Sequencing of the complete mitochondrial genomes of eight freshwater snail species exposes pervasive paraphyly within the Viviparidae family (Caenogastropoda)

<div><p>Phylogenetic relationships among snails (Caenogastropoda) are still unresolved, and many taxonomic categories remain non-monophyletic. Paraphyly has been reported within a large family of freshwater snails, Viviparidae, where the taxonomic status of several species remains questionable. As many endemic Chinese viviparid species have become endangered during the last few decades, this presents a major obstacle for conservation efforts. Mitochondrial genomes (mitogenomes) carry a large amount of data, so they can often provide a much higher resolution for phylogenetic analyses in comparison to the traditionally used molecular markers. To help resolve their phylogenetic relationships, the complete mitogenomes of eight Chinese viviparid snails, <i>Viviparus chui</i>, <i>Cipangopaludina chinensis</i>, <i>C</i>. <i>ussuriensis</i>, <i>C</i>. <i>dianchiensis</i> (endangered), <i>Margarya melanioides</i> (endangered), <i>M</i>. <i>monodi</i> (critically endangered), <i>Bellamya quadrata</i> and <i>B</i>. <i>aeruginosa</i>, were sequenced and compared to almost all of the available caenogastropod mitogenomes. Viviparidae possess the largest mitogenomes (16 392 to 18 544 bp), exhibit the highest A+T bias (72.5% on average), and some exhibit unique gene orders (a rearrangement of the standard MYCWQGE box), among the Caenogastropoda. Apart from the Vermetidae family and Cerithioidea superfamily, which possessed unique gene orders, the remaining studied caenogastropod mitogenomes exhibited highly conserved gene order, with minimal variations. Maximum likelihood and Bayesian inference analyses, used to reconstruct the phylogenetic relationships among 49 almost complete (all 37 genes) caenogastropod mitogenomes, produced almost identical tree topologies. Viviparidae were divided into three clades: a) <i>Margarya</i> and <i>Cipangopaludina</i> (except <i>C</i>. <i>ussuriensis</i>), b) <i>Bellamya</i> and <i>C</i>. <i>ussuriensis</i>, c) <i>Viviparus chui</i>. Our results present evidence that some <i>Cipangopaludina</i> species (<i>dianchiensis</i> and <i>cathayensis</i>) should be renamed into the senior genus <i>Margarya</i>. The phylogenetic resolution obtained in this study is insufficient to fully resolve the relationships within the ‘b’ clade, but if <i>C</i>. <i>chinensis</i> proves to be a valid representative of the genus, <i>C</i>. <i>ussuriensis</i> may have to be reassigned a different genus (possibly <i>Bellamya</i>, or even a new genus). Non-monophyly also remains pervasive among the higher (above the family-level) Caenogastropod taxonomic classes. Gene order distance matrix produced a different phylogenetic signal from the nucleotide sequences, which indicates a limited usability of this approach for inferring caenogastropod phylogenies. As phenotypic homoplasy appears to be widespread among some viviparid genera, in order to effectively protect the rapidly diminishing endemic Viviparid populations in China, further detailed molecular phylogenetic studies are urgently needed to resolve the taxonomic status of several species.</p></div

Mitochondrial genome sequences retrieved from GenBank for this study.

<p>Mitochondrial genome sequences retrieved from GenBank for this study.</p

Mitochondrial genome sequences retrieved from GenBank for this study.

<p>Mitochondrial genome sequences retrieved from GenBank for this study.</p

Sampling and specimen details.

<p>Sampling and specimen details.</p

Phylogenetic relationships and gene order.

<p>Phylogenetic relationships among the majority of the available caenogastropod mitogenomic sequences (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0181699#pone.0181699.t002" target="_blank">Table 2</a>) based on nucleotide sequences of all 37 genes. <i>Haliotis rubra</i> and <i>Pleuropoma jana</i> were used as outgroups. Numbers on the nodes correspond to ML bootstrap support (left, expressed as 0–100) and BI posterior probabilities (right, expressed as 0–1.0). Only values below 100/1.0 are depicted. Gene order is shown next to each sequence.</p