Abstract
BackgroundReconstructing the higher relationships of pulmonate gastropods has been difficult. The use of morphology is problematic due to high homoplasy. Molecular studies have suffered from low taxon sampling. Forty-eight complete mitochondrial genomes are available for gastropods, ten of which are pulmonates. Here are presented the new complete mitochondrial genomes of the ten following species of pulmonates: Salinator rhamphidia (Amphiboloidea); Auriculinella bidentata, Myosotella myosotis, Ovatella vulcani, and Pedipes pedipes (Ellobiidae); Peronia peronii (Onchidiidae); Siphonaria gigas (Siphonariidae); Succinea putris (Stylommatophora); Trimusculus reticulatus (Trimusculidae); and Rhopalocaulis grandidieri (Veronicellidae). Also, 94 new pulmonate-specific primers across the entire mitochondrial genome are provided, which were designed for amplifying entire mitochondrial genomes through short reactions and closing gaps after shotgun sequencing.ResultsThe structural features of the 10 new mitochondrial genomes are provided. All genomes share similar gene orders. Phylogenetic analyses were performed including the 10 new genomes and 17 genomes from Genbank (outgroups, opisthobranchs, and other pulmonates). Bayesian Inference and Maximum Likelihood analyses, based on the concatenated amino-acid sequences of the 13 protein-coding genes, produced the same topology. The pulmonates are paraphyletic and basal to the opisthobranchs that are monophyletic at the tip of the tree. Siphonaria, traditionally regarded as a basal pulmonate, is nested within opisthobranchs. Pyramidella, traditionally regarded as a basal (non-euthyneuran) heterobranch, is nested within pulmonates. Several hypotheses are rejected, such as the Systellommatophora, Geophila, and Eupulmonata. The Ellobiidae is polyphyletic, but the false limpet Trimusculus reticulatus is closely related to some ellobiids.ConclusionsDespite recent efforts for increasing the taxon sampling in euthyneuran (opisthobranchs and pulmonates) molecular phylogenies, several of the deeper nodes are still uncertain, because of low support values as well as some incongruence between analyses based on complete mitochondrial genomes and those based on individual genes (18S, 28S, 16S, CO1). Additional complete genomes are needed for pulmonates (especially for Williamia, Otina, and Smeagol), as well as basal heterobranchs closely related to euthyneurans. Increasing the number of markers for gastropod (and more broadly mollusk) phylogenetics also is necessary in order to resolve some of the deeper nodes -although clearly not an easy task. Step by step, however, new relationships are being unveiled, such as the close relationships between the false limpet Trimusculus and ellobiids, the nesting of pyramidelloids within pulmonates, and the close relationships of Siphonaria to sacoglossan opisthobranchs. The additional genomes presented here show that some species share an identical mitochondrial gene order due to convergence.
Highlights
Reconstructing the higher relationships of pulmonate gastropods has been difficult
Analyses based on individual gene sequences all provide similar relationships (Figure 1) [2,3,4,5]: pulmonates are monophyletic, but they include a few taxa not traditionally classified as pulmonates (Acochlidia, traditionally regarded as opisthobranchs, and Glacidorbidae and Pyramidelloidea, traditionally regarded as basal heterobranchs); opisthobranchs are paraphyletic, basal to pulmonates; the false limpet Siphonaria, traditionally regarded as a pulmonate, is in some cases found to be more closely related to opisthobranchs than pulmonates
Analyses based on complete mitochondrial genomes provide different phylogenetic relationships, at least for the deep nodes [6,7,8]: pulmonates are paraphyletic, basal to the monophyletic opisthobranchs; Siphonaria is nested within the opisthobranchs
Summary
Reconstructing the higher relationships of pulmonate gastropods has been difficult. The use of morphology is problematic due to high homoplasy. Molecular studies have been based on few individual genes, essentially 18S, 28S, 16S, and COI data [2,3,4,5], or few complete mitochondrial genomes [6,7,8]. Analyses based on individual gene sequences all provide similar relationships (Figure 1) [2,3,4,5]: pulmonates are monophyletic, but they include a few taxa not traditionally classified as pulmonates (Acochlidia, traditionally regarded as opisthobranchs, and Glacidorbidae and Pyramidelloidea, traditionally regarded as basal heterobranchs); opisthobranchs are paraphyletic, basal to pulmonates; the false limpet Siphonaria, traditionally regarded as a pulmonate, is in some cases found to be more closely related to opisthobranchs than pulmonates. Analyses based on complete mitochondrial genomes provide different phylogenetic relationships, at least for the deep nodes [6,7,8]: pulmonates are paraphyletic, basal to the monophyletic opisthobranchs; Siphonaria is nested within the opisthobranchs. Ten genomes became available in nine different publications between 1995 and 2006; since 2008, 38 genomes became available, 30 of which appeared in only four publications [6,8,10,11], a few papers with only one genome were published
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