School of Natural Sciences, University of California, 5200 North Lake Road, Merced, CA 95343, USA.
BMC Evol Biol. 2011 Oct 10;11:295. doi: 10.1186/1471-2148-11-295.
Reconstructing 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.
The 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.
Despite 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.
重建腹足纲软体动物的高级关系一直很困难。由于高度同形性，形态学的应用存在问题。分子研究受到低分类群采样的影响。目前已有 48 个完整的软体动物线粒体基因组，其中 10 个是腹足纲。本文介绍了以下 10 种腹足纲软体动物的新的完整线粒体基因组：Salinator rhamphidia（两栖类）； Auriculinella bidentata、Myosotella myosotis、Ovatella vulcani 和 Pedipes pedipes（Ellobiidae）；Peronia peronii（Onchidiidae）；Siphonaria gigas（Siphonariidae）；Succinea putris（Stylommatophora）；Trimusculus reticulatus（Trimusculidae）；和 Rhopalocaulis grandidieri（Veronicellidae）。此外，还提供了 94 个新的腹足纲特异性引物，这些引物跨越整个线粒体基因组，旨在通过短反应扩增整个线粒体基因组，并在散弹测序后闭合缺口。
提供了 10 个新线粒体基因组的结构特征。所有基因组都具有相似的基因排列。进行了基于贝叶斯推断和最大似然法的系统发育分析，包括 10 个新基因组和 17 个来自 Genbank 的基因组（外群、后鳃类和其他腹足纲）。基于 13 个蛋白质编码基因的氨基酸序列串联分析，产生了相同的拓扑结构。腹足纲是并系的，位于后鳃类的顶端，是单系的。传统上被认为是基础腹足纲的 Siphonaria 嵌套在后鳃类中。传统上被认为是基础（非后鳃类）后鳃类的 Pyramidella 嵌套在腹足纲中。几个假设被否定了，例如 Systellommatophora、Geophila 和 Eupulmonata。Ellobiidae 是多系的，但假石鳖 Trimusculus reticulatus 与一些 Ellobiidae 密切相关。
尽管最近在增加后鳃类（后鳃类和腹足纲）分子系统发育中的分类群采样方面做了很多努力，但由于支持值较低，以及基于完整线粒体基因组和基于单个基因（18S、28S、16S、CO1）的分析之间存在一些不一致，一些较深的节点仍然不确定。腹足纲（特别是 Williamia、Otina 和 Smeagol）以及与后鳃类密切相关的基础后鳃类需要更多的完整基因组。为了解决一些较深的节点，还需要增加腹足类（以及更广泛的软体动物）系统发育学的标记数量——尽管这显然不是一项容易的任务。然而，一步一步地，新的关系正在被揭示，例如假石鳖 Trimusculus 和 Ellobiidae 之间的密切关系、Pyramidelloids 嵌套在腹足纲中以及 Siphonaria 与 sacoglossan 后鳃类的密切关系。本文介绍的额外基因组表明，由于趋同，一些物种具有相同的线粒体基因排列。
