The Gastric Phenotype in the Cypriniform Loaches: A Case of Reinvention?

The stomach, which is characterized by acid peptic digestion in vertebrates, has been lost secondarily multiple times in the evolution of the teleost fishes. The Cypriniformes are largely seen as an agastric order; however, within the superfamily Cobitoidea, the closely related sister groups Nemacheilidae and Balitoridae have been identified as gastric families. The presence of these most recently diverged gastric families in an otherwise agastric clade indicates that either multiple (>2–3) loss events occurred with the Cyprinidae, Catostomidae and Cobitidae, or that gastric reinvention arose in a recent ancestor of the Nemacheilidae/Balitoridae sister clade. In the present study, the foregut regions of Cobitidae, Nemacheilidae/Balitoridae and the ancestral Botiidae family members were examined for the presence of gastric glands and gastric proton pump (Atp4a) α subunit expression by histology and immunohistochemistry respectively. Atp4a gene expression was assessed by reverse transcriptase-polymerase chain reaction (RT-PCR). Gastric glands expressing apical H+/K+-ATPase α subunit and isolated partial sequences of atp4a, identified using degenerate primers showing clear orthology to other vertebrate atp4a sequences, were detected in representative species from Nemacheilidae/ Balitoridae and Botiidae, but not Cobitidae (Misgurnus anguillicaudatus). In summary, we provide evidence for an uninterrupted gastric evolutionary lineage in the Cobitoidea, making it highly improbable that the stomach was reinvented in the Nemacheilidae/Balitoridae clade consistent with Dollo’s principle. These results also indicate that the gastric trait may be present elsewhere in the Cobitoidea.

脊椎动物的胃以酸性胃蛋白酶消化为核心特征，但其在真骨鱼类（teleost fishes）的演化历程中曾多次次生丢失。鲤形目（Cypriniformes）通常被视为无胃目；然而在鳅超科（Cobitoidea）内，亲缘关系紧密的姊妹群条鳅科（Nemacheilidae）与爬鳅科（Balitoridae）已被鉴定为有胃类群。在整体以无胃为特征的演化支中，这些最晚分化的有胃类群的存在，提示两种演化假说：一是鲤科（Cyprinidae）、胭脂鱼科（Catostomidae）与鳅科（Cobitidae）先后发生了至少2~3次胃丢失事件；二是胃结构在条鳅科/爬鳅科姊妹演化支的近期祖先中发生了次生重建。本研究通过组织学与免疫组织化学方法，分别检测了鳅科、条鳅科/爬鳅科以及Botiidae科（Botiidae）祖先类群物种的前肠区域是否存在胃腺与胃质子泵（Atp4a）α亚基的表达；同时采用逆转录聚合酶链式反应（RT-PCR）评估了Atp4a基因的表达水平。研究在条鳅科/爬鳅科与Botiidae科的代表性物种中，检测到了表达顶端H+/K+-ATP酶α亚基的胃腺，以及通过简并引物扩增得到、与其他脊椎动物atp4a序列具有明确直系同源关系的atp4a部分序列；但在鳅科物种泥鳅（Misgurnus anguillicaudatus）中未发现此类特征。综上，本研究为鳅超科内存在连续的胃演化谱系提供了实证，这使得条鳅科/爬鳅科演化支中胃结构次生重建的可能性极低，契合多洛法则（Dollo’s principle）。本研究结果同时提示，鳅超科的其他类群中或许也存在胃结构。