Identification of novel double-stranded RNA elements in developing antibiotic resistance in the agricultural environment

农业环境中抗生素耐药性发展中新型双链RNA元件的鉴定

• 批准号: NE/N019288/1
• 负责人: Igor Morozov
• 金额: $ 22.69万
• 依托单位: Coventry University
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2016
• 资助国家: 英国
• 起止时间: 2016 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=NE%2FN019288%2F1
• 关键词: Identification; novel; double; stranded; RNA

Many types of antibiotics (AB) which are used in humans (e.g. chloramphenicol and its derivatives) are also used in farms (e.g. thiamphenicol, a methyl-sulfonyl derivative of chloramphenicol) either to treat or prevent disease. They have a similar antibacterial spectrum and may significantly increase a possibility that clinical pathogens will develop cross-resistance to drugs used in human medicine. The intestinal microbiota is the epicentre but underexplored source for antibiotic resistance (AR) emergence in response to the selective pressure of AB. The vast majority of bacteria cannot be cultured in laboratory conditions and this limits our knowledge of the potential AR determinants these species may possess and express in a community-dependent manner.Metagenomics, for identification of encoded metabolic pathways present in bacterial populations, has revealed many novel, possibly dormant genes in microbial communities as well as a large discrepancy between predicted and detected resistance genes. While metagenomics has primarily focused on analysis of DNA as the source of genomic information, RNA can also serve as genetic material. Recent high-throughput RNA-sequencing (RNA-Seq) analyses of bacterial systems have made two critical discoveries. Firstly, expression of the bacterial genome is extensively regulated by non-coding (nc) RNAs, including antisense (as) RNAs (a class of ncRNA that are encoded on the opposite strand of their target genes). asRNAs regulate gene expression via RNA-RNA interactions, thus leading to modulation of mRNA translation and stability. asRNAs were firstly found on mobile elements (phages (bacterial viruses), plasmids or transposons) which are used for horizontal transfer (HT) of AR genes between different bacteria. However, the mechanism by which these asRNAs regulate expression and possibly acquisition and spread of genes involved in AR is unknown. Secondly, the microbial metatranscriptome contains double stranded (ds) RNA sequences, derived from uncharacterised phages which do not match to the corresponding DNA. Intriguingly, these dsRNAs have coding potential for a large proportion of novel proteins of unknown function.The use of AB in medicine and agriculture triggers a number of adaptation responses in bacterial communities. Dynamics and mechanisms underlying such functional changes in microbiomes in response to AB are still elusive. This may involve activation of expression of a number of genes relevant to resistance (e.g. transposases, proteases or efflux pump genes), HT or mobilisation of genes and non-coding DNA/RNA elements on mobile structures. Thus, key questions are: How do as-metatranscriptomes respond to antibiotic treatments? Does the animal gut microflora contain dsRNAs that do not correspond to their DNA metagenomes? If yes, then what roles do these dsRNAs play in this ecosystem? Does this uncharacterised genetic information play a role in adaptation responses, including AR?Our aim therefore is to undertake RNA-Seq of dsRNAs extracted from animal faecal samples in order to identify dsRNAs metatranscriptome in response to antibiotic therapy. The results will lead to identification of an unexplored array of novel genetic information, including non-coding regulatory elements and new open reading frames relevant to AR mechanisms. This in turn, will transform our view on the role novel dsRNAs play in the development and regulation of AR in bacterial communities. The results will also lead to detecting novel or dormant pathways which are regulated by dsRNAs for the production of secondary metabolites with low susceptibility to resistance and discovery of novel genetic information involved in development, transmission and regulation of AR. This ground breaking research project has the potential to provide a paradigm shift in the understanding of transmission and regulation of AR originated from environment and direct the future strategic development of novel antimicrobials.

用于人类的许多类型的抗生素（AB）（例如氯霉素及其衍生物）也用于农场（例如甲砜霉素，氯霉素的甲磺酰基衍生物）以治疗或预防疾病。它们具有相似的抗菌谱，并且可能显著增加临床病原体对人类医学中使用的药物产生交叉耐药性的可能性。肠道微生物群是抗生素耐药性（AR）出现的震中，但未充分探索的来源，以响应AB的选择性压力。绝大多数细菌不能在实验室条件下培养，这限制了我们对这些物种可能拥有的潜在AR决定因素的了解，并以社区依赖的方式表达。宏基因组学，用于鉴定细菌种群中存在的编码代谢途径，揭示了许多新的，可能是微生物群落中的休眠基因，以及预测和检测到的抗性基因之间的巨大差异。虽然宏基因组学主要集中在分析DNA作为基因组信息的来源，但RNA也可以作为遗传物质。最近对细菌系统的高通量RNA测序（RNA-Seq）分析有两个关键发现。首先，细菌基因组的表达受到非编码（nc）RNA的广泛调节，包括反义（as）RNA（一类在其靶基因的相反链上编码的ncRNA）。asRNA通过RNA-RNA相互作用调节基因表达，从而导致mRNA翻译和稳定性的调节。asRNA首先在移动的元件（细菌病毒、质粒或转座子）上发现，这些元件用于AR基因在不同细菌之间的水平转移（HT）。然而，这些asRNA调节AR相关基因表达以及可能的获得和传播的机制尚不清楚。其次，微生物元转录组含有双链（ds）RNA序列，其来源于与相应DNA不匹配的未表征的RNA。有趣的是，这些双链RNA具有编码大部分未知功能的新型蛋白质的潜力。AB在医学和农业中的应用引发了细菌群落的许多适应性反应。微生物组响应AB的这种功能变化的动力学和机制仍然难以捉摸。这可能涉及与抗性相关的许多基因（例如转座酶、蛋白酶或外排泵基因）的表达激活、HT或基因和移动的结构上的非编码DNA/RNA元件的移动。因此，关键的问题是：作为元转录组如何对抗生素治疗作出反应？动物肠道菌群是否含有与其DNA宏基因组不对应的dsRNA？如果是，那么这些dsRNA在这个生态系统中扮演什么角色？这些未被表征的遗传信息是否在包括AR在内的适应性反应中发挥作用？因此，我们的目标是进行从动物粪便样品中提取的dsRNA的RNA-Seq，以鉴定响应于抗生素治疗的dsRNA元转录组。这些结果将导致识别一系列未探索的新遗传信息，包括非编码调控元件和与AR机制相关的新开放阅读框架。这反过来将改变我们对新型dsRNA在细菌群落中AR的发育和调节中所起作用的看法。这些结果还将导致检测由dsRNA调节的新的或休眠的途径，用于产生对抗性敏感性低的次级代谢产物，并发现参与AR的发育、传递和调节的新的遗传信息。这一开创性的研究项目有可能为理解源自环境的AR的传播和调控提供范式转变，并指导新型抗菌剂的未来战略开发。

项目成果

Microbiota of the gut: Antibiotic-Induced Dysbiosis and the Adverse Effects on Human Health

肠道微生物群：抗生素引起的菌群失调及其对人类健康的不利影响

• 发表时间: 2018
• 期刊: Scientific Journal of Gastroenterology & Hepatology
• 作者: Lamaudière M.T.F.