Insights gained from sequencing Australian non-invasive and invasive Streptococcus pyogenes isolates.

Epidemiological data have indicated that invasive infections caused by the Gram-positive cocci Streptococcus pyogenes (group A streptococcus, GAS) have increased in many Australian states over the past two decades. In July 2022, invasive GAS (iGAS) infections became nationally notifiable in Australia via public-health agencies. Surveillance for S. pyogenes infections has been sporadic within the state of New South Wales (NSW). This has led to a lack of genetic data on GAS strains in circulation, particularly for non-invasive infections, which are the leading cause of GAS's burden on the Australian healthcare system. To address this gap, we used whole-genome sequencing to analyse the genomes of 318 S. pyogenes isolates collected within two geographical regions of NSW. Invasive isolates were collected in 2007-2017, whilst non-invasive isolates were collected in 2019-2021. We found that at least 66 different emm-types were associated with clinical disease within NSW. There was no evidence of any Australian-specific clones in circulation. The M1UK variant of the emm1 global pandemic clone (M1global) has been detected in our isolates from 2013 onwards. We detected antimicrobial-resistance genes (mainly tetM, ermA or ermB genes) in less than 10 % of our 318 isolates, which were more commonly associated with non-invasive infections. Superantigen virulence gene carriage was reasonably proportionate between non-invasive and invasive infection isolates. Our study adds rich data on the genetic makeup of historical S. pyogenes infections within Australia. Ongoing surveillance of invasive and non-invasive GAS infections within NSW by whole-genome sequencing is warranted to inform on outbreaks, antimicrobial resistance and vaccine coverage.

Nocardia species distribution and antimicrobial susceptibility within Australia.

BACKGROUND: Nocardia is a ubiquitous saprophyte capable of causing human disease. Disease is primarily respiratory or cutaneous, usually acquired via inhalation or inoculation. Under the influence of environmental and host factors, Nocardia incidence and species distribution demonstrate geographical variation. AIMS: To examine for differences in Nocardia incidence within Western Australia (WA) and analyse species distribution in the context of prior published studies. To analyse antibiogram data from a nationwide passive antimicrobial resistance surveillance program. METHODS: Retrospective extraction of laboratory data for Western Australian Nocardia isolates over a 21-year period. Analysis of Nocardia antimicrobial susceptibility testing data submitted to the Australian Passive Antimicrobial Resistance Surveillance (APAS) program between 2005 and 2022. RESULTS: Nine hundred sixty WA isolates were identified, giving an annual incidence of 3.03 per 100 000 population with apparent latitudinal variation. The four most common species identified within WA and amongst APAS isolates were N. nova, N. cyriacigeorgica, N. brasiliensis and N. farcinica. APAS data demonstrated that all species exhibited high rates of susceptibility to linezolid (100%) and trimethoprim-sulfamethoxazole (98%). Amikacin (>90% susceptibility for all species except N. transvalensis) was the next most active parenteral agent, superior to both carbapenems and third-generation cephalosporins. Susceptibility to oral antimicrobials (other than linezolid) demonstrated significant interspecies variation. CONCLUSIONS: We demonstrate geographical variation in the distribution of Nocardia incidence. Four species predominate in the Australian setting, and nationwide data confirm a high in vitro susceptibility to trimethoprim-sulphamethoxazole and linezolid, justifying their ongoing role as part of first-line empiric therapy.

Epidemiology of Group A Streptococcal bacteraemia in Hunter New England Local Health District, 2008 to 2019.

Invasive Group A Streptococcal infection (iGAS) is an uncommon but serious infection with Streptococcus pyogenes in a normally sterile body site. Manifestations include bacteraemia, necrotising fasciitis and toxic shock syndrome with attendant serious morbidity and mortality. An increasing incidence of iGAS has been observed in some regions of Australia. iGAS became a nationally notifiable condition from 1 July 2021. To determine if regional incidence has increased, and to identify priority populations, we undertook a retrospective data analysis of Group A Streptococcal (GAS) bacteraemia cases in Hunter New England Local Health District (HNELHD), New South Wales, Australia, from 1 January 2008 to 31 December 2019, as identified by NSW Health Pathology, John Hunter Hospital. A total of 486 cases were identified (age-standardised rate: 4.05 cases per 100,000 population per year). Incidence in HNELHD gradually increased over the study period (adjusted incidence rate ratio: 1.04; 95% confidence interval: 1.01-1.07) and was significantly higher in children under 5 years of age; in adults over 70 years of age; in males; and in First Nations peoples. A significant peak occurred in 2017 (9.00 cases per 100,000 population), the cause of which remains unclear. GAS bacteraemia is uncommon but severe, and incidence in HNELHD has slowly increased. Public health and clinical guidelines must address the needs of priority populations, which include young children, older adults and First Nations peoples. Routine surveillance and genomic analysis will help improve our understanding of iGAS and inform best public health management.

Gauge against the machine: revisiting quality for multi-targeted serology platforms.

Diagnosis of infections of public health significance, such as leptospirosis, often present challenges for laboratories. To counter common challenges and ensure quality driven health responses, rigorous validation and verification processes are required. Despite such rigor, however, can one be certain laboratory reports are truly reflective of infection, given the risk of rare, but potentially very significant quality oversights? Here we present a real-world scenario where diagnosis of leptospirosis cases was compromised over a 6-year period despite quality measures suggesting a well performing serological assay. A subsequent investigation revealed this was attributed to the programming of an automated microtitration plate analyser, evading detection by both quality control and external quality assurance processes. The quality oversight provides insight into potential limitations in quality processes in multi-targeted serological platforms.

Circulation of enterovirus D68 (EV-D68) causing respiratory illness in New South Wales, Australia, between August 2018 and November 2019.

The incidence of enterovirus D68 (EV-D68) in New South Wales, Australia, is unknown. As part of a state-wide surveillance program, enterovirus positive diagnostic specimens were assessed from patients presenting to hospitals with respiratory and meningitis syndromes from August 2018 to November 2019. Diagnostic enterovirus positive samples were collected from 339 patients and re-extracted followed by targeted PCR across the whole EV-D68 genome (7.4 kb). Obtained amplicons (n=208) were sequenced using Illumina sequencing technology and the phylogenetic relationships analysed relative to EV-D68 Fermon strain. We identified EV-D68 in 31 patients, both children (n=27) and adults (n=4). Phylogenetically, the majority (n=30) were from subclade B3, the same as that causing outbreaks of EV-D68 across the USA and Europe during 2018. These data strengthen the importance of having an active enterovirus surveillance network.

Severe community-acquired pneumonia due to Streptococcus pyogenes in the Newcastle area.

BACKGROUND: An apparent increase in the incidence of severe community-acquired pneumonia (CAP) caused by Streptococcus pyogenes (group A Streptococcus - GAS) was observed during 2017 in the Newcastle area. The study was undertaken to establish whether there was a true increase in severe S. pyogenes pneumonia and to explore its epidemiology and clinical features. METHODS: The study was a retrospective descriptive study of S. pyogenes pneumonia set in two tertiary referral hospitals in Newcastle, a large regional city, during the period 2007 to 2018. Subjects were adults identified as having S. pyogenes pneumonia by searching a database of severe CAP (defined as requiring intensive care unit [ICU] admission) for the period 2007-2018. Laboratory records were also searched for sterile site isolates of S. pyogenes to identify patients not requiring ICU admission. RESULTS: There were 13 cases of S. pyogenes CAP identified during the study period, of whom 12 (92%) required ICU admission. S. pyogenes accounted for 12/728 (1.6%) cases of severe CAP during the study period. The severity of S. pyogenes pneumonia was high despite a mean patient age of 48 years and 7/13 (54%) having no significant past medical history. The mortality rate was 2/13 (15%). Viral co-infection was found in 6/12 (50%) of patients tested. Overall 7/12 (58%) of the patients with severe S. pyogenes CAP during the study period presented in the winter or spring of 2017. CONCLUSIONS: Streptococcus pyogenes is a rare cause of severe CAP in the Newcastle area, but there was a marked increase in frequency observed during the 2017 influenza season. Further study of the epidemiology of invasive GAS (iGAS) disease in Newcastle is warranted to identify emerging trends in this severe infection.

Is prolonged incubation required for optimal recovery of Burkholderia cepacia complex in sputum from cystic fibrosis patients? Data versus dogma.

Cystic fibrosis (CF) expert groups globally recommend using selective medium for isolation of Burkholderia cepacia complex (BCC) from respiratory specimens of CF patients. However, there is no consensus available for optimal duration of incubation and recommendations are variable. The purpose of our study was to compare the difference in recovery of BCC in CF samples at 48 hours versus 7 days when inoculated on Burkholderia cepacia selective agar. A total of 307 consecutive clinical respiratory specimens from our local CF unit were studied prospectively (August 2017 to December 2017). All specimens were inoculated on Burkholderia cepacia medium, containing polymyxin B, gentamicin and ticarcillin. In our laboratory, these plates are routinely incubated for 48 hours as per the manufacturer's recommendation. However, for this study all plates with no growth at 48 hours were further incubated for total of 7 days at 35°C in O2. Plates were read daily to look for any growth. Microbial identification was performed using MALDI-TOF Vitek MS (database V3.0). Of the 307 CF respiratory specimens cultured, 177 (58%) were from paediatric and 130 (42%) were from adult patients; 155 (50%) specimens were sputum, 148 (48%) were cough swabs and four (1%) were bronchoalveolar lavage (BAL). All specimens from adults were sputum except one BAL. Thirteen (4%) cultures from eight adult and five paediatric specimens grew BCC. The majority (294, 96%) of specimens had no growth when incubated for 7 days. All 13 positive isolates recovered within 48 hours and there were no additional positive isolates found beyond 48 hours of incubation. We conclude from our analysis that prolonged incubation is not warranted for recovery of BCC in CF specimens if selective medium containing gentamicin and polymyxin is used. By adopting this approach of non-extended incubation, the burden of work on laboratory personnel can be significantly reduced and much faster turnaround time for CF cultures achieved. Our study confirms the results of recently published data on this point and challenges the prevailing dogma of utility of extended incubation for BCC isolation. For devising consensus statements for microbiology laboratories on this issue, CF societies and expert groups should consider reviewing data from the recent studies.

Next generation sequencing of human enterovirus strains from an outbreak of enterovirus A71 shows applicability to outbreak investigations.

BACKGROUND: The most recent documented Australian outbreak of enterovirus A71 (EV-A71) occurred in Sydney from 2012 to 2013. Over a four-month period more than 100 children presented to four paediatric hospitals with encephalitic presentations including fever and myoclonic jerks. The heterogeneous presentations included typical encephalomyelitis, and cardiopulmonary complications. OBJECTIVES: To characterise the genomes of enterovirus strains circulating during the 2013 Sydney EV-A71 outbreak and determine their phylogeny, phylogeography and association between genome and clinical phenotype. STUDY DESIGN: We performed an analysis of enterovirus (EV) positive specimens from children presenting to hospitals in the greater Sydney region of Australia during the 2013 outbreak. We amplified near full-length genomes of EV, and used next generation sequencing technology to sequence the virus. We used phylogenetic/phylogeographic analysis to characterize the outbreak viruses. RESULTS: We amplified and sequenced 23/63 (37 %) genomes, and identified the majority (61 %) as EV-A71. The EV-A71 sequences showed high level sequence homology to C4a genogroups of EV-A71 circulating in China and Vietnam during 2012-13. Phylogenetic analysis showed EV-A71 strains associated with more severe symptoms, including encephalitis or cardiopulmonary failure, grouped together more closely than those from patients with hand, foot and mouth disease. Amongst the non-EV-A71 sequences were five other EV subtypes (representing enterovirus subtypes A and B), reflecting the diversity of EV co-circulation within the community. CONCLUSIONS: This is the first Australian study investigating the near full-length genome of EV strains identified during a known outbreak of EV-A71. EV-A71 sequences were very similar to strains circulating in Asia during the same time period. Whole genome sequencing offers additional information over routine diagnostic testing such as characterisation of emerging recombinant strains and inform vaccine design.

Prevalence of hepatitis B and hepatitis C infection from a population-based study in Southern India.

OBJECTIVES: This study aims to estimate the prevalence of hepatitis B (HBV) and C (HCV) in the population through field-screening camps conducted by Chennai Liver Foundation, in the southern state of Tamil Nadu, India. This is the largest population-based study from Tamil Nadu. PATIENTS AND METHODS: A total of 75 camps were conducted across 14 districts of Tamil Nadu (2014-2017). Screening was done by rapid point-of-care assays (SD-bioline tests) and confirmed by enzyme-linked immunosorbent assay (Monolisa tests). Those tested negative were offered first dose of HBV vaccine. Positive patients with HBV count of more than 2000 IU/ml or HCV-RNA positive on quantitative analysis were treated. RESULTS: A total of 18 589 people were screened, with HBV infection detected in 303 (prevalence 1.63%) and HCV infection in 56 (prevalence 0.3%), with significant variation among districts. Males contributed to about three-fourths of detected HBV [233/303 (77%)] or HCV [41/56 (73%)] infection. Screening detected a higher overall HBV/HCV infection rate in rural [203 (2.52%) infections in 8047 people] than in urban [156 (1.47%) infections in 10 542 people] areas (P<0.0001). Slum areas had a HBV prevalence of 5%. In a dialysis unit, all patients were found to have either HBV/HCV infection. A total of 162/303 (54%) people with HBV and 27/56 (48%) with HCV infection were treated, and 7704 people received the first dose of HBV vaccine. CONCLUSION: The prevalence of HBV was 1.63% and HCV was 0.30% in Tamil Nadu. Three-fourths of HBV/HCV infected people were males. Prevalence of HBV/HCV was higher in rural areas. Slum area and dialysis unit had high HBV and HCV prevalence.

Does microbial colonisation of a neck drain predispose to surgical site infection: clean vs clean-contaminated procedures.

PURPOSE: The study was designed to assess the difference in microbiological colonisation and growth that may occur in drains, in the setting of clean-contaminated compared to clean head and neck surgery. METHODS: A prospective observational cohort study was performed. Surgical drain tips upon removal were sent for bacterial culture and the culture results were compared between clean-contaminated and clean procedures using mixed effects logistic regression. In all statistical analyses, a priori, p < 0.05 (two-tailed) was calculated to indicate statistical significance. RESULTS: One hundred and ten drains were examined in both clean-contaminated and clean procedures. Drains from clean-contaminated procedures had a significantly longer time in situ (11 vs 5 days, p < 0.001). Overall, significant evidence was seen for an association between procedure type and drain growth rates: 68% of clean-contaminated procedures; and 45% of clean procedures. Although not statistically significant, there was an increase in normal skin flora contaminated drains in clean-contaminated procedures (41 vs 25%). Rates of pathogenic skin organisms (15 vs 16%) and pathogenic oropharyngeal organisms (2.9 vs 0%) were similar for clean-contaminated vs clean procedure patients. CONCLUSION: This preliminary study demonstrated a higher rate of microbial contamination of neck drains that were placed during procedures that involved continuity with the upper aero-digestive tract and neck. Retrograde migration of skin flora along the drain is common but of no clinical significance. Similar rates of pathogenic microbial growth have been demonstrated thus far. However, selection of nosocomial pathogens due to extended antibiotic prophylaxis may pose a risk for infection. LEVEL OF EVIDENCE: 1b.