Whole genome sequence analysis indicates recent diversification of mammal-associated Campylobacter fetus and implicates a genetic factor associated with H2S production

cknowledgements We like to thank Emma Yee (U.S. Department of Agriculture) for the generation of sequence data, we thank James Bono (U.S. Department of Agriculture) for the generation of PacBio RS reads and thank Dr. Brian Brooks and Dr. John Devenish (Canadian Food Inspection Agency) for providing C. fetus strains and for critical review of this manuscript. Funding Publication charges for this article have been funded by Utrecht University, the Netherlands.Peer reviewedPublisher PD

Projector 2: contig mapping for efficient gap-closure of prokaryotic genome sequence assemblies

With genome sequencing efforts increasing exponentially, valuable information accumulates on genomic content of the various organisms sequenced. Projector 2 uses (un)finished genomic sequences of an organism as a template to infer linkage information for a genome sequence assembly of a related organism being sequenced. The remaining gaps between contigs for which no linkage information is present can subsequently be closed with direct PCR strategies. Compared with other implementations, Projector 2 has several distinctive features: a user-friendly web interface, automatic removal of repetitive elements (repeat-masking) and automated primer design for gap-closure purposes. Moreover, when using multiple fragments of a template genome, primers for multiplex PCR strategies can also be designed. Primer design takes into account that, in many cases, contig ends contain unreliable DNA sequences and repetitive sequences. Closing the remaining gaps in prokaryotic genome sequence assemblies is thereby made very efficient and virtually effortless. We demonstrate that the use of single or multiple fragments of a template genome (i.e. unfinished genome sequences) in combination with repeat-masking results in mapping success rates close to 100%. The web interface is freely accessible at

Campylobacter fetus Subspecies Contain Conserved Type IV Secretion Systems on Multiple Genomic Islands and Plasmids

Acknowledgments We like to thank Dr. John Devenish and Dr. Brian Brooks (Canadian Food Inspection Agency) for providing strains. We thank Nathaniel Simon and Mary Chapman for the generation of Illumina MiSeq reads and we thank James Bono for the generation of PacBio RS reads. Funding: The authors have no support or funding to report.Peer reviewedPublisher PD

Living in Cold Blood: Arcobacter, Campylobacter, and Helicobacter in Reptiles

Species of the Epsilonproteobacteria genera Arcobacter, Campylobacter, and Helicobacter are commonly associated with vertebrate hosts and some are considered significant pathogens. Vertebrate-associated Epsilonproteobacteria are often considered to be largely confined to endothermic mammals and birds. Recent studies have shown that ectothermic reptiles display a distinct and largely unique Epsilonproteobacteria community, including taxa which can cause disease in humans. Several Arcobacter taxa are widespread amongst reptiles and often show a broad host range. Reptiles carry a large diversity of unique and novel Helicobacter taxa, which apparently evolved in an ectothermic host. Some species, such as Campylobacter fetus, display a distinct intraspecies host dichotomy, with genetically divergent lineages occurring either in mammals or reptiles. These taxa can provide valuable insights in host adaptation and co-evolution between symbiont and host. Here, we present an overview of the biodiversity, ecology, epidemiology, and evolution of reptile-associated Epsilonproteobacteria from a broader vertebrate host perspective

Global transcriptional landscape and promoter mapping of the gut commensal Bifidobacterium breve UCC2003

Background: Bifidobacterium breve represents a common member of the infant gut microbiota and its presence in the gut has been associated with host well being. For this reason it is relevant to investigate and understand the molecular mechanisms underlying the establishment, persistence and activities of this gut commensal in the host environment. Results: The assessment of vegetative promoters in the bifidobacterial prototype Bifidobacterium breve UCC2003 was performed employing a combination of RNA tiling array analysis and cDNA sequencing. Canonical −10 (TATAAT) and −35 (TTGACA) sequences were identified upstream of transcribed genes or operons, where deviations from this consensus correspond to transcription level variations. A Random Forest analysis assigned the −10 region of B. breve promoters as the element most impacting on the level of transcription, followed by the spacer length and the 5’-UTR length of transcripts. Furthermore, our transcriptome study also identified rho-independent termination as the most common and effective termination signal of highly and moderately transcribed operons in B. breve. Conclusion: The present study allowed us to identify genes and operons that are actively transcribed in this organism during logarithmic growth, and link promoter elements with levels of transcription of essential genes in this organism. As homologs of many of our identified genes are present across the whole genus Bifidobacterium, our dataset constitutes a transcriptomic reference to be used for future investigations of gene expression in members of this genus

Bordetella pertussis population dynamics and phylogeny in Japan after adoption of acellular pertussis vaccines

Bordetella pertussis, the causative agent of whooping cough, has experienced a resurgence in the past 15 years, despite the existence of both whole-cell and acellular vaccines. Here, we performed whole genome sequencing analysis of 149 clinical strains, provided by the National Institute of Infectious Diseases (NIID), Japan, isolated in 1982-2014, after Japan became the first country to adopt acellular vaccines against B. pertussis. Additionally, we sequenced 39 strains provided by the Konan Kosei Hospital in Aichi prefecture, Japan, isolated in 2008-2013. The genome sequences afforded insight into B. pertussis genome variability and population dynamics in Japan, and revealed that the B. pertussis population in Japan was characterized by two major clades that divided more than 40 years ago. The pertactin gene was disrupted in about 20 % of the 149 NIID isolates, by either a deletion within the signal sequence (ΔSS) or the insertion of IS element IS481 (prn :: IS481). Phylogeny suggests that the parent clones for these isolates originated in Japan. Divergence dating traced the first generation of the pertactin-deficient mutants in Japan to around 1990, and indicated that strains containing the alternative pertactin allele prn2 may have appeared in Japan around 1974. Molecular clock data suggested that observed fluctuations in B. pertussis population size may have coincided with changes in vaccine usage in the country. The continuing failure to eradicate the disease warrants an exploration of novel vaccine compositions

Comparative Genomics of Campylobacter fetus from Reptiles and Mammals Reveals Divergent Evolution in Host-Associated Lineages

Acknowledgments The authors like to thank Brian Brooks and John Devenish (Canadian Food Inspection Agency) for providing strains and valuable suggestions.Peer reviewedPublisher PD

Genomics and pathotypes of the many faces of Escherichia coli

Escherichia coli is the most researched microbial organism in the world. Its varied impact on human health, consisting of commensalism, gastrointestinal disease, or extraintestinal pathologies, has generated a separation of the species into at least eleven pathotypes (also known as pathovars). These are broadly split into two groups, intestinal pathogenic E. coli (InPEC) and extraintestinal pathogenic E. coli (ExPEC). However, components of E. coli's infinite open accessory genome are horizontally transferred with substantial frequency, creating pathogenic hybrid strains that defy a clear pathotype designation. Here, we take a birds-eye view of the E. coli species, characterizing it from historical, clinical, and genetic perspectives. We examine the wide spectrum of human disease caused by E. coli, the genome content of the bacterium, and its propensity to acquire, exchange, and maintain antibiotic resistance genes and virulence traits. Our portrayal of the species also discusses elements that have shaped its overall population structure and summarizes the current state of vaccine development targeted at the most frequent E. coli pathovars. In our conclusions, we advocate streamlining efforts for clinical reporting of ExPEC, and emphasize the pathogenic potential that exists throughout the entire species

Development of Kaptive databases for Vibrio parahaemolyticus O- and K-antigen genotyping.

Vibrio parahaemolyticus is an important food-borne human pathogen and presents immunogenic surface polysaccharides, which can be used to distinguish problematic and disease-causing lineages. V. parahaemolyticus is divided in 16 O-serotypes (O-antigen) and 71 K-serotypes (K-antigen). Agglutination tests are still the gold standard for serotyping, but many V. parahaemolyticus isolates are not typable by agglutination. An alternative for agglutination tests is genotyping using whole-genome sequencing data, by which K- and O- genotypes have been curated and identified previously for other clinically relevant organisms with the software tool Kaptive. In this study, V. parahaemolyticus isolates were serotyped and sequenced, and all known and several novel O- and K-loci were identified. We developed Kaptive databases for all O- and K-loci after manual curation of the loci. In our study, we could genotype the O- and K-loci of 98 and 93 % of the genomes, respectively, with a Kaptive confidence score higher than 'none'. The newly developed Kaptive databases with the identified V. parahaemolyticus O- and K-loci can be used to identify the O- and K-genotypes of V. parahaemolyticus isolates from genome sequences

Supervised Lowess normalization of comparative genome hybridization data – application to lactococcal strain comparisons

Background: Array-based comparative genome hybridization (aCGH) is commonly used to determine the genomic content of bacterial strains. Since prokaryotes in general have less conserved genome sequences than eukaryotes, sequence divergences between the genes in the genomes used for an aCGH experiment obstruct determination of genome variations (e.g. deletions). Current normalization methods do not take into consideration sequence divergence between target and microarray features and therefore cannot distinguish a difference in signal due to systematic errors in the data or due to sequence divergence. Results: We present supervised Lowess, or S-Lowess, an application of the subset Lowess normalization method. By using a predicted subset of array features with minimal sequence divergence between the analyzed strains for the normalization procedure we remove systematic errors from dual-dye aCGH data in two steps: (1) determination of a subset of conserved genes (i.e. likely conserved genes, LCG); and (2) using the LCG for subset Lowess normalization. Subset Lowess determines the correction factors for systematic errors in the subset of array features and normalizes all array features using these correction factors. The performance of S-Lowess was assessed on aCGH experiments in which differentially labeled genomic DNA fragments of Lactococcus lactis IL1403 and L. lactis MG1363 strains were hybridized to IL1403 DNA microarrays. Since both genomes are sequenced and gene deletions identified, the success rate of different aCGH normalization methods in detecting these deletions in the MG1363 genome were determined. S-Lowess detects 97% of the deletions, whereas other aCGH normalization methods detect up to only 60% of the deletions. Conclusion: S-Lowess is implemented in a user-friendly web-tool. We demonstrate that it outperforms existing normalization methods and maximizes detection of genomic variation (e.g. deletions) from microbial aCGH data.