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In Australia, gonococcal isolates are monitored for antimicrobial susceptibilities. In Western Australia (WA), gonorrhoea notification rates increased by 63 % between 2013 and 2016, with the steepest increase occurring between 2015 and 2016, before stabilizing at this higher baseline between 2017 and 2020. This increased prevalence was associated with antimicrobial-susceptible (AMS) lineages. To understand the provenance of these isolates causing gonorrhoea in WA, whether they were introduced or expanded from endogenous lineages, 741 isolates were collected in 2017 and characterized by both iPLEX typing and whole genome sequencing (WGS). Antibiograms and genocoding of the isolates revealed that AMS isolates were most prevalent in the remote regions, while the urban/rural regions were characterized by antimicrobial-resistant (AMR) isolates. iPLEX typing identified 78 iPLEX genotypes (WA-1 to WA-78) of which 20 accounted for over 88 % of isolates. WA-10 was the most frequently identified genotype in the urban/rural regions whilst WA-29 was the most frequently identified genotype in the remote regions. Genotypes WA-38, WA-52 and WA-13 accounted for 81 % (n=36/44) of the azithromycin-resistant N. gonorrhoeae (AziR) isolates. A representative isolate of each iPLEX genotype and AMR biotype was whole genome sequenced and analysed using MLST, NG-MAST and NG-STAR, and the novel core genome clustering Ng_cgc_400 typing scheme. Five predominant Bayesian population groups (termed BPG-1 to 5) were identified in the study collection. BPG-1 and BPG-2 were associated with AMS isolates from the remote regions. BPG-1 and BPG-2 were shown to be unique to the remote regions based on a minimum spanning tree against 4000 international isolates. AMS isolates in urban/rural regions were dominated by international lineages. AziR and Cef DS (decreased susceptibility to ceftriaxone) was concentrated in three urban/rural genomic groups (BPG-3, 4 and 5). Azithromycin minimum inhibitory concentrations (0.5–16 mg l−1) correlated with the accumulation of mtrR mutations or/and the fraction of 23S rRNA C2611T mutated copies. The majority of isolates in BPG-3, 4 and 5 could be correlated with known AMR lineages circulating globally and nationally. In conclusion, the surge in AMS isolates in WA in 2017 was due to importation of international AMS lineages into urban/rural regions, whilst the local AMS lineages persisted largely in the remote regions. Bridging between the urban/rural and remote regions was relatively rare, but continued surveillance is required to prevent ingress of AMR strains/lineages into the remote regions of WA.

From 1 January to 31 December 2021, forty-eight institutions around Australia participated in the Australian Staphylococcus aureus Surveillance Outcome Programme (ASSOP). The aim of ASSOP 2021 was to determine the proportion of Staphylococcus aureus bacteraemia (SAB) isolates in Australia that were antimicrobial resistant, with particular emphasis on susceptibility to methicillin and on characterisation of the molecular epidemiology of the methicillin-resistant isolates. A total of 2,928 SAB episodes were reported, of which 78.4% were community-onset. Overall, 16.9% of S. aureus isolates were methicillin resistant. The 30-day all-cause mortality associated with methicillin-resistant SAB was 15.0%, which was not significantly different from the 14.4% all-cause mortality associated with methicillin-susceptible SAB (p = 0.7). With the exception of the β-lactams and erythromycin, antimicrobial resistance in methicillin-susceptible S. aureus was rare. However, in addition to the β-lactams, approximately 36% of methicillin-resistant S. aureus (MRSA) were resistant to ciprofloxacin; 30% to erythromycin; 15% to tetracycline; 16% to gentamicin; and 3% to co-trimoxazole. When applying the European Committee on Antimicrobial Susceptibility Testing (EUCAST) breakpoints, teicoplanin resistance was detected in three S. aureus isolates. Resistance to vancomycin or linezolid was not detected. Resistance to non-β-lactam antimicrobials was largely attributable to the healthcare-associated MRSA (HA-MRSA) clone ST22-IV [2B] (EMRSA-15), and the community-associated MRSA (CA-MRSA) clone ST45-V [5C25] which has acquired multiple antimicrobial resistance determinants including ciprofloxacin, erythromycin, clindamycin, gentamicin and tetracycline. The ST22-IV [2B] (EMRSA-15) clone is the predominant HA-MRSA clone in Australia. Nonetheless, 85% of methicillin-resistant SAB episodes were due to CA-MRSA clones. Although polyclonal, approximately 68% of CA-MRSA clones were characterised as ST93-IV [2B] (Queensland clone); ST45-V [5C25]; ST5-IV [2B]; ST1-IV [2B]; ST30-IV [2B]; and ST97-IV [2B]. As CA-MRSA is well established in the Australian community, it is important to monitor antimicrobial resistance patterns in community- and healthcare-associated SAB as this information will guide therapeutic practices in treating S. aureus bacteraemia.

From 1 January to 31 December 2021, forty-eight institutions around Australia participated in the Australian Enterococcal Surveillance Outcome Programme (AESOP). The aim of AESOP 2021 was to determine the proportion of enterococcal bacteraemia isolates in Australia that were antimicrobial resistant, and to characterise the molecular epidemiology of the Enterococcus faecium isolates. Of the 1,297 unique episodes of enterococcal bacteraemia investigated, 94.4% were caused by either E. faecalis (54.1%) or E. faecium (40.3%). Ampicillin resistance was detected in one E. faecalis isolate and in 89.3% of E. faecium isolates. Vancomycin non-susceptibility was not detected in E. faecalis but was detected in 37.9% of E. faecium. Overall, 39.6% of E. faecium harboured the vanA and/or vanB genes. For the vanA/vanB positive E. faecium isolates, 35.8% harboured the vanA gene and 64.2% the vanB gene. Although the percentage of vancomycin-resistant E. faecium bacteraemia isolates was significantly lower than that reported in the 2020 AESOP report (presumably due to the COVID-19 elective surgery restrictions placed on hospitals), it remains substantially higher than that recorded in most European countries. Isolates of E. faecium consisted of 73 multi-locus sequence types (STs); 77.2% of isolates were classified into seven major STs each containing more than ten isolates. All major STs belonged to clonal cluster (CC) 17, a major hospital-adapted polyclonal E. faecium cluster. The major STs (ST17, ST1424, ST796, ST78, ST80, ST1421 and ST555) were found across most regions of Australia. The predominant ST was ST17 which was identified in all regions except the Northern Territory. Overall, 46.5% of isolates belonging to the seven major STs harboured the vanA or vanB gene. The AESOP 2021 has shown that enterococcal bacteraemia episodes in Australia are frequently caused by polyclonal ampicillin-resistant high-level gentamicin resistant vanA- or vanB-positive E. faecium which have limited treatment options.

Background Staphylococcus aureus is a common cause of bacteremia, yet the epidemiology and predictors of poor outcome remain inadequately defined in childhood. Methods ISAIAH (Invasive Staphylococcus aureus Infections and Hospitalizations in children) is a prospective, cross-sectional study of S. aureus bacteremia (SAB) in children hospitalized in Australia and New Zealand over 24 months (2017–2018). Results Overall, 552 SABs were identified (incidence 4.4/100 000/year). Indigenous children, those from lower socioeconomic areas and neonates were overrepresented. Although 90-day mortality was infrequent, one-third experienced the composite of: length of stay >30 days (26%), intensive care unit admission (20%), relapse (4%), or death (3%). Predictors of mortality included prematurity (adjusted odds ratio [aOR],16.8; 95% confidence interval [CI], 1.6–296.9), multifocal infection (aOR, 22.6; CI, 1.4–498.5), necrotizing pneumonia (aOR, 38.9; CI, 1.7–1754.6), multiorgan dysfunction (aOR, 26.5; CI, 4.1–268.8), and empiric vancomycin (aOR, 15.7; CI, 1.6–434.4); while infectious diseases (ID) consultation (aOR, 0.07; CI .004–.9) was protective. Neither MRSA nor vancomycin trough targets impacted survival; however, empiric vancomycin was associated with nephrotoxicity (OR, 3.1; 95% CI 1.3–8.1). Conclusions High SAB incidence was demonstrated and for the first time in a pediatric setting, necrotizing pneumonia and multifocal infection were predictors of mortality, while ID consultation was protective. The need to reevaluate pediatric vancomycin trough targets and limit unnecessary empiric vancomycin exposure to reduce poor outcomes and nephrotoxicity is highlighted. One in 3 children experienced considerable SAB morbidity; therefore, pediatric inclusion in future SAB comparator trials is paramount to improve outcomes.

From 1 January to 31 December 2020, forty-nine institutions around Australia participated in the Australian Enterococcal Sepsis Outcome Programme (AESOP). The aims of AESOP 2020 were to determine the proportion of enterococcal bacteraemia isolates in Australia that were antimicrobial-resistant, and to characterise the molecular epidemiology of the E. faecium isolates. Of the 1,230 unique episodes of enterococcal bacteraemia investigated, 93.9% were caused by either E. faecalis (54.2%) or E. faecium (39.7%). Ampicillin resistance was not detected in E. faecalis but was detected in 88.2% of E. faecium . Vancomycin non-susceptibility was detected in 0.2% of E. faecalis and 32.6% of E. faecium . Overall, 35.2% of E. faecium harboured vanA and/or vanB genes. For the vanA/B positive E. faecium isolates, 38.8% harboured the vanA gene, 60.6% the vanB gene, and 0.6% harboured both vanA and vanB . Although the percentage of E. faecium bacteraemia isolates was significantly lower than that detected in the 2019 AESOP (presumably due to the COVID-19 elective surgery restrictions placed on hospitals), it remains substantially higher than that recorded in most European countries. The E. faecium isolates detected consisted of 71 multilocus sequence types (STs), with 81.7% of these isolates classified into eight major STs each containing ten or more isolates. All major STs belonged to clonal cluster 17 (CC17), a major hospital-adapted polyclonal E. faecium cluster. The major STs (ST17, ST1424, ST80, ST796, ST78, ST1421, ST555 and ST117) were found across most regions of Australia. The predominant clone was ST17, which was identified in all regions except the Northern Territory. Overall, 40.9% of isolates belonging to the eight major STs harboured the vanA or vanB gene. The AESOP 2020 has shown enterococcal bacteraemia episodes in Australia are frequently caused by polyclonal ampicillin-resistant high-level gentamicin-resistant vanA - or vanB -positive E. faecium which have limited treatment options.

Clonal complex 398 (CC398) livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) has been reported worldwide in a variety of food-animal species. Although CC398 is synonymous with LA-MRSA, community-associated MRSA (CA-MRSA) variants have emerged, including the Panton-Valentine leukocidin (PVL)-positive ST398-V and ST398 single-locus variant ST1232-V, and the PVL-negative ST398-V clones. Using comparative genomic analysis, we determined whether ten CC398 MRSA bacteraemia episodes recently identified in Australia were due to LA-MRSA or CA-MRSA CC398. Isolates were sourced from the Australian Group on Antimicrobial Resistance S. aureus surveillance programme and episodes occurred across Australia. Whole-genome sequencing (WGS) and phylogenetic comparison of the ten CC398 bacteraemia isolates with previously published CC398 MRSA whole-genome sequences identified that the Australian CC398 isolates were closely related to the human-associated II-GOI clade and the livestock-associated IIa clade. The identified CC398 MRSA clones were: PVL-positive ST1232-V (5C25), PVL-negative community-associated ST398-V (5C25) and livestock-associated ST398-V (5C25). Our findings demonstrate the importance of using WGS and comparing the sequences with international sequences to distinguish between CC398 CA-MRSA and LA-MRSA and to determine the isolates' origin. Furthermore, our findings suggest that CC398 CA-MRSA has become established in the Australian community and that ST398-V (5C25) LA-MRSA is now widespread in Australian piggeries. Our study emphasises the need for national One Health antimicrobial resistance surveillance programmes to assist in monitoring the ongoing epidemiology of MRSA and other clinically significant antimicrobial-resistant organisms.

Sequence type 1 (ST1) methicillin-resistant Staphylococcus aureus (MRSA) type IV[2B] has become one of the most common community-associated MRSA clones in Australia. We report the complete genome sequence of one of the earliest isolated Australian S. aureus ST1-MRSA-IV strains, WBG8287, isolated from an Indigenous Australian patient living in the remote Kimberley Region of Western Australia.

Sequence type 1 (ST1) methicillin-resistant Staphylococcus aureus (MRSA) SCCmec IV[2B] has become one of the most common community-associated MRSA clones in Australia. We report the complete genome sequence of one of the earliest isolated Australian S. aureus ST1-MRSA-IV strains, WBG8287, isolated from an Indigenous Australian patient living in the remote Kimberley region of Western Australia.

Initially reported in Western Australia in the 1980s, community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) has become a major cause of S. aureus infections globally. We report the complete genome sequences of three of the earliest CA-MRSA strains isolated from remote Australian Indigenous communities in the Kimberley region of Western Australia.

BackgroundAmoxicillin plus ceftriaxone combination therapy is now standard of care for enterococcal endocarditis. Due to amoxicillin instability in infusion devices, benzylpenicillin plus ceftriaxone may be substituted to facilitate outpatient parenteral antimicrobial therapy (OPAT) delivery, despite lack of guideline endorsement.ObjectivesTo assess the clinical efficacy of benzylpenicillin plus ceftriaxone for the management of enterococcal endovascular infections, in addition to assessing this combination’s in vitro synergy.Patients and methodsRetrospective cohort study assessing unplanned readmissions, relapses and mortality for 20 patients with endovascular Enterococcus faecalis infections treated with benzylpenicillin plus ceftriaxone delivered via OPAT. For a subset of isolates, synergism for both amoxicillin and benzylpenicillin in combination with ceftriaxone was calculated using a chequerboard method.ResultsPatients had endovascular infections of native cardiac valves (n = 11), mechanical or bioprosthetic cardiac valves (n = 7), pacemaker leads (n = 1) or left ventricular assistant devices (n = 1). The median duration of OPAT was 22 days, and the most frequent antimicrobial regimen was benzylpenicillin 14 g/day via continuous infusion and ceftriaxone 4 g once daily via short infusion. Rates of unplanned readmissions were high (30%), although rates of relapsed bacteraemia (5%) and 1 year mortality (15%) were comparable to the published literature. Benzylpenicillin less frequently displayed a synergistic interaction with ceftriaxone when compared with amoxicillin (3 versus 4 out of 6 isolates).ConclusionsLower rates of synergistic antimicrobial interaction and a significant proportion of unplanned readmissions suggest clinicians should exercise caution when treating enterococcal endovascular infection utilizing a combination of benzylpenicillin and ceftriaxone via OPAT.

Background Amoxicillin plus ceftriaxone combination therapy is now standard of care for enterococcal endocarditis. Due to amoxicillin instability in infusion devices, benzylpenicillin plus ceftriaxone may be substituted to facilitate outpatient parenteral antimicrobial therapy (OPAT) delivery, despite lack of guideline endorsement. Objectives To assess the clinical efficacy of benzylpenicillin plus ceftriaxone for the management of enterococcal endovascular infections, in addition to assessing this combination’s in vitro synergy. Patients and methods Retrospective cohort study assessing unplanned readmissions, relapses and mortality for 20 patients with endovascular Enterococcus faecalis infections treated with benzylpenicillin plus ceftriaxone delivered via OPAT. For a subset of isolates, synergism for both amoxicillin and benzylpenicillin in combination with ceftriaxone was calculated using a chequerboard method. Results Patients had endovascular infections of native cardiac valves (n = 11), mechanical or bioprosthetic cardiac valves (n = 7), pacemaker leads (n = 1) or left ventricular assistant devices (n = 1). The median duration of OPAT was 22 days, and the most frequent antimicrobial regimen was benzylpenicillin 14 g/day via continuous infusion and ceftriaxone 4 g once daily via short infusion. Rates of unplanned readmissions were high (30%), although rates of relapsed bacteraemia (5%) and 1 year mortality (15%) were comparable to the published literature. Benzylpenicillin less frequently displayed a synergistic interaction with ceftriaxone when compared with amoxicillin (3 versus 4 out of 6 isolates). Conclusions Lower rates of synergistic antimicrobial interaction and a significant proportion of unplanned readmissions suggest clinicians should exercise caution when treating enterococcal endovascular infection utilizing a combination of benzylpenicillin and ceftriaxone via OPAT.

Fosfomycin is a broad-spectrum antibiotic that targets UDP-Nacetylglucosamine enolpyruvyl transferase (MurA), an important enzyme in the early stages of peptidoglycan biosynthesis. In Escherichia coli, fosfomycin resistance can occur through the presence of fos genes (typically fosA3) which encode fosfomycininactivating enzymes (FosA, FosC2), or by mutations in proteins important for the uptake of fosfomycin (CyaA, GlpT, PtsI, UhpA, UhpT) or for its action (MurA)

Background The global emergence of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) has seen the dominance of specific clones in different regions around the world with the PVL-positive ST93-IV as the predominant CA-MRSA clone in Australia. In this study we applied a genome-wide association study (GWAS) approach on a collection of Australian ST93-IV MRSA genomes to identify genetic traits that may have assisted the ongoing transmission of ST93-IV in Australia. We also compared the genomes of ST93-IV bacteraemia and non-bacteraemia isolates to identify potential virulence factors associated with bacteraemia.Results Based on single nucleotide polymorphism phylogenetics we identified two distinct ST93-IV clades circulating concurrently in Australia. One of the clades contained isolates primarily isolated in the northern regions of Australia whilst isolates in the second clade were distributed across the country. Analyses of the ST93-IV genome plasticity over a 15-year period (2002-2017) revealed an observed gain in accessory genes amongst the clone’s population. The GWAS analysis on the bacteraemia isolates identified two genes that have also previously been associated to this kind of infection. Conclusions The emergence of a ST93-IV clade containing additional virulence genes may explain the high prevalence of ST93-IV infections amongst the indigenous population living in the northern regions of Australia. In summary, this study has shown ST93-IV is evolving with multiple additional genes possibly contributing to its dominance in the Australian community.

From 1 January to 31 December 2019, thirty-nine institutions around Australia participated in the Australian Enterococcal Sepsis Outcome Programme (AESOP). The aim of AESOP 2019 was to determine the proportion of enterococcal bacteraemia isolates in Australia that were antimicrobial resistant, and to characterise the molecular epidemiology of the E. faecium isolates. Of the 1,361 unique episodes of bacteraemia investigated, 95.2% were caused by either E. faecalis (51.4%) or E. faecium (43.8%). Ampicillin resistance was not detected in E. faecalis but was detected in 91.1% of E. faecium. Vancomycin non-susceptibility was detected in 0.1% of E. faecalis and in 41.8% of E. faecium. Overall, 45.4% of E. faecium harboured vanA and/or vanB genes. For the vanA/vanB positive E. faecium isolates, 49.1% harboured vanA genes only and 50.6% vanB genes; 0.3% harboured both vanA and vanB genes. The percentage of E. faecium bacteraemia isolates resistant to vancomycin in Australia is substantially higher than that seen in most European countries. E. faecium consisted of 78 multilocus sequence types (STs), of which 75.0% of isolates were classified into six major STs containing ten or more isolates. All major STs belong to clonal cluster (CC) 17, a major hospital-adapted polyclonal E. faecium cluster. The predominant STs (ST1424, ST17, ST796, ST80, ST1421, and ST78) were found across most regions of Australia. The most prevalent clone was ST1424, which was identified in all regions except the Northern Territory and Western Australia. Overall, 51.4% of isolates belonging to the six predominant STs harboured vanA or vanB genes. In 2019, AESOP has shown that enterococcal bacteraemias in Australia are frequently caused by polyclonal ampicillin-resistant high-level gentamicin-resistant vanA or vanB E. faecium which have limited treatment options.

From 1 January to 31 December 2019, 39 institutions around Australia participated in the Australian Staphylococcus aureus Sepsis Outcome Programme (ASSOP). The aim of ASSOP 2019 was to determine the proportion of Staphylococcus aureus bacteraemia (SAB) isolates in Australia that are antimicrobial resistant, with particular emphasis on susceptibility to methicillin and on characterising the molecular epidemiology of the methicillin-resistant isolates. A total of 3,157 S. aureus bacteraemia episodes were reported, of which 79.8% were community-onset. 18.5% of S. aureus were methicillin resistant. The 30-day all-cause mortality associated with methicillin-resistant SAB was 14.0%, which was not significantly different from the 14.3% mortality associated with methicillin-susceptible SAB (p = 0.9). With the exception of the β-lactams and erythromycin, antimicrobial resistance in methicillin-susceptible S. aureus was rare. However, in addition to the β-lactams, approximately 36% of methicillin-resistant S. aureus (MRSA) were resistant to ciprofloxacin, 34% to erythromycin, 13% to tetracycline, 9% to gentamicin and 4% to co-trimoxazole. When applying the EUCAST breakpoints, teicoplanin resistance was detected in two S. aureus isolates. Resistance was not detected for vancomycin and linezolid. Resistance to non-beta-lactam antimicrobials was largely attributable to two healthcare-associated MRSA clones: ST22-IV [2B] (EMRSA-15) and ST239-III [3A] (Aus-2/3 EMRSA). ST22-IV [2B] (EMRSA-15) is the predominant healthcare-associated clone in Australia. Eighty percent of methicillin-resistant SAB, however, were due to community-associated clones. Although polyclonal, approximately 71.4% of community-associated clones were variously characterised as ST93-IV [2B] (Queensland CA-MRSA), ST5-IV [2B], ST45-VT [5C2&5], ST1-IV [2B], ST30-IV [2B], ST78-IV [2B] and ST8-IV [2B]. Community-associated MRSA (CA-MRSA), in particular the ST45-VT [5C2&5] clone, have acquired multiple antimicrobial resistance determinants including ciprofloxacin, erythromycin, clindamycin, gentamicin and tetracycline. The multiresistant ST45-VT [5C2&5] clone accounted for 12.7% of CA-MRSA. As CA-MRSA is well established in the Australian community, it is important that antimicrobial resistance patterns in community- and healthcare-associated SAB are monitored, as this information will guide therapeutic practices in treating S. aureus sepsis.

Background The global emergence of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) has seen the dominance of specific clones in different regions around the world with the PVL-positive ST93-IV as the predominant CA-MRSA clone in Australia. In this study we applied a genome-wide association study (GWAS) approach on a collection of Australian ST93-IV MRSA genomes to screen for genetic traits that might have assisted the ongoing transmission of ST93-IV in Australia. We also compared the genomes of ST93-IV bacteraemia and non-bacteraemia isolates to search for potential virulence genes associated with bacteraemia. Results Based on single nucleotide polymorphism phylogenetics we revealed two distinct ST93-IV clades circulating concurrently in Australia. One of the clades contained isolates primarily isolated in the northern regions of Australia whilst isolates in the second clade were distributed across the country. Analyses of the ST93-IV genome plasticity over a 15-year period (2002–2017) revealed an observed gain in accessory genes amongst the clone’s population. GWAS analysis on the bacteraemia isolates identified two gene candidates that have previously been associated to this kind of infection. Conclusions Although this hypothesis was not tested here, it is possible that the emergence of a ST93-IV clade containing additional virulence genes might be related to the high prevalence of ST93-IV infections amongst the indigenous population living in the northern regions of Australia. More importantly, our data also demonstrated that GWAS can reveal candidate genes for further investigations on the pathogenesis and evolution of MRSA strains within a same lineage.

Of 1033 Escherichia coli urinary tract infection isolates collected from females >12 years of age in Australia in 2019, only 2 isolates were resistant to fosfomycin with a minimum inhibitory concentration (MIC) of >256 mg/L. Despite having different multilocus sequence types, the two isolates harboured an identical plasmid-encoded fosA4 gene. The fosA4 gene has previously been identified in a single clinical E. coli isolate cultured in Japan in 2014. Each fosfomycin-resistant isolate harboured two conjugative plasmids that possessed an array of genes conferring resistance to aminoglycosides, β-lactams, macrolides, quinolones, sulfonamides and/or trimethoprim.

From 1 January to 31 December 2018, thirty-six institutions around Australia participated in the Australian Staphylococcus aureus Sepsis Outcome Programme (ASSOP). The aim of ASSOP 2018 was to determine the proportion of Staphylococcus aureus bacteraemia (SAB) isolates in Australia that are antimicrobial resistant, with particular emphasis on susceptibility to methicillin, and to characterise the molecular epidemiology of the methicillin-resistant isolates. A total of 2,673 S. aureus bacteraemia episodes were reported, of which 78.9% were community-onset. A total of 17.4% of S. aureus isolates were methicillin resistant. The 30-day all-cause mortality associated with methicillin-resistant SAB was 17.1% which was not significantly higher than the 13.6% mortality associated with methicillin-susceptible SAB (p = 0.1). With the exception of the β-lactams and erythromycin, antimicrobial resistance in methicillin-susceptible S. aureus was rare. However in addition to the β-lactams approximately 42% of methicillin-resistant S. aureus (MRSA) were resistant to erythromycin, 36% to ciprofloxacin and approximately 13% resistant to co-trimoxazole, tetracycline and gentamicin. When applying the EUCAST breakpoints teicoplanin resistance was detected in two S. aureus isolates. Resistance was not detected for vancomycin and linezolid. Resistance to non-beta-lactam antimicrobials was largely attributable to two healthcare-associated MRSA clones: ST22-IV [2B] (EMRSA-15) and ST239-III [3A] (Aus-2/3 EMRSA). The ST22-IV [2B] (EMRSA-15) clone is the predominant healthcare-associated clone in Australia. Seventy-eight percent of methicillin-resistant SAB episodes in 2018 were due to community-associated clones. Although polyclonal, approximately 76.3% of community-associated clones were characterised as ST93-IV [2B] (Queensland CA-MRSA), ST5-IV [2B], ST45-VT [5C2&5], ST1-IV [2B], ST30-IV [2B], ST78-IV [2B] and ST97-IV [2B]. Community-associated MRSA, in particular the ST45-VT [5C2&5] clone, has acquired multiple antimicrobial resistance determinants including ciprofloxacin, erythromycin, clindamycin, gentamicin and tetracycline. The ST45-VT [5C2&5] clone accounted for 11.7% of CA-MRSA. As CA-MRSA is well established in the Australian community, it is important that antimicrobial resistance patterns in community- and healthcare-associated SAB are monitored, as this information will guide therapeutic practices in treating S. aureus sepsis.

The worldwide incidence of neisserial infections, particularly gonococcal infections, is increasingly associated with antibiotic-resistant strains. In particular, extensively drug-resistant Neisseria gonorrhoeae strains that are resistant to third-generation cephalosporins are a major public health concern. There is a pressing clinical need to identify new targets for the development of antibiotics effective against Neisseria-specific processes. In this study, we report that the bacterial disulfide reductase DsbD is highly prevalent and conserved among Neisseria spp. and that this enzyme is essential for survival of N. gonorrhoeae. DsbD is a membrane-bound protein that consists of two periplasmic domains, n-DsbD and c-DsbD, which flank the transmembrane domain t-DsbD. In this work, we show that the two functionally essential periplasmic domains of Neisseria DsbD catalyze electron transfer reactions through unidirectional interdomain interactions, from reduced c-DsbD to oxidized n-DsbD, and that this process is not dictated by their redox potentials. Structural characterization of the Neisseria n- and c-DsbD domains in both redox states provides evidence that steric hindrance reduces interactions between the two periplasmic domains when n-DsbD is reduced, thereby preventing a futile redox cycle. Finally, we propose a conserved mechanism of electron transfer for DsbD and define the residues involved in domain–domain recognition. Inhibitors of the interaction of the two DsbD domains have the potential to be developed as anti-neisserial agents.

Neisseria meningitidis is the causative agent of invasive meningococcal disease (IMD). A recombinant vaccine called Bexsero® incorporates four subcapsular antigens (fHbp, NHBA, NadA and PorA) which are used to assign a Bexsero® antigen sequence type (BAST) to each meningococcal strain. The vaccine elicits an immune response against combinations of variants of these antigens which have been grouped into specific BAST profiles that have been shown to have different distributions within geographical locations thus potentially affecting the efficacy of the vaccine. In this study, invasive meningococcal disease isolates from the western seaboard of Australia (Western Australia; WA) were compared to those from the south-eastern seaboard (Victoria; VIC) from 2008 to 2012. Whole-genome sequencing (WGS) of 131 meningococci from VIC and 70 meningococci from WA were analysed for MLST, FetA and BAST profiling. Serogroup B predominated in both jurisdictions and a total of 10 MLST clonal complexes (cc) were shared by both states. Isolates belonging to cc22, cc103 and cc1157 were unique to VIC whilst isolates from cc60 and cc212 were unique to WA. Clonal complex 41/44 represented one-third of the meningococcal population in each state but the predominant ST was locally different: ST-6058 in VIC and ST-146 in WA. Of the 108 BAST profiles identified in this collection, only 9 BASTs were simultaneously observed in both states. A significantly larger proportion of isolates in VIC harboured alleles for the NHBA-2 peptide and fHbp-1, antigenic variants predicted to be covered by the Bexsero® vaccine. The estimate for vaccine coverage in WA (47.1% [95% CI: 41.1-53.1%]) was significantly lower than that in VIC (66.4% [95% CI: 62.3-70.5%]). In conclusion, the antigenic structure of meningococci causing invasive disease in two geographically distinct states of Australia differed significantly during the study period which may affect vaccine effectiveness and highlights the need for representative surveillance when predicting potential impact of meningococcal B vaccines.

Pathogenic meningococci have acquired a 24 kb capsule synthesis island (cps) by horizontal gene transfer which consists of a synthetic locus and associated capsule transport genes flanked by repetitive Regions D and D’. Regions D and D’ contain an intact gene encoding a UDP-galactose epimerase (galE1) and a truncated remnant (galE2), respectively. In this study, GalE protein alleles were shown to be either mono-functional, synthesising UDP-galactose (UDP-Gal), or bi-functional, synthesising UDP-Gal and UDP-galactosamine (UDP-GalNAc). Meningococci possessing a capsule null locus (cnl) typically possessed a single bi-functional galE. Separation of functionality between galE1 and galE2 alleles in meningococcal isolates was retained for all serogroups except serogroup E which has a synthetic requirement for UDP-GalNAc. The truncated galE2 remnant in Region D’ was also phylogenetically related to the bi-functional galE of the cnl locus suggesting common ancestry. A model is proposed in which the illegitimate recombination of the cps island into the galE allele of the cnl locus results in the formation of Region D’ containing the truncated galE2 locus and the capture of the cps island en bloc. The retention of the duplicated Regions D and D’ enables inversion of the synthetic locus within the cps island during bacterial growth.

Neisseria meningitidis (meningococcus) is the causative agent of invasive meningococcal disease that manifests as life-threatening septicaemia and/or meningitis. This review provides a brief overview of the prevention of the disease and also highlights the importance of whole genome sequencing (WGS) in detecting outbreaks of meningococci in Australia. The use of WGS in identifying the emergence of a penicillin-resistant cluster of meningococci is Western Australia is used as an example for advocating the implementation of WGS on the routine surveillance in Australia.

The decoration of the lipid A headgroups of the lipooligosaccharide (LOS) by the LOS phosphoethanolamine (PEA) transferase (LptA) in Neisseria spp. is central for resistance to polymyxin. The structure of the globular domain of LptA shows that the protein has five disulphide bonds, indicating that it is a potential substrate of the protein oxidation pathway in the bacterial periplasm. When neisserial LptA was expressed in Escherichia coli in the presence of the oxidoreductase, EcDsbA, polymyxin resistance increased 30-fold. LptA decorated one position of the E. coli lipid A headgroups with PEA. In the absence of the EcDsbA, LptA was degraded in E. coli. Neisseria spp. express three oxidoreductases, DsbA1, DsbA2 and DsbA3, each of which appear to donate disulphide bonds to different targets. Inactivation of each oxidoreductase in N. meningitidis enhanced sensitivity to polymyxin with combinatorial mutants displaying an additive increase in sensitivity to polymyxin, indicating that the oxidoreductases were required for multiple pathways leading to polymyxin resistance. Correlates were sought between polymyxin sensitivity, LptA stability or activity and the presence of each of the neisserial oxidoreductases. Only meningococcal mutants lacking DsbA3 had a measurable decrease in the amount of PEA decoration on lipid A headgroups implying that LptA stability was supported by the presence of DsbA3 but did not require DsbA1/2 even though these oxidoreductases could oxidise the protein. This is the first indication that DsbA3 acts as an oxidoreductase in vivo and that multiple oxidoreductases may be involved in oxidising the one target in N. meningitidis. In conclusion, LptA is stabilised by disulphide bonds within the protein. This effect was more pronounced when neisserial LptA was expressed in E. coli than in N. meningitidis and may reflect that other factors in the neisserial periplasm have a role in LptA stability.

We compared exemplar strains from two hypervirulent clonal complexes, strain NMB-CDC from ST-8/11 cc and strain MC58 from ST-32/269 cc, in host cell attachment and invasion. Strain NMB-CDC attached to and invaded host cells at a significantly greater frequency than strain MC58. Type IV pili retained the primary role for initial attachment to host cells for both isolates regardless of pilin class and glycosylation pattern. In strain MC58, the serogroup B capsule was the major inhibitory determinant affecting both bacterial attachment to and invasion of host cells. Removal of terminal sialylation of lipooligosaccharide (LOS) in the presence of capsule did not influence rates of attachment or invasion for strain MC58. However, removal of either serogroup B capsule or LOS sialylation in strain NMB-CDC increased bacterial attachment to host cells to the same extent. Although the level of inhibition of attachment by capsule was different between these strains, the regulation of the capsule synthesis locus by the two-component response regulator MisR, and the level of surface capsule determined by flow cytometry were not significantly different. However, the diplococci of strain NMB-CDC were shown to have a 1.89-fold greater surface area than strain MC58 by flow cytometry. It was proposed that the increase in surface area without changing the amount of anchored glycolipid capsule in the outer membrane would result in a sparser capsule and increase surface hydrophobicity. Strain NMB-CDC was shown to be more hydrophobic than strain MC58 using hydrophobicity interaction chromatography and microbial adhesion-to-solvents assays. In conclusion, improved levels of adherence of strain NMB-CDC to cell lines was associated with increased bacterial cell surface and surface hydrophobicity. This study shows that there is diversity in bacterial cell surface area and surface hydrophobicity within N. meningitidis which influence steps in meningococcal pathogenesis.

Neisseria meningitidis is the causative agent of invasive meningococcal disease (IMD). The BEXSERO® vaccine which is used to prevent serogroup B disease is composed of four sub-capsular protein antigens supplemented with an outer membrane vesicle. Since the sub-capsular protein antigens are variably expressed and antigenically variable amongst meningococcal isolates, vaccine coverage can be estimated by the meningococcal antigen typing system (MATS) which measures the propensity of the strain to be killed by vaccinated sera. Whole genome sequencing (WGS) which identifies the alleles of the antigens that may be recognised by the antibody response could represent, in future, an alternative estimate of coverage. In this study, WGS of 278 meningococcal isolates responsible for 62% of IMD in Western Australia from 2000–2014 were analysed for association of genetic lineage (sequence type [ST], clonal complex [cc]) with BEXSERO® antigen sequence type (BAST) and MATS to predict the annual vaccine coverage. A hyper-endemic period of IMD between 2000–05 was caused by cc41/44 with the major sequence type of ST-146 which was not predicted by MATS or BAST to be covered by the vaccine. An increase in serogroup diversity was observed between 2010–14 with the emergence of cc11 serogroup W in the adolescent population and cc23 serogroup Y in the elderly. BASTs were statistically associated with clonal complex although individual antigens underwent antigenic drift from the major type. BAST and MATS predicted an annual range of 44–91% vaccine coverage. Periods of low vaccine coverage in years post-2005 were not a result of the resurgence of cc41/44:ST-146 but were characterised by increased diversity of clonal complexes expressing BASTs which were not predicted by MATS to be covered by the vaccine. The driving force behind the diversity of the clonal complex and BAST during these periods of low vaccine coverage is unknown, but could be due to immune selection and inter-strain competition with carriage of non-disease causing meningococci.