Identification of commonly expressed exoproteins and proteolytic cleavage events by proteomic mining of clinically relevant UK isolates of Staphylococcus aureus

The range of exoproteins and core exoproteome of 14 Staphylococcus aureus isolates representing major lineages associated with asymptomatic carriage and clinical disease in the UK was identified by MS proteomics using a combined database incorporating sequences derived from 39 S. aureus genomes. In all, 632 different proteins were identified and, of these, only 52 (8 %) were found in all 14 isolates whereas 144 (23 %) were found in just a single isolate. Comparison of the observed mass of each protein (based on migration by SDS-PAGE) with its predicted mass (based on amino acid sequence) suggested that 95 % of the proteins identified were not subject to any major post-translational modification. Migration of 5 % of the proteins was not as expected: 1 % of the proteins migrated at a mass greater than predicted, while 4 % appeared to have undergone proteolytic cleavage; these included SsaA2, Aur, SspP, Ebh as well as BlaR1, MecR1, FsH, OatA and LtaS. Intriguingly, a truncated SasG was produced by a single CC8 USA300-like strain. The analysis provided evidence of the marked heterogeneity in protein expression by S. aureus in broth, while yielding a core but narrow common exoproteome

Staphylococcus argenteus transmission among healthy Royal Marines: a molecular epidemiology case-study

Objectives: During a prospective study of S. aureus carriage in Royal Marines recruits, six S. argenteus strains were identified in four recruits. As S. argenteus sepsis leads to mortality similar to S. aureus, we determined the potential for within same troop transmission, to evaluate future outbreak risk. Methods: We used whole-genome sequencing to characterise S. argenteus and investigate phylogenetic relationships between isolates. Results: S. argenteus strains (t5078, ST2250) were detected in 4/40 recruits in the same troop (training cohort) in weeks 1, 6 or 15 of training. No mec, tsst or LukPV genes were detected. We identified differences of 1-17 core SNPs between S. argenteus from different recruits. In two recruits, two S. argenteus strains were isolated; these could be distinguished by 2 and 15 core SNPs. Conclusions: The identification of S. argenteus within a single troop from the total recruit population suggests a common source for transmission, though high number of SNPs were identified, both within-host and within-cluster. The high number of SNPs between some isolates may indicate a common source of diverse isolates or a high level of S. argenteus mutation in carriage. S. argenteus is newly recognised species; and understanding of the frequency of genetic changes during transmission and transition from asymptomatic carriage to disease is required

Sampling and diversity of Escherichia coli from the enteric microbiota in patients with Escherichia coli bacteraemia

Objective The increase in Escherichia coli bloodstream infections mandates better characterisation of the relationship between commensal and invasive isolates. This study adopted a simple approach to characterize E. coli in the gut reservoir from patients with either E. coli or other Gram-negative bacteraemia, or those without bacteraemia, establishing strain collections suitable for genomic investigation. Enteric samples from patients in the three groups were cultured on selective chromogenic agar. Genetic diversity of prevailing E. coli strains in gut microbiota was estimated by RAPD-PCR. Results Enteric samples from E. coli bacteraemia patients yielded a median of one E. coli RAPD pattern (range 1–4) compared with two (range 1–5) from groups without E. coli bacteraemia. Of relevance to large-scale clinical studies, observed diversity of E. coli among hospitalised patients was not altered by sample type (rectal swab or stool), nor by increasing the colonies tested from 10 to 20. Hospitalised patients demonstrated an apparently limited diversity of E. coli in the enteric microbiota and this was further reduced in those with E. coli bacteraemia. The reduced diversity of E. coli within the gut during E. coli bacteraemia raises the possibility that dominant strains may outcompete other lineages in patients with bloodstream infection

Frequency of Transmission, asymptomatic shedding, and airborne spread of Streptococcus pyogenes among school children exposed to scarlet fever: a longitudinal multi-cohort molecular epidemiology contact tracing study

Background: Despite recommendations regarding prompt treatment of cases and enhanced hygiene measures, scarlet fever outbreaks increased in England between 2014-2018. Methods: We undertook a prospective, intensive contact tracing study in schools with consecutive scarlet fever cases to assess the impact of standard interventions on transmission of Streptococcus pyogenes between cases, classroom contacts, households, and classroom environments over 4 weeks using genome sequencing. Findings: Six classes, comprising 12 scarlet fever cases, 17 household contacts, and 278 classroom contacts were recruited. Prevalence of the outbreak strain in asymptomatic classroom contacts was high, increasing from 9.6% in week 1, to 26.9% in week 2, 23.9% in week 3, then 14.3% in week 4. Colonisation with non-outbreak strains was 0 - 7.5%. Genome sequencing showed clonality of isolates within each of six classes, confirming recent transmission accounted for high carriage. Of asymptomatic classroom contacts with S. pyogenes -positive throat swabs who were tested for transmissibility, 6/28 (21%) had positive cough plates and/or hand swabs, of whom three remained S. pyogenes -positive for 3 weeks. Only 1/60 surface swabs taken in 3 classrooms yielded S. pyogenes . In contrast, settle plates placed in elevated locations were S. pyogenes -positive in both classrooms tested. Interpretation: S. pyogenes transmission in schools is intense and may occur prior to, or in spite of reported treatment of cases, underlining a need for rapid case management. Despite guideline adherence, heavy shedding of S. pyogenes by small numbers of classroom contacts may perpetuate outbreaks, and airborne transmission has a plausible role in spread. Funding: Action Medical Research, UKRI, NIH

Staphylococcus aureus colonisation and acquisition of skin and soft tissue infection amongst Royal Marines recruits: A prospective cohort study

Objectives: Skin and soft tissue infections (SSTIs) are a serious health issue for military personnel. Of particular importance are those caused by MRSA and PVL-positive S. aureus (PVL-SA), as they have been associated with outbreaks of SSTIs. A prospective observational study was conducted in Royal Marines recruits to investigate the prevalence of PVL-SA carriage and any association with SSTIs. Methods: 1,012 RM recruits were followed through a 32-week training programme, with nose and throat swabs obtained at weeks 1, 6, 15 and 32. S. aureus isolates were characterised by antibiotic susceptibility testing, spa typing, presence of mecA/C and PVL genes. Retrospective review of the clinical notes for SSTI acquisition was conducted. Results: S. aureus colonisation decreased from week-1 to week-32 (41% to 26%, p<0.0001). Of 1,168 S. aureus isolates, 3/1168 (0.3%) were MRSA and 10/1168 (0.9%) PVL-positive (all MSSA) and 169/1168 (14.5%) were resistant to clindamycin. Isolates showed genetic diversity with 238 different spa types associated with 25 MLST clonal complexes. SSTIs were seen in 35% (351/989) of recruits with 3 training days lost per recruit. SSTI acquisition rate was reduced amongst persistent carriers (p<0.0283). Conclusions: Nose and throat carriage of MRSA and PVL-SA was low amongst recruits, despite a high incidence of SSTIs being reported particularly cellulitis. Carriage strains were predominantly MSSA with a marked diversity of genotypes. Persistent nose and/or throat carriage was not associated with SSTI acquisition. Putative person-to-person transmission within troops was identified based on spa typing requiring further research to confirm and explore potential transmission routes