Special Issue "Bacterial Pathogenomics: From Technology to Application"

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A special issue of Pathogens (ISSN 2076-0817).

Deadline for manuscript submissions: closed (31 January 2014)

Special Issue Editor

Guest Editor
Dr. Anthony Underwood (Website)

Bioinformatics Group | Applied Laboratory and Bio- Informatics Unit, Microbiology Services, Colindale, Public Health England 61 Colindale Avenue, London NW9 5EQ, UK
Interests: pathogen genomics; phylogenomics; virulence; microbiology; public health

Special Issue Information

Dear Colleagues,

The first bacterial genome required a large team and many person years to complete. With current high throughput massively-parallel sequencing technologies it is possible to generate a nearly complete bacterial genome in just a few days. This major revolution opens up huge opportunities, particularly in the ability to compare the whole genomes of many bacterial isolates. This might include trying to answer basic biological questions by looking at genome wide associations and attempting to link genetic features to phenotypic traits. However the questions being asked can also be much more applied, such as using whole genome phylogenies to look for evidence of transmission within a community or hospital or find the source of a disease outbreak.

In this special issue, we invite investigators to submit manuscripts that cover this broad topic. These can include everything from novel uses of current sequencing technology to investigate bacterial genomics, to cutting edge applications of whole genome sequencing to 'real world' scenarios.

We look forward to your contribution.

Dr. Anthony Underwood
Guest Editor

Submission

Manuscripts should be submitted online at www.mdpi.com by registering and logging in to this website. Once you are registered, click here to go to the submission form. Manuscripts can be submitted until the deadline. Papers will be published continuously (as soon as accepted) and will be listed together on the special issue website. Research articles, review articles as well as communications are invited. For planned papers, a title and short abstract (about 100 words) can be sent to the Editorial Office for announcement on this website.

Submitted manuscripts should not have been published previously, nor be under consideration for publication elsewhere (except conference proceedings papers). All manuscripts are refereed through a peer-review process. A guide for authors and other relevant information for submission of manuscripts is available on the Instructions for Authors page. Pathogens is an international peer-reviewed Open Access quarterly journal published by MDPI.

Please visit the Instructions for Authors page before submitting a manuscript. The Article Processing Charge (APC) for publication in this open access journal is 600 CHF (Swiss Francs). English correction and/or formatting fees of 250 CHF (Swiss Francs) will be charged in certain cases for those articles accepted for publication that require extensive additional formatting and/or English corrections.

Published Papers (15 papers)

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Research

Jump to: Review, Other

Open AccessArticle Transcriptional Profiling of a Cross-Protective Salmonella enterica serovar Typhimurium UK-1 dam Mutant Identifies a Set of Genes More Transcriptionally Active Compared to Wild-Type, and Stably Transcribed across Biologically Relevant Microenvironments
Pathogens 2014, 3(2), 417-436; doi:10.3390/pathogens3020417
Received: 1 March 2014 / Revised: 30 April 2014 / Accepted: 4 May 2014 / Published: 9 May 2014
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Abstract
Vaccination with Salmonella enterica serovar Typhimurium lacking DNA adenine methyltransferase confers cross-protective immunity against multiple Salmonella serotypes. The mechanistic basis is thought to be associated with the de-repression of genes that are tightly regulated when transiting from one microenvironment to another. This [...] Read more.
Vaccination with Salmonella enterica serovar Typhimurium lacking DNA adenine methyltransferase confers cross-protective immunity against multiple Salmonella serotypes. The mechanistic basis is thought to be associated with the de-repression of genes that are tightly regulated when transiting from one microenvironment to another. This de-repression provides a potential means for the production of a more highly expressed and stable antigenic repertoire capable of inducing cross-protective immune responses. To identify genes encoding proteins that may contribute to cross-protective immunity, we used a Salmonella Typhimurium DNA adenine methyltransferase mutant strain (UK-1 dam mutant) derived from the parental UK-1 strain, and assessed the transcriptional profile of the UK-1 dam mutant and UK-1 strain grown under conditions that simulate the intestinal or endosomal microenvironments encountered during the infective process. As expected, the transcriptional profile of the UK-1 dam mutant identified a set of genes more transcriptionally active when compared directly to UK-1, and stably transcribed in biologically relevant culture conditions. Further, 22% of these genes were more highly transcribed in comparison to two other clinically-relevant Salmonella serovars. The strategy employed here helps to identify potentially conserved proteins produced by the UK-1 dam mutant that stimulate and/or modulate the development of cross-protective immune responses toward multiple Salmonella serotypes. Full article
(This article belongs to the Special Issue Bacterial Pathogenomics: From Technology to Application)
Open AccessArticle Characterization of Antimicrobial Resistance Dissemination across Plasmid Communities Classified by Network Analysis
Pathogens 2014, 3(2), 356-376; doi:10.3390/pathogens3020356
Received: 27 November 2013 / Revised: 4 April 2014 / Accepted: 9 April 2014 / Published: 15 April 2014
Cited by 3 | PDF Full-text (2958 KB) | HTML Full-text | XML Full-text
Abstract
The global clustering of gene families through network analysis has been demonstrated in whole genome, plasmid, and microbiome analyses. In this study, we carried out a plasmidome network analysis of all available complete bacterial plasmids to determine plasmid associations. A blastp clustering [...] Read more.
The global clustering of gene families through network analysis has been demonstrated in whole genome, plasmid, and microbiome analyses. In this study, we carried out a plasmidome network analysis of all available complete bacterial plasmids to determine plasmid associations. A blastp clustering search at 100% aa identity cut-off and sharing at least one gene between plasmids, followed by a multilevel community network analysis revealed that a surprisingly large number of the plasmids were connected by one largest connected component (LCC), with dozens of community sub-groupings. The LCC consisted mainly of Bacilli and Gammaproteobacteria plasmids. Intriguingly, horizontal gene transfer (HGT) was noted between different phyla (i.e., Staphylococcus and Pasteurellaceae), suggesting that Pasteurellaceae can acquire antimicrobial resistance (AMR) genes from closely contacting Staphylococcus spp., which produce the external supplement of V-factor (NAD). Such community network analysis facilitate displaying possible recent HGTs like a class 1 integron, str and tet resistance markers between communities. Furthermore, the distribution of the Inc replicon type and AMR genes, such as the extended-spectrum ß-lactamase (ESBL) CTX-M or the carbapenemases KPC NDM-1, implies that such genes generally circulate within limited communities belonging to typical bacterial genera. Thus, plasmidome network analysis provides a remarkable discriminatory power for plasmid-related HGT and evolution. Full article
(This article belongs to the Special Issue Bacterial Pathogenomics: From Technology to Application)
Open AccessArticle Pseudomonas aeruginosa Genome Evolution in Patients and under the Hospital Environment
Pathogens 2014, 3(2), 309-340; doi:10.3390/pathogens3020309
Received: 30 November 2013 / Revised: 26 March 2014 / Accepted: 28 March 2014 / Published: 10 April 2014
Cited by 3 | PDF Full-text (1283 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
Pseudomonas aeruginosa is a Gram-negative environmental species and an opportunistic microorganism, establishing itself in vulnerable patients, such as those with cystic fibrosis (CF) or those hospitalized in intensive care units (ICU). It has become a major cause of nosocomial infections worldwide and [...] Read more.
Pseudomonas aeruginosa is a Gram-negative environmental species and an opportunistic microorganism, establishing itself in vulnerable patients, such as those with cystic fibrosis (CF) or those hospitalized in intensive care units (ICU). It has become a major cause of nosocomial infections worldwide and a serious threat to Public Health because of overuse and misuse of antibiotics that have selected highly resistant strains against which very few therapeutic options exist. Herein is illustrated the intraclonal evolution of the genome of sequential isolates collected in a single CF patient from the early phase of pulmonary colonization to the fatal outcome. We also examined at the whole genome scale a pair of genotypically-related strains made of a drug susceptible, environmental isolate recovered from an ICU sink and of its multidrug resistant counterpart found to infect an ICU patient. Multiple genetic changes accumulated in the CF isolates over the disease time course including SNPs, deletion events and reduction of whole genome size. The strain isolated from the ICU patient displayed an increase in the genome size of 4.8% with major genetic rearrangements as compared to the initial environmental strain. The annotated genomes are given in free access in an interactive web application WallGene  designed to facilitate large-scale comparative analysis and thus allowing investigators to explore homologies and syntenies between P. aeruginosa strains, here PAO1 and the five clinical strains described. Full article
(This article belongs to the Special Issue Bacterial Pathogenomics: From Technology to Application)
Open AccessArticle Genome Sequencing of Xanthomonas vasicola Pathovar vasculorum Reveals Variation in Plasmids and Genes Encoding Lipopolysaccharide Synthesis, Type-IV Pilus and Type-III Secretion Effectors
Pathogens 2014, 3(1), 211-237; doi:10.3390/pathogens3010211
Received: 13 December 2013 / Revised: 10 February 2014 / Accepted: 3 March 2014 / Published: 18 March 2014
Cited by 3 | PDF Full-text (2436 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
Xanthomonas vasicola pathovar vasculorum (Xvv) is the bacterial agent causing gumming disease in sugarcane. Here, we compare complete genome sequences for five isolates of Xvv originating from sugarcane and one from maize. This identified two distinct types of lipopolysaccharide synthesis [...] Read more.
Xanthomonas vasicola pathovar vasculorum (Xvv) is the bacterial agent causing gumming disease in sugarcane. Here, we compare complete genome sequences for five isolates of Xvv originating from sugarcane and one from maize. This identified two distinct types of lipopolysaccharide synthesis gene clusters among Xvv isolates: one is similar to that of Xanthomonas axonopodis pathovar citri (Xac) and is probably the ancestral type, while the other is similar to those of the sugarcane-inhabiting species, Xanthomonas sacchari. Four of six Xvv isolates harboured sequences similar to the Xac plasmid, pXAC47, and showed a distinct Type-IV pilus (T4P) sequence type, whereas the T4P locus of the other two isolates resembled that of the closely related banana pathogen, Xanthomonas campestris pathovar musacearum (Xcm). The Xvv isolate from maize has lost a gene encoding a homologue of the virulence effector, xopAF, which was present in all five of the sugarcane isolates, while xopL contained a premature stop codon in four out of six isolates. These findings shed new light on evolutionary events since the divergence of Xvv and Xcm, as well as further elucidating the relationships between the two closely related pathogens. Full article
(This article belongs to the Special Issue Bacterial Pathogenomics: From Technology to Application)
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Open AccessArticle Genomics-Based Exploration of Virulence Determinants and Host-Specific Adaptations of Pseudomonas syringae Strains Isolated from Grasses
Pathogens 2014, 3(1), 121-148; doi:10.3390/pathogens3010121
Received: 22 November 2013 / Revised: 20 January 2014 / Accepted: 22 January 2014 / Published: 28 January 2014
Cited by 3 | PDF Full-text (532 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
The Pseudomonas syringae species complex has recently been named the number one plant pathogen, due to its economic and environmental impacts, as well as for its role in scientific research. The bacterium has been repeatedly reported to cause outbreaks on bean, cucumber, [...] Read more.
The Pseudomonas syringae species complex has recently been named the number one plant pathogen, due to its economic and environmental impacts, as well as for its role in scientific research. The bacterium has been repeatedly reported to cause outbreaks on bean, cucumber, stone fruit, kiwi and olive tree, as well as on other crop and non-crop plants. It also serves as a model organism for research on the Type III secretion system (T3SS) and plant-pathogen interactions. While most of the current work on this pathogen is either carried out on one of three model strains found on dicot plants with completely sequenced genomes or on isolates obtained from recent outbreaks, not much is known about strains isolated from grasses (Poaceae). Here, we use comparative genomics in order to identify putative virulence-associated genes and other Poaceae-specific adaptations in several newly available genome sequences of strains isolated from grass species. All strains possess only a small number of known Type III effectors, therefore pointing to the importance of non-Type III secreted virulence factors. The implications of this finding are discussed. Full article
(This article belongs to the Special Issue Bacterial Pathogenomics: From Technology to Application)
Open AccessArticle Live Genomics for Pathogen Monitoring in Public Health
Pathogens 2014, 3(1), 93-108; doi:10.3390/pathogens3010093
Received: 26 September 2013 / Revised: 16 December 2013 / Accepted: 7 January 2014 / Published: 21 January 2014
Cited by 2 | PDF Full-text (580 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
Whole genome analysis based on next generation sequencing (NGS) now represents an affordable framework in public health systems. Robust analytical pipelines of genomic data provides in short laps of time (hours) information about taxonomy, comparative genomics (pan-genome) and single polymorphisms profiles. Pathogenic [...] Read more.
Whole genome analysis based on next generation sequencing (NGS) now represents an affordable framework in public health systems. Robust analytical pipelines of genomic data provides in short laps of time (hours) information about taxonomy, comparative genomics (pan-genome) and single polymorphisms profiles. Pathogenic organisms of interest can be tracked at the genomic level, allowing monitoring at one-time several variables including: epidemiology, pathogenicity, resistance to antibiotics, virulence, persistence factors, mobile elements and adaptation features. Such information can be obtained not only at large spectra, but also at the “local” level, such as in the event of a recurrent or emergency outbreak. This paper reviews the state of the art in infection diagnostics in the context of modern NGS methodologies. We describe how actuation protocols in a public health environment will benefit from a “streaming approach” (pipeline). Such pipeline would NGS data quality assessment, data mining for comparative analysis, searching differential genetic features, such as virulence, resistance persistence factors and mutation profiles (SNPs and InDels) and formatted “comprehensible” results. Such analytical protocols will enable a quick response to the needs of locally circumscribed outbreaks, providing information on the causes of resistance and genetic tracking elements for rapid detection, and monitoring actuations for present and future occurrences. Full article
(This article belongs to the Special Issue Bacterial Pathogenomics: From Technology to Application)
Open AccessArticle Genetic Diversity of Tick-Borne Rickettsial Pathogens; Insights Gained from Distant Strains
Pathogens 2014, 3(1), 57-72; doi:10.3390/pathogens3010057
Received: 8 November 2013 / Revised: 27 December 2013 / Accepted: 7 January 2014 / Published: 14 January 2014
Cited by 1 | PDF Full-text (1273 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
The ability to capture genetic variation with unprecedented resolution improves our understanding of bacterial populations and their ability to cause disease. The goal of the pathogenomics era is to define genetic diversity that results in disease. Despite the economic losses caused by [...] Read more.
The ability to capture genetic variation with unprecedented resolution improves our understanding of bacterial populations and their ability to cause disease. The goal of the pathogenomics era is to define genetic diversity that results in disease. Despite the economic losses caused by vector-borne bacteria in the Order Rickettsiales, little is known about the genetic variants responsible for observed phenotypes. The tick-transmitted rickettsial pathogen Anaplasma marginale infects cattle in tropical and subtropical regions worldwide, including Australia. Genomic analysis of North American A. marginale strains reveals a closed core genome defined by high levels of Single Nucleotide Polymorphisms (SNPs). Here we report the first genome sequences and comparative analysis for Australian strains that differ in virulence and transmissibility. A list of genetic differences that segregate with phenotype was evaluated for the ability to distinguish the attenuated strain from virulent field strains. Phylogenetic analyses of the Australian strains revealed a marked evolutionary distance from all previously sequenced strains. SNP analysis showed a strikingly reduced genetic diversity between these strains, with the smallest number of SNPs detected between any two A. marginale strains. The low diversity between these phenotypically distinct bacteria presents a unique opportunity to identify the genetic determinants of virulence and transmission. Full article
(This article belongs to the Special Issue Bacterial Pathogenomics: From Technology to Application)
Open AccessArticle Comparative Genomics Identifies a Potential Marker of Human-Virulent Anaplasma phagocytophilum
Pathogens 2014, 3(1), 25-35; doi:10.3390/pathogens3010025
Received: 27 November 2013 / Revised: 18 December 2013 / Accepted: 7 January 2014 / Published: 9 January 2014
Cited by 2 | PDF Full-text (2973 KB) | HTML Full-text | XML Full-text
Abstract
We have previously described a comparative genome analysis of nine strains of Anaplasma phagocytophilum that showed similarity between strains infecting humans and U.S. dogs and a more distant relationship with horse and ruminant strains. This suggested that it may be possible to [...] Read more.
We have previously described a comparative genome analysis of nine strains of Anaplasma phagocytophilum that showed similarity between strains infecting humans and U.S. dogs and a more distant relationship with horse and ruminant strains. This suggested that it may be possible to distinguish human-infective strains using simple DNA sequence-based diagnostic tests. This would be of epidemiologic significance in identifying and tracking the presence of virulent strains in tick vector populations. Further analysis identified a gene that was present in several strains, including U.S. Ap-variant 1 (ruminant), MRK (horse), and European sheep, but was deleted in strains infecting U.S. humans and dogs, suggesting that it could be a useful marker of human virulence. A simple PCR test was developed to identify the presence/absence of this gene. The PCR test discriminated A. phagocytophilum strains from clinically affected humans and U.S. dogs from the strains more distantly related in genome sequence. This warrants further testing of globally diverse A. phagocytophilum strains to examine world-wide conservation of this gene. Full article
(This article belongs to the Special Issue Bacterial Pathogenomics: From Technology to Application)
Open AccessArticle Whole-Proteome Analysis of Twelve Species of Alphaproteobacteria Links Four Pathogens
Pathogens 2013, 2(4), 627-635; doi:10.3390/pathogens2040627
Received: 1 August 2013 / Revised: 19 November 2013 / Accepted: 20 November 2013 / Published: 26 November 2013
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Abstract
Thousands of whole-genome and whole-proteome sequences have been made available through advances in sequencing technology, and sequences of millions more organisms will become available in the coming years. This wealth of genetic information will provide numerous opportunities to enhance our understanding of [...] Read more.
Thousands of whole-genome and whole-proteome sequences have been made available through advances in sequencing technology, and sequences of millions more organisms will become available in the coming years. This wealth of genetic information will provide numerous opportunities to enhance our understanding of these organisms including a greater understanding of relationships among species. Researchers have used 16S rRNA and other gene sequences to study the evolutionary origins of bacteria, but these strategies do not provide insight into the sharing of genes among bacteria via horizontal transfer. In this work we use an open source software program called pClust to cluster proteins from the complete proteomes of twelve species of Alphaproteobacteria and generate a dendrogram from the resulting orthologous protein clusters. We compare the results with dendrograms constructed using the 16S rRNA gene and multiple sequence alignment of seven housekeeping genes. Analysis of the whole proteomes of these pathogens grouped Rickettsia typhi with three other animal pathogens whereas conventional sequence analysis failed to group these pathogens together. We conclude that whole-proteome analysis can give insight into relationships among species beyond their phylogeny, perhaps reflecting the effects of horizontal gene transfer and potentially providing insight into the functions of shared genes by means of shared phenotypes. Full article
(This article belongs to the Special Issue Bacterial Pathogenomics: From Technology to Application)
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Open AccessArticle Comprehensive Analysis of Prokaryotes in Environmental Water Using DNA Microarray Analysis and Whole Genome Amplification
Pathogens 2013, 2(4), 591-605; doi:10.3390/pathogens2040591
Received: 2 August 2013 / Revised: 16 October 2013 / Accepted: 24 October 2013 / Published: 30 October 2013
Cited by 1 | PDF Full-text (1464 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
The microflora in environmental water consists of a high density and diversity of bacterial species that form the foundation of the water ecosystem. Because the majority of these species cannot be cultured in vitro, a different approach is needed to identify [...] Read more.
The microflora in environmental water consists of a high density and diversity of bacterial species that form the foundation of the water ecosystem. Because the majority of these species cannot be cultured in vitro, a different approach is needed to identify prokaryotes in environmental water. A novel DNA microarray was developed as a simplified detection protocol. Multiple DNA probes were designed against each of the 97,927 sequences in the DNA Data Bank of Japan and mounted on a glass chip in duplicate. Evaluation of the microarray was performed using the DNA extracted from one liter of environmental water samples collected from seven sites in Japan. The extracted DNA was uniformly amplified using whole genome amplification (WGA), labeled with Cy3-conjugated 16S rRNA specific primers and hybridized to the microarray. The microarray successfully identified soil bacteria and environment-specific bacteria clusters. The DNA microarray described herein can be a useful tool in evaluating the diversity of prokaryotes and assessing environmental changes such as global warming. Full article
(This article belongs to the Special Issue Bacterial Pathogenomics: From Technology to Application)
Open AccessArticle An Emerging Tick-Borne Disease of Humans Is Caused by a Subset of Strains with Conserved Genome Structure
Pathogens 2013, 2(3), 544-555; doi:10.3390/pathogens2030544
Received: 17 July 2013 / Revised: 29 August 2013 / Accepted: 2 September 2013 / Published: 10 September 2013
Cited by 7 | PDF Full-text (896 KB) | HTML Full-text | XML Full-text
Abstract
The prevalence of tick-borne diseases is increasing worldwide. One such emerging disease is human anaplasmosis. The causative organism, Anaplasma phagocytophilum, is known to infect multiple animal species and cause human fatalities in the U.S., Europe and Asia. Although long known to [...] Read more.
The prevalence of tick-borne diseases is increasing worldwide. One such emerging disease is human anaplasmosis. The causative organism, Anaplasma phagocytophilum, is known to infect multiple animal species and cause human fatalities in the U.S., Europe and Asia. Although long known to infect ruminants, it is unclear why there are increasing numbers of human infections. We analyzed the genome sequences of strains infecting humans, animals and ticks from diverse geographic locations. Despite extensive variability amongst these strains, those infecting humans had conserved genome structure including the pfam01617 superfamily that encodes the major, neutralization-sensitive, surface antigen. These data provide potential targets to identify human-infective strains and have significance for understanding the selective pressures that lead to emergence of disease in new species. Full article
(This article belongs to the Special Issue Bacterial Pathogenomics: From Technology to Application)
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Review

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Open AccessReview Leptospiral Pathogenomics
Pathogens 2014, 3(2), 280-308; doi:10.3390/pathogens3020280
Received: 18 January 2014 / Revised: 22 March 2014 / Accepted: 28 March 2014 / Published: 10 April 2014
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Abstract
Leptospirosis, caused by pathogenic spirochetes belonging to the genus Leptospira, is a zoonosis with important impacts on human and animal health worldwide. Research on the mechanisms of Leptospira pathogenesis has been hindered due to slow growth of infectious strains, poor transformability, [...] Read more.
Leptospirosis, caused by pathogenic spirochetes belonging to the genus Leptospira, is a zoonosis with important impacts on human and animal health worldwide. Research on the mechanisms of Leptospira pathogenesis has been hindered due to slow growth of infectious strains, poor transformability, and a paucity of genetic tools. As a result of second generation sequencing technologies, there has been an acceleration of leptospiral genome sequencing efforts in the past decade, which has enabled a concomitant increase in functional genomics analyses of Leptospira pathogenesis. A pathogenomics approach, by coupling of pan-genomic analysis of multiple isolates with sequencing of experimentally attenuated highly pathogenic Leptospira, has resulted in the functional inference of virulence factors. The global Leptospira Genome Project supported by the U.S. National Institute of Allergy and Infectious Diseases to which key scientific contributions have been made from the international leptospirosis research community has provided a new roadmap for comprehensive studies of Leptospira and leptospirosis well into the future. This review describes functional genomics approaches to apply the data generated by the Leptospira Genome Project towards deepening our knowledge of virulence factors of Leptospira using the emerging discipline of pathogenomics. Full article
(This article belongs to the Special Issue Bacterial Pathogenomics: From Technology to Application)
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Open AccessReview Genomic and Global Approaches to Unravelling How Hypermutable Sequences Influence Bacterial Pathogenesis
Pathogens 2014, 3(1), 164-184; doi:10.3390/pathogens3010164
Received: 4 December 2013 / Revised: 6 January 2014 / Accepted: 13 February 2014 / Published: 25 February 2014
Cited by 2 | PDF Full-text (889 KB) | HTML Full-text | XML Full-text
Abstract
Rapid adaptation to fluctuations in the host milieu contributes to the host persistence and virulence of bacterial pathogens. Adaptation is frequently mediated by hypermutable sequences in bacterial pathogens. Early bacterial genomic studies identified the multiplicity and virulence-associated functions of these hypermutable sequences. [...] Read more.
Rapid adaptation to fluctuations in the host milieu contributes to the host persistence and virulence of bacterial pathogens. Adaptation is frequently mediated by hypermutable sequences in bacterial pathogens. Early bacterial genomic studies identified the multiplicity and virulence-associated functions of these hypermutable sequences. Thus, simple sequence repeat tracts (SSRs) and site-specific recombination were found to control capsular type, lipopolysaccharide structure, pilin diversity and the expression of outer membrane proteins. We review how the population diversity inherent in the SSR-mediated mechanism of localised hypermutation is being unlocked by the investigation of whole genome sequences of disease isolates, analysis of clinical samples and use of model systems. A contrast is presented between the problematical nature of analysing simple sequence repeats in next generation sequencing data and in simpler, pragmatic PCR-based approaches. Specific examples are presented of the potential relevance of this localized hypermutation to meningococcal pathogenesis. This leads us to speculate on the future prospects for unravelling how hypermutable mechanisms may contribute to the transmission, spread and persistence of bacterial pathogens. Full article
(This article belongs to the Special Issue Bacterial Pathogenomics: From Technology to Application)
Open AccessReview Identifying Pathogenicity Islands in Bacterial Pathogenomics Using Computational Approaches
Pathogens 2014, 3(1), 36-56; doi:10.3390/pathogens3010036
Received: 30 November 2013 / Revised: 30 December 2013 / Accepted: 7 January 2014 / Published: 13 January 2014
Cited by 11 | PDF Full-text (353 KB) | HTML Full-text | XML Full-text
Abstract
High-throughput sequencing technologies have made it possible to study bacteria through analyzing their genome sequences. For instance, comparative genome sequence analyses can reveal the phenomenon such as gene loss, gene gain, or gene exchange in a genome. By analyzing pathogenic bacterial genomes, [...] Read more.
High-throughput sequencing technologies have made it possible to study bacteria through analyzing their genome sequences. For instance, comparative genome sequence analyses can reveal the phenomenon such as gene loss, gene gain, or gene exchange in a genome. By analyzing pathogenic bacterial genomes, we can discover that pathogenic genomic regions in many pathogenic bacteria are horizontally transferred from other bacteria, and these regions are also known as pathogenicity islands (PAIs). PAIs have some detectable properties, such as having different genomic signatures than the rest of the host genomes, and containing mobility genes so that they can be integrated into the host genome. In this review, we will discuss various pathogenicity island-associated features and current computational approaches for the identification of PAIs. Existing pathogenicity island databases and related computational resources will also be discussed, so that researchers may find it to be useful for the studies of bacterial evolution and pathogenicity mechanisms. Full article
(This article belongs to the Special Issue Bacterial Pathogenomics: From Technology to Application)
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Other

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Open AccessOpinion Culture-Independence for Surveillance and Epidemiology
Pathogens 2013, 2(3), 556-570; doi:10.3390/pathogens2030556
Received: 1 August 2013 / Revised: 3 September 2013 / Accepted: 5 September 2013 / Published: 24 September 2013
Cited by 2 | PDF Full-text (301 KB) | HTML Full-text | XML Full-text
Abstract
Culture-independent methods in microbiology (quantitative PCR (qPCR), sequencing, microarrays, direct from sample matrix assisted laser desorption/ionization time of flight mass spectroscopy (MALDI-TOF MS), etc.) are disruptive technology. Rather than providing the same results as culture-based methods more quickly, more cheaply or [...] Read more.
Culture-independent methods in microbiology (quantitative PCR (qPCR), sequencing, microarrays, direct from sample matrix assisted laser desorption/ionization time of flight mass spectroscopy (MALDI-TOF MS), etc.) are disruptive technology. Rather than providing the same results as culture-based methods more quickly, more cheaply or with improved accuracy, they reveal an unexpected diversity of microbes and illuminate dark corners of undiagnosed disease. At times, they overturn existing definitions of presumably well-understood infections, generating new requirements for clinical diagnosis, surveillance and epidemiology. However, current diagnostic microbiology, infection control and epidemiology rest principally on culture methods elegantly optimized by clinical laboratorians. The clinical significance is interwoven; the new methods are out of context, difficult to interpret and impossible to act upon. Culture-independent diagnostics and surveillance methods will not be deployed unless the reported results can be used to select specific therapeutics or infection control measures. To cut the knots surrounding the adoption of culture-independent methods in medical microbiology, culture-dependent methods should be supported by consistent culture-independent methods providing the microbial context. This will temper existing biases and motivate appropriate scrutiny of the older methods and results. Full article
(This article belongs to the Special Issue Bacterial Pathogenomics: From Technology to Application)

Planned Papers

The below list represents only planned manuscripts. Some of these manuscripts have not been received by the Editorial Office yet. Papers submitted to MDPI journals are subject to peer-review.

Type of Paper: Review
Title: From Reads to Whole Genome Sequencing, High-Throughput Sequencing in the Fight against Pathogens
Authors: Ségolène Caboche *, Christophe Audebert and David Hot
Affiliation: University of Lille_IFR142, France; Genes Diffusion, France and Institut Pasteur de Lille, France; *E-Mail: segolene.caboche@pasteur-lille.fr (S.G.)
Abstract: The recent progresses of High-Throughput Sequencing (HTS) technologies enable easy and cost-reduced access to whole genome sequencing (WGS) or re-sequencing. This is particularly true for small genome organisms such as bacteria and viruses. HTS associated with adapted, automatic and fast bioinformatic solutions in WGS promises an accurate and timely identification and characterization of pathogen agents. However, so far, it has only been used for posterior studies of circulating pathogen genomes and not yet as a tool of real time genomic identification to help physicians and clinicians in their medical protocol choice. In this review, the already established milestones and those yet to be completed toward this purpose will be described and discussed.
Keywords: HTS; WGS; pathogen outbreak; bioinformatics analysis; analysis pipeline; comparative genomic

Type of Paper: Article
Title: Whole Genome MLST of Staphylococci as a Tool for Clinical Microbiology
Author: Samuel K. Sheppard
Affiliation: Reader in Microbiology and Infectious Disease, Swansea University Senior Research Associate, University of Oxford, Wellcome Trust Fellow, UK; E-Mail: S.K.Sheppard@swansea.ac.uk (S.K.S.)
Abstract: Genome sequencing of bacterial pathogens can provide detailed information about strain variation in relation to important clinical phenotypes such as virulence or susceptibility to treatment, and enhance understanding of disease epidemiology. However the potential of this technology remains largely unrealized in clinical microbiology laboratories, in part because of difficulties in handling population genomic datasets. Here we present a rapid comparative genomics approach based on gene-by-gene analysis of 247 published Staphylococcus aureus and S. epidermidis genomes. We demonstrate the utility of this web accessible analysis platform. First, for basic population genomic analysis including investigation of population structure, variations in the core and accessory genome, and phylogenetic history of lineages – including the relatedness of MLST clonal complexes. Second, for finer scale investigation of clinically important genes such as mecA, which was present in 27% of S. epidermidis and ubiquitous in the S. aureus isolates, and toxin associated genes such as those encoding the PVL toxins and genes responsible for the synthesis of delta- and beta-toxins that were differentially present in the two species. This freely available gene-by-gene analysis pipeline provides a rapid and scalable tool for epidemiological and evolutionary analysis of Staphylococcus, and other bacterial pathogens, and offers the potential for further increasing the use of whole genome sequencing in clinical  microbiology.

Type of Paper: Article
Title: Integrons in the Commensal Flora as a Reservoir for Transmission of Antibiotic Resistance Genes to Pathogens
Author: Knut Rudi
Affiliation: Norwegian University of Life Sciences, Department of Chemistry, Biotechnology and Food Science, P.O. Box 5003, NO-1432 Aas, Norway; E-Mail: knut.rudi@umb.no (K.R.)
Abstract: Antibiotics have revolutionized treatment of bacterial infections. Unfortunately, the use of antibiotics has been over-emphasized, resulting in the spread of antibiotic resistance genes. Integrons residing in transposons or genome islands have served as a main vector for this spread. These integrons consist of several genes acquired from different microbes and are expressed by a strong promoter. The genes in the integrons can be related to both resistance to antibiotics and host-microbe interaction. Integrons in pathogenic bacteria have been extensively investigated, while integrons in the commensal flora have not yet gained much attention. This review will mainly focus on integrons in the commensal flora and the potential of their transfer to pathogens. Information on the integrons residing in the commensal flora can potentially aid in controlling the spread of antibiotic resistance genes to pathogens, and as a consequence the spread of multi-resistant infections. This review will give an appraisal on a very important, but often overlooked aspect of the microbiota as a reservoir for antibiotic resistant genes in integrons and genome plasticity of microbes. With the accumulated information on the role of commensals in genetic transfer of integrons, in the future, this knowledge can be used to curb the spread of antibiotic resistance genes.

Type of Paper: Article
Title: Comprehensive Analysis of Prokaryotes in Environmental Water Using Whole Genome Amplification and DNA Microarray
Author: Koichi Suzuki
Affiliation: Department of Microbiology, Leprosy Research Center, National Institute of Infectious Diseases, Tokyo, Japan; E-Mail: koichis@nih.go.jp (K.S.)
Abstract: Environmental water contains a microflora consisted of high density and diversity of bacterial species that form the foundation of the water ecosystem. Because the majority of these species cannot be cultured in vitro, an approach other than cultivation is needed to identify prokaryotes in environmental water. To establish a simplified protocol to detect prokaryotes in environmental water, a novel DNA microarray was developed. Multiple DNA probes were designed against each of the 97,927 sequences in the DNA Data Bank of Japan and mounted on a glass chip in duplicate. To evaluate this DNA microarray, one liter of environmental water samples were collected from seven sites in Japan and whole DNA was extracted and uniformly amplified using whole genome amplification (WGA). DNA was labeled with Cy3-conjugated 16S rRNA specific primers and subjected to microarray analysis. The microarray successfully identified indigenous and pathogenic bacteria, and environment-specific bacteria clusters. The DNA microarray described herein can be a useful tool in evaluating the diversity of prokaryotes and assessing environmental changes such as global warming.

Type of Paper: Review
Title: Live Genomics for Pathogen Monitoring in Public Health Systems
Authors: Giuseppe D'Auria 1,*, Maria Victoria Schneider 2 and Andrés Moya 1
Affiliations: 1 Joint Unit of Research in Genomics and Health Center for Public Health Research (FISABIO-CSISP), Valencia, Spain
2 The Genome Analysis Centre, Norwich Research Park, Norwich, UK; *E-Mail: dauria_giu@gva.es (G.D.A.)
Abstract: Whole genome analysis based on next generation sequencing represents now an affordable framework in public health systems. Robust analytical pipelines of genomic data provides in short laps of time (hours) information about taxonomy, comparative genomics (pan-genome) and single polymorphisms profiles. Thus, pathogenic organisms of interest could be tracked at genomic level monitoring all-at-one its expansion, pathogenicity, resistance to antibiotics, virulence and persistence factors, mobile elements, adaptation features, etc. not only in a world-wide context but, probably more useful, at a local context behind a recurrent or emergency outbreak event. In this paper we review the state of the art in infection diagnostics and propose an actuation protocol based on a “streaming approach” going from pathogens genome sequencing trough results formatting, comparative analysis and data mining. Thus, genomics data of target pathogens populate relational databases where automatic data-mining pipelines allows extracting differential data of interest such as virulence, resistance persistence factors, SNPs and InDels profiles. Such analytical protocol allows responding in short time to the needs of locally circumscribed outbreaks providing causes of resistance and genetic tracking elements for rapid detection and monitoring actuations for present or future occurrences.

Type of Paper: Review
Title: Genomic and Global Approaches to Unraveling How Hypermutable Sequences Influence Bacterial Pathogenesis
Authors: Fadil A. Bidmos and Christopher D. Bayliss *
Affiliation: Department of Genetics, University of Leicester, University Road, Leicester, LE1 7RH, UK; *E-Mail: cdb12@le.ac.uk (C.D.B.)
Abstract: Rapid adaptation to fluctuations in the host mileau contributes to the host persistence and virulence of bacterial pathogens. Adaptation is frequently mediated by hypermutable sequences in bacterial pathogens. Early bacterial genomic studies identified the multiplicity and potential for virulence-associated functions of these hypermutable sequences. Thus simple sequence repeat tracts and site-specific recombination were found to control capsular type, lipopolysaccharide structure, pilin diversity and expression of outer membrane proteins. We review how the population diversity inherent in these mechanisms of localised hypermutation is now being unlocked by investigation of model systems and clinical samples. A contrast is presented between the problematical nature of analysing simple sequence repeats in next generation sequencing data and in simpler, pragmatic PCR-based approaches. Specific examples are presented of the potential relevance of this localized hypermutation to meningococcal pathogenesis. This leads us to speculate on the future prospects for unraveling how hypermutable mechanisms may contribute to transmission, spread and persistence of bacterial pathogens.

Type of Paper: Article
Title:
Whole-Proteome and 16S rRNA Analyses of Pathogenic Strains of Alphaproteobacteria
Authors:
Yunyun Zhou 1, Douglas R. Call 1,2 and Shira L. Broschat 1,2,3,*
Affiliations:

1
School of Electrical Engineering and Computer Science, Washington State University, Pullman, WA, USA
2
Paul G. Allen School for Global Animal Health, Washington State University, Pullman, WA, USA
3 Department of Veterinary Microbiology and Pathology, Washington State University, Pullman, WA, USA; *E-Mail: cdb12@le.ac.uk (C.D.B.)
Abstract:
Thousands of whole-genome and whole-proteome sequences have been made available through advances in sequencing technology, and millions more will become available in the coming years. This wealth of genetic information does and will provide numerous opportunities for learning, among which is a greater understanding of relationships among species. Researchers have used 16S rRNA and other techniques to study the evolutionary origins of bacteria, but these techniques do not give insight into the sharing of genes among bacteria via horizontal transfer. In this work we use an open source software program called pClust to cluster proteins from whole proteomes of twelve Alphaproteobacteria and compare the results with their established 16S rRNA phylogeny. We obtain dendrograms using orthologous protein clusters and using a simple 16S rRNA technique. The results differ significantly only in one respect: the animal pathogen Rickettsia typhi is clustered with the three remaining animal pathogens using the whole proteomes rather than as a singlet cluster using the 16S rRNA genes. We conclude that whole-proteome analysis can give insight into relationships among species beyond their established phylogenies, perhaps reflecting the effects of horizontal gene transfer and potentially providing insight into the functions of shared genes by means of shared phenotypes.
Keywords:
whole-proteome sequences; alphaproteobacteria; bacterial pathogens; bacterial phenotypes; pClust

 

Type of Paper: Article
Title:
On the limits of bacterial lifestyle predictions based on genome content
Author(s):
Eudes Barbosa 1,3*, Richard Röttger 2 , Vasco Azevedo1 and Jan Baumbach 3
Affiliation:

1
Institute of Biological Sciences, Federal University of Minas Gerais, Belo Horizonte, Brazil
2 Max Planck Institute for Informatics, Saarbrücken, Germany
3 Department of Mathematics and Computer Science, University of Southern Denmark, Odense, Denmark; E-Mails: eudesgvb@ufmg.br (E.B.); vasco@icb.ufmg.br (V.A.); roettger@mpi-inf.mpg.de (R.R.); jbaumbac@imada.sdu.dk (J.B.)
Abstract:
Actinobacterial species occupy various niches in diverse habitats. Hence, we may assume the existence of lifestyle-specific genomic features. Here, we study the genomic repertoire of 89 actinobacteria classified into four life styles, the pathogenicity classes HP (human pathogens), AP (animal pathogens), OP (opportunistic pathogens), and NP (non-pathogen). Our aim is to investigate the level of their genome specificity by identifying class-specific gene sets. We hypothesize: (H1) Pathogens (HPs, APs, OPs) possess specific pathogenicity genes not existing in NPs. (H2) Animal and human pathogens cannot be distinguished due to an observation bias, i.e. the diversity of AP hosts, and the assumption that HPs might also be APs although never having been identified as such. Additionally, (H3) there is no intrinsic genomic characteristic of opportunistic pathogens (OPs) compared to HPs and APs, as the interplay between host and pathogen as well as small mutations might play a more dominant role. To study these hypotheses, we implemented a bioinformatics pipeline that combines evolutionary sequence analysis (BLAST and Transitivity Clustering) with statistical learning methods (R software package). Essentially, we present orthologous gene sets that computationally distinguish pathogens from NPs (H1). We further show a limited differentiating repertoire for HP from AP (H2), and for OP from HP&AP (H3). In conclusion, we give evidence and illustrate the limitation we face when trying to deduce a certain microbial lifestyle from genomic content, at least in the case of actinobacterial pathogenicity.
Keywords:
Bacterial lifestyle; Actinobacteria; Pathogenicity

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