Bacteriophages and plasmids in Streptococcus thermophilus

嗜热链球菌中的噬菌体和质粒

• 批准号: RGPIN-2014-05132
• 负责人: Moineau, Sylvain
• 金额: $ 5.17万
• 依托单位: Université Laval
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2014
• 资助国家: 加拿大
• 起止时间: 2014-01-01 至 2015-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=566973
• 关键词: Bacteriophages; plasmids; Streptococcus; thermophilus

The dairy industry is a significant part of our heritage and our economy. It is the third largest sector of the Canadian agri-food economy. In 2012, dairy farming generated $5.9 billion and sales from dairy processors equaled $14.7 billion. Overall, the dairy industry accounted for 15.1% of all processing sales in our nation’s food and beverage sector. Canadian dairy farm operators number over 22,000 and 480 dairy plants, nation-wide, employ over 24,000 additional workers. This makes the dairy sector the second largest employer in the food industry, producing more than 1050 cheese varieties. In 2012, Canadian cheese producers had their second best year, producing 408,197 tons of cheeses, an increase of 1.0% from 2011. The record of 408,647 tons was established in 2010. In 2012, yogurt production also had its second best year, with 312,633 tons (vs. 315,413 tons in 2011). There is no indication that this production trend will change. However, our dairy industry faces intense competition in national and international markets and must deliver high-quality products. Cheese and yogurt manufacture requires the inoculation of milk with carefully selected bacterial cells (starter culture) to properly control the fermentation and to obtain high-quality products. The starter culture is a combination of lactic acid-producing bacteria, from which Streptococcus thermophilus (ST) is one of the most important species. It is well documented that increased productivity within existing manufacturing facilities will lead to milk fermentation failures due to bacterial viruses (phages) that destroy the starter cultures. This natural phenomenon leads to low quality products and economic losses. Our yogurt and cheese factories are facing phage infection problems with their ST strains. This research program will characterize these phages to aid the development of phage-resistant ST strains. We will focus on three general areas. 1- PHAGE BIOLOGY. Our previous genomics research produced an in-depth analysis of two model phages, representing the two main groups of ST phages. We now plan to sequence the genomes of 50 additional ST phages from our collection to elucidate their evolution processes. We will also refine the core and pangenome of both phage groups. In addition, we have previously identified several structural proteins in these two phage groups. We intend now to investigate non-structural phage proteins, many of which have no known function. 2- PHAGE RESISTANCE. Since 2005, we have extensively studied CRISPR-Cas systems in ST, making several landmark discoveries. CRISPR-Cas is now one of the most active and competitive research areas in microbiology. We will continue to study this phage resistance system, especially the spacer acquisition step, and we will investigate other host factors involved in phage replication. 3- PLASMID BIOLOGY. We previously showed that some plasmids are surprisingly stable in ST. We will investigate this phenomenon using sequencing and transformation strategies. Plasmids also play a central role in our ST studies as tools for manipulating DNA. Because no genetic tool is available to modify the genome of virulent ST phages, we plan to construct a plasmid system for phage genome editing to study each phage gene/protein. This ambitious and state-of-the-art program will lead to the most comprehensive analysis of phage-host interactions in a Gram-positive bacterium. This research will provide fundamental knowledge to our Canadian dairy industry to give them a competitive edge. This knowledge will also be applicable to other bacterial strains susceptible to phage attack in industrial fermentations.

乳制品行业是我们的遗产和经济的重要组成部分。它是加拿大农业食品经济的第三大部门。2012年，奶牛养殖产生了59亿美元，乳制品加工商的销售额相当于147亿美元。总体而言，乳制品行业占我国食品和饮料行业所有加工销售额的15.1%。加拿大奶牛场经营者数量超过22，000和480乳品厂，全国范围内，雇用超过24，000额外的工人。这使得乳制品行业成为食品行业的第二大雇主，生产超过1050种奶酪。2012年，加拿大奶酪生产商迎来了第二好的一年，生产了408，197吨奶酪，比2011年增长了1.0%。2010年创造了408，647吨的记录。2012年，酸奶产量也是第二好的一年，产量为312，633吨（2011年为315，413吨）。没有迹象表明这种生产趋势会改变。然而，我们的乳制品行业面临着国内和国际市场的激烈竞争，必须提供高质量的产品。奶酪和酸奶的生产需要用精心挑选的细菌细胞（发酵剂）接种牛奶，以适当控制发酵并获得高质量的产品。发酵剂是一种产乳酸菌的组合，其中嗜热链球菌（ST）是最重要的菌种之一。有充分的证据表明，现有生产设施中生产率的提高将导致牛奶发酵失败，这是由于细菌病毒（ESTA）破坏了发酵剂。这种自然现象导致低质量的产品和经济损失。我们的酸奶和奶酪工厂正面临着ST菌株的噬菌体感染问题。这项研究计划将表征这些细菌，以帮助开发抗噬菌体ST菌株。我们将重点关注三个一般领域。1-噬菌体生物学。我们之前的基因组学研究对两种模型进行了深入分析，代表了两种主要的ST基因组。我们现在计划对我们收集的另外50个ST的基因组进行测序，以阐明它们的进化过程。我们还将完善两个噬菌体组的核心和泛基因组。此外，我们以前已经确定了这两个噬菌体群中的几个结构蛋白。我们现在打算研究非结构噬菌体蛋白，其中许多没有已知的功能。2-噬菌体抗性。自2005年以来，我们广泛研究了ST中的CRISPR-Cas系统，取得了几个里程碑式的发现。CRISPR-Cas现在是微生物学中最活跃和最具竞争力的研究领域之一。我们将继续研究这种噬菌体抗性系统，特别是间隔区获取步骤，我们将研究其他参与噬菌体复制的宿主因素。3-质粒生物学。我们以前表明，一些质粒是令人惊讶的稳定在ST。我们将调查这一现象使用测序和转化策略。Plasma作为操纵DNA的工具，在我们的ST研究中也发挥着核心作用。由于没有遗传工具可用于修改毒性ST的基因组，我们计划构建用于噬菌体基因组编辑的质粒系统以研究每个噬菌体基因/蛋白。这个雄心勃勃的最先进的计划将导致对革兰氏阳性细菌中噬菌体-宿主相互作用的最全面的分析。这项研究将为我们加拿大的乳制品行业提供基础知识，使他们具有竞争优势。这一知识也将适用于在工业发酵中易受噬菌体攻击的其他细菌菌株。