The potential of natural phage communities in biofilm degradation

生物膜是工业过程和航运业中成本高昂的生物污垢的原因，该项目旨在通过使用生物膜来减少生物膜。当微生物如细菌粘附到任何固体表面时，会引起生物膜，导致与细胞外基质和DNA相关的微生物群落定植在表面上。微生物生物膜对微生物剂的抵抗力比嗜热微生物更强，因此会引起问题，清除起来可能非常昂贵。在工业生产过程中，生物膜可能会导致堵塞，需要关闭和清洁系统，通过减少生物污染，该项目旨在提高生产率和可持续性。该项目将研究使用天然病毒（特别是噬菌体;感染细菌的病毒）群落作为抗菌剂的潜力，以防止生物膜形成（图1A）和破坏已建立的生物膜（图1B）。噬菌体（或噬菌体）是地球上最丰富的生物实体。溶解性噬菌体有能力杀死宿主细菌，而一些噬菌体则分解细胞外聚合物，这些聚合物可以形成将生物膜固定在一起的粘合剂，而无需进入宿主。任何地方发现细菌，噬菌体感染他们也被发现。该项目将比较效力噬菌体社区从不同的环境中，在防止和降解生物膜。这些环境将包括堆肥堆、池塘、河流、河口、海洋系统和污水处理厂。为此，学生将开发一种合成的混合物种生物膜作为模型，以测试不同噬菌体群落的功效。多物种生物膜的建立（图1B.i）将有助于理解环境噬菌体在微生物群落动态中的作用以及生物膜的不同元素如何对它们做出反应。开发生物膜模型将涉及选择共存以形成稳定生物膜的真核生物（即真菌）和细菌伴侣。将感染合成生物膜的成员的特异性噬菌体与天然噬菌体群落在生物膜预防和去除的功效方面进行比较。先前在沃里克大学分离的各种噬菌体是可用的，并且将进行其他针对生物膜成员的特异性噬菌体的分离。方法：最多150个单词开发高通量生物膜评估方法，例如使用带有钉盖或聚碳酸酯过滤器的96孔微量滴定板。细胞可以通过超声处理分离。钉板或过滤器将用于建立生物膜并评估不同的噬菌体群体以减少生物膜。将基于文献选择主要生物污损物种（3 - 6个物种，即代表性拟杆菌门、变形菌门和酵母菌门）的合成生物膜。生物膜物种结构将通过活/死Q-PCR和共聚焦显微镜进行表征。噬菌体分离和表征将使用噬菌斑试验对单个生物膜物种进行分离和计数噬菌体。通过酶标仪和流式细胞仪测定噬菌体的潜伏期、爆发大小和毒力。通过0.2um过滤将天然噬菌体群落与细菌和真核生物分离，然后使用蛋白浓缩离心管浓缩。浓缩的DNA将进行Illumina宏基因组测序，以检查不同的噬菌体群落。

Biofilms are the cause of costly biofouling in industrial processes and the shipping industry and this project aims to reduce biofilms with the use of phages. Biofilms are caused when microorganisms such as bacteria adhere to any solid surface, leading to a community of microorganisms colonising the surface that are associated with an extracellular matrix and DNA. Microbial biofilms are more resistant to microbial agents than planktonic microbes and therefore cause problems and can be very expensive to clear up. In industrial processes biofilms can cause blockages which can require shutdowns and cleaning regimes, by reducing biofouling this project aims to increase productivity and sustainability.This project will investigate the potential for using natural viral (particularly bacteriophage; viruses that infect bacteria) communities as an antimicrobial to prevent biofilm formation (Fig. 1A) and break up established biofilms (Fig. 1B). Bacteriophage (or phage) are the most abundant biological entities on earth. Lytic phage have the capacity to kill their host bacteria while some break down extracellular polymers that can form the adhesive that holds biofilms together, without entering their hosts. Anywhere bacteria are found, phage that infect them are also found.The project will compare the efficacy of phage communities from different environments at preventing and degrading biofilms. These environments will include compost heaps, ponds, rivers, estuaries, marine systems and sewage works. To this end the student will develop a synthetic mixed species biofilm as a model to test the efficacy of different phage communities. The establishment of a multi-species biofilm (Fig. 1B.i) will help to understand the roles of environmental phage in microbial community dynamics and how different elements of the biofilm react to them. Developing a biofilm model will involve selecting eukaryotic (i.e. fungi) and bacterial partners that co-exist to form a stable biofilm. Specific phage that infect members of the synthetic biofilm will be compared to the natural phage communities in terms of efficacy of biofilm prevention and removal. A variety of phage that have previously been isolated at the University of Warwick are available and the isolation of other specific phage to the biofilm members will be undertaken.Methodology: Maximum 150 wordsDevelop a high through-put biofilm assessment method, such as using 96 well microtitre plates with peg lids or polycarbonate filters. Cells can be detached by sonication. Peg plates or filters will be used to establish a biofilm and assess different phage populations for reducing biofilms. A synthetic biofilm of dominant biofouling species (3-6 species i.e. representative Bacteroidetes, Proteobacteria and yeast) will be selected based on the literature. Biofilm species structure will be characterised by live/dead Q-PCR and confocal microscopy.Phage isolation and characterisation will be carried out on individual biofilm species using plaque assays to isolate and enumerate phage. Phage latency period, burst size and virulence will be determined through plate reader assays and flow cytometry.Natural phage communities will be separated from bacteria and eukaryotes by 0.2um filtration, then concentrated using protein concentrating centrifuge tubes. Concentrated phages will have Illumina metagenomics sequencing carried out to examine different phage communities.