Evidence for mitochondrial heteroplasmy in the freshwater bug Aphelocheirus aestivalis (Fabricius, 1794) (Insecta: Heteroptera): a challenge for future studies based on mtDNA markers

The mitochondrial genome of Metazoa is predominantly maternally inherited, providing a valuable genetic marker in various fields such as phylogenetics and population genetics. However, mitochondrial heteroplasmy, where multiple copies of mitochondrial DNA (mtDNA) coexist within an individual or even cell, presents a challenge to the use of mtDNA markers. This phenomenon may arise from replication errors, mutagenic processes, or paternal leakage. In this study, we investigated mitochondrial heteroplasmy in the freshwater bug Aphelocheirus aestivalis (Fabricius, 1794) using both Sanger and Illumina sequencing assays. Our analyses confirmed the presence of diverse mtDNA variants within individual specimens collected from various locations across Europe. Signals of mitochondrial heteroplasmy were observed in chromatograms derived from both female and male individuals, indicating a prevalent phenomenon across A. aestivalis populations in general. Illumina sequencing of a single male specimen revealed 102 heteroplasmic sites across the protein coding sequences of mitogenome. Comparative analyses indicated a high similarity between determined COI sequences. Phylogenetic analyses unveiled two distinct clades, potentially originated from different mitogenomes. These findings suggest the widespread and heritable nature of mitochondrial heteroplasmy in A. aestivalis, highlighting its importance for future phylogenetic and phylogeographic studies.

后生动物（Metazoa）的线粒体基因组主要呈母系遗传，为系统发育学、群体遗传学等诸多研究领域提供了极具价值的遗传标记。然而，线粒体异质性——即单个个体乃至单个细胞内共存多种线粒体DNA（mtDNA）拷贝的现象——却为mtDNA标记的应用带来了挑战。该现象可由复制错误、诱变过程或父系渗漏引发。本研究针对淡水水生蝽类盛夏圆胸水蝽（Aphelocheirus aestivalis Fabricius, 1794）的线粒体异质性展开探究，采用桑格测序（Sanger sequencing）与伊卢米纳测序（Illumina sequencing）两种技术手段。分析结果证实，采自欧洲多地的个体标本中存在多样化的mtDNA变异体。雌性与雄性个体的测序峰图均检测到线粒体异质性信号，表明该现象在盛夏圆胸水蝽种群中普遍存在。对单只雄性个体的伊卢米纳测序结果显示，其线粒体基因组蛋白质编码序列中存在102个异质性位点。比较分析表明，本研究测定的细胞色素C氧化酶亚基I（cytochrome c oxidase subunit I, COI）序列具有较高相似性。系统发育分析揭示了两个独立的进化枝，其起源可能与不同的线粒体基因组相关。上述研究结果表明，线粒体异质性在盛夏圆胸水蝽中具有普遍性与可遗传性，凸显了其在未来系统发育与系统地理学研究中的重要意义。