Contribution of sample processing to gut microbiome analysis in the model Lepidoptera, silkworm Bombyx mori,Computational and Structural Biotechnology Journal

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Contribution of sample processing to gut microbiome analysis in the model Lepidoptera, silkworm Bombyx mori
Computational and Structural Biotechnology Journal ( IF 4.4 ) Pub Date : 2021-08-17 , DOI: 10.1016/j.csbj.2021.08.020
Nan Zhang ₁ , Jintao He ₁ , Xiaoqiang Shen ₁ , Chao Sun ₂ , Abrar Muhammad ₁ , Yongqi Shao _{1,

3}

Affiliation

Microbes that live inside insects play various roles in host biology, ranging from nutrient supplementation to host defense. Although Lepidoptera (butterflies and moths) are one of the most diverse insect taxa and important in natural ecosystems, their microbiotas are little-studied, and to understand their structure and function, it is necessary to identify potential factors that affect microbiome analysis. Using a model organism, the silkworm , we investigated the effects of different sample types (whole gut, gut content, gut tissue, starvation, or frass) and metagenomic DNA extraction methodologies (small-scale versus large-scale) on the composition and diversity of the caterpillar gut microbial communities. High-throughput 16S rRNA gene sequencing and computational analysis of the resulting data unraveled that DNA extraction has a large effect on the outcome of metagenomic analysis: significant biases were observed in estimates of community diversity and in the ratio between Gram-positive and Gram-negative bacteria. Furthermore, bacterial communities differed significantly among sample types. The gut content and whole gut samples differed least, both had a higher percentage of and species; whereas the frass and starvation samples differed substantially from the whole gut and were poor representatives of the gut microbiome. Thus, we recommend a small-scale DNA extraction methodology for sampling the whole gut under normal insect rearing conditions whenever possible, as this approach provides the most accurate assessment of the gut microbiome. Our study highlights that evaluation of the optimal sample-processing approach should be the first step taken to confidently assess the contributions of microbiota to Lepidoptera.

中文翻译：

样品处理对鳞翅目、家蚕模型肠道微生物组分析的贡献

生活在昆虫体内的微生物在宿主生物学中发挥着各种作用，从营养补充到宿主防御。尽管鳞翅目（蝴蝶和飞蛾）是最多样化的昆虫类群之一，并且在自然生态系统中很重要，但对其微生物群的研究却很少，为了了解其结构和功能，有必要确定影响微生物组分析的潜在因素。使用模式生物——蚕，我们研究了不同样本类型（全肠道、肠道内容物、肠道组织、饥饿或粪便）和宏基因组 DNA 提取方法（小规模与大规模）对组成和多样性的影响毛毛虫肠道微生物群落。高通量 16S rRNA 基因测序和所得数据的计算分析揭示了 DNA 提取对宏基因组分析的结果有很大影响：在群落多样性的估计以及革兰氏阳性和革兰氏阴性之间的比率中观察到显着偏差细菌。此外，不同样本类型的细菌群落存在显着差异。肠道内容物和全肠道样本差异最小，两者都含有较高比例的和物种；而粪便和饥饿样本与整个肠道有很大不同，不能很好地代表肠道微生物组。因此，我们建议尽可能在正常昆虫饲养条件下采用小规模 DNA 提取方法对整个肠道进行采样，因为这种方法可以提供对肠道微生物组最准确的评估。我们的研究强调，评估最佳样品处理方法应该是自信地评估微生物群对鳞翅目的贡献的第一步。

更新日期：2021-08-17

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