BACKGROUND Aspergillus spp. comprises a very diverse group of lower eukaryotes with a high relevance for industrial applications and clinical implications. These multinucleate species are often cultured for many generations in the laboratory, which can unknowingly propagate hidden genetic mutations. To assess the likelihood of such events, we studied the genome stability of aspergilli by using a combination of mutation accumulation (MA) lines and whole genome sequencing. RESULTS We sequenced the whole genomes of 30 asexual and 10 sexual MA lines of three Aspergillus species (A. flavus, A. fumigatus and A. nidulans) and estimated that each MA line accumulated mutations for over 4000 mitoses during asexual cycles. We estimated mutation rates of 4.2 × 10-11 (A. flavus), 1.1 × 10-11 (A. fumigatus) and 4.1 × 10-11 (A. nidulans) per site per mitosis, suggesting that the genomes are very robust. Unexpectedly, we found a very high rate of GC → TA transversions only in A. flavus. In parallel, 30 asexual lines of the non-homologous end-joining (NHEJ) mutants of the three species were also allowed to accumulate mutations for the same number of mitoses. Sequencing of these NHEJ MA lines gave an estimated mutation rate of 5.1 × 10-11 (A. flavus), 2.2 × 10-11 (A. fumigatus) and 4.5 × 10-11 (A. nidulans) per base per mitosis, which is slightly higher than in the wild-type strains and some ~ 5-6 times lower than in the yeasts. Additionally, in A. nidulans, we found a NHEJ-dependent interference of the sexual cycle that is independent of the accumulation of mutations. CONCLUSIONS We present for the first time direct counts of the mutation rate of filamentous fungal species and find that Aspergillus genomes are very robust. Deletion of the NHEJ machinery results in a slight increase in the mutation rate, but at a rate we suggest is still safe to use for biotechnology purposes. Unexpectedly, we found GC→TA transversions predominated only in the species A. flavus, which could be generated by the hepatocarcinogen secondary metabolite aflatoxin. Lastly, a strong effect of the NHEJ mutation in self-crossing was observed and an increase in the mutations of the asexual lines was quantified.

中文翻译：

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进化的曲霉种群的基因组测序揭示了健壮的基因组，黄曲霉的转化和非同源末端连接突变体的性畸变。

背景技术曲霉属。包含各种不同的低等真核生物，与工业应用和临床意义高度相关。这些多核物种通常在实验室中培养了好几代，可以在不知不觉中繁殖出隐藏的基因突变。为了评估此类事件的可能性，我们通过使用突变累积（MA）系和全基因组测序的组合研究了曲霉的基因组稳定性。结果我们对三种曲霉属物种（黄曲霉，烟曲霉和构巢曲霉）的30个无性和10个有性MA系的整个基因组进行了测序，并估计每个MA系在无性周期中积累了超过4000个有丝分裂的突变。我们估计每个有丝分裂每个位点的突变率分别为4.2×10-11（黄曲霉），1.1×10-11（烟曲霉）和4.1×10-11（构巢曲霉），这表明基因组非常健壮。出乎意料的是，我们仅在黄曲霉中发现了很高的GC→TA转化率。同时，还允许三个物种的30个无性末端连接（NHEJ）突变体的无性系积累相同数目有丝分裂的突变。这些NHEJ MA品系的测序估计每个有丝分裂每个碱基的突变率为5.1×10-11（黄曲霉），2.2×10-11（烟曲霉）和4.5×10-11（构巢曲霉）。它比野生型菌株略高，比酵母低约5-6倍。此外，在构巢曲霉中，我们发现了性周期的NHEJ依赖性干扰，其独立于突变的积累。结论我们首次提出了丝状真菌物种突变率的直接计数，并发现曲霉基因组非常健壮。删除NHEJ机器会导致突变率略有增加，但是我们建议仍可以安全地将其用于生物技术目的。出乎意料的是，我们发现GC→TA转化仅在黄曲霉菌种中占主导地位，而黄曲霉菌可能是由肝致癌物次生代谢产物黄曲霉毒素产生的。最后，观察到NHEJ突变在自交中有很强的作用，并且定量了无性系突变的增加。我们发现GC→TA的转化仅在黄曲霉菌中占主导地位，这可能是由肝癌原次级代谢产物黄曲霉毒素产生的。最后，观察到NHEJ突变在自交中有很强的作用，并且定量了无性系突变的增加。我们发现GC→TA的转化仅在黄曲霉菌中占主导地位，这可能是由肝癌原次级代谢产物黄曲霉毒素产生的。最后，观察到NHEJ突变在自交中有很强的作用，并且定量了无性系突变的增加。