Progress of analytical tools and techniques for human gut microbiome research

Massive DNA sequencing studies have expanded our insights and understanding of the ecological and functional characteristics of the gut microbiome. Advanced sequencing technologies allow us to understand the close association of the gut microbiome with human health and critical illnesses. In the future, analyses of the gut microbiome will provide key information associating with human individual health, which will help provide personalized health care for diseases. Numerous molecular biological analysis tools have been rapidly developed and employed for the gut microbiome researches; however, methodological differences among researchers lead to inconsistent data, limiting extensive share of data. It is therefore very essential to standardize the current methodologies and establish appropriate pipelines for human gut microbiome research. Herein, we review the methods and procedures currently available for studying the human gut microbiome, including fecal sample collection, metagenomic DNA extraction, massive DNA sequencing, and data analyses with bioinformatics. We believe that this review will contribute to the progress of gut microbiome research in the clinical and practical aspects of human health.     

The Human Mycobiome and its Impact on Health and Disease

Current Fungal Infection Reports - Until recently, the human microbiome was synonymous with the bacterial community. Fortunately, recent studies have shown that the human microbiome is complex and...     

Public human microbiome data are dominated by highly developed countries

The importance of sampling from globally representative populations has been well established in human genomics. In human microbiome research, however, we lack a full understanding of the global distribution of sampling in research studies. This information is crucial to better understand global patterns of microbiome-associated diseases and to extend the health benefits of this research to all populations. Here, we analyze the country of origin of all 444,829 human microbiome samples that are available from the world’s 3 largest genomic data repositories, including the Sequence Read Archive (SRA). The samples are from 2,592 studies of 19 body sites, including 220,017 samples of the gut microbiome. We show that more than 71% of samples with a known origin come from Europe, the United States, and Canada, including 46.8% from the US alone, despite the country representing only 4.3% of the global population. We also find that central and southern Asia is the most underrepresented region: Countries such as India, Pakistan, and Bangladesh account for more than a quarter of the world population but make up only 1.8% of human microbiome samples. These results demonstrate a critical need to ensure more global representation of participants in microbiome studies.

The importance of sampling from globally representative populations has been well established in human genomics, but what about the microbiome? This study shows that metadata from almost half a million samples reveals worldwide human microbiome research is skewed heavily in favor of Europe and North America and excludes large but less developed nations in Asia and Africa.     

An emerging field: Post-translational modification in microbiome

Post-translational modifications (PTMs) play an essential role in most biological processes. PTMs on human proteins have been extensively studied. Studies on bacterial PTMs are emerging, which demonstrate that bacterial PTMs are different from human PTMs in their types, mechanisms and functions. Few PTM studies have been done on the microbiome. Here, we reviewed several studied PTMs in bacteria including phosphorylation, acetylation, succinylation, glycosylation, and proteases. We discussed the enzymes responsible for each PTM and their functions. We also summarized the current methods used to study microbiome PTMs and the observations demonstrating the roles of PTM in the microbe-microbe interactions within the microbiome and their interactions with the environment or host. Although new methods and tools for PTM studies are still needed, the existing technologies have made great progress enabling a deeper understanding of the functional regulation of the microbiome. Large-scale application of these microbiome-wide PTM studies will provide a better understanding of the microbiome and its roles in the development of human diseases.     

Ökosystem Darm

Ecosystem gut The analysis of the microbiome opened a new chapter in human biology. The composition of the gut microbiome was associated with a variety of human diseases. After an analytic research phase, scientists now search for possibilities of therapeutic interventions. The high complexity of the system and the relative lack of prospective studies make it difficult to differentiate causal relationships from mere associations. Targeted health-promoting modulations of the gut microbiome are still difficult.     

Experimental design and quantitative analysis of microbial community multiomics

Studies of the microbiome have become increasingly sophisticated, and multiple sequence-based, molecular methods as well as culture-based methods exist for population-scale microbiome profiles. To link the resulting host and microbial data types to human health, several experimental design considerations, data analysis challenges, and statistical epidemiological approaches must be addressed. Here, we survey current best practices for experimental design in microbiome molecular epidemiology, including technologies for generating, analyzing, and integrating microbiome multiomics data. We highlight studies that have identified molecular bioactives that influence human health, and we suggest steps for scaling translational microbiome research to high-throughput target discovery across large populations.     

Proceedings from the 3rd International Conference on Microbiome Engineering

The human microbiome has been inextricably linked to multiple facets of human physiology. From an engineering standpoint, the ability to precisely control the composition and activity of the microbiome holds great promise for furthering our understanding of disease etiology and for new avenues of therapeutic and diagnostic agents. While the field of microbiome research is still in its infancy, growing engineering efforts are emerging to enable new studies in the microbiome and to rapidly translate these findings to microbiome-based interventions. At the 3rd International Conference on Microbiome Engineering, leading experts in the field presented state-of-the-art work in microbiome engineering, discussing probiotics, prebiotics, engineered microbes, microbially derived biomolecules, and bacteriophage.     

人体肠道病毒组学研究进展

人体微生物组计划开展近10年来,大量的研究显示人体微生物通过各种方式深刻地影响着人体健康。人体肠道内丰富多样的病毒构成了肠道病毒组,是人体微生物组的重要组成部分,和人体健康密切相关。本文综述了近些年国际上人体肠道病毒组研究的最新进展,分别从人体肠道病毒组的组成特征、肠道病毒组-细菌组-人体间的相互作用及其对人体健康的影响、病毒组研究的技术策略及挑战等方面进行了论述,探讨了肠道病毒组在人体疾病预防和治疗领域应用的可行性。     

Rapidly expanding knowledge on the role of the gut microbiome in health and disease

The human gut is colonized by a wide diversity of micro-organisms, which are now known to play a key role in the human host by regulating metabolic functions and immune homeostasis. Many studies have indicated that the genomes of our gut microbiota, known as the gut microbiome or our “other genome” could play an important role in immune-related, complex diseases, and growing evidence supports a causal role for gut microbiota in regulating predisposition to diseases. A comprehensive analysis of the human gut microbiome is thus important to unravel the exact mechanisms by which the gut microbiota are involved in health and disease. Recent advances in next-generation sequencing technology, along with the development of metagenomics and bioinformatics tools, have provided opportunities to characterize the microbial communities. Furthermore, studies using germ-free animals have shed light on how the gut microbiota are involved in autoimmunity. In this review we describe the different approaches used to characterize the human microbiome, review current knowledge about the gut microbiome, and discuss the role of gut microbiota in immune homeostasis and autoimmunity. Finally, we indicate how this knowledge could be used to improve human health by manipulating the gut microbiota. This article is part of a Special Issue entitled: From Genome to Function.     

非人灵长类肠道微生物研究进展与展望

肠道微生物组被誉为动物的“第二套基因组”,与动物的个体发育、营养获取、生理功能、免疫调节等重要活动密切相关。非人灵长类在生态位、社会结构、地理分布以及进化上与人类相近,开展其肠道微生物研究不仅有助于了解灵长类的生态、保护和进化,而且对深入了解肠道微生物在人类进化中所发挥的作用也具有重要的参考价值。本文总结了影响非人灵长类肠道微生物变化的因素,包括系统发育、觅食、栖息地破碎化、年龄和性别、圈养方式以及社群生活,并探讨了肠道微生物研究在非人灵长类生态、行为、保护以及适应性进化方面的应用。未来,非人灵长类肠道微生物研究将为灵长类生态、进化和人类健康的研究提供新的视角,为灵长类的保护提供新的理论基础和研究方法。     

Micro-coevolution of host genetics with gut microbiome in three Chinese ethnic groups

Understanding the micro-coevolution of the human gut microbiome with host genetics is challenging but essential in both evolutionary and medical studies. To gain insight into the interactions between host genetic variation and the gut microbiome, we analyzed both the human genome and gut microbiome collected from a cohort of 190 students in the same boarding college and representing 3 ethnic groups, Uyghur, Kazakh, and Han Chinese. We found that differences in gut microbiome were greater between genetically distinct ethnic groups than those genetically closely related ones in taxonomic composition, functional composition, enterotype stratification, and microbiome genetic differentiation. We also observed considerable correlations between host genetic variants and the abundance of a subset of gut microbial species. Notably, interactions between gut microbiome species and host genetic variants might have coordinated effects on specific human phenotypes. Bacteroides ovatus, previously reported to modulate intestinal immunity, is significantly correlated with the host genetic variant rs12899811 (meta-P = 5.55 × 10⁻⁵), which regulates the VPS33B expression in the colon, acting as a tumor suppressor of colorectal cancer. These results advance our understanding of the micro-coevolution of the human gut microbiome and their interactive effects with host genetic variation on phenotypic diversity.     

Composition and stability of the vervet monkey milk microbiome

The human milk microbiome is vertically transmitted to offspring during the postnatal period and has emerged as a critical driver of infant immune and metabolic development. Despite this importance in humans, the milk microbiome of nonhuman primates remains largely unexplored. This dearth of comparative work precludes our ability to understand how species‐specific differences in the milk microbiome may differentially drive maternal effects and limits how translational models can be used to understand the role of vertically transmitted milk microbes in human development. Here, we present the first culture‐independent data on the milk microbiome of a nonhuman primate. We collected milk and matched fecal microbiome samples at early and late lactation from a cohort of captive lactating vervet monkeys (N = 15). We found that, similar to humans, the vervet monkey milk microbiome comprises a shared community of taxa that are universally present across individuals. However, unlike in humans, this shared community is dominated by the genera Lactobacillus, Bacteroides, and Prevotella. We also found that, in contrast to previous culture‐dependent studies in humans, the vervet milk microbiome exhibits greater alpha‐diversity than the gut microbiome across lactation. Finally, we did not find support for the translocation of microbes from the gut to the mammary gland within females (i.e., “entero‐mammary pathway”). Taken together, our results show that the vervet monkey milk microbiome is taxonomically diverse, distinct from the gut microbiome, and largely stable. These findings demonstrate that the milk microbiome is a unique substrate that may selectively favor the establishment and persistence of particular microbes across lactation and highlights the need for future experimental studies on the origin of microbes in milk.     

植物多糖对肠道微生态的作用研究进展

近年来,随着微生态学的发展,越来越多的学者认识到肠道微生态平衡对人体健康的重要意义。许多研究表明植物多糖作为肠道微生态调节剂,具有调节肠道菌群比例、促进肠黏膜修复、增强肠黏膜分泌型免疫球蛋白表达及调节细胞因子水平等作用。除此以外,植物多糖具有来源广泛、成本低廉、作用效果好等优势。本文归纳了近年来报道的具有肠道微生态调节功能的植物多糖及其研究方法和作用机制等,希望为今后的研究重点和发展方向提供参考。     

Our microbial selves: what ecology can teach us

Advances in DNA sequencing have allowed us to characterize microbial communities--including those associated with the human body--at a broader range of spatial and temporal scales than ever before. We can now answer fundamental questions that were previously inaccessible and use well-tested ecological theories to gain insight into changes in the microbiome that are associated with normal development and human disease. Perhaps unsurprisingly, the ecosystems associated with our body follow trends identified in communities at other sites and scales, and thus studies of the microbiome benefit from ecological insight. Here, we assess human microbiome research in the context of ecological principles and models, focusing on diversity, biological drivers of community structure, spatial patterning and temporal dynamics, and suggest key directions for future research that will bring us closer to the goal of building predictive models for personalized medicine.     

Contrasting Strategies: Human Eukaryotic Versus Bacterial Microbiome Research

Most discussions of human microbiome research have focused on bacterial investigations and findings. Our target is to understand how human eukaryotic microbiome research is developing, its potential distinctiveness, and how problems can be addressed. We start with an overview of the entire eukaryotic microbiome literature (578 papers), show tendencies in the human‐based microbiome literature, and then compare the eukaryotic field to more developed human bacterial microbiome research. We are particularly concerned with problems of interpretation that are already apparent in human bacterial microbiome research (e.g. disease causality, probiotic interventions, evolutionary claims). We show where each field converges and diverges, and what this might mean for progress in human eukaryotic microbiome research. Our analysis then makes constructive suggestions for the future of the field.     

Metaproteomic and Metabolomic Approaches for Characterizing the Gut Microbiome

The gut microbiome has been shown to play a significant role in human healthy and diseased states. The dynamic signaling that occurs between the host and microbiome is critical for the maintenance of host homeostasis. Analyzing the human microbiome with metaproteomics, metabolomics, and integrative multi‐omics analyses can provide significant information on markers for healthy and diseased states, allowing for the eventual creation of microbiome‐targeted treatments for diseases associated with dysbiosis. Metaproteomics enables functional activity information to be gained from the microbiome samples, while metabolomics provides insight into the overall metabolic states affecting/representing the host–microbiome interactions. Combining these functional ‐omic platforms together with microbiome composition profiling allows for a holistic overview on the functional and metabolic state of the microbiome and its influence on human health. Here the benefits of metaproteomics, metabolomics, and the integrative multi‐omic approaches to investigating the gut microbiome in the context of human health and diseases are reviewed.     

Inter-Individual Differences in the Oral Bacteriome Are Greater than Intra-Day Fluctuations in Individuals

Given the advent of massively parallel DNA sequencing, human microbiome is analyzed comprehensively by metagenomic approaches. However, the inter- and intra-individual variability and stability of the human microbiome remain poorly characterized, particularly at the intra-day level. This issue is of crucial importance for studies examining the effects of microbiome on human health. Here, we focused on bacteriome of oral plaques, for which repeated, time-controlled sampling is feasible. Eighty-one supragingival plaque subjects were collected from healthy individuals, examining multiple sites within the mouth at three time points (forenoon, evening, and night) over the course of 3 days. Bacterial composition was estimated by 16S rRNA sequencing and species-level profiling, resulting in identification of a total of 162 known bacterial species. We found that species compositions and their relative abundances were similar within individuals, and not between sampling time or tooth type. This suggests that species-level oral bacterial composition differs significantly between individuals, although the number of subjects is limited and the intra-individual variation also occurs. The majority of detected bacterial species (98.2%; 159/162), however, did not fluctuate over the course of the day, implying a largely stable oral microbiome on an intra-day time scale. In fact, the stability of this data set enabled us to estimate potential interactions between rare bacteria, with 40 co-occurrences supported by the existing literature. In summary, the present study provides a valuable basis for studies of the human microbiome, with significant implications in terms of biological and clinical outcomes.     

An Improved Methodology to Overcome Key Issues in Human Fecal Metagenomic DNA Extraction

Microbes are ubiquitously distributed in nature, and recent culture-independent studies have highlighted the significance of gut microbiota in human health and disease. Fecal DNA is the primary source for the majority of human gut microbiome studies. However, further improvement is needed to obtain fecal metagenomic DNA with sufficient amount and good quality but low host genomic DNA contamination. In the current study, we demonstrate a quick, robust, unbiased,and cost-effective method for the isolation of high molecular weight(23 kb) metagenomic DNA(260/280 ratio 1.8) with a good yield(55.8 ± 3.8 ng/mg of feces). We also confirm that there is very low human genomic DNA contamination(eubacterial: human genomic DNA marker genes = 2~(27.9):1) in the human feces. The newly-developed method robustly performs for fresh as well as stored fecal samples as demonstrated by 16 S r RNA gene sequencing using 454 FLX+.Moreover, 16 S r RNA gene analysis indicated that compared to other DNA extraction methods tested, the fecal metagenomic DNA isolated with current methodology retains species richnessand does not show microbial diversity biases, which is further confirmed by q PCR with a known quantity of spike-in genomes. Overall, our data highlight a protocol with a balance between quality,amount, user-friendliness, and cost effectiveness for its suitability toward usage for cultureindependent analysis of the human gut microbiome, which provides a robust solution to overcome key issues associated with fecal metagenomic DNA isolation in human gut microbiome studies.     

核心微生物组的研究及利用现状

随着分子生物学和生物信息学的飞速发展,新一代测序技术可以轻松地检测不同样本中复杂的微生物分类单元。面对这些复杂而大量的微生物组数据带来的分析挑战,利用核心微生物组的方法来描述和分析样本中的核心微生物组和关键种是近年来新的研究热点,这些结果将揭示与宿主健康、生长和生产等密切相关的微生物种类,有助于深入认识微生物与宿主间的相互关系,深刻理解微生物对宿主的影响作用,更好地理解微生物组在自然生态系统中的功能。本文阐述了核心微生物组的定义、研究方法、与动植物的关系等方面的研究及利用现状,为更好地利用核心微生物组解决环境、人类健康和农业生产问题提供思路。     

Discovering functional small molecules in the gut microbiome

The human microbiome has emerged as a source of bacterially produced, functional small molecules that help regulate health and disease, and their discovery and annotation has become a popular research topic. Identifying these molecules provides an essential step in unraveling the molecular mechanisms underlying biological outcomes. The relevance of specific bacterial members of the microbiome has been demonstrated in a variety of correlative studies, and there are many possible paths from these correlations to the responsible metabolites. Herein, we summarize two studies that have recently identified gut microbiome metabolites that modulate immune responses or promote physical activity. Aside from the deep insights gained, these studies provide blueprints for successfully uncovering the molecules and mechanisms that control important physiological pathways.