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Streptococcus mitisis a common oral commensal and an opportunistic pathogen that causes bacteremia and infective endocarditis; however, the species has received little attention compared to other pathogenic streptococcal species. Effective and easy-to-use molecular typing tools are essential for understanding bacterial population diversity and biology, but schemes specific forS. mitisare not currently available. We therefore developed a multilocus sequence typing (MLST) scheme and defined sequence clusters or lineages ofS. mitisusing a comprehensive global data set of 322 genomes (148 publicly available and 174 newly sequenced). We used internal 450-bp sequence fragments of seven housekeeping genes (accA,gki,hom,oppC,patB,rlmN, andtsf) to define the MLST scheme and derived the globalS. mitissequence clusters using the PopPUNK clustering algorithm. We identified an initial set of 259 sequence types (STs) and 258 global sequence clusters. The schemes showed high concordance (100%), capturing extensiveS. mitisdiversity with strains assigned to multiple unique STs and global sequence clusters. The tools also identified extensive within- and between-hostS. mitisgenetic diversity among isolates sampled from a cohort of healthy individuals, together with potential transmission events, supported by both phylogeny and pairwise single nucleotide polymorphism (SNP) distances. Our novel molecular typing and strain clustering schemes forS. mitisallow for the integration of new strain data, are electronically portable at the PubMLST database (https://pubmlst.org/smitis), and offer a standardized approach to understanding the population structure ofS. mitis. These robust tools will enable new insights into the epidemiology ofS. mitiscolonization, disease and transmission.

BACKGROUNDRespiratory syncytial virus (RSV) is the leading cause of hospitalisation associated with acute respiratory infection in infants and young children, with substantial disease burden globally. The impact of additional respiratory pathogens on RSV disease severity is not completely understood.OBJECTIVESThe objective of this study was to explore the associations between RSV disease severity and the presence of other respiratory pathogens.METHODSNasopharyngeal swabs were prospectively collected from two infant cohorts: a prospective longitudinal birth cohort study and an infant cross-sectional study recruiting infants RESULTSRSV was detected in 433 infants. Nearly one in four of the infants (24%) harboured at least one additional non-RSV respiratory virus, with human rhinovirus being the most frequently detected (15% of the infants), followed by seasonal coronaviruses (4%). In this cohort, RSV-infected infants harbouring any other virus tended to be older (median age: 4.3 vs. 3.7 months) and were more likely to require intensive care and mechanical ventilation than those who did not.Moraxella, Streptococcus, andHaemophilusspecies were the most frequently identified target bacteria, together found in 392 (91%) of the 433 infants (S. pneumoniaein 51% of the infants andH. influenzaein 38%). The strongest contributors to severity of presentation were younger age and the co-detection ofHaemophilusspecies alongside RSV. Across all age groups in both cohorts, detection ofHaemophilusspecies was associated with higher overall severity, as captured by ReSVinet scores, and specifically with increased rates of hospitalisation and respiratory distress. In contrast, presence ofMoraxellaspecies was associated with lower ReSVinet scores and reduced need for intensive care and mechanical ventilation. Infants with and withoutStreptococcusspecies (orS. pneumoniaein particular) had similar clinical outcomes. No specific RSV strain was associated with co-detection of other pathogens.CONCLUSIONOur findings provide strong evidence for associations between RSV disease severity and the presence of additional respiratory viruses and bacteria. The associations, while not indicating causation, are of potential clinical relevance. Awareness of coexisting microorganisms could inform therapeutic and preventive measures to improve the management and outcome of RSV-infected infants.

SummaryBackgroundThe Invasive Respiratory Infection Surveillance (IRIS) Consortium was established to assess the impact of the COVID-19 pandemic on invasive diseases caused byStreptococcus pneumoniae, Haemophilus influenzae, Neisseria meningitidisandStreptococcus agalactiae. Here we analyse the incidence and distribution of disease during the first two years of the pandemic.MethodsLaboratories in 30 countries/territories representing five continents submitted case data from 2018-2021 to private projects within databases in PubMLST. The impact of COVID-19 containment measures on the overall number of cases was analysed, and changes in disease distributions by patient age and serotype/group were examined. Interrupted time series analyses quantified the impact of pandemic response measures and their relaxation on disease rates, and autoregressive integrated moving average models estimated effect sizes and forecasted counterfactual trends by hemisphere.FindingsOverall, 116,841 cases were analysed: 76,481 (2018-2019, pre-pandemic) plus 40,360 (2020-2021, pandemic). During the pandemic there was a significant reduction in the risk of disease caused byS pneumoniae(risk ratio: 0.47; 95% confidence interval: 0.40-0.55),H influenzae(0.51; 0.40-0.66) andN meningitidis(0.26; 0.21-0.31), whereas no significant changes were observed for the non-respiratory-transmitted pathogenS agalactiae(1.02; 0.75-1.40). No major changes in the distribution of cases were observed when stratified by patient age or serotype/group. An estimated 36,289 (17,145-55,434) cases of invasive bacterial disease were averted during the first two years of the pandemic among IRIS participating countries/territories.InterpretationCOVID-19 containment measures were associated with a sustained decrease in the incidence of invasive disease caused byS pneumoniae, H influenzaeandN meningitidisduring the first two years of the pandemic, but cases began to increase in some countries as pandemic restrictions were lifted.Research in contextEvidence before this studyEarly in the COVID-19 pandemic the IRIS Consortium reported a significant reduction in invasive disease due to respiratory-transmitted bacterial pathogens, which was associated with the implementation of COVID-19 stringency measures and changes in human social behaviour. All 26 countries/territories participating in IRIS at the time experienced a significant reduction in infections between January and May 2020, compared with the previous two years. In particular,S pneumoniaeinfections decreased by 68% at four weeks after COVID-19 containment measures were imposed, and by 82% at eight weeks.Added value of this studyThese new data from the expanded IRIS Consortium across 30 countries/territories demonstrated a sustained reduction in invasive disease throughout the first two years of the COVID-19 pandemic. Using time series modelling, we estimated that over 36,000 cases of invasive bacterial disease were averted in 2020-2021 among the countries participating in IRIS; however, minor increases in disease in the latter half of 2021 require close monitoring to understand the nature of re-emerging cases.Implications of all the available evidenceFuture epidemics and pandemics will occur, and we need to understand not only the pathogen that is directly responsible for the pandemic, but also that population-level responses to an epidemic or pandemic more broadly affect overall human health and other microbes. IRIS provides evidence for the effects of such public health responses on severe invasive bacterial infections across many countries. Moreover, these IRIS data provide a better understanding of microbial transmission, will inform vaccine development and implementation, and can contribute to healthcare service planning and provision of policies.

Bacterial genomics is making an increasing contribution to the fields of medicine and public health microbiology. Consequently, accurate species identification of bacterial genomes is an important task, particularly as the number of genomes stored in online databases increases rapidly and new species are frequently discovered. Existing database entries require regular re-evaluation to ensure that species annotations are consistent with the latest species definitions. We have developed an automated method for bacterial species identification that is an extension of ribosomal multilocus sequence typing (rMLST). The method calculates an 'rMLST nucleotide identity' (rMLST-NI) based on the nucleotides present in the protein-encoding ribosomal genes derived from bacterial genomes. rMLST-NI was used to validate the species annotations of 11839 publicly available

Brucellosis poses a significant burden to human and animal health worldwide. Robust and harmonized molecular epidemiological approaches and population studies that include routine disease screening are needed to efficiently track the origin and spread of Brucella strains. Core genome multilocus sequence typing (cgMLST) is a powerful genotyping system commonly used to delineate pathogen transmission routes for disease surveillance and control. Except for Brucella melitensis, cgMLST schemes for Brucella species are currently not established. Here, we describe a novel cgMLST scheme that covers multiple Brucella species. We first determined the phylogenetic breadth of the genus using 612 Brucella genomes. We selected 1,764 genes that were particularly well conserved and typeable in at least 98% of these genomes. We tested the new scheme on 600 genomes and found high agreement with the whole-genome-based single nucleotide polymorphism (SNP) analysis. Next, we applied the scheme to reanalyze the genome of Brucella strains from epidemiologically linked outbreaks. We demonstrated the applicability of the new scheme for high-resolution typing required in outbreak investigations as previously reported with whole-genome SNP methods. We also used the novel scheme to define the global population structure of the genus using 1,322 Brucella genomes. Finally, we demonstrated the possibility of tracing distribution of Brucella strains by performing cluster analysis of cgMLST profiles and found nearly identical cgMLST profiles in different countries. Our results show that sequencing depth of more than 40-fold is optimal for allele calling with this scheme. In summary, this study describes a novel Brucella-wide cgMLST scheme that is applicable in Brucella molecular epidemiology and helps in accurately tracking and thus controlling the sources of infection. The scheme is publicly accessible and should represent a valuable resource for laboratories with limited computational resources and bioinformatics expertise.

Objective:Serogroup W and Y invasive meningococcal disease increased globally from 2000 onwards. Responding to a rapid increase in serogroup W clonal complex 11 (W:cc11) invasive meningococcal disease, the UK replaced an adolescent booster dose of meningococcal C conjugate vaccine with quadrivalent MenACWY conjugate vaccine in 2015. By 2018, the vaccine coverage in the eligible school cohorts aged 14 to 19years was 84%. We assessed the impact of the MenACWY vaccination programme on meningococcal carriage. Methods:An observational study of culture-defined oropharyngeal meningococcal carriage prevalence before and after the start of the MenACWY vaccination programme in UK school students, aged 15 to 19years, using two cross-sectional studies: 2014 to 2015 “UKMenCar4” and 2018 “Be on the TEAM” (ISRCTN75858406). Results:A total of 10625 participants preimplementation and 13438 postimplementation were included. Carriage of genogroups C, W, and Y (combined) decreased from 2.03 to 0.71% (OR 0.34 [95% CI 0.27–0.44], p.those expressing capsule) decreased for genogroup W by 53.8% (95% CI -5.0 – 79.8, p=0.016) and for genogroup Y by 30.1% (95% CI 8.946·3, p=0.0025). Discussion:The UK MenACWY vaccination programme reduced carriage acquisition of genogroup and serogroup Y and W meningococci and sustained low levels of genogroup C carriage. These data support the use of quadrivalent MenACWY conjugate vaccine for indirect (herd) protection.

Expansion of quinolone-resistant Neisseria meningitidis clone ChinaCC4821-R1-C/B from sequence type (ST) 4821 clonal complex (CC4821) caused a serogroup shift from serogroup A to serogroup C invasive meningococcal disease (IMD) in China. To determine the relationship among globally distributed CC4821 meningococci, we analyzed whole-genome sequence data from 173 CC4821 meningococci isolated from 4 continents during 1972-2019. These meningococci clustered into 4 sublineages (1-4); sublineage 1 primarily comprised of IMD isolates (41/50, 82%). Most isolates from outside China (40/49, 81.6%) formed a distinct sublineage, the Europe-USA cluster, with the typical strain designation B:P1.17-6,23:F3-36:ST-3200(CC4821), harboring mutations in penicillin-binding protein 2. These data show that the quinolone-resistant clone ChinaCC4821-R1-C/B has expanded to other countries. The increasing distribution worldwide of serogroup B CC4821 raises the concern that CC4821 has the potential to cause a pandemic that would be challenging to control, despite indirect evidence that the Trumenba vaccine might afford some protection.

Neisseria gonorrhoeae (the gonococcus) is the causative agent of the sexually-transmitted infection gonorrhoea, and has developed resistance to all classes of antimicrobials. In gonococci, plasmids can mediate high-level antimicrobial resistance (AMR) to tetracyclines and ß-lactams. Plasmids can spread through bacterial populations by transformation and conjugation, resulting in the rapid dissemination of traits. Characterisation of plasmids, including understanding their distribution in bacterial populations, is therefore key to understanding bacterial evolution, and in particular the spread of AMR. N. gonorrhoeae can harbour three plasmids, conjugative (pConj), ß-lactamase (pbla) and cryptic (pCryp). Using genomic and phylogenetic analyses, we show that plasmids are widespread in a large collection of gonococcal isolates from 56 countries. We found that variants of pConj (which can mediate tetracycline resistance) and pbla expressing TEM-135 ß-lactamase are associated with distinct gonococcal lineages. Furthermore, AMR plasmids are significantly more prevalent in gonococci from less wealthy countries. Over 94% of gonococci possess the cryptic plasmid (pCryp), and its absence can be correlated with the presence of a novel chromosomal Type IV secretion system. Our results reveal the extent of plasmid-mediated AMR in the gonococcus, particularly in less wealthy countries, where diagnostic and therapeutic options can be limited, and highlight the risk of their global spread.

Clostridioides difficile remains a key cause of healthcare-associated infection, with multi-drug-resistant (MDR) lineages causing high mortality (≥20%) outbreaks. Cephalosporin treatment is a long-established risk factor, and antimicrobial stewardship a key control. A mechanism underlying raised cephalosporin MICs has not been identified in C. difficile, but among other species resistance is often acquired via amino acid substitutions in cell wall transpeptidases (penicillin binding proteins, PBPs). Here, we investigated five C. difficile transpeptidases (PBP1-5) for recent substitutions. Previously published genome assemblies (n=7096) were obtained, representing sixteen geographically widespread lineages, including healthcare-associated MDR ST1(027), ST3(001) and ST17(018). Recent amino acid substitutions were found within PBP1 (n=50) and PBP3 (n=48), ranging from 1-10 substitutions per genome. β-lactam MICs were measured for closely related pairs of wild-type and PBP substituted isolates separated by 20-273 SNPs. Recombination-corrected, dated phylogenies were constructed to date substitution acquisition. Key substitutions such as PBP3 V497L and PBP1 T674I/N/V emerged independently across multiple lineages. They were associated with extremely high cephalosporin MICs; 1-4 doubling dilutions >wild-type up to ≤1506μg/ml. Substitution patterns varied by lineage and clade, showed geographic structure, and notably occurred post-1990, coincident with the acquisition of gyrA/B substitutions conferring fluoroquinolone resistance. In conclusion, recent PBP1 and PBP3 substitutions are associated with raised cephalosporin MICs in C. difficile. The co-occurrence of resistance to cephalosporins and fluoroquinolones hinders attempts to understand their relative importance in the dissemination of epidemic lineages. Further controlled studies of cephalosporin and fluoroquinolone stewardship are needed to determine their relative effectiveness in outbreak control.IMPORTANCEFluoroquinolone and cephalosporin prescribing in healthcare settings have triggered outbreaks of high-mortality, multi-drug resistant C. difficile infection. Here, we identify a mechanism of acquired cephalosporin resistance in C. difficile, comprising amino acid substitutions in two cell-wall transpeptidase enzymes (penicillin binding proteins). The higher the number of substitutions, the greater the impact on phenotype. Dated phylogenies revealed that resistance to both cephalosporins and fluoroquinolones was co-acquired immediately before clinically important, outbreak strains emerged. PBP substitutions were geographically structured within genetic lineages, suggesting adaptation to local antimicrobial prescribing. Antimicrobial stewardship of cephalosporins and fluoroquinolones is an effective means of C. difficile outbreak control. Genetic changes conferring resistance likely impart a ‘fitness-cost’ after antibiotic withdrawal. Our study identifies a mechanism that may explain the contribution of cephalosporin stewardship to resolving outbreak conditions. However, due to the co-occurrence of cephalosporin and fluoroquinolone resistance, further work is needed to determine the relative importance of each.

Background Gonorrhea, caused by the bacterium Neisseria gonorrhoeae, is a globally prevalent sexually transmitted infection. The dynamics of gonococcal population biology have been poorly defined due to a lack of resolution in strain typing methods. Methods In this study, we assess how the core genome can be used to improve our understanding of gonococcal population structure compared with current typing schemes. Results A total of 1668 loci were identified as core to the gonococcal genome. These were organized into a core genome multilocus sequence typing scheme (N gonorrhoeae cgMLST v1.0). A clustering algorithm using a threshold of 400 allelic differences between isolates resolved gonococci into discrete and stable core genome groups, some of which persisted for multiple decades. These groups were associated with antimicrobial genotypes and non-overlapping NG-STAR and NG-MAST sequence types. The MLST-STs were more widely distributed among core genome groups. Conclusions Clustering with cgMLST identified globally distributed, persistent, gonococcal lineages improving understanding of the population biology of gonococci and revealing its population structure. These findings have implications for the emergence of antimicrobial resistance in gonococci and how this is associated with lineages, some of which are more predisposed to developing antimicrobial resistance than others.

Horizontal gene transfer (HGT) can allow traits that have evolved in one bacterial species to transfer to another. This has potential to rapidly promote new adaptive trajectories such as zoonotic transfer or antimicrobial resistance. However, for this to occur requires gaps to align in barriers to recombination within a given time frame. Chief among these barriers is the physical separation of species with distinct ecologies in separate niches. Within the genus Campylobacter, there are species with divergent ecologies, from rarely isolated single-host specialists to multihost generalist species that are among the most common global causes of human bacterial gastroenteritis. Here, by characterizing these contrasting ecologies, we can quantify HGT among sympatric and allopatric species in natural populations. Analyzing recipient and donor population ancestry among genomes from 30 Campylobacter species, we show that cohabitation in the same host can lead to a six-fold increase in HGT between species. This accounts for up to 30% of all SNPs within a given species and identifies highly recombinogenic genes with functions including host adaptation and antimicrobial resistance. As described in some animal and plant species, ecological factors are a major evolutionary force for speciation in bacteria and changes to the host landscape can promote partial convergence of distinct species through HGT.

BackgroundThe re-emergence of whooping cough and geographic disparities in vaccine escape or antimicrobial resistance dynamics, underline the importance of a unified definition of Bordetella pertussis strains. Understanding of the evolutionary adaptations of Bordetella pathogens to humans and animals requires comparative studies with environmental bordetellae.MethodsWe have set-up a unified library of Bordetella genomes by merging previously existing Oxford and Pasteur databases, importing genomes from public repositories, and developing harmonized genotyping schemes. We developed a genus-wide cgMLST genotyping scheme and incorporated a previous B. pertussis cgMLST scheme. Specific schemes were developed to define antigenic, virulence and macrolide resistance profiles. Genomic sequencing of 83 French B. bronchiseptica isolates and of B. tumulicola, B. muralis and B. tumbae type strains was performed.ResultsThe public library currently includes 2,581 Bordetella isolates and their provenance data, and 2,084 genomes. The “classical Bordetella” (B. bronchiseptica, B. parapertussis and B. pertussis), which form a single genomic species (B. bronchiseptica genomic species, BbGS), were overrepresented (n=2,382). The phylogenetic analysis of Bordetella genomes associated the three novel species B. tumulicola, B. muralis and B. tumbae in a clade with B. petrii and revealed 18 yet undescribed species. A sister lineage of the classical bordetellae, provisionally named Bbs lineage II, was uncovered and may represent a novel species (average nucleotide identity with BbGS strains: ∼95%). It comprised strain HT200 from India, two strains of ‘genogroup 6’ from the USA and six clinical isolates from France; this lineage lacked ptxP and its fim2 gene was divergent. Within B. pertussis, vaccine antigen sequence types marked important phylogenetic subdivisions, and macrolide resistance markers (23S_rRNA allele 13 and fhaB3) confirmed the current restriction of this phenotype in China with few exceptions.ConclusionsThe genomic platform provides an expandable resource for unified genotyping of Bordetella strains and will facilitate collective evolutionary and epidemiological understanding of the re-emergence of whooping cough and other Bordetella infections.Data summaryBordetella genomes list and accession numbers: Supplementary Table S4Bordetella genus phylogeny dataset (92 isolates):https://bigsdb.pasteur.fr/cgi-bin/bigsdb/bigsdb.pl?db=pubmlst_bordetella_isolates&page=query&project_list=23&submit=1B. bronchiseptica phylogeny dataset (213 isolates):https://bigsdb.pasteur.fr/cgi-bin/bigsdb/bigsdb.pl?db=pubmlst_bordetella_isolates&page=query&project_list=24&submit=1B. pertussis phylogeny (124 isolates):https://bigsdb.pasteur.fr/cgi-bin/bigsdb/bigsdb.pl?db=pubmlst_bordetella_isolates&page=query&project_list=25&submit=1iTOL interactive trees: https://itol.embl.de/shared/1l7Fw0AvKOoCF

Clostridium perfringens is a spore-forming anaerobic pathogen responsible for a variety of histotoxic and intestinal infections in humans and animals. High-resolution genotyping aiming to identify bacteria at strain level has become increasingly important in modern microbiology to understand pathogen transmission pathways and to tackle infection sources. This study aimed at establishing a publicly available genome-wide multilocus sequence-typing (MLST) scheme for C. perfringens. A total of 1,431 highly conserved core genes (1.34 megabases; 50% of the reference genome genes) were indexed for a core genome-based MLST (cgMLST) scheme for C. perfringens. The scheme was applied to 282 ecologically and geographically diverse genomes, showing that the genotyping results of cgMLST were highly congruent with the core genome-based single-nucleotide-polymorphism typing in terms of resolution and tree topology. In addition, the cgMLST provided a greater discrimination than classical MLST methods for C. perfringens. The usability of the scheme for outbreak analysis was confirmed by reinvestigating published outbreaks of C. perfringens-associated infections in the United States and the United Kingdom. In summary, a publicly available scheme and an allele nomenclature database for genomic typing of C. perfringens have been established and can be used for broad-based and standardized epidemiological studies. IMPORTANCE: Global epidemiological surveillance of bacterial pathogens is enhanced by the availability of standard tools and sharing of typing data. The use of whole-genome sequencing has opened the possibility for high-resolution characterization of bacterial strains down to the clonal and subclonal levels. Core genome multilocus sequence typing is a robust system that uses highly conserved core genes for deep genotyping. The method has been successfully and widely used to describe the epidemiology of various bacterial species. Nevertheless, a cgMLST typing scheme for Clostridium perfringens is currently not publicly available. In this study, we (i) developed a cgMLST typing scheme for C. perfringens, (ii) evaluated the performance of the scheme on different sets of C. perfringens genomes from different hosts and geographic regions as well as from different outbreak situations, and, finally, (iii) made this scheme publicly available supported by an allele nomenclature database for global and standard genomic typing.

Mycobacterium bovis (M. bovis) is a causative agent of bovine tuberculosis, a significant source of morbidity and mortality in the global cattle industry. The Randomised Badger Culling Trial was a field experiment carried out between 1998 and 2005 in the South West of England. As part of this trial, M. bovis isolates were collected from contemporaneous and overlapping populations of badgers and cattle within ten defined trial areas. We combined whole genome sequences from 1,442 isolates with location and cattle movement data, identifying transmission clusters and inferred rates and routes of transmission of M. bovis. Most trial areas contained a single transmission cluster that had been established shortly before sampling, often contemporaneous with the expansion of bovine tuberculosis in the 1980s. The estimated rate of transmission from badger to cattle was approximately two times higher than from cattle to badger, and the rate of within-species transmission considerably exceeded these for both species. We identified long distance transmission events linked to cattle movement, recurrence of herd breakdown by infection within the same transmission clusters and superspreader events driven by cattle but not badgers. Overall, our data suggests that the transmission clusters in different parts of South West England that are still evident today were established by long-distance seeding events involving cattle movement, not by recrudescence from a long-established wildlife reservoir. Clusters are maintained primarily by within-species transmission, with less frequent spill-over both from badger to cattle and cattle to badger., Author summary Bovine tuberculosis (bTB), predominantly caused by Mycobacterium bovis, is a significant cause of sickness and death amongst cattle globally. Due to its broad host range which includes a variety of wildlife species such as the Eurasian badger, control measures for bTB are often both ineffective and expensive. One method used to attempt to control the spread of bTB in Great Britain is to cull badgers in areas adjacent to infected herds. To assess the effect of badger culling on the prevalence of bTB in nearby herds, the Randomised Badger Culling Trial (RBCT) was set up by the British government and ran between 1998 and 2005. Here we use 1,442 whole genome sequences obtained from isolates collected as part of the RBCT to describe the population structure of Mycobacterium bovis in the trial areas, identify putative transmission clusters and estimate the directionality of transmission, and integrate the genomic data with associated cattle movement data to identify examples of recurrence, superspreading and long-distance transmission. We found that the transmission clusters identified were strongly associated with RBCT trial area and that, whilst badger to cattle transmission was broadly more common than vice versa, the majority of transmission was occurring within the herds or badger populations. Additionally, molecular dating of the transmission clusters showed that the clusters were likely seeded during the 1980s, a period during which the prevalence of bTB in Great Britain increased markedly. Our study provides novel insights into the transmission of bTB between cattle and badgers in Great Britain.

Horizontal gene transfer (HGT) can allow traits that have evolved in one bacterial species to transfer to another. This has potential to rapidly promote new adaptive trajectories such as zoonotic transfer or antimicrobial resistance. However, for this to occur requires gaps to align in barriers to recombination within a given time frame. Chief among these barriers is the physical separation of species with distinct ecologies in separate niches. Within the genus

Horizontal gene transfer (HGT) can allow traits that have evolved in one bacterial species to transfer to another. This has potential to rapidly promote new adaptive trajectories such as zoonotic transfer or antimicrobial resistance. However, for this to occur requires gaps to align in barriers to recombination within a given time frame. Chief among these barriers is the physical separation of species with distinct ecologies in separate niches. Within the genus Campylobacter there are species with divergent ecologies, from rarely isolated single host specialists to multi-host generalist species that are among the most common global causes of human bacterial gastroenteritis. Here, by characterising these contrasting ecologies, we can quantify HGT among sympatric and allopatric species in natural populations. Analysing recipient and donor population ancestry among genomes from 30 Campylobacter species we show that cohabitation in the same host can lead to a 6-fold increase in HGT between species. This accounts for up to 30% of all SNPs within a given species and identifies highly recombinogenic genes with functions including host adaptation and antimicrobial resistance. As described in some animal and plant species, ecological factors are a major evolutionary force for speciation in bacteria and changes to the host landscape can promote partial convergence of distinct species through HGT.

Whole-genome sequencing (WGS) has been established for bacterial subtyping and is regularly used to study pathogen transmission, to investigate outbreaks, and to perform routine surveillance. Core-genome multilocus sequence typing (cgMLST) is a bacterial subtyping method that uses WGS data to provide a high-resolution strain characterization. This study aimed at developing a novel cgMLST scheme for Bacillus anthracis, a notorious pathogen that causes anthrax in livestock and humans worldwide. The scheme comprises 3,803 genes that were conserved in 57 B. anthracis genomes spanning the whole phylogeny. The scheme has been evaluated and applied to 584 genomes from 50 countries. On average, 99.5% of the cgMLST targets were detected. The cgMLST results confirmed the classical canonical single-nucleotide-polymorphism (SNP) grouping of B. anthracis into major clades and subclades. Genetic distances calculated based on cgMLST were comparable to distances from whole-genome-based SNP analysis with similar phylogenetic topology and comparable discriminatory power. Additionally, the application of the cgMLST scheme to anthrax outbreaks from Germany and Italy led to a definition of a cutoff threshold of five allele differences to trace epidemiologically linked strains for cluster typing and transmission analysis. Finally, the association of two clusters of B. anthracis with human cases of injectional anthrax in four European countries was confirmed using cgMLST. In summary, this study presents a novel cgMLST scheme that provides high-resolution strain genotyping for B. anthracis. This scheme can be used in parallel with SNP typing methods to facilitate rapid and harmonized interlaboratory comparisons, essential for global surveillance and outbreak analysis. The scheme is publicly available for application by users, including those with little bioinformatics knowledge.

Background: The incidence of invasive meningococcal disease in the UK decreased by approximately four times from 1999 to 2014, with reductions in serogroup C and serogroup B disease. Lower serogroup C invasive meningococcal disease incidence was attributable to implementation of the meningococcal serogroup C conjugate vaccine in 1999, through direct and indirect protection, but no vaccine was implemented against serogroup B disease. UK Meningococcal Carriage surveys 1–3 (UKMenCar1–3), conducted in 1999, 2000, and 2001, were essential for understanding the impact of vaccination. To investigate the decline in invasive meningococcal disease incidence, we did a large oropharyngeal carriage survey in 2014–15, immediately before the changes to meningococcal vaccines in the UK national immunisation schedule. Methods: UKMenCar4 was a cross-sectional survey in adolescents aged 15–19 years who were enrolled from schools and colleges geographically local to one of 11 UK sampling centres between Sept 1, 2014, and March 30, 2015. Participants provided an oropharyngeal swab sample and completed a questionnaire on risk factors for carriage, including social behaviours. Samples were cultured for putative Neisseria spp, which were characterised with serogrouping and whole-genome sequencing. Data from this study were compared with the results from the UKMenCar1–3 surveys (1999–2001). Findings: From the 19 641 participants (11 332 female, 8242 male, 67 not stated) in UKMenCar4 with culturable swabs and completed risk-factor questionnaires, 1420 meningococci were isolated, with a carriage prevalence of 7·23% (95% CI 6·88–7·60). Carriage prevalence was substantially lower in UKMenCar4 than in the previous surveys: carriage prevalence was 16·6% (95% CI 15·89–17·22; 2306/13 901) in UKMenCar1 (1999), 17·6% (17·05–18·22; 2873/16 295) in UKMenCar2 (2000), and 18·7% (18·12–19·27; 3283/17 569) in UKMenCar3 (2001). Carriage prevalence was lower for all serogroups in UKMenCar4 than in UKMenCar1–3, except for serogroup Y, which was unchanged. The prevalence of carriage-promoting social behaviours decreased from 1999 to 2014–15, with individuals reporting regular cigarette smoking decreasing from 2932 (21·5%) of 13 650 to 2202 (11·2%) of 19 641, kissing in the past week from 6127 (44·8%) of 13 679 to 7320 (37·3%) of 19 641, and attendance at pubs and nightclubs in the past week from 8436 (62·1%) of 13 594 to 7662 (39·0%) of 19 641 (all p Interpretation: We show that meningococcal carriage prevalence in adolescents sampled nationally during a low incidence period (2014–15) was less than half of that in an equivalent population during a high incidence period (1999–2001). Disease and carriage caused by serogroup C was well controlled by ongoing vaccination. The prevalence of behaviours associated with carriage declined, suggesting that public health policies aimed at influencing behaviour might have further reduced disease. Funding: Wellcome Trust, UK Department of Health, and National Institute for Health Research.

Chickens are the most common birds on Earth and colibacillosis is among the most common diseases affecting them. This major threat to animal welfare and safe sustainable food production is difficult to combat because the etiological agent, avian pathogenic Escherichia coli (APEC), emerges from ubiquitous commensal gut bacteria, with no single virulence gene present in all disease-causing isolates. Here, we address the underlying evolutionary mechanisms of extraintestinal spread and systemic infection in poultry. Combining population scale comparative genomics and pangenome-wide association studies, we compare E. coli from commensal carriage and systemic infections. We identify phylogroup-specific and species-wide genetic elements that are enriched in APEC, including pathogenicity-associated variation in 143 genes that have diverse functions, including genes involved in metabolism, lipopolysaccharide synthesis, heat shock response, antimicrobial resistance and toxicity. We find that horizontal gene transfer spreads pathogenicity elements, allowing divergent clones to cause infection. Finally, a Random Forest model prediction of disease status (carriage vs. disease) identifies pathogenic strains in the emergent ST-117 poultry-associated lineage with 73% accuracy, demonstrating the potential for early identification of emergent APEC in healthy flocks.

Significance StatementGenetic exchange among bacteria shapes the microbial world. From the acquisition of antimicrobial resistance genes to fundamental questions about the nature of bacterial species, this powerful evolutionary force has preoccupied scientists for decades. However, the mixing of genes between species rests on a paradox. On one hand, promoting adaptation by conferring novel functionality, on the other potentially introducing disharmonious gene combinations (negative epistasis) that will be selected against. Taking an interdisciplinary approach to analyse natural populations of the enteric bacteriaCampylobacter, an ideal example of long-range admixture, we demonstrate that genes can independently transfer across species boundaries and rejoin in epistasis in a recipient genome. This challenges conventional ideas and highlights the possibility of single step evolution by saltation.AbstractRecombination of short DNA fragments via horizontal gene transfer (HGT) can both introduce beneficial alleles and create genomic disharmony through negative epistasis. For non-core (accessory) genes, the negative epistatic cost is likely to be minimal because the incoming genes have not co-evolved with the recipient genome. By contrast, for the core genome, interspecific recombination is expected to be rare because disruptive allelic replacement is likely to introduce negative epistasis. Why then is homologous recombination common in the core of bacterial genomes? To understand this enigma we take advantage of an exceptional model system, the common enteric pathogensCampylobacter jejuniandCampylobacter coli, that are known for very high magnitude interspecies gene flow in the core genome. As expected, HGT does indeed disrupt co-adapted allele pairings (negative epistasis). However, multiple HGT events enable recovery of the genome’s co-adaption between introgressing alleles, even in core metabolism genes (e.g., formate dehydrogenase). These findings demonstrate that, even for complex traits, genetic coalitions can be decoupled, transferred and independently reinstated in a new genetic background – facilitating transition between fitness peaks. In this example, the two-step recombinational process is associated withC. colithat are adapted to the agricultural niche.

Objectives: The genomic epidemiology of group b streptococcus (GBS) isolates from the Rotunda maternity hospital, Dublin, 2008-2017, was investigated. Methods: Whole genome sequences of isolates (invasive, n = 114; non-invasive, n = 76) from infants and women were analysed using the PubMLST database (https://pubmlst.org/sagalactiae/). Results: Serotypes III (36%), Ia (18%), V (17%), II (11%) and Ib, (9%) and sequence types (ST) 17 (23%), ST-23 (14%), ST-1 (12%) and ST-19 (7%) were most common. Core genome MLST (cgMLST) differentiated isolates of the same ST, grouped STs into five lineages congruent with known clonal complexes and identified known mother-baby pairs and suspected linked infant cases. Clonal complex (CC) 17 accounted for 40% and 22% of infant and maternal invasive cases, respectively and 21% of non-invasive isolates. CC23 and CC19 were associated with maternal disease (30%) and carriage (24%), respectively. Erythromycin (26%) and clindamycin (18%) resistance increased over the study period and was associated with presence of the erm(B) gene (55%), CC1 (33%) and CC19 (24%). A multi-resistant integrative conjugative element incorporated in the PI-1 locus was detected in CC17, an ST-12 and ST-23 isolate confirming the global dissemination of this element. All isolates possessed one or more pilus islands. Genes encoding other potential protective proteins including Sip, C5a peptidase and Srr1 were present in 100%, 99.5% and 65.8% of isolates, respectively. The srr2 gene was unique to CC17. Conclusions: The PubMLST.org website provides a valuable framework for genomic GBS surveillance to inform on local and global GBS epidemiology, preventive and control measures.

Chickens are the most common birds on Earth and colibacillosis is among the most common diseases affecting them. This major threat to animal welfare and safe sustainable food production is difficult to combat because the etiological agent, avian pathogenic Escherichia coli (APEC), emerges from ubiquitous commensal gut bacteria, with no single virulence gene present in all disease-causing isolates. Here, we address the underlying evolutionary mechanisms of extraintestinal spread and systemic infection in poultry. Combining population scale comparative genomics and pangenome-wide association studies, we compare E. coli from commensal carriage and systemic infections. We identify phylogroup-specific and species-wide genetic elements that are enriched in APEC, including pathogenicity-associated variation in 143 genes that have diverse functions, including genes involved in metabolism, lipopolysaccharide synthesis, heat shock response, antimicrobial resistance and toxicity. We find that horizontal gene transfer spreads pathogenicity elements, allowing divergent clones to cause infection. Finally, a Random Forest model prediction of disease status (carriage vs. disease) identifies pathogenic strains in the emergent ST-117 poultry-associated lineage with 73% accuracy, demonstrating the potential for early identification of emergent APEC in healthy flocks., It is unclear how gut-dwelling E. coli bacteria often emerge to cause systemic infection in chickens. Here, Mageiros et al. use population genomics and pangenome-wide association studies to identify genetic elements associated with pathogenicity in avian E. coli.

As microbial genomics makes increasingly important contributions to clinical and public health microbiology, the interpretation of whole-genome sequence data by nonspecialists becomes essential. In the absence of capsule-based vaccines, two protein-based vaccines have been used for the prevention of invasive serogroup B meningococcal disease (IMD) since their licensure in 2013 and 2014. These vaccines have different components and different levels of coverage of meningococcal variants. Hence, decisions regarding which vaccine to use in managing serogroup B IMD outbreaks require information about the index case isolate, including (i) the presence of particular vaccine antigen variants, (ii) the expression of vaccine antigens, and (iii) the likely susceptibility of its antigen variants to antibody-dependent bactericidal killing., As microbial genomics makes increasingly important contributions to clinical and public health microbiology, the interpretation of whole-genome sequence data by nonspecialists becomes essential. In the absence of capsule-based vaccines, two protein-based vaccines have been used for the prevention of invasive serogroup B meningococcal disease (IMD) since their licensure in 2013 and 2014. These vaccines have different components and different levels of coverage of meningococcal variants. Hence, decisions regarding which vaccine to use in managing serogroup B IMD outbreaks require information about the index case isolate, including (i) the presence of particular vaccine antigen variants, (ii) the expression of vaccine antigens, and (iii) the likely susceptibility of its antigen variants to antibody-dependent bactericidal killing. To obtain this information requires a multitude of laboratory assays, impractical in real-time clinical settings, where the information is most urgently needed. To facilitate assessment for public health and clinical purposes, we synthesized genomic and experimental data from published sources to develop and implement the Meningococcal Deduced Vaccine Antigen Reactivity (MenDeVAR) Index, which is publicly available on PubMLST (https://pubmlst.org). Using whole-genome sequences or individual gene sequences obtained from IMD isolates or clinical specimens, the MenDeVAR Index provides rapid evidence-based information on the presence and possible immunological cross-reactivity of different meningococcal vaccine antigen variants. The MenDeVAR Index enables practitioners who are not genomics specialists to assess the likely reactivity of vaccines for individual cases, outbreak management, or the assessment of public health vaccine programs. The MenDeVAR Index has been developed in consultation with, but independently of, both the 4CMenB (Bexsero; GSK) and rLP2086 (Trumenba; Pfizer, Inc.) vaccine manufacturers.

BackgroundStreptococcus pneumoniae, Haemophilus influenzae and Neisseria meningitidis are leading causes of invasive diseases including bacteraemic pneumonia and meningitis, and of secondary infections post-viral respiratory disease. They are typically transmitted via respiratory droplets. We investigated rates of invasive disease due to these pathogens during the early phase of the COVID-19 pandemic.MethodsLaboratories in 26 countries across six continents submitted data on cases of invasive disease due to S pneumoniae, H influenzae and N meningitidis from 1 January 2018 to 31 May 2020. Weekly cases in 2020 vs 2018-2019 were compared. Streptococcus agalactiae data were collected from nine laboratories for comparison to a non-respiratory pathogen. The stringency of COVID-19 containment measures was quantified by the Oxford COVID-19 Government Response Tracker. Changes in population movements were assessed by Google COVID-19 Community Mobility Reports. Interrupted time series modelling quantified changes in rates of invasive disease in 2020 relative to when containment measures were imposed.FindingsAll countries experienced a significant, sustained reduction in invasive diseases due to S pneumoniae, H influenzae and N meningitidis, but not S agalactiae, in early 2020, which coincided with the introduction of COVID-19 containment measures in each country. Similar impacts were observed across most countries despite differing stringency in COVID-19 control policies. There was no evidence of a specific effect due to enforced school closures.InterpretationThe introduction of COVID-19 containment policies and public information campaigns likely reduced transmission of these bacterial respiratory pathogens, leading to a significant reduction in life-threatening invasive diseases in many countries worldwide.

The expansion of quinolone-resistant Neisseria meningitidis clone ChinaCC4821-R1-C/B from ST-4821 clonal complex (cc4821) caused a serogroup shift from serogroup A to C in invasive meningococcal disease (IMD) in China. To establish the relationship among globally distributed cc4821 meningococci, we analysed whole genome sequence data from 173 cc4821 meningococci isolated in four continents from 1972-2019. These meningococci clustered into four sub-lineages (1-4), with sub-lineage 1 primarily comprising serogroup C IMD isolates (82%, 41/50). Most isolates from outside China formed a distinct sub-lineage (81.6%, 40/49, the Europe-USA cluster), with the typical strain designation B:P1.17-6,23:F3-36:ST-3200(cc4821) and harbouring mutations in penicillin-binding protein 2. These data show that the quinolone-resistant clone ChinaCC4821-R1-C/B has expanded to other countries. The increasing global distribution of B:cc4821 meningococci raises concern that cc4821 has the potential to cause a global pandemic and, this would be challenging to control though there is indirect evidence that Trumenba® vaccine might afford some protection.