Once in a while, a conceptual or technical breakthrough changes the way we approach science. In microbiology, Leeuwenhoek’s lenses, Koch’s solid growth medium, Fleming’s penicillin, and Woese’s recognition of the 16S rRNA sequence as a universal chronometer all changed the way discoveries were made, each providing a new way to study or understand microbial life. Following Woese’s groundbreaking insight, Pace applied the polymerase chain reaction to the 16S rRNA gene, making it possible to characterize the molecular diversity of microorganisms in natural environments without culturing bias.(1) The cascade of publications that followed made it clear that microbiologists were ignorant about the staggering diversity that had eluded detection by the culture-based methods that had dominated microbiology research since Koch’s 19th century innovation. Also made clear was the fact that many of the as-yet unculturable organisms were deeply divergent from those known from culture, presenting an opportunity to redefine the limits of microbial life. Pace then proposed that it was time to extract and clone DNA directly from environmental samples to conduct genomic analysis on the cornucopia of organisms that could not be cultured. This approach eventually became known as metagenomics—the analysis of collective genomes extracted from an environment. For the past 20 years, metagenomics research has primarily focused on sequence-based information and only a small proportion has been dedicated to functional metagenomics, which involves expressing DNA extracted from an environmental sample in a surrogate host to discover new metabolites and proteins.(2−4) Functional metagenomics has been plagued by the hurdle of heterologous gene expression, and consequently, progress has been slow. Recently, Sugimoto et al.(5) presented a game-changing approach to functional metagenomics that has the potential to revolutionize discovery. Sugimoto et al. offer two innovations in the search for gene function and new secondary metabolites encoded in microbial genomes. First, they present a major step forward in the characterization of biosynthetic pathways directly from sequencing reads, rather than from metagenomic assemblies, allowing them to avoid the fragmentation that often arises from variable coverage of shotgun community sequencing. Working directly with sequencing reads also allowed them to explore new taxonomic and functional genomic space and mitigated the uneven sampling of previously sequenced or culturable organisms (biased toward the Western, heathy human gut). Their new algorithm, designated MetaBGC, identifies biosynthetic gene cluster (BGC) reads from shotgun metagenomes with systematically tested and tuned sequence-scoring models that work in read-length sized chunks. Reads are binned for targeted assembly enabling reconstitution of entire pathways. The authors first focus on pathways containing type II polyketide synthases (T2PKSs) from a variety of human microbiome databases and show impressive results, even at very low coverage and abundance. Importantly, their method facilitates easy separation of true-positive and false-positive BGC bins. Looking across 3203 human metagenome samples from all major human body sites, the authors identify many new T2PKS BGCs, many from organisms not found in reference databases. BGC abundance was then correlated to body site, taxonomy of the organism of origin, and metatranscriptome information to provide further insight into the diversity and distribution of chemistry in the human microbiome. Sugimoto et al. focus on two novel BGC pathways for expression and chemical structure elucidation. The first, BGC3 from the oral microbiome, was found in a strain that had not yet been cultured. Another innovation in the work is that the authors opted to synthesize the entire BGC, inserting strong promoters and optimizing codon usage for expression in Streptomyces, and expressed the pathway in the heterologous host Staphylococcus albus. The resulting suite of molecules, dubbed metamycins, show strong inhibitory activity against Gram-positive isolates, especially those isolated from the human oral cavity (Figure 1). Transcriptomics analysis showed that the metamycin BGC is expressed under physiological conditions in human plaque during early biofilm formation. A second novel pathway, BGC6 from the gut microbiome, was also found in a cultured isolate. The authors amplified the BGC from Blautia wexlerae and expressed it in Bacillus subtilis to discover wexrubicin, a tetracyclic anthracycline. Unlike other anthracyclines (e.g., doxyrubicin), wexrubicin displays no toxicity in human cell assays. Figure 1. Innovation in metagenomics from Donia and colleagues uncovers new chemical diversity in the microbiome. New molecules, metamycin C and D, are active against bacteria sourced from the human microbiome at levels similar to that of the antibiotic tetracycline. MIC-A is the minimum inhibitory concentration on solid media. The discoveries reported by Sugimoto et al. advance natural products chemistry and drug discovery from uncultured organisms to a new level by making it more routine to access the biosynthetic machinery of the microbiome. But the work is equally impactful on microbial ecology. By returning to the source of the metagenome and examining expression of the pathways discovered in vitro, Sugimoto et al. have been able to propose ecological roles for the newly discovered molecules in the microbiome of origin. The metamycin biosynthetic pathway is expressed in the oral microbiome during initial biofilm formation, and the metamycins target Gram-positive bacteria in the human oral microbiome. The contributions to both chemical discovery and ecology make this research a turning point in understanding the diversity and roles of microbial metabolites in the microbial communities in which they are produced. Despite advances in molecular biology and sequencing over the past four decades, our understanding of microbiomes and the genes and molecules underlying microbial interaction networks remains in its infancy. The astounding diversity of microbial life begets exponential chemical complexity in the communities that comprise it, inviting microbiologists and chemists to unravel the structures, activities, and ecological roles of the cornucopia of compounds encoded in these communities’ metagenomes. We have learned that the intricate networks underpinning microbial community interactions form miniature versions of Darwin’s tangled bank, but work has been stymied by the inaccessibility of the tangible products of many functions encoded in metagenomes. Just as microscopy, culturing, antibiotics, and culture-independent analyses of microbes have all revolutionized microbiology, Sugimoto et al.’s advance will accelerate understanding microbiome functions in many biological processes, including global nutrient cycling, agriculture, and human health. The authors are thankful for Army Research Office MURI Grant W911NF1910269. The authors declare no competing financial interest. This article references 5 other publications.

中文翻译：

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从元基因组到分子：功能元基因组学的创新揭示了人类微生物组中的隐藏化学。

有时，概念或技术上的突破会改变我们对待科学的方式。在微生物学上，Leeuwenhoek的镜片，Koch的固体生长培养基，Fleming的青霉素以及Woese将16S rRNA序列识别为通用天文钟表，都改变了发现的方式，每一个都为研究或理解微生物生活提供了新的途径。遵循Woese的开创性见解，Pace将聚合酶链反应应用于16S rRNA基因，从而可以表征自然环境中微生物的分子多样性而无需培养偏见。（1）随后的一系列出版物明确表明微生物学家是无知的自从科赫19世纪创新以来，基于微生物的研究一直主导着基于文化的方法，而这些方法却难以发现。还明确的事实是，许多尚未培养的生物与文化已知的生物有很大差异，为重新定义微生物生命的极限提供了机会。佩斯随后提出，是时候直接从环境样品中提取和克隆DNA，对无法培养的生物的聚宝盆进行基因组分析了。这种方法最终被称为宏基因组学-分析从环境中提取的集体基因组。在过去的20年中，宏基因组学的研究主要集中在基于序列的信息上，只有一小部分致力于功能宏基因组学，这涉及在替代宿主中表达从环境样品中提取的DNA以发现新的代谢产物和蛋白质。（2-4）功能宏基因组学受到异源基因表达障碍的困扰，因此进展缓慢。Sugimoto等人（5）最近提出了一种功能宏基因组学改变游戏规则的方法，它有可能彻底改变发现。Sugimoto等。在寻找基因功能和微生物基因组中编码的新的次生代谢产物方面提供了两项创新。首先，它们代表了直接从测序读物而不是宏基因组学组件表征生物合成途径方面迈出的重要一步，从而使它们避免了因shot弹枪社区测序的可变覆盖而经常造成的碎片化。直接与测序读片一起工作还使他们能够探索新的分类学和功能基因组空间，并减轻了先前测序或可培养生物的不均匀采样（偏向于西部，健康的人类肠道）。他们的新算法称为MetaBGC，它通过系统测试和调整的序列计分模型（可用于读取长度大小的片段），识别identifies弹枪基因组中的生物合成基因簇（BGC）读数。读段被分装以进行有针对性的组装，从而可以重建整个途径。作者首先关注了来自各种人类微生物组数据库的包含II型聚酮化合物合酶（T2PKS）的途径，即使在极低的覆盖率和丰度下也显示出令人印象深刻的结果。重要的是，他们的方法有助于轻松区分真阳性和假阳性BGC分档。作者查看了来自所有主要人体部位的3203个人类元基因组样本，确定了许多新的T2PKS BGC，其中许多来自参考数据库中找不到的生物。然后将BGC的丰度与人体部位，起源生物的分类学和转录组信息相关联，以进一步了解人类微生物组中化学的多样性和分布。Sugimoto等。专注于表达和化学结构阐明的两个新颖的BGC途径。第一种是来自口腔微生物组的BGC3，是在尚未培养的菌株中发现的。这项工作的另一个创新之处是，作者选择了合成整个BGC，插入强启动子并优化密码子用法以在BGC中表达。许多来自参考数据库中找不到的生物。然后将BGC的丰度与人体部位，起源生物的分类学和转录组信息相关联，以进一步了解人类微生物组中化学的多样性和分布。Sugimoto等。专注于表达和化学结构阐明的两个新颖的BGC途径。第一种是来自口腔微生物组的BGC3，是在尚未培养的菌株中发现的。这项工作的另一个创新之处是，作者选择了合成整个BGC，插入强启动子并优化密码子用法以在BGC中表达。许多来自参考数据库中找不到的生物。然后将BGC的丰度与人体部位，起源生物的分类学和转录组信息相关联，以进一步了解人类微生物组中化学的多样性和分布。Sugimoto等。专注于表达和化学结构阐明的两个新颖的BGC途径。第一种是来自口腔微生物组的BGC3，是在尚未培养的菌株中发现的。这项工作的另一个创新之处是，作者选择了合成整个BGC，插入强启动子并优化密码子用法以在BGC中表达。以及转录组信息，以进一步了解人类微生物组中化学的多样性和分布。Sugimoto等。专注于表达和化学结构阐明的两个新颖的BGC途径。第一种是来自口腔微生物组的BGC3，是在尚未培养的菌株中发现的。这项工作的另一个创新之处在于，作者选择了合成整个BGC，插入强启动子并优化密码子用法以在BGC中表达。和转录组信息，以进一步了解人类微生物组中化学的多样性和分布。Sugimoto等。专注于表达和化学结构阐明的两个新颖的BGC途径。第一种是来自口腔微生物组的BGC3，是在尚未培养的菌株中发现的。这项工作的另一个创新之处是，作者选择了合成整个BGC，插入强启动子并优化密码子用法以在BGC中表达。链霉菌，并在异源宿主金黄色葡萄球菌中表达了该途径。所得的分子分子称为“间霉素”，对革兰氏阳性分离株，特别是从人口腔分离出的分离株，显示出较强的抑制活性（图1）。转录组学分析表明，在生物膜早期形成过程中，斑霉素的BGC在生理条件下在人斑中表达。在培养的分离物中还发现了第二种新途径，即肠道微生物组的BGC6。作者扩增了来自Blautia wxlerae的BGC，并将其在枯草芽孢杆菌中表达发现四环蒽环霉素。与其他蒽环类抗生素（例如强力霉素）不同，韦柔比星在人体细胞试验中未显示毒性。图1. Donia及其同事的宏基因组学创新发现了微生物组中新的化学多样性。新分子，新霉素C和D，以与抗生素四环素相似的水平对源自人类微生物组的细菌具有活性。MIC-A是在固体培养基上的最低抑制浓度。Sugimoto等报道的发现。通过使日常使用微生物组的生物合成机制更加常规化，可以将未培养生物的天然产物化学和药物开发提高到一个新水平。但是这项工作对微生物生态学同样具有影响。Sugimoto等人通过返回到元基因组的来源并检查了体外发现的途径的表达。能够为起源的微生物组中新发现的分子提出生态作用。在最初的生物膜形成过程中，口腔细菌组中表达了表达菌素的生物合成途径，并且该真菌素靶向人口腔微生物组中的革兰氏阳性细菌。对化学发现和生态学的贡献使本研究成为了解微生物代谢产物在其产生的微生物群落中的多样性和作用的转折点。尽管在过去的四十年中分子生物学和测序技术取得了进步，但我们对微生物组以及微生物相互作用网络背后的基因和分子的了解仍处于起步阶段。令人震惊的微生物生命多样性在组成其的社区中引起了指数级的化学复杂性，从而邀请微生物学家和化学家揭示这些社区的基因组中编码的聚宝盆的结构，活性和生态作用。我们已经了解到，支撑微生物群落相互作用的复杂网络形成了达尔文缠结银行的微型版本，但是由于元基因组中编码的许多功能的有形产物无法获得而使工作陷入了困境。正如微生物的显微镜检查，培养，抗生素和非培养分析都彻底改变了微生物学一样，Sugimoto等人的研究进展将加速人们对许多生物过程中微生物组功能的理解，包括全球营养循环，农业和人类健康。作者感谢陆军研究办公室MURI Grant W911NF1910269。作者宣称没有竞争性的经济利益。本文引用了其他5个出版物。