davidianus can be used as an ecological indicator for environmental evaluation.?2?To understand whether significant morphological differences are present among different geographical populations of A.davidianus,and whether these populations could be divided into different subspecies.?3?To investigate genetic diversity,distribution and evolutionary history of A.davidianus populations,results from which could inform designations of conservation areas for A.davidianus in situ protection.Methods:?1?We carried out field surveys of A.davidianus in 35 counties of Guizhou and tested water quality using 13 parameters of 39 rivers with A.davidianus historical records over the past 30 years.Mann-Whitney U test was then conducted to determine the water quality factors associated with A.davidianus distribution in the past 5 years.?2?We collected swab samples in 27 A.davidianus captive breeding farms,facilitated by the Fisheries Bureau of 16 counties–the authorities in charge of A.davidianus farms.After obtaining the approval,we collected morphometric data on36 A.davidianus from 4 different geographic populations in 7 farms and carried out cluster analysis and Proncipal component analysis using SPSS16.0 software to investigate morphological variations of A.davidianus according to their geographic distributions.?3?We collected 215 tissue samples of A.davidianus from 27 farms in16 counties and conducted genetic lab work,including sequencing of mtDNA COI to understand the phylogeny and haplotypes of this species,and studies on genetic diversity,distribution patterns and evolutionary process of A.davidianus.Results:?1?Dissolved solids,conductivity and salinity were found to be the significant determinants of A.davidianus distribution in recent five years?P<0.01?,as well as Alkalinity,total hardness,pH and salinity?P<0.05?.There was no A.davidianus distribution in the recent five years in rivers with dissolved solids greater than 415 mg/L,conductivity greater than 639 us/cm,salinity greater than 0.31 ppt and total hardness greater than 150 mg/L.A.davidianus could hence be used as one of the environmental indicator.?2?The results of morphological cluster analysis and the principal component analysis showed that populations in Guiding,Songtao and Jiangkou were similar in morphology,but differed significantly in four morphological?P<0.01?and one morphological?P<0.05?in Pingtang.Overall,greater morphological differences of populations were found between river systems?i.e.Yangtze River and Pearl River?than within the same river?i.e.Yangtze River?.The differential analysis result indicated that variations among different populations of A.davidianus in Guizhou could not reach subspecies level.?3?We sequenced 457 bp mtDNA of COI gene towards 215 samples,detected 437 conservation loci and 20variation loci.Results of phylogenetic tree and haplotype network diagram showed that the A.davidianus samples were classified into Clade A and B.Clade A constituted samples mainly from Guizhou?90%?and Clade B constituted samples mainly from Shaanxi?85%?.?4?There were 5 haplotypes in Clade A and 4 haplotypes in Clade B,and the genetic variations between the two populations differed significantly(F_CT=0.93798,P<0.001).Also,the haplotype diversity?0.574?and the nucleotide diversity?0.00209?of Clade A were both much higher than that of Clade B?which is0.130 and 0.00124,respectively?.There were 4 haplotypes in Fanjingshan and only 1haplotype in Pingtang.?5?Analysis of molecular variance?AMOVA?indicated that the genetic differentiation among Guizhou population and Shaanxi population was occupied 93.8%.Guizhou population and Shaanxi population has been divided into two different population.?6?The divergence time for Guizhou population and Shaanxi populations is between 3.125 to 4.5 million years and the divergence time among populations within Guizhou is between 0.25 to 2 million years.The population from Fanjingshan had significant variations in haplotypes,making it the key region for A.davidianus divergence in Guizhou.Conclusion:?1?Eutrophication,chemical fertilizer,pesticides and inorganic pollutants may be one of reasons driving wild A.davidianus into extinction.As A.davidianus are sensitive to environment,this species is an important indicator for environmental quality.?2?Differential analysis showed that the variations of A.davidianus populations did not divide them into subspecies.?3?Fanjingshan is one of the shelters for Asian species in the changing environment and A.davidianus could represent the genetic diversity with high in southern and low in northern of Asian species;A.davidianus in Fanjingshan population has significant variations in haplotypes,and that of Pingtang is featured by one haplotype,so we suggest that Fanjingshan could be a administrate unit for managing genetic diversity,and Pingtang could be a guard unit for protecting genetic uniqueness and in-situ conservation for this species to maintain its genetic diversity.