Biology subjects

Ware, V. C.

Publications and source records attributed to Ware, V. C..

3 recordsLinked to original sources

bioRxiv2023

Identification of a new antiphage system in Mycobacterium phage Butters

During lysogeny temperate phages establish a truce with the bacterial host. In this state, the phage genome (prophage) is maintained within the host environment. Consequently, many prophages have evolved systems to protect the host from heterotypic viral attack. This phenomenon of prophages mediating defense of their host against competitor phages is widespread among temperate mycobacteriophages. We previously showed that the Mycobacterium phage Butters prophage encodes a two-component system (gp30/31) that inhibits infection from a subset of mycobacteriophages that include PurpleHaze, but not Island3. Here we show that Butters gp57r is both necessary and sufficient to inhibit infection by Island3 and other phages. Gp57r acts post-DNA injection and its antagonism results in the impairment of Island3 DNA amplification. Gp57r inhibition of Island3 is absolute with no defense escape mutants. However, mutations mapping to minor tail proteins allow PurpleHaze to overcome gp57r defense. Gp57r has a HEPN domain which is present in many proteins involved in inter-genomic conflicts, suggesting that gp57r may inhibit heterotypic phage infections via its HEPN domain. We also show that Butters gp57r has orthologues in clinical isolates of Mycobacterium abscessus sp. including the phage therapy candidate strain GD91 which was found to be resistant to the panel of phages tested. It is conceivable that this GD91 orthologue of gp57r may mediate resistance to the subset of phages tested. Challenges of this nature underscore the importance of elucidating mechanisms of antiphage systems and mutations that allow for escape from inhibition. IMPORTANCEThe evolutionary arms race between phages and their bacteria host is ancient. During lysogeny, temperate phages establish a ceasefire with the host where they do not kill the host but derive shelter from it. Within the phenomenon of prophage-mediated defense, some temperate phages contribute genes that make their host more fit and resistant to infections by other phages. This arrangement has significance for both phage and bacterial evolutionary dynamics. Further, the prevalence of such antiphage systems poses a challenge to phage therapy. Thus, studies aimed at elucidating antiphage systems will further our understanding of phage-bacteria evolution as well as help with efforts to engineer therapeutic phages that circumvent antiphage systems.

microbiology↗

bioRxiv2022

Characterization of Novel Recombinant Mycobacteriophage derived from Homologous Recombination between two Temperate Phages

Comparative analyses of mycobacteriophage genomes reveals extensive genetic diversity in genome organization and gene content, contributing to widespread mosaicism. We previously reported that the prophage of mycobacteriophage Butters (cluster N) provides defense against infection by Island3 (subcluster I1). To explore the anti-Island3 defense mechanism, we attempted to isolate Island3 defense escape mutants on a Butters lysogen, but only uncovered phages with recombinant genomes comprised of regions of Butters and Island3 arranged from left arm to right arm as Butters-Island3-Butters (BIBs). Recombination occurs within two distinct homologous regions that encompass lysin A, lysin B, and holin genes in one segment, and RecE and RecT genes in the other. Structural genes of mosaic BIB genomes are contributed by Butters while the immunity cassette is derived from Island3. Consequently, BIBs are morphologically identical to Butters (as shown by transmission electron microscopy) but are homoimmune with Island3. A reverse experiment where an Island3 lysogen was infected with Butters yielded Butters phages and no recombinants, demonstrating directionality to the recombination phenomenon. Recombinant phages overcome antiphage defense and silencing of the lytic cycle. We leverage this observation to propose a stratagem to generate novel phages for therapeutic use. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=169 SRC="FIGDIR/small/520664v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@19a817dorg.highwire.dtl.DTLVardef@ab5f65org.highwire.dtl.DTLVardef@1041cc0org.highwire.dtl.DTLVardef@fbb92a_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2020

Mycobacterium phage Butters-encoded proteins contribute to host defense against viral attack

A diverse set of prophage-mediated mechanisms protecting bacterial hosts from infection has been recently uncovered within Cluster N mycobacteriophages. In that context, we unveil a novel defense mechanism in Cluster N prophage Butters. By using bioinformatics analyses, phage plating efficiency experiments, microscopy, and immunoprecipitation assays, we show that Butters genes located in the central region of the genome play a key role in the defense against heterotypic viral attack. Our study suggests that a two component system articulated by interactions between protein products of genes 30 and 31 confers defense against heterotypic phage infection by PurpleHaze or Alma, but is insufficient to confer defense against attack by the heterotypic phage Island3. Therefore, based on heterotypic phage plating efficiencies on the Butters lysogen, additional prophage genes required for defense are implicated. IMPORTANCEMany sequenced bacterial genomes including pathogenic bacteria contain prophages. Some prophages encode defense systems that protect their bacterial host against heterotypic viral attack. Understanding the mechanisms undergirding these defense systems will be critical to development of phage therapy that circumvents these defenses. Additionally, such knowledge will help engineer phage-resistant bacteria of industrial importance.

microbiology↗