Morphology,Molecular Identification And Complete Mitochondrial Genome Of Bithyniidae

Bithyniidae belonging to Mollusca,Gastropoda,and Mesogastropoda is widely distributed in the freshwater of Asia,Europe,Africa,Australia and North America.Many of them are intermediate hosts of the parasite which is harmful for human health.There were few studies on the classification of Bithyniidae in China.They were mostly classified based on shell morphology.However,owing to the shell plasticity or individual differences during different period of life history,it always leads to inaccurately identification of Bithyniidae species.The samples studied in this paper were mainly collected from Jiangxi province and surrounding areas,and deposited in the Institute of Zoology,Chinese Academy of Sciences.On the basis of collected samples of the family Bithyniidae,we studied the morphological characteristics of four species,and further carried out the molecular identification.These results laid a foundation for the systematic classification of the Bithyniidae family in China.The main results are as follows:1.We studied the morphological characteristics of four species of Bithyniidae:Parafossarulus eximius(Frauenfeld,1864),Parafossarulus striatulus(Benson,1842),Parafossarulus sinensis(Neumayr,1883),and Alocinma longicornis(Benson,1842).The results showed that there were significant differences on shell morphology(such as:shell height,shell width,spiral height,body height,etc),operculum morphology,and radula characteristics among the four species.These features are of significant for Bithyniidae classification.2.Phylogenetic relationships of Bithyniidae species were analyzed based on three molecular markers,i.e.mitochondrial genes of COI,16S rRNA,nuclear 28S rRNA gene,and combined COI+16S rRNA genes.The results showed that mitochondrial genes could be more accurate to identify species.Especielly,the classification based on COI gene was almost consistent with the morphology results.Therefore,COI gene can be used as DNA barcode for species identification.However,the nuclear 28S rRNA gene was relatively conservative,so it could not distinguish Bithyniidae species.3.According to the specimens deposited in the Institute of Zoology,Chinese Academy of Sciences and literature,species of Bithyniidae were sorted.A total of 25 species of Bithyniidae were described,including one species of the genus Alocinma,14 species of the genus Bithynia,and 10 species of the genus Parafossarulus.4.Mitochondrial genomes of two freshwater snails were sequenced and analyzed.The mitogenome of Parafossarulus eximius was 15,574 bp in length.The contents of each base were:34.5%A,34.7%T,17.5%G,and 13.9%C.The mitogenome of Radix plicatula was 13,751 bp in length(GenBank number:MN175602).The contents of each base were 30.7%A,39.6%T,15.7%G,and 13.9%C.They both had 37 genes,containing 13 protein-coding genes,22 tRNA genes,and 2 rRNA genes.Phylogenetic analysis based on 12 PCGs showed that the family Lymnaeidae was monophyletic,and R.plicatula and R.auricularia had a close relationship.P.eximius of the family Bithyniidae was sister to the family Pomatiopsidae.