Metagenomic analysis of cellulose degrading communities

纤维素降解群落的宏基因组分析

• 批准号: NE/H007563/1
• 负责人: Alan McCarthy
• 金额: $ 49.36万
• 依托单位: University of Liverpool
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2010
• 资助国家: 英国
• 起止时间: 2010 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=NE%2FH007563%2F1
• 关键词: Metagenomic; analysis; cellulose; degrading; communities

The carbon cycle in freshwater lake systems comprises two main phases: primary production (photosynthesis) and biodegradation. Sediments and the lower regions of the water column are often anoxic, particularly in summer, and here anaerobic microorganisms degrade the cellulosic phytoplankton biomass, principally to carbon dioxide and methane. In the environment, microorganisms function as communities or consortia from which the isolation and cultivation of individual species is both difficult and a poor representation of the inherent complexity, particularly for anaerobes. In landfill sites, cellulosic waste is the principal source of carbon that is ultimately converted again to carbon dioxide and methane, and in this managed environment it is the activity of anaerobic microorganisms that is entirely responsible for the stabilisation of waste. Cellulose is the principal component of biomass on earth and its degradation and recycling is an important driver of the carbon cycle. Yet the number of species known to be capable of colonising and degrading native crystalline cellulose is limited, and due at least in part to the inadequacies of conventional microbiological methods that rely on the isolation of strains and their cultivation in the laboratory. One alternative is to analyse DNA and RNA extracted directly from environmental samples, and we have applied this molecular ecological approach to the cellulose-degrading community of freshwater lakes and landfill sites. Our strategy has been to suspend cotton (cellulose) baits directly in lakes and in validated landfill leachate microcosms, to enrich and specifically target microorganisms that truly colonise and degrade cellulose in situ. We have indeed been able to publish evidence for the presence of novel species implicated in cellulose degradation in both environments, and more importantly, demonstrate that their occurrence and distribution is quantitatively significant. We now want to exploit the recent advances in high throughput affordable DNA sequencing technology (pyrosequencing) to analyse the metagenomes (total gene content) of cellulose colonised in, and subsequently retrieved from, the environment. We will primarily use RNA as our template, both for identification of the species present and to focus on genes that are actually being expressed in the biofilm and responsible for degradation of the cellulose substrate. This first description of the colonised cellulose expressed gene pool will enable us to design the tools that we will then apply to the identification of larger genetic fragments containing the combinations of ordered genes responsible for adsorption to and degradation of cellulose, and enabling the identification of any entities that are truly novel. In some cases, it may be possible to express these genes and overproduce cellulases for characterisation but in tandem we will also attempt to isolate and cultivate the species responsible for their production in the envrionment. It remains inconceivable that our knowledge of the species responsible for cellulose degradation in the natural aquatic and managed landfill environments is so superficial, but in addition to redressing this imbalance, we hope to identify new sources of cellulases with commercial potential, particularly in the field of second generation biofuel production.

淡水湖泊系统的碳循环包括两个主要阶段：初级生产（光合作用）和生物降解。沉积物和水柱的较低区域通常是缺氧的，特别是在夏季，这里的厌氧微生物降解纤维素浮游植物生物量，主要是二氧化碳和甲烷。在环境中，微生物作为群落或聚生体起作用，从其中分离和培养单个物种既困难又不能很好地代表固有的复杂性，特别是对于厌氧菌。在垃圾填埋场，纤维素废物是碳的主要来源，最终再次转化为二氧化碳和甲烷，在这种受管理的环境中，厌氧微生物的活动完全负责废物的稳定。纤维素是地球上生物质的主要成分，其降解和回收是碳循环的重要驱动力。然而，已知能够定殖和降解天然结晶纤维素的物种的数量是有限的，并且至少部分是由于依赖于菌株的分离及其在实验室中的培养的常规微生物学方法的不足。一种替代方法是分析直接从环境样品中提取的DNA和RNA，我们已经将这种分子生态学方法应用于淡水湖泊和垃圾填埋场的纤维素降解社区。我们的策略是将棉花（纤维素）诱饵直接悬浮在湖泊和经验证的垃圾渗滤液微生态系统中，以富集和特异性靶向真正在原位定植和降解纤维素的微生物。我们确实能够发表证据表明，在这两种环境中存在与纤维素降解有关的新物种，更重要的是，证明它们的发生和分布具有定量意义。我们现在希望利用高通量负担得起的DNA测序技术（焦磷酸测序）的最新进展来分析定殖在环境中并随后从环境中回收的纤维素的宏基因组（总基因含量）。我们将主要使用RNA作为我们的模板，既用于鉴定存在的物种，又专注于实际上在生物膜中表达并负责纤维素底物降解的基因。这第一次描述的殖民纤维素表达的基因库将使我们能够设计的工具，我们将应用于鉴定较大的遗传片段，包含有序的基因的组合，负责吸附和降解纤维素，并使任何实体的鉴定是真正的新。在某些情况下，可能会表达这些基因并过量产生纤维素酶以进行表征，但同时我们也将尝试分离和培养在环境中负责其生产的物种。令人难以置信的是，我们对天然水生和管理垃圾填埋场环境中负责纤维素降解的物种的了解是如此肤浅，但除了纠正这种不平衡之外，我们希望确定具有商业潜力的纤维素酶的新来源，特别是在第二代生物燃料生产领域。

项目成果

Encyclopedia of Metagenomics

宏基因组学百科全书

• DOI: 10.1007/978-1-4614-6418-1_738-1
• 发表时间: 2013
• 作者: Rooks D

Viromic Analysis of Wastewater Input to a River Catchment Reveals a Diverse Assemblage of RNA Viruses.

• DOI: 10.1128/msystems.00025-18
• 发表时间: 2018-05
• 期刊: mSystems
• 影响因子: 6.4
• 作者: Adriaenssens EM;Farkas K;Harrison C;Jones DL;Allison HE;McCarthy AJ
• 通讯作者: McCarthy AJ

Metagenomic approaches to the discovery of cellulases.

• DOI: 10.1016/b978-0-12-415931-0.00020-3
• 发表时间: 2012
• 期刊: Methods in enzymology
• 作者: D. J. Rooks;J. McDonald;A. Mccarthy

The microbial ecology of anaerobic cellulose degradation in municipal waste landfill sites: evidence of a role for fibrobacters.

• DOI: 10.1111/j.1462-2920.2011.02688.x
• 发表时间: 2012-04
• 期刊: Environmental microbiology
• 影响因子: 5.1
• 作者: J. McDonald;James N. I. Houghton;D. J. Rooks;H. Allison;A. Mccarthy

Lignocellulose-Degrading Microbial Communities in Landfill Sites Represent a Repository of Unexplored Biomass-Degrading Diversity.

• DOI: 10.1128/msphere.00300-17
• 发表时间: 2017-07
• 期刊: mSphere
• 影响因子: 4.8
• 作者: Ransom-Jones E;McCarthy AJ;Haldenby S;Doonan J;McDonald JE
• 通讯作者: McDonald JE