Department of Microbiology & Immunology, Faculty of Medicine, McGill University, Montreal, Quebec, Canada.
Department of Microbiology & Immunology, McGill Genome Centre, Faculty of Medicine, McGill University, Montreal, Quebec, Canada.
Gut Microbes. 2021 Jan-Dec;13(1):1-15. doi: 10.1080/19490976.2021.1903289.
Changes in bacterial diversity in the human gut have been associated with many conditions, despite not always reflecting changes in bacterial activity. Methods linking bacterial identity to function are needed for improved understanding of how bacterial communities adapt and respond to their environment, including the gut. Here, we optimized bioorthogonal non-canonical amino acid tagging (BONCAT) for the gut microbiota and combined it with fluorescently activated cell sorting and sequencing (FACS-Seq) to identify the translationally active members of the community. We then used this novel technique to compare with other bulk community measurements of activity and viability: relative nucleic acid content and membrane damage. The translationally active bacteria represent about half of the gut microbiota, and are not distinct from the whole community. The high nucleic acid content bacteria also represent half of the gut microbiota, but are distinct from the whole community and correlate with the damaged subset. Perturbing the community with xenobiotics previously shown to alter bacterial activity but not diversity resulted in stronger changes in the distinct physiological fractions than in the whole community. BONCAT is a suitable method to probe the translationally active members of the gut microbiota, and combined with FACS-Seq, allows for their identification. The high nucleic acid content bacteria are not necessarily the protein-producing bacteria in the community; thus, further work is needed to understand the relationship between nucleic acid content and bacterial metabolism in the human gut. Considering physiologically distinct subsets of the gut microbiota may be more informative than whole-community profiling.
尽管肠道中细菌多样性的变化并不总是反映细菌活性的变化，但与许多疾病有关。为了更好地理解细菌群落如何适应和响应其环境，包括肠道环境，需要将细菌身份与功能联系起来的方法。在这里，我们优化了用于肠道微生物群的生物正交非天然氨基酸标记（BONCAT），并将其与荧光激活细胞分选和测序（FACS-Seq）相结合，以鉴定群落中具有翻译活性的成员。然后，我们使用这项新技术与其他群体测量的活性和生存能力进行比较：相对核酸含量和膜损伤。翻译活性细菌约占肠道微生物群的一半，与整个群落没有区别。高核酸含量的细菌也占肠道微生物群的一半，但与整个群落不同，与受损亚群相关。用先前被证明可以改变细菌活性而不改变多样性的外源性物质扰乱群落，与整个群落相比，明显的生理部分发生了更强的变化。BONCAT 是一种合适的方法，可以探测肠道微生物群中的翻译活性成员，并且与 FACS-Seq 相结合，可以对其进行鉴定。高核酸含量的细菌不一定是群落中产生蛋白质的细菌；因此，需要进一步研究来了解人类肠道中核酸含量与细菌代谢之间的关系。考虑肠道微生物群的生理上不同的子集可能比整个群落分析更具信息量。
