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Klumpp J.,ETH Zurich | Fouts D.E.,J. Craig Venter Institute | Sozhamannan S.,GoldBelt Raven LLC
Briefings in Functional Genomics | Year: 2013

Emerging and reemerging bacterial infectious diseases are a major public health concern worldwide. The role of bacteriophages in the emergence of novel bacterial pathogens by horizontal gene transfer was highlighted by the May 2011 Escherichia coli O104:H4 outbreaks that originated in Germany and spread to other European countries. This outbreak also highlighted the pivotal role played by recent advances in functional genomics in rapidly deciphering the virulence mechanism elicited by this novel pathogen and developing rapid diagnostics and therapeutics. However, despite a steady increase in the number of phage sequences in the public databases, boosted by the next-generation sequencing technologies, few functional genomics studies of bacteriophages have been conducted. Our definition of 'functional genomics' encompasses a range of aspects: phage genome sequencing, annotation and ascribing functions to phage genes, prophage identification in bacterial sequences, elucidating the events in various stages of phage life cycle using genomic, transcriptomic and proteomic approaches, defining the mechanisms of host takeover including specific bacterial-phage protein interactions and identifying virulence and other adaptive features encoded by phages and finally, using prophage genomic information for bacterial detection/diagnostics. Given the breadth and depth of this definition and the fact that some of these aspects (especially phage-encoded virulence/adaptive features) have been treated extensively in other reviews, we restrict our focus only on certain aspects. These include phage genome sequencing and annotation, identification of prophages in bacterial sequences and genetic characterization of phages, functional genomics of the infection process and finally, bacterial identification using genomic information. © The Author 2013. Published by Oxford University Press. All rights reserved.

Carol C.,Henry M. Jackson Foundation for the Advancement of Military Medicine, Inc | Carol C.,Naval Medical Research Center Frederick | Matthew H.,Henry M. Jackson Foundation for the Advancement of Military Medicine, Inc | Matthew H.,Naval Medical Research Center Frederick | And 15 more authors.
PLoS ONE | Year: 2015

Historically, cholera outbreaks have been linked to V. cholerae O1 serogroup strains or its derivatives of the O37 and O139 serogroups. A genomic study on the 2010 Haiti cholera outbreak strains highlighted the putative role of non O1/non-O139 V . cholerae in causing cholera and the lack of genomic sequences of such strains from around the world. Here we address these gaps by scanning a global collection of V. cholerae strains as a first step towards understanding the population genetic diversity and epidemic potential of non O1/ non-O139 strains. Whole Genome Mapping (Optical Mapping) based bar coding produces a high resolution, ordered restriction map, depicting a complete view of the unique chromosomal architecture of an organism. To assess the genomic diversity of non-O1/non-O139 V. cholerae, we applied a Whole Genome Mapping strategy on a well-defined and geographically and temporally diverse strain collection, the Sakazaki serogroup type strains. Whole Genome Map data on 91 of the 206 serogroup type strains support the hypothesis that V. cholerae has an unprecedented genetic and genomic structural diversity. Interestingly, we discovered chromosomal fusions in two unusual strains that possess a single chromosome instead of the two chromosomes usually found in V. cholerae. We also found pervasive chromosomal rearrangements such as duplications and indels in many strains. The majority of Vibrio genome sequences currently in public databases are unfinished draft sequences. The Whole Genome Mapping approach presented here enables rapid screening of large strain collections to capture genomic complexities that would not have been otherwise revealed by unfinished draft genome sequencing and thus aids in assembling and finishing draft sequences of complex genomes. Furthermore, Whole Genome Mapping allows for prediction of novel V. cholerae non-O1/non-O139 strains that may have the potential to cause future cholera outbreaks.

Adams B.L.,U.S. Army | Finch A.S.,U.S. Army | Hurley M.M.,U.S. Army | Sarkes D.A.,GoldBelt Raven LLC | Stratis-Cullum D.N.,U.S. Army
Advanced Materials | Year: 2013

The first-ever peptide biomaterial discovery using an unconstrained engineered bacterial display technology is reported. Using this approach, we have developed genetically engineered peptide binders for a bulk aluminum alloy and use molecular dynamics simulation of peptide conformational fluctuations to demonstrate sequence-dependent, structure-function relationships for metal and metal oxide interactions. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Utter B.,Rockefeller University | Deutsch D.R.,Rockefeller University | Schuch R.,Rockefeller University | Schuch R.,ContraFect | And 11 more authors.
PLoS ONE | Year: 2014

In Staphylococcus aureus, the disease impact of chromosomally integrated prophages on virulence is well described. However, the existence of extra-chromosomal prophages, both plasmidial and episomal, remains obscure. Despite the recent explosion in bacterial and bacteriophage genomic sequencing, studies have failed to specifically focus on extrachromosomal elements. We selectively enriched and sequenced extra-chromosomal DNA from S. aureus isolates using Roche-454 technology and uncovered evidence for the widespread distribution of multiple extra-chromosomal prophages (ExPΦs) throughout both antibiotic-sensitive and -resistant strains. We completely sequenced one such element comprised of a 43.8 kbp, circular ExPΦ (designated FBU01) from a vancomycin-intermediate S. aureus (VISA) strain. Assembly and annotation of ΦBU01 revealed a number of putative virulence determinants encoded within a bacteriophage immune evasion cluster (IEC). Our identification of several potential ExPΦs and mobile genetic elements (MGEs) also revealed numerous putative virulence factors and antibiotic resistance genes. We describe here a previously unidentified level of genetic diversity of stealth extra-chromosomal elements in S. aureus, including phages with a larger presence outside the chromosome that likely play a prominent role in pathogenesis and strain diversity driven by horizontal gene transfer (HGT).

Fouts D.E.,J. Craig Venter Institute | Klumpp J.,ETH Zurich | Bishop-Lilly K.A.,Henry M. Jackson Foundation for the Advancement of Military Medicine, Inc | Bishop-Lilly K.A.,Biological Defense Research Directorate | And 16 more authors.
Virology Journal | Year: 2013

Background: Vibrio cholerae O139 Bengal is the only serogroup other than O1 implicated in cholera epidemics. We describe the isolation and characterization of an O139 serogroup-specific phage, vB-VchP-VchO139-I (φVchO139-I) that has similar host range and virion morphology as phage vB-VchP-JA1 (φJA1) described previously. We aimed at a complete molecular characterization of both phages and elucidation of their genetic and structural differences and assessment of their genetic relatedness to the N4-like phage group. Methods. Host-range analysis and plaque morphology screening were done for both φJA1 and φVchO139-I. Both phage genomes were sequenced by a 454 and Sanger hybrid approach. Genomes were annotated and protein homologies were determined by Blast and HHPred. Restriction profiles, PFGE patterns and data on the physical genome structure were acquired and phylogenetic analyses were performed. Results: The host specificity of φJA1 has been attributed to the unique capsular O-antigen produced by O139 strains. Plaque morphologies of the two phages were different; φVchO139-I produced a larger halo around the plaques than φJA1. Restriction profiles of φJA1 and φVchO139-I genomes were also different. The genomes of φJA1 and φVchO139-I consisted of linear double-stranded DNA of 71,252 and 70,938 base pairs. The presence of direct terminal repeats of around 1974 base pairs was demonstrated. Whole genome comparison revealed single nucleotide polymorphisms, small insertions/deletions and differences in gene content. Both genomes had 79 predicted protein encoding sequences, of which only 59 were identical between the two closely related phages. They also encoded one tRNA-Arg gene, an intein within the large terminase gene, and four homing endonuclease genes. Whole genome phylogenetic analyses of φJA1 and φVchO139-I against other sequenced N4-like phages delineate three novel subgroups or clades within this phage family. Conclusions: The closely related phages feature significant genetic differences, in spite of being morphologically identical. The phage morphology, genetic organization, genomic content and large terminase protein based phylogeny support the placement of these two phages in the Podoviridae family, more specifically within the N4-like phage group. The physical genome structure of φJA1 could be demonstrated experimentally. Our data pave the way for potential use of φJA1 and φVchO139-I in Vibrio cholerae typing and control. © 2013 Fouts et al.; licensee BioMed Central Ltd.

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