A decade of genomic history for healthcare-associated Enterococcus faecium in the United Kingdom and Ireland.

Vancomycin-resistant Enterococcus faecium (VREfm) is an important cause of healthcare-associated infections worldwide. We undertook whole-genome sequencing (WGS) of 495 E. faecium bloodstream isolates from 2001-2011 in the United Kingdom and Ireland (UK&I) and 11 E. faecium isolates from a reference collection. Comparison between WGS and multilocus sequence typing (MLST) identified major discrepancies for 17% of isolates, with multiple instances of the same sequence type (ST) being located in genetically distant positions in the WGS tree. This confirms that WGS is superior to MLST for evolutionary analyses and is more accurate than current typing methods used during outbreak investigations. E. faecium has been categorized as belonging to three clades (Clades A1, hospital-associated; A2, animal-associated; and B, community-associated). Phylogenetic analysis of our isolates replicated the distinction between Clade A (97% of isolates) and Clade B but did not support the subdivision of Clade A into Clade A1 and A2. Phylogeographic analyses revealed that Clade A had been introduced multiple times into each hospital referral network or country, indicating frequent movement of E. faecium between regions that rarely share hospital patients. Numerous genetic clusters contained highly related vanA-positive and -negative E. faecium, which implies that control of vancomycin-resistant enterococci (VRE) in hospitals also requires consideration of vancomycin-susceptible E. faecium Our findings reveal the evolution and dissemination of hospital-associated E. faecium in the UK&I and provide evidence for WGS as an instrument for infection control.

The history, genome and biology of NCTC 30: a non-pandemic Vibrio cholerae isolate from World War One.

The sixth global cholera pandemic lasted from 1899 to 1923. However, despite widespread fear of the disease and of its negative effects on troop morale, very few soldiers in the British Expeditionary Forces contracted cholera between 1914 and 1918. Here, we have revived and sequenced the genome of NCTC 30, a 102-year-old Vibrio cholerae isolate, which we believe is the oldest publicly available live V. cholerae strain in existence. NCTC 30 was isolated in 1916 from a British soldier convalescent in Egypt. We found that this strain does not encode cholera toxin, thought to be necessary to cause cholera, and is not part of V. cholerae lineages responsible for the pandemic disease. We also show that NCTC 30, which predates the introduction of penicillin-based antibiotics, harbours a functional ß-lactamase antibiotic resistance gene. Our data corroborate and provide molecular explanations for previous phenotypic studies of NCTC 30 and provide a new high-quality genome sequence for historical, non-pandemic V. cholerae.

The extant World War 1 dysentery bacillus NCTC1: a genomic analysis.

BACKGROUND: Shigellosis (previously bacillary dysentery) was the primary diarrhoeal disease of World War 1, but outbreaks still occur in military operations, and shigellosis causes hundreds of thousands of deaths per year in developing nations. We aimed to generate a high-quality reference genome of the historical Shigella flexneri isolate NCTC1 and to examine the isolate for resistance to antimicrobials. METHODS: In this genomic analysis, we sequenced the oldest extant Shigella flexneri serotype 2a isolate using single-molecule real-time (SMRT) sequencing technology. Isolated from a soldier with dysentery from the British forces fighting on the Western Front in World War 1, this bacterium, NCTC1, was the first isolate accessioned into the National Collection of Type Cultures. We created a reference sequence for NCTC1, investigated the isolate for antimicrobial resistance, and undertook comparative genetics with S flexneri reference strains isolated during the 100 years since World War 1. FINDINGS: We discovered that NCTC1 belonged to a 2a lineage of S flexneri, with which it shares common characteristics and a large core genome. NCTC1 was resistant to penicillin and erythromycin, and contained a complement of chromosomal antimicrobial resistance genes similar to that of more recent isolates. Genomic islands gained in the S flexneri 2a lineage over time were predominately associated with additional antimicrobial resistances, virulence, and serotype conversion. INTERPRETATION: This S flexneri 2a lineage is a well adapted pathogen that has continued to respond to selective pressures. We have created a valuable historical benchmark for shigellae in the form of a high-quality reference sequence for a publicly available isolate. FUNDING: The Wellcome Trust.

NCTC3000: a century of bacterial strain collecting leads to a rich genomic data resource.

The National Collection of Type Cultures (NCTC) was founded on 1 January 1920 in order to fulfil a recognized need for a centralized repository for bacterial and fungal strains within the UK. It is among the longest-established collections of its kind anywhere in the world and today holds approximately 6000 type and reference bacterial strains - many of medical, scientific and veterinary importance - available to academic, health, food and veterinary institutions worldwide. Recently, a collaboration between NCTC, Pacific Biosciences and the Wellcome Sanger Institute established the NCTC3000 project to long-read sequence and assemble the genomes of up to 3000 NCTC strains. Here, at the beginning of the collection's second century, we introduce the resulting NCTC3000 sequence read datasets, genome assemblies and annotations as a unique, historically and scientifically relevant resource for the benefit of the international bacterial research community.

Complete genome sequences of Streptococcus pyogenes type strain reveal 100%-match between PacBio-solo and Illumina-Oxford Nanopore hybrid assemblies.

We present the first complete, closed genome sequences of Streptococcus pyogenes strains NCTC 8198T and CCUG 4207T, the type strain of the type species of the genus Streptococcus and an important human pathogen that causes a wide range of infectious diseases. S. pyogenes NCTC 8198T and CCUG 4207T are derived from deposit of the same strain at two different culture collections. NCTC 8198T was sequenced, using a PacBio platform; the genome sequence was assembled de novo, using HGAP. CCUG 4207T was sequenced and a de novo hybrid assembly was generated, using SPAdes, combining Illumina and Oxford Nanopore sequence reads. Both strategies yielded closed genome sequences of 1,914,862 bp, identical in length and sequence identity. Combining short-read Illumina and long-read Oxford Nanopore sequence data circumvented the expected error rate of the nanopore sequencing technology, producing a genome sequence indistinguishable to the one determined with PacBio. Sequence analyses revealed five prophage regions, a CRISPR-Cas system, numerous virulence factors and no relevant antibiotic resistance genes. These two complete genome sequences of the type strain of S. pyogenes will effectively serve as valuable taxonomic and genomic references for infectious disease diagnostics, as well as references for future studies and applications within the genus Streptococcus.

Draft Genome Sequences of 64 Type Strains of 50 Species and 25 Subspecies of the Genus Staphylococcus Rosenbach 1884.

Members of the genus Staphylococcus have been isolated from humans, animals, and the environment. Accurate identification with whole-genome sequencing requires access to data derived from type strains. We provide sequence data for type strains of 64 taxa in the genus that at the time of this writing have standing in the nomenclature.

Complete Whole-Genome Sequences of Two Raoultella terrigena Strains, NCTC 13097 and NCTC 13098, Isolated from Human Cases.

Raoultella terrigena is a bacterial species associated with soil and aquatic environments; however, sporadic cases of opportunistic disease in humans have been reported. Here, we report the first two complete genome sequences from clinical strains isolated from human sources that have been deposited in the National Collection of Type Cultures (NCTC).

Complete Whole-Genome Sequence of Haemophilus haemolyticus NCTC 10839.

Haemophilus haemolyticus is a Gram-negative bacterium that is a commensal of the respiratory tract in humans. Here, we report the complete genome sequence available for Haemophilus haemolyticus strain NCTC 10839, which was originally isolated from the nasopharynx of a child.

Whole-Genome Sequences of Five Strains of Kocuria rosea, NCTC2676, NCTC7514, NCTC7512, NCTC7528, and NCTC7511.

Kocuria rosea is a Gram-positive coccus found in the environment and within normal human skin microbiota, and more recently, it has been potentially implicated as an opportunistic pathogen. Here, we describe the genome sequences of five strains of K. rosea (NCTC2676, NCTC7514, NCTC7512, NCTC7528, and NCTC7511).

Assessment of the stability of cell-surface components of microorganisms by MALDI-TOF-MS following preservation on lenticule discs.

Strains representing the species Campylobacter coli, Escherichia coli, Listeria monocytogenes, Pseudomonas aeruginosa, Salmonella enterica, and Staphylococcus aureus were randomly selected to assess the consistency of cells preserved on lenticule discs to those archived in traditional freeze-dried ampoules. Each matched pair was cultured using identical conditions and analysed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MS) to profile the surface-associated molecules of the cells. In addition, the cytosolic/membrane-bound proteins of C. coli and S. aureus strains were further analysed by surface-enhanced laser desorption/ionization time-of-flight MS. The mass spectral profiles in all cases showed a high degree of concordance between cells preserved by both methods and suggest that the properties of cells preserved on lenticule disc are consistent with those archived by the traditional method of freeze-drying.

Mycobacterial biomaterials and resources for researchers.

There are many resources available to mycobacterial researchers, including culture collections around the world that distribute biomaterials to the general scientific community, genomic and clinical databases, and powerful bioinformatics tools. However, many of these resources may be unknown to the research community. This review article aims to summarize and publicize many of these resources, thus strengthening the quality and reproducibility of mycobacterial research by providing the scientific community access to authenticated and quality-controlled biomaterials and a wealth of information, analytical tools and research opportunities.

Genome-based characterization of hospital-adapted Enterococcus faecalis lineages.

Vancomycin-resistant Enterococcus faecalis (VREfs) is an important nosocomial pathogen(1,2). We undertook whole genome sequencing of E. faecalis associated with bloodstream infection in the UK and Ireland over more than a decade to determine the population structure and genetic associations with hospital adaptation. Three lineages predominated in the population, two of which (L1 and L2) were nationally distributed, and one (L3) geographically restricted. Genome comparison with a global collection identified that L1 and L3 were also present in the USA, but were genetically distinct. Over 90% of VREfs belonged to L1-L3, with resistance acquired and lost multiple times in L1 and L2, but only once followed by clonal expansion in L3. Putative virulence and antibiotic resistance genes were over-represented in L1, L2 and L3 isolates combined, versus the remainder. Each of the three main lineages contained a mixture of vancomycin-resistant and -susceptible E. faecalis (VSEfs), which has important implications for infection control and antibiotic stewardship.

Complete Genome Sequence of Neisseria weaveri Strain NCTC13585.

Neisseria weaveri is a commensal organism of the canine oral cavity and an occasional opportunistic human pathogen which is associated with dog bite wounds. Here we report the first complete genomic sequence of the N. weaveri NCTC13585 (CCUG30381) strain, which was originally isolated from a patient with a canine bite wound.

Complete Genome Sequence of Plesiomonas shigelloides Type Strain NCTC10360.

Plesiomonas shigelloides is a Gram-negative rod within the Enterobacteriaceae family. It is a gastrointestinal pathogen of increasing notoriety, often associated with diarrheal disease. P. shigelloides is waterborne, and infection is often linked to the consumption of seafood. Here, we describe the first complete genome for P. shigelloides type strain NCTC10360.

Genome-based characterization of hospital-adapted Enterococcus faecalis lineages.

Vancomycin-resistant Enterococcus faecalis (VREfs) is an important nosocomial pathogen1,2. We undertook whole genome sequencing of E. faecalis associated with bloodstream infection in the UK and Ireland over more than a decade to determine the population structure and genetic associations with hospital adaptation. Three lineages predominated in the population, two of which (L1 and L2) were nationally distributed, and one (L3) geographically restricted. Genome comparison with a global collection identified that L1 and L3 were also present in the USA, but were genetically distinct. Over 90% of VREfs belonged to L1-L3, with resistance acquired and lost multiple times in L1 and L2, but only once followed by clonal expansion in L3. Putative virulence and antibiotic resistance genes were over-represented in L1, L2 and L3 isolates combined, versus the remainder. Each of the three main lineages contained a mixture of vancomycin-resistant and -susceptible E. faecalis (VSEfs), which has important implications for infection control and antibiotic stewardship.

Complete Whole-Genome Sequence of Salmonella enterica subsp. enterica Serovar Java NCTC5706.

Salmonellae are a significant cause of morbidity and mortality globally. Here, we report the first complete genome sequence for Salmonella enterica subsp. enterica serovar Java strain NCTC5706. This strain is of historical significance, having been isolated in the pre-antibiotic era and was deposited into the National Collection of Type Cultures in 1939.

Global phylogeography and evolutionary history of Shigella dysenteriae type 1.

Together with plague, smallpox and typhus, epidemics of dysentery have been a major scourge of human populations for centuries(1). A previous genomic study concluded that Shigella dysenteriae type 1 (Sd1), the epidemic dysentery bacillus, emerged and spread worldwide after the First World War, with no clear pattern of transmission(2). This is not consistent with the massive cyclic dysentery epidemics reported in Europe during the eighteenth and nineteenth centuries(1,3,4) and the first isolation of Sd1 in Japan in 1897(5). Here, we report a whole-genome analysis of 331 Sd1 isolates from around the world, collected between 1915 and 2011, providing us with unprecedented insight into the historical spread of this pathogen. We show here that Sd1 has existed since at least the eighteenth century and that it swept the globe at the end of the nineteenth century, diversifying into distinct lineages associated with the First World War, Second World War and various conflicts or natural disasters across Africa, Asia and Central America. We also provide a unique historical perspective on the evolution of antibiotic resistance over a 100-year period, beginning decades before the antibiotic era, and identify a prevalent multiple antibiotic-resistant lineage in South Asia that was transmitted in several waves to Africa, where it caused severe outbreaks of disease.

The Murray collection of pre-antibiotic era Enterobacteriacae: a unique research resource.

Studies of historical isolates inform on the evolution and emergence of important pathogens and phenotypes, including antimicrobial resistance. Crucial to studying antimicrobial resistance are isolates that predate the widespread clinical use of antimicrobials. The Murray collection of several hundred bacterial strains of pre-antibiotic era Enterobacteriaceae is an invaluable resource of historical strains from important pathogen groups. Studies performed on the Collection to date merely exemplify its potential, which will only be realised through the continued effort of many scientific groups. To enable that aim, we announce the public availability of the Murray collection through the National Collection of Type Cultures, and present associated metadata with whole genome sequence data for over half of the strains. Using this information we verify the metadata for the collection with regard to subgroup designations, equivalence groupings and plasmid content. We also present genomic analyses of population structure and determinants of mobilisable antimicrobial resistance to aid strain selection in future studies. This represents an invaluable public resource for the study of these important pathogen groups and the emergence and evolution of antimicrobial resistance.