From molecules to dynamic biological communities

Microbial ecology is flourishing, and in the process, is making contributions to how the ecology and biology of large organisms is understood. Ongoing advances in sequencing technology and computational methods have enabled the collection and analysis of vast amounts of molecular data from diverse biological communities. While early studies focused on cataloguing microbial biodiversity in environments ranging from simple marine ecosystems to complex soil ecologies, more recent research is concerned with community functions and their dynamics over time. Models and concepts from traditional ecology have been used to generate new insight into microbial communities, and novel system-level models developed to explain and predict microbial interactions. The process of moving from molecular inventories to functional understanding is complex and challenging, and never more so than when many thousands of dynamic interactions are the phenomena of interest. We outline the process of how epistemic transitions are made from producing catalogues of molecules to achieving functional and predictive insight, and show how those insights not only revolutionize what is known about biological systems but also about how to do biology itself. Examples will be drawn primarily from analyses of different human microbiota, which are the microbial consortia found in and on areas of the human body, and their associated microbiomes (the genes of those communities). Molecular knowledge of these microbiomes is transforming microbiological knowledge, as well as broader aspects of human biology, health and disease.     

Impact of Next Generation Sequencing Techniques in Food Microbiology

Understanding the Maxam-Gilbert and Sanger sequencing as the first generation, in recent years there has been an explosion of newly-developed sequencing strategies, which are usually referred to as next generation sequencing (NGS) techniques. NGS techniques have high-throughputs and produce thousands or even millions of sequences at the same time. These sequences allow for the accurate identification of microbial taxa, including uncultivable organisms and those present in small numbers. In specific applications, NGS provides a complete inventory of all microbial operons and genes present or being expressed under different study conditions. NGS techniques are revolutionizing the field of microbial ecology and have recently been used to examine several food ecosystems. After a short introduction to the most common NGS systems and platforms, this review addresses how NGS techniques have been employed in the study of food microbiota and food fermentations, and discusses their limits and perspectives. The most important findings are reviewed, including those made in the study of the microbiota of milk, fermented dairy products, and plant-, meat- and fish-derived fermented foods. The knowledge that can be gained on microbial diversity, population structure and population dynamics via the use of these technologies could be vital in improving the monitoring and manipulation of foods and fermented food products. They should also improve their safety.     

Symbiont genomics,our new tangled bank

Microbial symbionts inhabit the soma and surfaces of most multicellular species and instigate both beneficial and harmful infections. Despite their ubiquity, we are only beginning to resolve major patterns of symbiont ecology and evolution. Here, we summarize the history, current progress, and projected future of the study of microbial symbiont evolution throughout the tree of life. We focus on the recent surge of data that whole-genome sequencing has introduced into the field, in particular the links that are now being made between symbiotic lifestyle and molecular evolution. Post-genomic and systems biology approaches are also emerging as powerful techniques to investigate host–microbe interactions, both at the molecular level of the species interface and at the global scale. In parallel, next-generation sequencing technologies are allowing new questions to be addressed by providing access to population genomic data, as well as the much larger genomes of microbial eukaryotic symbionts and hosts. Throughout we describe the questions that these techniques are tackling and we conclude by listing a series of unanswered questions in microbial symbiosis that can potentially be addressed with the new technologies.     

Microbiome analysis of dairy cows fed pasture or total mixed ration diets

Understanding rumen microbial ecology is essential for the development of feed systems designed to improve livestock productivity, health and for methane mitigation strategies from cattle. Although rumen microbial communities have been studied previously, few studies have applied next-generation sequencing technologies to that ecosystem. The aim of this study was to characterize changes in microbial community structure arising from feeding dairy cows two widely used diets: pasture and total mixed ration (TMR). Bacterial, archaeal and protozoal communities were characterized by terminal restriction fragment length polymorphism of the amplified SSU rRNA gene and statistical analysis showed that bacterial and archaeal communities were significantly affected by diet, whereas no effect was observed for the protozoal community. Deep amplicon sequencing of the 16S rRNA gene revealed significant differences in the bacterial communities between the diets and between rumen solid and liquid content. At the family level, some important groups of rumen bacteria were clearly associated with specific diets, including the higher abundance of the Fibrobacteraceae in TMR solid samples and members of the propionate-producing Veillonelaceae in pasture samples. This study will be relevant to the study of rumen microbial ecology and livestock feed management.     

Next-generation sequencing data interpretation: enhancing reproducibility and accessibility

Areas of life sciences research that were previously distant from each other in ideology, analysis practices and toolkits, such as microbial ecology and personalized medicine, have all embraced techniques that rely on next-generation sequencing instruments. Yet the capacity to generate the data greatly outpaces our ability to analyse it. Existing sequencing technologies are more mature and accessible than the methodologies that are available for individual researchers to move, store, analyse and present data in a fashion that is transparent and reproducible. Here we discuss currently pressing issues with analysis, interpretation, reproducibility and accessibility of these data, and we present promising solutions and venture into potential future developments.     

微生物分子生态学研究方法的新进展

环境中微生物的群落结构及多样性和微生物的功能及代谢机理是微生物生态学的研究热点,长期以来,由于受到研究技术的限制,对微生物的群落结构和多样性的认识还不全面,微生物的功能及代谢机理方面了解也很少.随着高通量测序、基因芯片等新技术的不断更新,微生物分子生态学的研究方法和研究途径也在不断变化.高通量测序技术改变了微生物多样性、宏基因组学和宏转录组学的研究方法,GeoChip高密度覆盖海量已知功能的基因探针于单张芯片,能快速确定微生物和已知功能基因的存在与否.总结和比较了目前最新的研究手段,并归纳了这些方法的适用性和优缺点.     

Microbes in food processing technology

Abstract: There is an increasing understanding that the microbial quality of a certain food is the result of a chain of events. It is clear that the microbial safety of food can only be guaranteed when the overall processing, including the production of raw materials, distribution and handling by the consumer are taken into consideration. Therefore, the microbiological quality assurance of foods is not only a matter of control, but also of a careful design of the total process chain. Food industry has now generally adapted quality assurance systems and is implementing the Hazard Analysis Critical Control Point (HACCP) concept. Rapid microbiological monitoring systems should be used in these cases. There is a need for rapid and simple microbiological tests which can be adapted to the technology and logistics of specific production processes. Traditional microbiological methods generally do not meet these high requirements. This paper discusses the tests, based on molecular biological principles, to detect and identify microbes in food-processing chains. Tests based on DNA technology are discussed, including in vitro DNA amplification like the polymerase chain reaction (PCR) method and identifications based on RFLP, RAPD and DNA fingerprinting analysis. PCR-haled methodology can be used for the rapid detection of microbes in food manufacturing environments. In addition, DNA fingerprinting methods are suitable for investigating sources and routes of microbial contamination in the food cycle.     

Meeting report: the terabase metagenomics workshop and the vision of an Earth microbiome project

Between July 18(th) and 24(th) 2010, 26 leading microbial ecology, computation, bioinformatics and statistics researchers came together in Snowbird, Utah (USA) to discuss the challenge of how to best characterize the microbial world using next-generation sequencing technologies. The meeting was entitled "Terabase Metagenomics" and was sponsored by the Institute for Computing in Science (ICiS) summer 2010 workshop program. The aim of the workshop was to explore the fundamental questions relating to microbial ecology that could be addressed using advances in sequencing potential. Technological advances in next-generation sequencing platforms such as the Illumina HiSeq 2000 can generate in excess of 250 billion base pairs of genetic information in 8 days. Thus, the generation of a trillion base pairs of genetic information is becoming a routine matter. The main outcome from this meeting was the birth of a concept and practical approach to exploring microbial life on earth, the Earth Microbiome Project (EMP). Here we briefly describe the highlights of this meeting and provide an overview of the EMP concept and how it can be applied to exploration of the microbiome of each ecosystem on this planet.     

单分子测序技术及应用研究进展

从DNA双螺旋结构的发现开始,生命科学研究进入分子水平,在20世纪70年代出现的测序技术为破译遗传密码作出了巨大贡献.近几年出现的单分子测序技术,可以在单个分子水平读取核苷酸序列,也被称为第三代测序技术,主要代表有HeliScope、Nanopore和PacBio等.与传统的第一代和第二代测序技术相比,第三代测序能够产生更长的碱基读长,能直接对RNA进行测序,无需逆转录,测序速度极快,同时其中某些技术所涉及的设备可以小型化,可便携至野外现场测序.第三代测序技术在生命科学基础理论研究及生物医学临床实践中,具有广泛的应用.本文重点介绍了各种单分子测序技术的原理、优缺点,及其应用研究进展.     

Next-generation sequencing reveals significant bacterial diversity of botrytized wine

While wine fermentation has long been known to involve complex microbial communities, the composition and role of bacteria other than a select set of lactic acid bacteria (LAB) has often been assumed either negligible or detrimental. This study served as a pilot study for using barcoded amplicon next-generation sequencing to profile bacterial community structure in wines and grape musts, comparing the taxonomic depth achieved by sequencing two different domains of prokaryotic 16S rDNA (V4 and V5). This study was designed to serve two goals: 1) to empirically determine the most taxonomically informative 16S rDNA target region for barcoded amplicon sequencing of wine, comparing V4 and V5 domains of bacterial 16S rDNA to terminal restriction fragment length polymorphism (TRFLP) of LAB communities; and 2) to explore the bacterial communities of wine fermentation to better understand the biodiversity of wine at a depth previously unattainable using other techniques. Analysis of amplicons from the V4 and V5 provided similar views of the bacterial communities of botrytized wine fermentations, revealing a broad diversity of low-abundance taxa not traditionally associated with wine, as well as atypical LAB communities initially detected by TRFLP. The V4 domain was determined as the more suitable read for wine ecology studies, as it provided greater taxonomic depth for profiling LAB communities. In addition, targeted enrichment was used to isolate two species of Alphaproteobacteria from a finished fermentation. Significant differences in diversity between inoculated and uninoculated samples suggest that Saccharomyces inoculation exerts selective pressure on bacterial diversity in these fermentations, most notably suppressing abundance of acetic acid bacteria. These results determine the bacterial diversity of botrytized wines to be far higher than previously realized, providing further insight into the fermentation dynamics of these wines, and demonstrate the utility of next-generation sequencing for wine ecology studies.     

Resolving functional diversity in relation to microbial community structure in soil: exploiting genomics and stable isotope probing

The microbial ecology of soil still presents a challenge to microbiologists attempting to establish the ways in which bacteria and fungi actively metabolise substrates, link into food webs and recycle plant and animal remains and provide essential nutrients for plants. Extraction and in situ analysis of rRNA has enabled identification of active taxa, and detection of mRNA has provided an insight into the expression of key functional genes in soil. Recent advances in genomic analysis and stable isotope probing are the first steps in resolving the linkage between structure and function in microbial communities.     

全球变化背景下内蒙古草原土壤微生物多样性维持机制研究进展

全球变化对人类环境的影响是近几十年世界广泛关注的热点之一。内蒙古草原不仅是我国重要的牲畜和饲料生产基地, 而且有着不可替代的生态系统功能。土壤微生物是地球上多样性最高的生物类群, 在驱动碳氮循环等多种生态系统过程中发挥着至关重要的作用。由于研究技术的限制和群落结构复杂等原因, 土壤微生物生态学研究还处于描述性阶段, 理论研究还很缺乏。鉴于此, 利用分子生物学技术尤其是新一代测序技术, 从理论层面上系统地研究全球变化背景下我国北方草地微生物多样性的维持机制具有重要意义。本文在比较各种环境变化对土壤微生物群落的相对影响的基础上, 分析全球变化对微生物多样性影响的物理化学和生态学机制, 并对未来内蒙古草原微生物多样性的重点研究领域进行了展望, 包括: (1)加强全球变化多因素综合研究; (2)加强微生物多样性维持的生态学机制的研究; (3)加强地上与地下多样性关联机制的研究; (4)加强全球大尺度多生态系统的整合研究。     

环境DNA技术在地下生态学中的应用

地下生态过程是生态系统结构、功能和过程研究中最不确定的因素。由于技术和方法的限制,作为"黑箱"的地下生态系统已经成为限制生态学发展的瓶颈,也是未来生态学发展的主要方向。环境DNA技术,是指从土壤等环境样品中直接提取DNA片段,然后通过DNA测序技术来定性或定量化目标生物,以确定目标生物在生态系统中的分布及功能特征。环境DNA技术已成功用于地下生态过程的研究。目前,环境DNA技术在土壤微生物多样性及其功能方面的研究相对成熟,克服了土壤微生物研究中不能培养的问题,可以有效地分析土壤微生物的群落组成、多样性及空间分布,尤其是宏基因组学技术的发展,使得微生物生态功能方面的研究成为可能;而且,环境DNA技术已经在土壤动物生态学的研究中得到了初步应用,可快速分析土壤动物的多样性及其分布特征,更有效地鉴定出未知的或稀少的物种,鉴定土壤动物类群的幅度较宽;部分研究者通过提取分析土壤中DNA片段信息对生态系统植物多样性及植物分类进行了研究,其结果比传统的植物分类及物种多样性测定更精确,改变了以往对植物群落物种多样性模式的理解。同时,环境DNA技术克服传统根系研究方法中需要洗根、分根、只能测定单物种根系的局限,降低根系研究中细根区分的误差,并探索性地用于细根生物量的研究。主要综述了基于环境DNA技术的分子生物学方法在土壤微生物多样性及功能、土壤动物多样性、地下植物多样性及根系生态等地下生态过程研究中的应用进展。环境DNA技术对于以土壤微生物、土壤动物及地下植物根系为主体的地下生态学过程的研究具有革命性意义,并展现出良好的应用前景。可以预期,分子生物学技术与传统的生态学研究相结合将成为未来地下生态学研究的一个发展趋势。     

The role of ecological theory in microbial ecology

Microbial ecology is currently undergoing a revolution, with repercussions spreading throughout microbiology, ecology and ecosystem science. The rapid accumulation of molecular data is uncovering vast diversity, abundant uncultivated microbial groups and novel microbial functions. This accumulation of data requires the application of theory to provide organization, structure, mechanistic insight and, ultimately, predictive power that is of practical value, but the application of theory in microbial ecology is currently very limited. Here we argue that the full potential of the ongoing revolution will not be realized if research is not directed and driven by theory, and that the generality of established ecological theory must be tested using microbial systems.     

猪microRNA组学研究进展

MicroRNA(miRNA)是一类长约22 nt的非编码小RNA,广泛存在于各种生物中,调节生物体生长、发育和凋亡等过程。研究表明,miRNA在猪肌肉、脂肪、生殖系统以及免疫系统等的发育过程中发挥着重要的调控作用。此外,高通量的新一代测序技术在猪miRNA的挖掘和差异表达研究中发挥着巨大的作用。文章综述了高通量的新一代测序技术在挖掘猪miRNA中的应用以及一些miRNA在猪脂肪代谢、肌肉发育、卵母细胞成熟和B、T淋巴细胞发育中的调控作用,旨在为猪miRNA的研究提供参考,为利用miRNA调控和改善猪肉品质、生长性能、繁殖性能以及免疫机能提供理论基础和研究思路。     

基于第二代测序技术的基因资源挖掘

转录组研究一直是生命科学研究的一个重要方向,在第二代测序技术问世以前,已经产生了一些行之有效的转录组研究方法,但这些方法存在一定的局限性。第二代测序技术的出现不仅使转录组研究很快进入了高速发展期,同时也为遗传资源的挖掘提供了一套全新的技术平台。本文简要介绍了第二代测序技术的化学原理和特性,重点阐述了利用第二代测序技术进行转录组测序,从而在此基础上挖掘遗传资源的研究。     

微生物生态学理论框架

微生物是生态系统的重要组成部分,直接或间接地参与所有的生态过程。微生物生态学是基于微生物群体的科学,利用微生物群体DNA/RNA等标志物,重点研究微生物群落构建、组成演变、多样性及其与环境的关系,在生态学理论的指导和反复模型拟合下由统计分析得出具有普遍意义的结论。其研究范围从基因尺度到全球尺度。分子生物学技术的发展,使人们可以直接从基因水平上考查其多样性,从而使得对微生物空间分布格局及其成因的深入研究成为可能。进而可以从方法学探讨微生物生物多样性、分布格局、影响机制及其对全球变化的响应等。在微生物生态学研究中,群落构建与演化、分布特征(含植物-微生物相互关系)、执行群体功能的机理(生物地球化学循环等)、对环境变化的响应与反馈机理是今后需要关注的重点领域。概述了微生物生态学的概念,并初步提出其理论框架,在对比宏观生态学基础理论和模型的基础上,分析微生物多样性的研究内容、研究方法和群落构建的理论机制,展望了今后研究的重点领域。