High-throughput mapping of the phage resistance landscape in E. coli

Abstract

Bacteriophages (phages) are critical players in the dynamics and function of microbial communities and drive processes as diverse as global biogeochemical cycles and human health. Phages tend to be predators finely tuned to attack specific hosts, even down to the strain level, which in turn defend themselves using an array of mechanisms. However, to date, efforts to rapidly and comprehensively identify bacterial host factors important in phage infection and resistance have yet to be fully realized. Here, we globally map the host genetic determinants involved in resistance to 14 phylogenetically diverse double-stranded DNA phages using two model Escherichia coli strains (K-12 and BL21) with known sequence divergence to demonstrate strain-specific differences. Using genome-wide lossof-function and gain-of-function genetic technologies, we are able to confirm previously described phage receptors as well as uncover a number of previously unknown host factors that confer resistance to one or more of these phages. We uncover differences in resistance factors that strongly align with the susceptibility of K-12 and BL21 to specific phage. We also identify both phage-specific mechanisms, such as the unexpected role of cyclic-di-GMP in host sensitivity to phage N4, and more generic defenses, such as the overproduction of colanic acid capsular polysaccharide that defends against a wide array of phages. Our results indicate that host responses to phages can occur via diverse cellular mechanisms. Our systematic and high-throughput genetic workflow to characterize phage-host interaction determinants can be extended to diverse bacteria to generate datasets that allow predictive models of how phage-mediated selection will shape bacterial phenotype and evolution. The results of this study and future efforts to map the phage resistance landscape will lead to new insights into the coevolution of hosts and their phage, which can

Genes Influencing Phage Host Range in Staphylococcus aureus on a Species-Wide Scale

“…Two recent studies used single gene knockout, overexpression, and transcriptional suppression methods as well as global transcriptional profiling to identify phage resistance determinants in Escherichia coli ( 80 ) and Enterococcus faecalis ( 61 ). Unlike these previous studies, our findings are not limited to one or a few genetic backgrounds, making them more widely applicable to the species and its underlying evolution.…”

A Spontaneous rapZ Mutant Impairs Infectivity of Lytic Bacteriophage vB_EcoM_JS09 against Enterotoxigenic Escherichia coli

“…The amino acid mutations of RapZ in S. aureus were P48A and H267D (31). Afterwards, (32). These further demonstrate that phage infection caused the occurrence of phage-resistant bacteria by mutations in the rapZ gene.…”

Systematic exploration ofEscherichia coliphage-host interactions with the BASEL phage collection

“…This process has been studied in detail for the archetype of all Autographiviridae, phage T7, though several open questions remain. The lateral tail fibers of this phage contact a receptor in rough LPS of E. coli K-12 that is not fully understood, possibly because several alternative and overlapping sugar motifs in the K-12 LPS core can be targeted [73][74][75] (Fig 10A). Conformational changes in the stubby tail tube triggered by this receptor interaction then initiate the injection of the phage genome from the virion [75,76].…”

“…Bacteriophage N4 is the archetype of Enquatrovirus phages that are hallmarked by using a large, virion-encapsidated RNA polymerase for the transcription of their early genes [33], and our new Enquatrovirus isolate AlfredRasser (Bas67) is a very close relative of N4 ( Fig 10E). Based primarily on genetic evidence, phage N4 is thought to initiate infections by contacting the host's ECA with its lateral tail fibers [73,85,86] (Fig 10A). We indeed confirmed a remarkable dependence of N4 and AlfredRasser on wecB ( Fig 10C) and detected homology between the glycan deacetylase domain of the N4 lateral tail fiber and a lateral tail fiber protein of Vequintavirinae which also seem to use the ECA as their primary receptor (see above, Figs 9 and 10C).…”

“…We reproduced the dependence of our lambdavir variant on LamB and, as reported previously, found that an intact inner core of the LPS was required for infectivity [73 and literature cited therein] (Fig 11F and S5 Table). Myoviruses P1 and P2 were both shown to require a rough LPS phenotype to contact their receptors in the LPS core, but the exact identity of these receptors has not been unraveled [40,50,73,95]. While the molecular details of the infection process after adsorption have not been well studied for P1, previous work showed that an interaction of the P2 lateral tail fibers with the LPS core of K-12 strains triggers penetration of the outer membrane by the tail tip [40,50,96] (Fig 11D).…”