Collaborative Research: Prophages and how they manipulate model microbiomes

动物的肠道被一群微生物定居，肠道微生物群会影响健康和福祉。肠道中最丰富的微生物是细菌，它们具有新陈代谢活性，而且大多数对动物宿主友好或无害。像大多数有机体一样，细菌也会被病毒感染，这些病毒被称为噬菌体或噬菌体。已知这些噬菌体的一种版本可以感染细菌，并作为先知整合到它们的DNA中，保持在被认为是休眠状态。然而，由于它们携带的基因或它们整合的位置，前驱体可能会影响细菌的行为，并可能增加它们对抗生素的抗药性。尽管人们越来越多地认识到动物微生物群如何影响健康，但人们对前驱噬菌体如何影响动物微生物群的结构和功能知之甚少。该项目将利用两个模型系统，一个是无脊椎动物脊索动物Ciona robusta，另一个是脊椎动物斑马鱼Danio rerio，以确定先知如何影响动物微生物群。该项目将结合遗传学、预测建模和实验方法来研究原噬菌体对动物微生物群的生物学和生态学影响。这项研究的数据将有助于理解动物肠道内形成动态平衡的因素。该项目的其他更广泛的影响包括培训和指导学生，其中一些学生来自代表性不足的社区，从事微生物学、生物信息学和免疫学的研究。调查人员将让K-12年级的学生参与诸如“Skype-a-Science”和为女孩举办的海洋夏令营等活动。此外，调查人员还将在科学节、博物馆和农贸市场向普通公众进行宣传。大多数动物都与细菌形成了重要的联系。噬菌体(或称“噬菌体”)是感染这些细菌的病毒，众所周知，通过裂解，“裂解”噬菌体重组细菌群落，而“温和”噬菌体可以作为“前驱体”整合到细菌基因组中，将其宿主转化为“溶原体”。整合到细菌基因组中可能会影响附近的基因，而其他病毒基因或调节区可能会影响细菌的生理和适应能力，这一过程被称为“溶原性转化”。然而，与我们对细菌在动物微生物群中的不同角色的显著提高相比，前驱噬菌体对这些群落的影响在很大程度上仍然被忽视。这一提议将检验这样一种假设，即前驱体塑造其细菌宿主的生物学，并推动动物微生物群的结构和功能变化。利用细菌基因组测序和分子遗传学的结合，同时结合生态建模和实验方法，研究人员将在两个强大的模式系统-被囊状乔纳罗布斯塔和斑马鱼Danio rerio的微生物群中研究溶原菌的生物学和生态学。具体地说，研究人员将表征模式微生物培养的异养生物基因组中前驱体的分布和位置，通过有针对性的前驱体缺失突变来确定前驱体对细菌宿主特性(游泳、生物膜形成、生长/生理)的影响，并研究前驱体在体内和硅胶中塑造定植和影响肠道微生物群稳定性的作用。这项研究可能揭示前驱直接影响微生物功能的一般原理，导致动物微生物的重大和微小变化。这一奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

The animal gut is colonized by a community of microbes, the gut microbiome, that can impact health and well-being. The most abundant microbes in the gut are bacteria, and they are metabolically active and most of them are friendly or benign to the animal host. Like most organisms, bacteria also get infected by viruses, and these are known as bacteriophages or phages. One version of these phages is known to infect bacteria and integrate into their DNA as prophages, remaining in what is thought to be a dormant state. However, because of the genes they carry or the location of their integration, prophages can impact the behavior of bacteria and can increase their resistance to antibiotics. Despite the increased recognition of how animal microbiomes impact health, little is known about how prophages influence the structure and function of animal microbiomes. This project will leverage two model systems, an invertebrate chordate, Ciona robusta, and a vertebrate, the zebrafish Danio rerio to identify how prophages affect animal microbiomes. The project will integrate genetic, predictive modeling, and experimental approaches to study the biology and ecology of prophage influence on animal microbiomes. Data from this study will contribute to the understanding of factors shaping homeostasis within the gut of animals. Other broader impacts of this project include training and mentoring of students, some from underrepresented communities, in research on microbiology, bioinformatics, and immunology. The investigators will engage K-12 students with activities such as “Skype-a-Scientist” and an oceanography camp for girls. Additionally, the investigators will perform outreach to the general public at science festivals, museums, and farmer markets. Most animals form essential relationships with bacteria. Bacteriophages (or ‘phages’) are viruses that infect these bacteria, and it is well established that through lysis, ‘lytic’ phages restructure bacterial communities while ‘temperate’ phages can integrate into bacterial genomes as ‘prophages,’ transforming their hosts into ‘lysogens’. Integration into bacterial genomes can influence nearby genes, while other viral genes or regulatory regions can influence bacterial physiology and fitness in a process referred to as ‘lysogenic conversion.’ However, compared to our significantly improved understanding of the diverse roles of bacteria in animal microbiomes, the impact of prophages on these communities remains largely overlooked. This proposal will test the hypothesis that prophages shape the biology of their bacterial hosts and drive structural and functional changes in animal microbiomes. Utilizing a combination of bacterial genome sequencing and molecular genetics while integrating ecological modeling and experimental approaches, the investigators will study the biology and ecology of lysogens in the microbiomes of two powerful model systems, the tunicate Ciona robusta, and the zebrafish Danio rerio. Specifically, the investigators will characterize prophage distribution and location in genomes of cultivated heterotrophs from model microbiomes, determine the influence of prophages on bacterial host properties (swimming, biofilm formation, growth / physiology) via targeted prophage deletion mutagenesis, and examine the role of prophages in shaping colonization and influencing the stability of the gut microbiome in vivo and in silico. This research may reveal general principles by which prophages directly affect microbial function, leading to both major and minor changes in animal microbiomes.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.