Cryptic Species Of China Coelomactra Antiquata Based On Complete Mitochondrial Genomic Evidence

The complete mitochondrial genomes of three species in Mactridae-Coelomactra antiquata (including two geographic stocks:Rizhao C. antiquata, Zhangzhou C. antiquata), Mactra veneriformis and M. chinensis were obtained by using mtDNA as a template with the combination of long amplification polymerase chain reaction (Long-PCR), conventional PCR, cloning and and primer walking sequencing technologies. The differences of mtDNA structure, codon utilization, nucleotide and amino acid sequences, rRNA, tRNA and non-coding regions (NCR) between the RZ and ZZ C. antiquata were also analyzed. The evidence of the cryptic species in C. antiquata was presented with the reference of mtDNA difference between M. veneriformis and M. chinensis. The results are as follows:1. The mtDNA of RZ C. antiquata and ZZ C. antiquata is circular molecule,17384bp and17199bp in length, respectively. The former contains35genes and the latter contains36genes. All encoding genes are located in the heavy strand. The mtDNA both consists of12PCGs (atp8gene is absent) and two rRNA genes. zz-mtDNA(22tRNAs) has one more tRNA gene (tRNAMet) than rz-mtDNA (21tRNA genes). The NCR length of rz-mtDNA and zz-mtDNA is2187bp and1089bp, respectively. The major non-coding region (MNR) of rz-mtDNA is1638bp in length and contains a99bpxll tandem repeat region (1089bp). The mtDNA of ZZ C. antiquata contains two MNRs:the MNR (882bp in length and no tandem repeats are found) and a species-specific non-coding region (SNR,400bp in length).2. The mtDNA of M. veneriformis and M. chinensis is circular molecule,16853bp and17285bp in length. The former contains33genes and the latter contains35genes. The mtDNA both consists of12PCGs and two rRNA genes, however, atp8and two tRNASer genes are absent. All encoding genes are located in the heavy strand. Mve-mtDNA has one more tRNA gene (tRNALys) than Mch-mtDNA. The lengths of Mve-mtDNA and Mch-mtDNA non-coding regions (NCR) are1978bp and2231bp. Meanwhile, the MNR is 700bp and699bp in length, another NCR is at the same location. Mve-mtDNA has a664bp NCR which contains a125bpx5.3tandem repeat region. Mch-mtDNA has a1075bp SNR that containing a125bpx8.1tandem repeat region.3. The nucleotide differences analysis of12PCGs was presented with the reference of difference between M. veneriformis and M. chinensis (ZZ C. antiquata, RZ C. antiquata, M. veneriformis and M. chinensis belong to the same family-Mactridae), revels that the difference between rz-mtDNA and zz-mtDNA are greater than those between M. veneriformis and M. chinensis. The differences of nucleotides and amino acids coded by mitochondrial genes between ZZ and RZ C. antiquata are greater than or equal to the values of three Meretrix species in Bivalvia (M. meretrix, M. lusoria and M. petechialis). The ratios of total genetic distance of the coxl and lrRNA in RZS and ZZS stocks to that of each stock are both over10, which reveal that the difference between ZZ and RZ C. antiquata has reached intraspecies difference level. This proves that ZZ C. antiquata is a cryptic species of Coelomactra.4. The mitochondrial genome (mtDNA) of four Mactridae clams species are sequenced for the first time, the composition and structure of mtDNA was analyzed. It fills the research gap of complete mitochondrial genomes from Mactridae. With the reference of mtDNA difference between M. veneriformis and M. chinensis, combined with single gene differences of other species of bivalve, the difference between rz-mtDNA and zz-mtDNA was analyzed. This study considered that Zhangzhou C. antiquata is a cryptic species of Coelomactra, and it should be named. We should strengthen management of this excellent germplasm to ensure the restoration and sustainable usage.