Abstract:

Objective To identify Pomacea spp. based on the polymorphism of mitochondria DNA cytochrome c oxidase subunit I gene (COⅠ) amplified from Pomacea collected from southwest Yunnan Province. Methods The samples of Pomacea snails were collected from Jinghong City and Mengla County of Xishuangbanna, Simao District and Ximeng County of Pu’er, Gengma County of Lincang, Ruili City of Dehong, Yunnan Province, from November 18, 2016 to May 8, 2017, morphological observation was performed. DNA was extracted and COⅠgene was amplified and sequenced. The phylogenetic tree and genetic distances among the snail samples compared with known species of Pomacea were constructed using the neighbour-joining method with MEGA software (version 6.06). Results A total of 200 Pomacea snails were collected. It is difficult to distinguish species by morphological identification. 129 COⅠsequences were amplified, 123 sequences belonged to Pomacea canaliculata, 6 sequences belonged to P. insularum. The final length of amplified sequence was 621 bp, there were 76 nucleotide variation sites and 76 simplified information sites, without single variation, insertion and deletion sites, and without base conversion and transmutation. The average contents of A, G, T and C were 23.1%, 20.5%, 39.8% and 16.7%, respectively. The DNA sequence for 207 amino acids with 203 conserved (98.1%). The results showed that COⅠ gene sequence fell into 4 unique haplotypes based on the single mutations. HAP-1 haplotype was the predominant one (71.3%, 92/129), followed by HAP-3(17.8%, 23/129), HAP-2 (6.2%, 8/129） and HAP-4 (4.7%, 6/129). The sequence data showed that HAP-1, HAP-2, HAP-3 shared closer genetic distance to P. canaliculata ranged from 0.000 to 0.052 (with 0 or 31 nucleotide differences), further genetic distance to Pila conica with distances ranged from 0.214 to 0.239 (with 115-126 nucleotide differences). The genetic distances between HAP-4 and P. canaliculate, P. insularum, Pila conica were 0.100, 0.060 and 0.223, and with nucleotide differences of 57, 35 and 119, respectively. The genetic distances among the four HAPs ranged from 0.021-0.100 with nucleotide differences of 13-57. Among them, HAP-1 is closer to HAP-2 (0.021), HAP-4 is further away from HAP-1, HAP-2 and HAP-3 (0.100). The genetic distance between HAP-3 and HAP-1/HAP-2 was 0.052. HAP-1, HAP-2 and HAP-3 shared 98%-100% sequence identity to Pomacea canaliculate (GenBank accession AB433769, KT313034、EU523129), and HAP-4 shared 99% sequence identity to P. insularum (EF514942). Conclusion The species of Pomacea snails in southwest Yunnan Province are P. canaliculata and P. insularum based on COⅠgene polymorphism with certain genetic diversity amongst the acquired haplotypes of P. canaliculata identified in this study.

Key words: Pomacea canaliculata, Pomacea insularum, Cytochrome c oxidase subunit I gene, Genetic diversity, Yunnan Province