Search

Objectives: There is clinical uncertainty over the optimal treatment for penicillin-susceptible Staphylococcus aureus (PSSA) infections. Furthermore, there is concern that phenotypic penicillin susceptibility testing methods are not reliably able to detect some blaZ-positive S. aureus., Methods: Nine S. aureus isolates, including six genetically diverse strains harbouring blaZ, were sent in triplicate to 34 participating laboratories from Australia (n = 14), New Zealand (n = 6), Canada (n = 12), Singapore (n = 1) and Israel (n = 1). We used blaZ PCR as the gold standard to assess susceptibility testing performance of CLSI (P10 disc) and EUCAST (P1 disc) methods. Very major errors (VMEs), major error (MEs) and categorical agreement were calculated., Results: Twenty-two laboratories reported 593 results according to CLSI methodology (P10 disc). Nineteen laboratories reported 513 results according to the EUCAST (P1 disc) method. For CLSI laboratories, the categorical agreement and calculated VME and ME rates were 85% (508/593), 21% (84/396) and 1.5% (3/198), respectively. For EUCAST laboratories, the categorical agreement and calculated VME and ME rates were 93% (475/513), 11% (84/396) and 1% (3/198), respectively. Seven laboratories reported results for both methods, with VME rates of 24% for CLSI and 12% for EUCAST., Conclusions: The EUCAST method with a P1 disc resulted in a lower VME rate compared with the CLSI methods with a P10 disc. These results should be considered in the context that among collections of PSSA isolates, as determined by automated MIC testing, less than 10% harbour blaZ. Furthermore, the clinical relevance of phenotypically susceptible, but blaZ-positive S. aureus, remains unclear., (© The Author(s) 2023. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

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., (© The Author(s) 2021. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Objectives: Implementation of EUCAST susceptibility testing in an Australian hospital laboratory demonstrated higher rates of aminopenicillin and amoxicillin/clavulanate resistance in Haemophilus influenzae than previously recognized. This study aimed to better define the variability in the detection of β-lactam resistance based on EUCAST and CLSI disc diffusion (DD) methodology, by comparison with the recommended reference method, broth microdilution (BMD), and by concordance with genomic analysis., Methods: A total of 100 random H. influenzae isolates were assessed for ampicillin and amoxicillin/clavulanate susceptibility by EUCAST and CLSI DD and BMD. WGS was used to analyse the ftsI gene of a subset of isolates with β-lactam resistance, other than that due to isolated β-lactamase production., Results: Of the 100 isolates, 32 were categorized as either β-lactamase negative, ampicillin resistant (BLNAR) (n = 18) or β-lactamase positive, amoxicillin/clavulanate resistant (BLPACR) (n = 14) by EUCAST DD. All 18 EUCAST BLNAR isolates were genotypically confirmed by WGS. Five of 18 BLNAR isolates were concordant by CLSI DD, 12 by EUCAST BMD and 4 by CLSI BMD. Nine of 14 EUCAST BLPACR isolates were confirmed by WGS; the remaining 5 were 1 mm below the EUCAST DD breakpoint. Only one isolate was detected as BLPACR by CLSI DD. Group III mutations associated with high-level ampicillin resistance were identified in 10/32 isolates., Conclusions: The EUCAST DD susceptibility method is more reliable than either CLSI or BMD for the detection of genotypically defined BLNAR resistance. However, accurate categorization of amoxicillin/clavulanate resistance remains problematic. Continuous and reproducible surveillance of resistance is needed; for this to be possible, robust susceptibility methods are required., (© The Author(s) 2020. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Background: There is increasing knowledge of antimicrobial usage in children yet limited availability of nationally representative paediatric-specific data on antimicrobial resistance., Objectives: Paediatric data from this national surveillance programme are presented to explore differences between childhood and adult bloodstream infections and antimicrobial resistance surveillance., Methods: Using information collected from a prospective coordinated antimicrobial resistance surveillance programme, children ≤18 years and adults >18 years with a positive blood culture for Staphylococcus aureus, Enterococcus spp. or Gram-negative spp. presenting to one of 34 Australian hospitals during 2013-16 were evaluated. Consistent methodologies for key sepsis pathogens were employed and a comparative analysis between children and adults was conducted., Results: There are stark contrasts between children and adults in this national antimicrobial resistance (AMR) data set. Notable differences include lower rates of AMR, different clinical and molecular phenotypes and lower mortality amongst children. The burden of Gram-negative resistance is disproportionately experienced in children, with higher odds of death with an ESBL versus non-ESBL bacteraemia in comparison with adults., Conclusions: These data support that children are not just 'little adults' in the AMR era, and analyses by age group are important to detect differences in antibiotic susceptibility, clinical phenotype and genetic virulence factors. Antimicrobial surveillance incorporated into routine laboratory practice is vital to inform an array of wider applications including antimicrobial guidelines, stewardship and direction for prioritization of novel antimicrobial development., (© The Author(s) 2020. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Background: Early identification of MRSA by diagnostic medical microbiology laboratories enables improved antimicrobial choice and outcomes. The Cepheid Xpert® MRSA/SA BC test rapidly identifies Staphylococcus aureus bloodstream infections through spa gene detection and methicillin resistance via mecA gene detection. Recent emergence of S. aureus with deletions in the spa gene has resulted in false-negative results for this test, leading to misidentification of infections with this organism, particularly MRSA ST45., Objectives: To investigate the emergence and prevalence of ST45 MRSA in New South Wales (NSW), Australia., Methods: WGS read data from six NSW hospitals were collected for 131 ST45 MRSA isolates and analysed., Results: Of the 131 ST45 MRSA investigated, 88.5% (116/131) contained a deletion in the spa gene that appeared to have arisen once in approximately 2010 followed by clonal expansion. Given the successful establishment of this 'spa-deletion' MRSA clone, the Cepheid Xpert® MRSA/SA BC test became unreliable for confirming S. aureus bacteraemia in NSW. Subsequently, the algorithm used by this test has been updated and evaluated to take into account the presence of S. aureus with either a spa deletion or SCCmec target variations., Conclusions: This study highlighted the applied use of WGS for assessing diagnostic assays and informing necessary changes to ensure the viability of the Cepheid Xpert® MRSA/SA BC test in the context of the new 'spa-deletion' MRSA clone. It demonstrated how continued surveillance through WGS can reveal evolutionary events that may impact diagnostic assays, allowing corrective modifications to be made in real time., (© The Author(s) 2020. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Neisseria spp. possess four genogroups of filamentous prophages, termed Nf1 to 4. A filamentous bacteriophage from the Nf1 genogroup termed meningococcal disease-associated phage (MDA φ) is associated with clonal complexes of Neisseria meningitidis that cause invasive meningococcal disease. Recently, we recovered an isolate of Neisseria gonorrhoeae (ExNg63) from a rare case of gonococcal meningitis, and found that it possessed a region with 90% similarity to Nf1 prophages, specifically, the meningococcal MDA φ. This led to the hypothesis that the Nf1 prophage may be more widely distributed amongst the genus Neisseria. An analysis of 92 reference genomes revealed the presence of intact Nf1 prophages in the commensal species, Neisseria lactamica and Neisseria cinerea in addition to the pathogen N. gonorrhoeae. In N. gonorrhoeae, Nf1 prophages had a restricted distribution but were present in all representatives of MLST ST1918. Of the 160 phage integration sites identified, only one common insertion site was found between one isolate of N. gonorrhoeae and N. meningitidis. There was an absence of any obvious conservation of the receptor for prophage entry, PilE, suggesting that the phage may have been obtained by natural transformation. An examination of the restriction modification systems and mutated mismatch repair systems with prophage presence suggested that there was no obvious preference for these hosts. A timed phylogeny inferred that N. meningitidis was the donor of the Nf1 prophages in N. lactamica and N. gonorrhoeae. Further work is required to determine whether Nf1 prophages are active and can act as accessory colonization factors in these species., (© The Author(s) 2020. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

Background: There are few epidemiological data available to inform a national response to community-acquired methicillin-resistant Staphylococcus aureus (MRSA) in Papua New Guinea (PNG)., Methods: We performed a cross-sectional survey to determine the pattern of MRSA nasal colonization and the diversity of circulating MRSA clones among adults and adolescents in Madang Province, PNG., Results: S. aureus nasal colonization was confirmed in 44 (17.1%) of 257 participants. Four (9.1%) isolates were methicillin resistant. Resistance to other antimicrobial agents was uncommon. Detailed molecular typing of three MRSA isolates demonstrated multiple MRSA clones in this community, of which two carried the Panton-Valentin leukocidin-associated virulence genes., Conclusions: MRSA is likely to account for a clinically important proportion of staphylococcal disease in PNG. There are multiple MRSA clones in PNG. Ongoing surveillance of community and invasive isolates is a critical component of an effective response to the challenge of community-acquired MRSA in this and many other resource-limited contexts., (© The Author 2017. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Objectives: To investigate the genetic context associated with the emergence of vanA VRE in Australia., Methods: The whole genomes of 18 randomly selected vanA -positive Enterococcus faecium patient isolates, collected between 2011 and 2013 from hospitals in four Australian capitals, were sequenced and analysed., Results: In silico typing and transposon/plasmid assembly revealed that the sequenced isolates represented (in most cases) different hospital-adapted STs and were associated with a variety of different Tn 1546 variants and plasmid backbone structures., Conclusions: The recent emergence of vanA VRE in Australia was polyclonal and not associated with the dissemination of a single 'dominant' ST or vanA -encoding plasmid. Interestingly, the factors contributing to this epidemiological change are not known and future studies may need to consider investigation of potential community sources., (© The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Background: In Western Australia (WA), clonal complex 5, ST835, community-associated (CA) MRSA is isolated almost exclusively from aged care facilities. In WA four different staphylococcal cassette chromosome (SCC) mec (SCCmec) elements have been identified in this ST, indicating high genetic activity in the SCCmec region., Objectives: To investigate the SCC region of ST835 CA-MRSA WA MRSA-40 and determine the distribution of an SCCsorbitol element found within the region., Results: The SCC region contained a composite island, SCCmec WA MRSA-40 -CI, that was composed of three elements, ΨSCCpls, SCCsorbitol and SCCmecV T (5C2&5). This is the first time that a sorbitol operon has been reported in an SCC element., Conclusions: Generation of SCCmec WA MRSA-40 -CI has involved multiple genetic events and recombination with CoNS has occurred during evolution of the SCC elements. While Staphylococcus aureus is renowned for its ability to utilize mobile genetic elements to disseminate antimicrobial resistance, the SCC region of WA MRSA-40 shows that this clone has also utilized SCC elements to acquire extra virulence and possibly adapt to a niche environment., (© The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Staphylococcus aureus is a common cause of hospital, community and livestock-associated infections and is increasingly resistant to multiple antimicrobials. A significant proportion of antimicrobial-resistance genes are plasmid-borne, but only a minority of S. aureus plasmids encode proteins required for conjugative transfer or Mob relaxase proteins required for mobilisation. The pWBG749 family of S. aureus conjugative plasmids can facilitate the horizontal transfer of diverse antimicrobial-resistance plasmids that lack Mob genes. Here we reveal that these mobilisable plasmids carry copies of the pWBG749 origin-of-transfer (oriT) sequence and that these oriT sequences facilitate mobilisation by pWBG749. Sequences resembling the pWBG749 oriT were identified on half of all sequenced S. aureus plasmids, including the most prevalent large antimicrobial-resistance/virulence-gene plasmids, pIB485, pMW2 and pUSA300HOUMR. oriT sequences formed five subfamilies with distinct inverted-repeat-2 (IR2) sequences. pWBG749-family plasmids encoding each IR2 were identified and pWBG749 mobilisation was found to be specific for plasmids carrying matching IR2 sequences. Specificity of mobilisation was conferred by a putative ribbon-helix-helix-protein gene smpO. Several plasmids carried 2-3 oriT variants and pWBG749-mediated recombination occurred between distinct oriT sites during mobilisation. These observations suggest this relaxase-in trans mechanism of mobilisation by pWBG749-family plasmids is a common mechanism of plasmid dissemination in S. aureus., (© The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Objectives: To describe a family of conjugative plasmids isolated from colonizing community Staphylococcus aureus and determine their ability to mobilize unrelated antimicrobial resistance/virulence plasmids, not encoding mobilization functions., Methods: Plasmid pWBG749 was labelled with Tn551 (pWBG749e) to enable laboratory manipulation. Plasmid pWBG749e was conjugated into S. aureus of seven different lineages that harboured unrelated plasmids and mobilization experiments were performed. Plasmids were screened by EcoRI restriction and hybridization with probes prepared from unique pWBG749 conjugation genes., Results: Conjugative plasmids pWBG745, pWBG748 and pWBG749 belong to the same conjugative-plasmid family as the vancomycin resistance plasmid pBRZ01. Plasmid pWBG749e mobilized five unrelated plasmids. Mobilized plasmid pWBG744 is a pIB485-family plasmid that was also found in international S. aureus., Conclusions: Plasmid pWBG749e can mobilize unrelated S. aureus plasmids whose means of dissemination have not previously been understood., (© The Author 2014. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Background: It is uncertain whether particular clones causing invasive community-onset methicillin-resistant and methicillin-sensitive Staphylococcus aureus (cMRSA/cMSSA) infection differ in virulence., Methods: Invasive cMRSA and cMSSA cases were prospectively identified. Principal component analysis was used to derive an illness severity score (ISS) from clinical data, including 30-day mortality, requirement for intensive hospital support, the presence of bloodstream infection, and hospital length of stay. The mean ISS for each S. aureus clone (based on MLST) was compared with its DNA microarray-based genotype., Results: Fifty-seven cMRSA and 50 cMSSA infections were analyzed. Ten clones caused 82 (77%) of these infections and had an ISS calculated. The enterotoxin gene cluster (egc) and the collagen adhesin (cna) gene were found in 4 of the 5 highest-ranked clones (ST47-MSSA, ST30-MRSA-IV[2B], ST45-MSSA, and ST22-MRSA-IV[2B]) compared with none and 1 of the lowest 5 ranked clones, respectively. cMSSA clones caused more severe infection than cMRSA clones. The lukF/lukS Panton-Valentine leukocidin (PVL) genes did not directly correlate with the ISS, being present in the second, fourth, and 10th most virulent clones., Conclusions: The clinical severity of invasive cMRSA and cMSSA infection is likely to be attributable to the isolates' entire genotype rather than a single putative virulence determinant such as PVL.

Background: There are concerns about reduced efficacy of vancomycin in patients with Staphylococcus aureus bacteremia (SAB), especially when the minimum inhibitory concentration (MIC) nears the upper limit of the susceptible range., Methods: We examined the relationship between antibiotic treatment, 30-day mortality, and microbiologic parameters in a large Australasian cohort of patients with SAB., Results: We assessed 532 patients with SAB from 8 hospitals. All patients with methicillin-resistant S. aureus (MRSA) bacteremia were treated with vancomycin, and patients with methicillin-susceptible S. aureus (MSSA) bacteremia received either flucloxacillin or vancomycin. Increasing vancomycin MIC was associated with increased mortality in vancomycin-treated patients. However, even in patients with MSSA bacteremia treated with flucloxacillin, mortality was also higher if the vancomycin Etest MIC of their isolate was >1.5 μg/mL, compared with those with lower MIC isolates (26.8% vs 12.2%; P < .001). After adjustment in a multivariate model, age, hospital-onset SAB and vancomycin MIC were independently associated with mortality, but methicillin resistance and antibiotic choice were not., Conclusions: We have confirmed an association between higher vancomycin MIC and increased mortality in patients with SAB, but surprisingly this relationship was not related to the antibiotic treatment received, suggesting that the use of vancomycin per se is not responsible for the poorer outcome.

Objectives: Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) was first reported in remote regions of Western Australia (WA) in 1992 and is now the predominant MRSA isolated in the State. To gain insights into the emergence of CA-MRSA, 2146 people living in 11 remote WA communities were screened for colonization with S. aureus., Methods: Antibiogram analysis, contour-clamped homogeneous electric field electrophoresis, multilocus sequence typing, Panton-Valentine leucocidin determinant detection and accessory genetic regulator typing were performed to characterize the isolates. MRSA was further characterized by staphylococcal cassette chromosome mec typing., Results: The S. aureus population consisted of 13 clonal complexes and two Singleton lineages together with 56 sporadic isolates. Five lineages contained MRSA; however, these were not the predominant methicillin-susceptible S. aureus (MSSA) lineages. There was greater diversity amongst the MSSA while the MRSA appeared to have emerged clonally following acquisition of the staphylococcal cassette chromosome mec. Three MRSA lineages were considered to have been endemic in the communities and have subsequently become predominant lineages of CA-MRSA in the wider WA community. People colonized with MSSA tended to harbour clones of a different genetic lineage at each anatomical site while people colonized with MRSA tended to harbour clones of the same lineage at each site. Overall, the isolates were resistant to few antimicrobials., Conclusions: Although the evidence suggests that in WA CA-MRSA strains arose in remote communities and have now disseminated into the wider community, there is no evidence that they arose from the predominant MSSA clones in these communities.

Objectives: Methicillin-resistant staphylococci (MRS) isolates from healthy and diseased cats and dogs were characterized by staphylococcal cassette chromosome mec (SCCmec), multilocus sequence typing (MLST) and cassette chromosome recombinase gene (ccrAB) sequencing., Methods: PCR-directed SCCmec typing was carried out for all MRS isolates and two Staphylococcus aureus and two Staphylococcus epidermidis strains were analysed by MLST. Strains belonging to SCCmec type III and IV were sequenced for their ccrAB gene of allotypes 3 and 2, respectively., Results: Five types of SCCmec, types I, III, IV, IV (paediatric) and V SCCmec, were found. The S. aureus strains belonged to sequence type (ST) 239 and the two S. epidermidis belonged to ST43 and ST60 respectively. High sequence conservation was observed for the ccrAB gene of allotypes 2 and 3., Conclusions: MRS isolates from cats and dogs demonstrate a similar diversity of SCCmec types to those found in human staphylococci and ST239-MRSA-III, a widely dispersed strain in human hospitals, was identified in diseased dogs.

We developed a standardized DNA sequence-based approach for the accurate and timely identification of medically important fungi by sequencing polymerase chain reaction (PCR) products with a rapid automated capillary electrophoresis system. A simple DNA extraction method and PCR amplification using universal fungal primers was used to amplify ribosomal DNA from a range of clinical isolates and reference strains. The entire internal transcribed spacer (ITS) 1-5.8S-ITS2 ribosomal DNA region was sequenced using automated dye termination sequencing for 89 clinical isolates. These had previously been identified by traditional methods and included 12 ascomycetous yeast species, three basidiomycetous yeast species, eight dermatophyte species and two thermally dimorphic fungi, Scedosporium prolificans and S. apiospermum. Furthermore, 21 reference strains representing 19 different Candida species, Geotrichum candidum and Malassezia furfur were also sequenced as part of this study and were used either as standards for sequence-based comparisons, or as assay controls. Sequence-based identification was compared to traditional identification in a blinded manner. Of the clinical isolates tested, 88/89 had DNA sequences that were highly homologous to those of reference strains accessioned in GenBank, and 87/89 gave a sequence-based identification result that correlated with the traditional identification. In contrast to relatively slow conventional methods of identification, a sequence-based identification from a pure culture can be obtained within 24 h of a DNA extraction carried out after a minimal period of culture growth. We conclude that this approach is rapid, and may be a more accurate cost-effective alternative than most phenotypic methods for identification of many medically important fungi frequently encountered in a routine diagnostic microbiology laboratory.

The mecA gene that encodes methicillin resistance in Staphylococcus aureus may be regulated by the mecR1 and mecI genes, and this region has been referred to as the mec complex. An analysis of these regulatory genes in 35 epidemic methicillin-resistant S. aureus (MRSA) isolated in England and Australia has found that they contain three classes of mec complex. Firstly, the Class A mec complex with complete mecR1 and mecI genes. Secondly, a new variant of Class A, the Class A1 mec complex, with a 166 bp deletion in the membrane-spanning domain of mecR1 and a complete mecI gene. Thirdly, the Class B mec complex, in which the penicillin-binding domain of mecR1 and the whole mecI gene are deleted by the insertion of a partial sequence of IS1272. Seven MRSA isolated in England and Australia over different time periods had the Class A mec complex. However, the isolates did not have closely related pulsed-field gel electrophoresis (PFGE) patterns. The Class A1 mec complex was found in 12 Australian isolates and the English epidemic MRSA, EMRSA-1. All these organisms were isolated in the early 1980s and had closely related PFGE patterns. The Class B mec complex region was found in nine EMRSA and seven Australian MRSA isolated over the period from the 1970s to 2000. These isolates had related PFGE patterns. The mecA region was also compared in the isolates and all but two of the isolates had an XbaI restriction site. These results support the global spread of epidemic clones and confirm the close relationship between the Australian and English MRSA strains.