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Abstract Streptococcus dysgalactiae subspecies equisimilis (SDSE) and Streptococcus pyogenes share skin and throat niches with extensive genomic homology and horizontal gene transfer (HGT) possibly underlying shared disease phenotypes. It is unknown if cross-species transmission interaction occurs. Here, we conduct a genomic analysis of a longitudinal household survey in remote Australian First Nations communities for patterns of cross-species transmission interaction and HGT. Collected from 4547 person-consultations, we analyse 294 SDSE and 315 S. pyogenes genomes. We find SDSE and S. pyogenes transmission intersects extensively among households and show that patterns of co-occurrence and transmission links are consistent with independent transmission without inter-species interference. We identify at least one of three near-identical cross-species mobile genetic elements (MGEs) carrying antimicrobial resistance or streptodornase virulence genes in 55 (19%) SDSE and 23 (7%) S. pyogenes isolates. These findings demonstrate co-circulation of both pathogens and HGT in communities with a high burden of streptococcal disease, supporting a need to integrate SDSE and S. pyogenes surveillance and control efforts.

ABSTRACT Bacterial species often consist of strains with variable gene content, collectively referred to as the pangenome. Variations in the genetic makeup of strains can alter bacterial physiology and fitness. To define biologically relevant genes of a genome, genome-wide transposon mutant libraries have been used to identify genes essential for survival or virulence in a given strain. Such phenotypic studies have been conducted in four different genotypes of the human pathogen Streptococcus pyogenes, yet challenges exist in comparing results across studies conducted in different genetic backgrounds and conditions. To advance genotype to phenotype inferences across different S. pyogenes strains, we built a pangenome database of 249 S. pyogenes reference genomes. We systematically re-analyzed publicly available transposon sequencing datasets from S. pyogenes using a transposon sequencing-specific analysis pipeline, Transit. Across four genetic backgrounds and nine phenotypic conditions, 355 genes were essential for survival, corresponding to ~24% of the core genome. Clusters of Orthologous Genes (COG) categories related to coenzyme and lipid transport and growth functions were overrepresented as essential. Finally, essential operons across S. pyogenes genotypes were defined, with an increased number of essential operons detected under in vivo conditions. This study provides an extendible database to which new studies can be added, and a searchable html-based resource to direct future investigations into S. pyogenes biology.IMPORTANCEStreptococcus pyogenes is a human-adapted pathogen occupying restricted ecological niches. Understanding the essentiality of genes across different strains and experimental conditions is important to direct research questions and efforts to prevent the large burden of disease caused by S. pyogenes. To this end we systematically reanalyzed transposon sequencing studies in S. pyogenes using transposon sequencing-specific methods, integrating them into an extendible meta-analysis framework. This provides a repository of gene essentiality in S. pyogenes which was used to highlight specific genes of interest and for the community to guide future phenotypic studies.

Abstract Streptococcus dysgalactiae subsp. equisimilis (SDSE) is an emerging cause of human infection with invasive disease incidence and clinical manifestations comparable to the closely related species, Streptococcus pyogenes. Through systematic genomic analyses of 501 disseminated SDSE strains, we demonstrate extensive overlap between the genomes of SDSE and S. pyogenes. More than 75% of core genes are shared between the two species with one third demonstrating evidence of cross-species recombination. Twenty-five percent of mobile genetic element (MGE) clusters and 16 of 55 SDSE MGE insertion regions were shared across species. Assessing potential cross-protection from leading S. pyogenes vaccine candidates on SDSE, 12/34 preclinical vaccine antigen genes were shown to be present in >99% of isolates of both species. Relevant to possible vaccine evasion, six vaccine candidate genes demonstrated evidence of inter-species recombination. These findings demonstrate previously unappreciated levels of genomic overlap between these closely related pathogens with implications for streptococcal pathobiology, disease surveillance and prevention.

Abstract In northern Australia, a region with limited access to healthcare and a substantial population living remotely, antibiotic resistance adds to the complexity of treating infections. Focussing on Escherichia coli urinary tract infections (UTIs) and Staphylococcus aureus skin & soft tissue infections (SSTIs) captured by a northern Australian antibiotic resistance surveillance system, we used logistic regression to investigate predictors of a subsequent resistant isolate during the same infection episode. We also investigated predictors of recurrent infection. Our analysis included 98,651 E. coli isolates and 121,755 S. aureus isolates from 70,851 patients between January 2007 and June 2020. Following an initially susceptible E. coli UTI, subsequent recovery of a cefazolin (8%) or ampicillin (13%) -resistant isolate during the same infection episode was more common than a ceftriaxone-resistant isolate (2%). For an initially susceptible S. aureus SSTI, subsequent recovery of a methicillin-resistant isolate (8%) was more common than a trimethoprim-sulfamethoxazole-resistant isolate (2%). For UTIs and SSTIs, prior infection with a resistant pathogen was a strong predictor of both recurrent infection and resistance in future infection episodes. This multi-centre study demonstrates an association between antibiotic resistance and an increased likelihood of recurrent infection. Particularly in remote areas, a patient’s past antibiograms should guide current treatment choices since recurrent infection will most likely be at least as resistant as previous infection episodes. Using population-level surveillance data in this way can also help clinicians decide if they should switch antibiotics for patients with ongoing symptoms, while waiting for diagnostic results.

Abstract The Staphylococcus aureus Network Adaptive Platform (SNAP) trial is a multifactorial Bayesian adaptive platform trial that aims to improve the way that S. aureus bloodstream infection, a globally common and severe infectious disease, is treated. In a world first, the SNAP trial will simultaneously investigate the effects of multiple intervention modalities within multiple groups of participants with different forms of S. aureus bloodstream infection. Here, we formalise the trial structure, modelling approach, and decision rules that will be used for the SNAP trial. By summarising the statistical principles governing the design, our hope is that the SNAP trial will serve as an adaptable template that can be used to improve comparative effectiveness research efficiency in other disease areas. Trial registration NCT05137119 . Registered on 30 November 2021.

A variant of group A Streptococcus serotype M1 (UK) has been increasingly reported and can be differentiated from the global variant by its overexpression of the superantigen SpeA. Here, Davies et al probe the mechanism behind enhanced SpeA expression and demonstrate that a SNP in the 5’ leader sequence of ssrA is responsible for this virulence phenotype.

Abstract Background Chronic hepatitis B is a public health concern in Aboriginal communities in the Northern Territory of Australia with prevalence almost four times the non-Aboriginal population. Infection is suspected to mainly occur in early life, however, the mode of transmission and vaccine effectiveness is not known in this population. WHO has set a target for hepatitis B elimination by 2030; elimination in this disproportionately affected population in Australia will require understanding of the modes of transmission and vaccine effectiveness. Methods We conducted the study at four very remote Aboriginal communities. We approached mothers who had chronic hepatitis B and had given birth between 1988 and 2013 for consent. We obtained hepatitis B serology, immunisation and birth details from the medical record. If both mother and child had hepatitis B viral DNA detected, we performed viral whole genome sequencing. Results We approached 45 women for consent, of whom 23 agreed to participate. We included 20 mothers and 38 of their children. Of the 20 included mothers, 5 (25%) had children who were hepatitis B immune by exposure and 3 (15%) had children with evidence of chronic hepatitis B infection at the time of assessment. Hepatitis B immunoglobulin (HBIg) had been given at birth in 29/38 (76.3, 95% CI 59.8–88.6) children, and 26 children (68.4, 95% CI 51.3–82.5) were fully vaccinated. Of the 3 children who had chronic hepatitis B, all had received HBIg at birth and two were fully vaccinated. Of the 5 who were immune by exposure, 4 had received HBIg at birth and one was fully vaccinated. Whole genome sequencing revealed one episode of definite mother to child transmission. There was also one definite case of horizontal transmission. Conclusions Chronic hepatitis B in this context is a sensitive issue, with a high proportion of women refusing consent. Although uncommon, there is ongoing transmission of hepatitis B to Aboriginal children in remote northern Australia despite vaccination, and this is likely occurring by both vertical and horizontal routes. Prevention will require ongoing investment to overcome the many barriers experienced by this population in accessing care.

Abstract Background For decades, the research community has called for participant information sheets/consent forms (PICFs) to be improved. Recommendations include simplifying content, reducing length, presenting information in layers and using multimedia. However, there are relatively few studies that have evaluated health consumers’ (patients/carers) perspectives on the type and organisation of information, and the level of detail to be included in a PICF to optimise an informed decision to enter a trial. We aimed to elicit consumers’ views on a layered approach to consent that provides the key information for decision-making in a short PICF (layer 1) with additional optional information that is accessed separately (layer 2). We also elicited consumers’ views on the optimal content and layout of the layered consent materials for a large and complex Bayesian adaptive platform trial (the SNAP trial). Methods We conducted a qualitative multicentre study (4 focus groups and 2 semi-structured interviews) involving adolescent and adult survivors of Staphylococcus aureus bloodstream infection (22) and their carers (2). Interview transcripts were examined using inductive thematic analysis. Results Consumers supported a layered approach to consent. The primary theme that emerged was the value of agency; the ability to exert some control over the amount of information read before the consent form is signed. Three other themes emerged; the need to prioritise participants’ information needs; the importance of health literacy; the importance of information about a trial’s benefits (over its risks) for decision-making and the interplay between the two. Conclusions Our findings suggest that consumers may challenge the one-size-fits-all approach currently applied to the development of PICFs in countries like Australia. Consumers supported a layered approach to consent that offers choice in the amount of information to be read before deciding whether to enter a trial. A 3-page PICF was considered sufficient for decision-making for the SNAP trial, provided that further information was available and accessible.

There is increasing evidence for metabolic processes mediating antimicrobial resistance. Here, the authors present a mechanism of sulfamethoxazole resistance in Group A Streptococcus that is dependent on acquiring end products of the host folate biosynthesis pathway.

Abstract Background The incidence of Gram-negative bacteraemia is rising globally and remains a major cause of morbidity and mortality. The majority of patients with Gram-negative bacteraemia initially receive intravenous (IV) antibiotic therapy. However, it remains unclear whether patients can step down to oral antibiotics after appropriate clinical response has been observed without compromising outcomes. Compared with IV therapy, oral therapy eliminates the risk of catheter-associated adverse events, enhances patient quality of life and reduces healthcare costs. As current management of Gram-negative bacteraemia entails a duration of IV therapy with limited evidence to guide oral conversion, we aim to evaluate the clinical efficacy and economic impact of early stepdown to oral antibiotics. Methods This is an international, multicentre, randomised controlled, open-label, phase III, non-inferiority trial. To be eligible, adult participants must be clinically stable / non-critically ill inpatients with uncomplicated Gram-negative bacteraemia. Randomisation to the intervention or standard arms will be performed with 1:1 allocation ratio. Participants randomised to the intervention arm (within 72 h from index blood culture collection) will be immediately switched to an oral fluoroquinolone or trimethoprim-sulfamethoxazole. Participants randomised to the standard arm will continue to receive IV therapy for at least 24 h post-randomisation before clinical re-assessment and decision-making by the treating doctor. The recommended treatment duration is 7 days of active antibiotics (including empiric therapy), although treatment regimen may be longer than 7 days if clinically indicated. Primary outcome is 30-day all-cause mortality, and the key secondary outcome is health economic evaluation, including estimation of total healthcare cost as well as assessment of patient quality of life and number of quality-adjusted life years saved. Assuming a 30-day mortality of 8% in the standard and intervention arms, with 6% non-inferiority margin, the target sample size is 720 participants which provides 80% power with a one-sided 0.025 α-level after adjustment for 5% drop-out. Discussion A finding of non-inferiority in efficacy of oral fluoroquinolones or trimethoprim-sulfamethoxazole versus IV standard of care antibiotics may hypothetically translate to wider adoption of a more cost-effective treatment strategy with better quality of life outcomes. Trial registration ClinicalTrials.gov NCT05199324 . Registered 20 January 2022.

Background More than 50 million influenza infections and over 100,000 deaths from influenza occur annually. While Indigenous populations experience an inequitable influenza burden, the magnitude of this inequity has not previously been estimated on a global scale. This study compared rates of influenza-associated hospitalisation and mortality between Indigenous and non-Indigenous populations globally. Methods A systematic review and meta-analysis was conducted including literature published prior to 13 July 2021. Eligible articles either reported a rate ratio (RR) comparing laboratory-confirmed influenza-associated hospitalisation and/or mortality between an Indigenous population and a corresponding benchmark population, or reported sufficient information for this to be calculated using publicly available data. Findings were reported by country/region and pooled by country and period (pandemic/seasonal) when multiple studies were available using a random-effects model. The I2 statistic assessed variability between studies. Results Thirty-six studies (moderate/high quality) were included; all from high or high-middle income countries. The pooled influenza-associated hospitalisation RR (HRR) for indigenous compared to benchmark populations was 5·7 (95% CI: 2·7–12·0) for Canada, 5·2 (2.9–9.3) for New Zealand, and 5.2 (4.2–6.4) for Australia. Of the Australian studies, the pooled HRR for seasonal influenza was 3.1 (2·7–3·5) and for pandemic influenza was 6·2 (5·1–7·5). Heterogeneity was slightly higher among studies of pandemic influenza than seasonal influenza. The pooled mortality RR was 4.1 (3·0–5.7) in Australia and 3·3 (2.7–4.1) in the United States. Conclusions Ethnic inequities in severe influenza persist and must be addressed by reducing disparities in the underlying determinants of health. Influenza surveillance systems worldwide should include Indigenous status to determine the extent of the disease burden among Indigenous populations. Ethnic inequities in pandemic influenza illustrate the need to prioritise Indigenous populations in pandemic response plans.

Abstract Background Convalescent plasma has been widely used to treat COVID-19 and is under investigation in numerous randomized clinical trials, but results are publicly available only for a small number of trials. The objective of this study was to assess the benefits of convalescent plasma treatment compared to placebo or no treatment and all-cause mortality in patients with COVID-19, using data from all available randomized clinical trials, including unpublished and ongoing trials (Open Science Framework, https://doi.org/10.17605/OSF.IO/GEHFX ). Methods In this collaborative systematic review and meta-analysis, clinical trial registries (ClinicalTrials.gov, WHO International Clinical Trials Registry Platform), the Cochrane COVID-19 register, the LOVE database, and PubMed were searched until April 8, 2021. Investigators of trials registered by March 1, 2021, without published results were contacted via email. Eligible were ongoing, discontinued and completed randomized clinical trials that compared convalescent plasma with placebo or no treatment in COVID-19 patients, regardless of setting or treatment schedule. Aggregated mortality data were extracted from publications or provided by investigators of unpublished trials and combined using the Hartung–Knapp–Sidik–Jonkman random effects model. We investigated the contribution of unpublished trials to the overall evidence. Results A total of 16,477 patients were included in 33 trials (20 unpublished with 3190 patients, 13 published with 13,287 patients). 32 trials enrolled only hospitalized patients (including 3 with only intensive care unit patients). Risk of bias was low for 29/33 trials. Of 8495 patients who received convalescent plasma, 1997 died (23%), and of 7982 control patients, 1952 died (24%). The combined risk ratio for all-cause mortality was 0.97 (95% confidence interval: 0.92; 1.02) with between-study heterogeneity not beyond chance (I2 = 0%). The RECOVERY trial had 69.8% and the unpublished evidence 25.3% of the weight in the meta-analysis. Conclusions Convalescent plasma treatment of patients with COVID-19 did not reduce all-cause mortality. These results provide strong evidence that convalescent plasma treatment for patients with COVID-19 should not be used outside of randomized trials. Evidence synthesis from collaborations among trial investigators can inform both evidence generation and evidence application in patient care.

The immunology of Indigenous populations is generally understudied outside the context of diseases that are prevalent in these communities. Here the authors identify prevalence of influenza CD8+ T cell epitopes in an Indigenous Australian population expressing the susceptibility allomorph HLA A*24:02 and validate immunodominance of some of these epitopes in mice.

CD8+ T cells are a pivotal part of the immune response to viruses, playing a key role in disease outcome and providing long-lasting immunity to conserved pathogen epitopes. Understanding CD8+ T cell immunity in humans is complex due to CD8+ T cell restriction by highly polymorphic Human Leukocyte Antigen (HLA) proteins, requiring T cell epitopes to be defined for different HLA allotypes across different ethnicities. Here we evaluate strategies that have been developed to facilitate epitope identification and study immunogenic T cell responses. We describe an immunopeptidomics approach to sequence HLA-bound peptides presented on virus-infected cells by liquid chromatography with tandem mass spectrometry (LC-MS/MS). Using antigen presenting cell lines that stably express the HLA alleles characteristic of Indigenous Australians, this approach has been successfully used to comprehensively identify influenza-specific CD8+ T cell epitopes restricted by HLA allotypes predominant in Indigenous Australians, including HLA-A*24:02 and HLA-A*11:01. This is an essential step in ensuring high vaccine coverage and efficacy in Indigenous populations globally, known to be at high risk from influenza disease and other respiratory infections.

Abstract Objectives To determine if lopinavir/ritonavir +/- hydroxychloroquine will reduce the proportion of participants who survive without requiring ventilatory support, 15 days after enrolment, in adult participants with non-critically ill SARS-CoV-2 infection. Trial design ASCOT is an investigator-initiated, multi-centre, open-label, randomised controlled trial. Participants will have been hospitalised with confirmed COVID-19, and will be randomised 1:1:1:1 to receive lopinavir /ritonavir, hydroxychloroquine, both or neither drug in addition to standard of care management. Participants Participants will be recruited from >80 hospitals across Australia and New Zealand, representing metropolitan and regional centres in both public and private sectors. Admitted patients will be eligible if aged ≥ 18 years, have confirmed SARS-CoV-2 by nucleic acid testing in the past 12 days and are expected to remain an inpatient for at least 48 hours from the time of randomisation. Potentially eligible participants will be excluded if admitted to intensive care or requiring high level respiratory support, are currently receiving study drugs or their use is contraindicated due to allergy, drug interaction or comorbidities (including baseline QTc prolongation of 470ms for women or 480ms for men), or death is anticipated imminently. Intervention and comparator Participants will be randomised 1:1:1:1 to: Group 1: standard of care; Group 2: lopinavir (400mg) / ritonavir (100mg) twice daily for 10 days in tablet form; Group 3: hydroxychloroquine (800mg) 4x200mg administered 12 hours apart on Day 1, followed by 400mg twice a day for 6 days; Group 4: lopinavir /ritonavir plus hydroxychloroquine. Main outcomes Proportion of participants alive and not having required intensive respiratory support (invasive or non-invasive ventilation) at 15 days after enrolment. A range of clinical and virological secondary outcomes will also be evaluated. Randomisation The randomisation schedule will be generated by an independent statistician. Randomisation will be stratified by site and will be in permuted blocks of variable block size. The randomised sequence allocation will only be accessible to the data management group, and site investigators will have individual participant allocation provided through a web-based trial enrolment platform. Blinding (masking) This is an open-label study, with researchers assessing the laboratory outcomes blinded to treatment allocation. No unblinding procedures relating to potential adverse effects are therefore required. Numbers to be randomised (sample size) We assumed that 5% of participants receiving standard of care would meet the primary outcome, aimed to evaluate whether interventions could lead to a relative risk of 0.5, assuming no interaction between intervention arms. This corresponds to a required sample size of 610 per arm, with a 5% two-sided significance level (alpha) and 80% power. The total sample size therefore is planned to be 2440. Trial Status ASCOT protocol version 3, May 5, 2020. Recruitment opened April 4, 2020 and is ongoing, with planned completion of enrolment July 31, 2021. Trial registration Australian New Zealand Clinical Trials Registry ( ACTRN12620000445976 ). Prospectively registered April 6, 2020. Full protocol The full protocol is attached as an additional file, accessible from the Trials website (Additional file 1). In the interest in expediting dissemination of this material, the familiar formatting has been eliminated; this Letter serves as a summary of the key elements of the full protocol.

Abstract Background Exotoxins are important virulence factors in Staphylococcus aureus. Clindamycin, a protein synthesis inhibitor antibiotic, is thought to limit exotoxin production and improve outcomes in severe S. aureus infections. However, randomised prospective data to support this are lacking. Methods An open-label, multicentre, randomised controlled trial (RCT) will compare outcome differences in severe S. aureus infection between standard treatment (flucloxacillin/cefazolin in methicillin-susceptible S. aureus; and vancomycin/daptomycin in methicillin-resistant S. aureus) and standard treatment plus an additional clindamycin given for 7 days. We will include a minimum of 60 participants (both adult and children) in the pilot study. Participants will be enrolled within 72 h of an index culture. Severe infections will include septic shock, necrotising pneumonia, or multifocal and non-contiguous skin and soft tissue/osteoarticular infections. Individuals who are immunosuppressed, moribund, with current severe diarrhoea or Clostridiodes difficile infection, pregnant, and those with anaphylaxis to β-lactams or lincosamides will be excluded. The primary outcomes measure is the number of days alive and free (1 or 0) of systemic inflammatory response syndrome (SIRS) within the first 14 days post randomisation. The secondary outcomes measure will include all-cause mortality at 14, 42, and 90 days, time to resolution of SIRS, proportion with microbiological treatment failure, and rate of change of C-reactive protein over time. Impacts of inducible clindamycin resistance, strain types, methicillin susceptibility, and presence of various exotoxins will also be analysed. Discussion This study will assess the effect of adjunctive clindamycin on patient-centred outcomes in severe, toxin-mediated S. aureus infections. The pilot study will provide feasibility for a much larger RCT. Trial registration Australian New Zealand Clinical Trials Registry, ACTRN12617001416381p. Registered on 6 October 2017.

Abstract Background This is the first review of literature and synthesis of data on community onset methicillin resistant Staphylococcus aureus (CO-MRSA) infections in Australia. Incidence of CO-MRSA varies considerably in Australia, depending on geographic and demographic factors. Methods Data for the rates of MRSA infections were collected from articles identified using PubMed, Scopus, the grey literature and data from State and Federal Government Surveillance Systems. We synthesized data and developed a framework for how data was selected, collated, linked, organized and interpreted. Results The results of our literature search demonstrates considerable gaps in the reporting of CO-MRSA in Australia. Consequently, total incidences were under reported; however the available data suggests the incidence varied between 44 (Tasmania) and 388 (southern Northern Territory) cases per 100,000 person years. Hospitalised cases of CO-MRSA varied between 3.8 (regional Victoria) and 329 (southern Northern Territory). Taking the median percentage of infections by site for all regions available, skin and soft tissue infections (SSTIs) consisted of 56% of hospitalized CO-MRSA, compared with bacteremias, which represented 14%. No region had a complete data set of CO-MRSA infections treated in out-patient settings and so incidences were underestimates. Nevertheless, estimates of the incidence of CO-MRSA treated outside hospitals varied between 11.3 (Melbourne) and 285 (Northern Territory) per 100,000 person-years. These infections were chiefly SSTIs, although urinary tract infections were also noted. Incidences of CO-MRSA blood-stream infections and outpatient skin and soft tissue infections have been increasing with time, except in Tasmania. CO-MRSA is observed to affect people living in remote areas and areas of socioeconomic disadvantage disproportionately. Conclusions We generated the first estimates of the incidence of CO-MRSA infections in Australia and identified stark regional differences in the nature and frequency of infections. Critically, we demonstrate that there has been a lack of consistency in reporting CO-MRSA and a general dearth of data. The only government in Australia that requires reporting of CO-MRSA is the Tasmanian, where the infection was least prevalent. Some regions of Australia have very high incidences of CO-MRSA. To improve surveillance and inform effective interventions, we recommend a standardized national reporting system in Australia that reports infections at a range of infection sites, has broad geographic coverage and consistent use of terminology. We have identified limitations in the available data that hinder understanding the prevalence of CO-MRSA.

Staphylococcus aureusS. aureus

ABSTRACT A recent clinical report has linked Streptococcus pyogenes β-lactam antibiotic resistance to mutation in the penicillin binding protein (PBP) PBP2x. To determine whether this is an isolated case or reflects a broader prevalence of mutations that might confer reduced β-lactam susceptibility, we investigated the relative frequency of PBP sequence variation within a global database of 9,667 S. pyogenes isolates. We found that mutations in S. pyogenes PBPs (PBP2x, PBP1a, PBP1b, and PBP2a) occur infrequently across this global database, with fewer than 3 amino acid changes differing between >99% of the global population. Only 4 of the 9,667 strains contained mutations near transpeptidase active sites of PBP2x or PBP1a. The reported PBP2x T553K substitution was not identified. These findings are in contrast to those of 2,520 S. pneumococcus sequences where PBP mutations are relatively frequent and are often located in key β-lactam binding pockets. These data, combined with the general lack of penicillin resistance reported in S. pyogenes worldwide, suggests that extensive, unknown constraints restrict S. pyogenes PBP sequence plasticity. Our findings imply that while heavy antibiotic pressure may select for mutations in the PBPs, there is currently no evidence of such mutations becoming fixed in the S. pyogenes population or that mutations are being sequentially acquired in the PBPs. IMPORTANCE β-Lactam antibiotics are the first-line therapeutic option for Streptococcus pyogenes infections. Despite the global high prevalence of S. pyogenes infections and widespread use of β-lactams worldwide, reports of resistance to β-lactam antibiotics, such as penicillin, have been incredibly rare. Recently, β-lactam resistance, as defined by clinical breakpoints, was detected in two clinical S. pyogenes isolates with accompanying mutations in the active site of the penicillin binding protein PBP2x, raising concerns that β-lactam resistance will become more widespread. We screened a global database of S. pyogenes genome sequences to investigate the frequency of PBP mutations, identifying that PBP mutations are uncommon relative to those of Streptococcus pneumoniae. These findings support clinical observations that β-lactam resistance is rare in S. pyogenes and suggest that there are considerable constraints on S. pyogenes PBP sequence variation.

Abstract Background Prostatic abscess is a rare complication of acute bacterial prostatitis and is most commonly caused by Enterobacteriaceae. We report on a case of prostatic abscess caused by Staphylococcus aureus and conduct a review of the literature. Case presentation We present a case of S. aureus prostatic abscess that was successfully treated with a combination of antibiotic and surgical therapy. The isolate was non–multidrug-resistant, methicillin-resistant Staphylococcus aureus and was genotyped as clonal complex 5, an emerging regional clone that is trimethoprim resistant and Panton-Valentine leukocidin positive. This current case report is the first to describe the use of clindamycin step-down therapy. A literature review identified a further 39 cases of S. aureus prostatic abscesses, of which 26 were methicillin resistant. Conclusions S. aureus is an uncommon cause of prostatic abscess. Optimal management includes both antibiotic therapy and surgical drainage. Our use of clindamycin as step-down therapy was guided by its excellent prostatic penetration.

ABSTRACT The evolution and global transmission of antimicrobial resistance have been well documented for Gram-negative bacteria and health care-associated epidemic pathogens, often emerging from regions with heavy antimicrobial use. However, the degree to which similar processes occur with Gram-positive bacteria in the community setting is less well understood. In this study, we traced the recent origins and global spread of a multidrug-resistant, community-associated Staphylococcus aureus lineage from the Indian subcontinent, the Bengal Bay clone (ST772). We generated whole-genome sequence data of 340 isolates from 14 countries, including the first isolates from Bangladesh and India, to reconstruct the evolutionary history and genomic epidemiology of the lineage. Our data show that the clone emerged on the Indian subcontinent in the early 1960s and disseminated rapidly in the 1990s. Short-term outbreaks in community and health care settings occurred following intercontinental transmission, typically associated with travel and family contacts on the subcontinent, but ongoing endemic transmission was uncommon. Acquisition of a multidrug resistance integrated plasmid was instrumental in the emergence of a single dominant and globally disseminated clade in the early 1990s. Phenotypic data on biofilm, growth, and toxicity point to antimicrobial resistance as the driving force in the evolution of ST772. The Bengal Bay clone therefore combines the multidrug resistance of traditional health care-associated clones with the epidemiological transmission of community-associated methicillin-resistant S. aureus (MRSA). Our study demonstrates the importance of whole-genome sequencing for tracking the evolution of emerging and resistant pathogens. It provides a critical framework for ongoing surveillance of the clone on the Indian subcontinent and elsewhere. IMPORTANCE The Bengal Bay clone (ST772) is a community-associated and multidrug-resistant Staphylococcus aureus lineage first isolated from Bangladesh and India in 2004. In this study, we showed that the Bengal Bay clone emerged from a virulent progenitor circulating on the Indian subcontinent. Its subsequent global transmission was associated with travel or family contact in the region. ST772 progressively acquired specific resistance elements at limited cost to its fitness and continues to be exported globally, resulting in small-scale community and health care outbreaks. The Bengal Bay clone therefore combines the virulence potential and epidemiology of community-associated clones with the multidrug resistance of health care-associated S. aureus lineages. This study demonstrates the importance of whole-genome sequencing for the surveillance of highly antibiotic-resistant pathogens, which may emerge in the community setting of regions with poor antibiotic stewardship and rapidly spread into hospitals and communities across the world.

Background: Pediatric Staphylococcus aureus bacteraemia is one of the leading causes of community-acquired blood-stream infection in the developed world; however, our understanding of management practices by treating clinicians is limited.Methods: The authors designed a web-based clinician survey with support from the Australian and New Zealand Pediatric Infectious Diseases group, of the Australasian Society of Infectious Diseases. Clinicians were presented with three pediatric cases of varying severity. Antibiotic choice, durations of intravenous and oral therapy and research priorities for pediatric S. aureus bacteraemia trials were gauged.Results and Conclusion: Large variation in antibiotic prescribing amongst clinicians is demonstrated and increased, corresponding with escalating case complexity and persisting MRSA bacteraemia. Most clinicians chose defining optimal duration of therapy for S. aureus bacteraemia as their top clinical trial priority. These findings highlight the importance of prioritizing pediatric S. aureus bacteraemia clinical trials, to inform guidelines and best practice management.

Plasticity and clonal population structure in bacterial genomes can hinder traditional SNP-based genetic association studies. Here, Corander and colleagues present a method to identify variable-length sequence elements enriched in a phenotype of interest, and demonstrate its use in human pathogens.

Chlamydia trachomatis isolates causing a blinding disease (trachoma) form a single lineage that is different from the lineages causing urogenital infections. Here, Andersson et al. show however that trachoma isolates from Australia are more closely related to urogenital strains than to other trachoma isolates.

Background: In Australia, community-associated methicillin-resistant Staphylococcus aureus (MRSA) lineage sequence type (ST) 93 has rapidly risen to dominance since being described in the early 1990s. We examined 459 ST93 genome sequences from Australia, New Zealand, Samoa, and Europe to investigate the evolutionary history of ST93, its emergence in Australia and subsequent spread overseas.Results: Comparisons with other S. aureus genomes indicate that ST93 is an early diverging and recombinant lineage, comprising of segments from the ST59/ST121 lineage and from a divergent but currently unsampled Staphylococcal population. However, within extant ST93 strains limited genetic diversity was apparent with the most recent common ancestor dated to 1977 (95% highest posterior density 1973–1981). An epidemic ST93 population arose from a methicillin-susceptible progenitor in remote Northern Australia, which has a proportionally large Indigenous population, with documented overcrowded housing and a high burden of skin infection. Methicillin-resistance was acquired three times in these regions, with a clade harboring a staphylococcal cassette chromosome mec (SCCmec) IVa expanding and spreading to Australia’s east coast by 2000. We observed sporadic and non-sustained introductions of ST93-MRSA-IVa to the United Kingdom. In contrast, in New Zealand, ST93-MRSA-IVa was sustainably transmitted with clonal expansion within the Pacific Islander population, who experience similar disadvantages as Australian Indigenous populations.Conclusion: ST93 has a highly recombinant genome including portions derived from an early diverging S. aureus population. Our findings highlight the need to understand host population factors in the emergence and spread of antimicrobial resistant community pathogens.

For infectious pathogens such as Staphylococcus aureus and Streptococcus pneumoniae, some hosts may carry the pathogen and transmit it to others, yet display no symptoms themselves. These asymptomatic carriers contribute to the spread of disease but go largely undetected and can therefore undermine efforts to control transmission. Understanding the natural history of carriage and its relationship to disease is important for the design of effective interventions to control transmission. Mathematical models of infectious diseases are frequently used to inform decisions about control and should therefore accurately capture the role played by asymptomatic carriers. In practice, incorporating asymptomatic carriers into models is challenging due to the sparsity of direct evidence. This absence of data leads to uncertainty in estimates of model parameters and, more fundamentally, in the selection of an appropriate model structure. To assess the implications of this uncertainty, we systematically reviewed published models of carriage and propose a new model of disease transmission with asymptomatic carriage. Analysis of our model shows how different assumptions about the role of asymptomatic carriers can lead to different conclusions about the transmission and control of disease. Critically, selecting an inappropriate model structure, even when parameters are correctly estimated, may lead to over- or under-estimates of intervention effectiveness. Our results provide a more complete understanding of the role of asymptomatic carriers in transmission and highlight the importance of accurately incorporating carriers into models used to make decisions about disease control.

ABSTRACT Nosocomial outbreaks of vancomycin-resistant Enterococcus faecium (VREfm) are thought to occur by transmission of VREfm between patients, predicting that infection control interventions will limit cross-transmission. Despite implementation of such strategies, the incidence of VREfm infections continues to rise. We aimed to use genomics to better understand the epidemiology of E. faecium within a large hospital and investigate the reasons for failure of infection control strategies. Whole-genome sequencing was performed on 61 E. faecium (36 VREfm) isolates, predominately from blood cultures collected at a single hospital between 1998 and 2009, and on five vanB-positive anaerobic commensal bacteria isolated from human feces. Phylogenomic analysis and precise mapping of the vanB gene, which contains the Tn1549 transposon, showed that at least 18 of the 36 VREfm isolates had acquired the transposon via independent insertion events, indicating de novo generation of VREfm rather than cross-transmission. Furthermore, Tn1549 sequences found in 15 of the 36 VREfm isolates were the same as the Tn1549 sequence from one of the gut anaerobes. National and international comparator E. faecium isolates were phylogenetically interspersed with isolates from our hospital, suggesting that our findings might be globally representative. These data demonstrate that VREfm generation within a patient is common, presumably occurring in the human bowel during antibiotic therapy, and help explain our inability to reduce VREfm infections. A recommendation from our findings is that infection control practices should include screening patients for specific hospital clones of vancomycin-susceptible E. faecium rather than just VREfm. IMPORTANCE Enterococcus faecium is an increasingly important human pathogen causing predominantly antibiotic-resistant infections in hospitalized patients. Large amounts of health care funding are spent trying to control antibiotic-resistant bacteria in hospitals globally, yet in many institutions around the world, vancomycin-resistant E. faecium (VREfm) infections continue to rise. The new findings from this study help explain the failures of our current approaches to controlling vanB VREfm in health care institutions. Given the importance of this bacterium as a cause of hospital-acquired infections and the difficulties faced by infection control units in trying to prevent colonization in their institutions, the novel findings from this study provide evidence that a new approach to controlling VREfm in hospitals is required. In particular, more attention should be given to understanding the epidemiology of hospital-adapted vancomycin-susceptible E. faecium, and patients at higher risk for de novo generation of VREfm need to be identified and optimally managed.

Pregnancy may be associated with risk of developing tuberculosis (TB) in those infected with Mycobacterium tuberculosis (Mtb). The perinatal period could provide opportunities for targeted screening and treatment. This study aims to synthesise published literature on Mtb infection in pregnancy, relating to prevalence, natural history, test performance, cascade of care, and treatment. We searched Ovid MEDLINE, Embase+Embase Classic, Web of Science, and Cochrane Central Register of Controlled Trials (CENTRAL) on October 3, 2023, and 47 studies met the inclusion criteria. The prevalence of Mtb infection was as high as 57.0% in some populations, with rates increasing with maternal age and in women from high TB-incidence settings. Five studies quantified perinatal progression from Mtb infection to active TB disease, with two demonstrating increased risk compared to non-pregnant populations (IRR 1.3-1.4 during pregnancy and IRR 1.9-2 postpartum). Concordance between Tuberculin Skin Test (TST) and Interferon Gamma-Release Assay (IGRA) ranged from 49.4%-96.3%, with k-values of 0.19-0.56. High screening adherence was reported, with 62.0-100.0% completing antenatal TST and 81.0-100.0% of those positive having chest radiograph. Four studies of TB preventative treatment (TPT) did not find a significant association with serious adverse events. The antenatal period could provide opportunities for contextualised Mtb infection screening and treatment. As women with increased age and from high TB-incidence settings demonstrate higher prevalence and risk of disease, this cohort should be prioritised. TPT appears safe and feasible; however, further studies are needed to optimise algorithms, ensuring pregnant and postpartum women can make evidence-informed decisions for effective TB prevention.

Introduction Skin is important in Australian Aboriginal culture informing kinship and identity. In many remote Aboriginal communities, scabies and impetigo are very common. Untreated skin infections are painful, itchy and frequently go untreated due to under-recognition and lack of awareness of their potential serious complications. We hypothesise that the skin infection burden in remote Aboriginal communities can be reduced by implementing streamlined training and treatment pathways integrated with environmental health and health promotion activities, tested in the See, Treat, Prevent (SToP skin sores and scabies) trial.Methods and analysis SToP will evaluate a skin control programme using a stepped-wedge, cluster randomised trial design with three intervention components (the ‘SToP activities’): (1) seeing skin infections (development of training resources implemented within a community dermatology model); (2) treating skin infections (employing the latest evidence for impetigo, and scabies treatment); and (3) preventing skin infections (embedded, culturally informed health promotion and environmental health activities). Four community clusters in the remote Kimberley region of Western Australia will participate. Following baseline data collection, two clusters will be randomly allocated to the SToP activities. At 12 months, the remaining two clusters will transition to the SToP activities. The primary outcome is the diagnosis of impetigo in children (5–9 years) at school-based surveillance. Secondary outcome measures include scabies diagnosis, other child health indicators, resistance to cotrimoxazole in circulating pathogenic bacteria, determining the economic burden of skin disease and evaluating the cost effectiveness of SToP activities.Ethics and dissemination This study protocol was approved by the health ethics review committees at the Child and Adolescent Health Service (Approval number RGS0000000584), the Western Australian Aboriginal Health Ethics Committee (Reference number: 819) and the University of Western Australia (Reference RA/4/20/4123). Study findings will be shared with community members, academic and medical communities via publications and presentations, and in reports to funders. Authorship for all publications based on this study will be determined in line with the Uniform Requirements for Manuscripts Submitted to Biomedical Journals published by the International Committee of Medical Journal Editors. Sharing results with organisations and communities who contributed to the study is paramount. The results of the SToP trial will be shared with participants in a suitable format, such as a single summary page provided to participants or presentations to communities, the Kimberly Aboriginal Health Planning Forum Research Subcommittee and other stakeholders as appropriate and as requested. Communication and dissemination will require ongoing consultation with Aboriginal communities to determine appropriate formats.Trial registration number ACTRN12618000520235.