Protracted transmission and persistence of ST80 vancomycin-resistant Enterococcus faecium clonal complex types CT2933, CT2932 and CT1916 in a large Irish hospital: a 39-month whole-genome sequencing study.

BACKGROUND: Vancomycin-resistant Enterococcus faecium (VREfm) are significant nosocomial pathogens. Sequence type (ST) 80 vanA-encoding VREfm predominate in Irish hospitals, but their transmission is poorly understood. AIMS: To investigate transmission and persistence of predominant complex type (CT) VREfm in two wards of an Irish hospital (H1) using whole-genome sequencing, and their intra- and inter-hospital dissemination. METHODS: Rectal screening (N = 330, September 2019 to December 2022) and environmental (N = 48, November 2022 to December 2022) E. faecium were investigated. Isolate relatedness was assessed by core-genome multi-locus sequence typing (cgMLST) and core-genome single nucleotide polymorphism (cgSNP) analysis. Likely transmission chains were identified using SeqTrack (https://graphsnp.fordelab.com/graphsnp) using cgSNP data and recovery location. Well-characterized E. faecium (N = 908) from seven Irish hospitals including H1 (June 2017 to July 2022) were also investigated. FINDINGS: Conventional MLST assigned isolates to nine STs (ST80, 82%). cgMLST identified three predominant ST80 CTs (CT2933, CT2932 and CT1916) (55% of isolates) of related isolates (≤20 allelic differences). cgSNP analysis differentiated these CTs into multiple distinct closely related genomic clusters (≤10 cgSNPs). Parisimonious network construction identified 55 likely inter- and intra-ward transmissions with epidemiological support between patients ≤30 days involving 73 isolates (≤10 cgSNPs) from seven genomic clusters. Numerous other likely transmissions over longer time periods without evident epidemiological links were identified, suggesting persistence and unidentified reservoirs contribute to dissemination. The three CTs predominated among E. faecium (N = 1286) in seven hospitals, highlighting inter-hospital spread without known epidemiological links. CONCLUSION: This study revealed the long-term intra- and inter-hospital dominance of three major CT ST80 VREfm lineages, widespread transmission and persistence, implicating unidentified reservoirs.

Immersion of debrided diabetic foot ulcer tissue in electrochemically generated pH neutral hypochlorous acid significantly reduces the microbial bioburden: whole-genome sequencing of Staphylococcus aureus, the most prevalent species recovered.

BACKGROUND: Diabetic foot ulcer infections (DFUIs) are the leading cause of lower-limb amputations, mediated predominantly by Staphylococcus aureus. pH-neutral electrochemically generated hypochlorous acid (anolyte) is a non-toxic, microbiocidal agent with significant potential for wound disinfection. AIMS: To investigate both the effectiveness of anolyte for microbial bioburden reduction in debrided ulcer tissues and the population of resident S. aureus. METHODS: Fifty-one debrided tissues from 30 people with type II diabetes were aliquoted by wet weight and immersed in 1- or 10-mL volumes of anolyte (200 parts per million) or saline for 3 min. Microbial loads recovered were determined in colony forming units/g (cfu/g) of tissue following aerobic, anaerobic and staphylococcal-selective culture. Bacterial species were identified and 50 S. aureus isolates from 30 tissues underwent whole-genome sequencing (WGS). FINDINGS: The ulcers were predominantly superficial, lacking signs of infection (39/51, 76.5%). Of the 42/51 saline-treated tissues yielding ≥105 cfu/g, a microbial threshold reported to impede wound-healing, only 4/42 (9.5%) were clinically diagnosed DFUIs. Microbial loads from anolyte-treated tissues were significantly lower than saline-treated tissues using 1 mL (1065-fold, 2.0 log) and 10 mL (8216-fold, 2.1 log) immersion volumes (P<0.0005). S. aureus was the predominant species recovered (44/51, 86.3%) and 50 isolates underwent WGS. All were meticillin susceptible and comprised 12 sequence types (STs), predominantly ST1, ST5 and ST15. Whole-genome multi-locus sequence typing identified three clusters of closely related isolates from 10 patients indicating inter-patient transmission. CONCLUSIONS: Short immersions of debrided ulcer tissue in anolyte significantly reduced microbial bioburden: a potential novel DFUI treatment.

An emerging Panton-Valentine leukocidin-positive CC5-meticillin-resistant Staphylococcus aureus-IVc clone recovered from hospital and community settings over a 17-year period from 12 countries investigated by whole-genome sequencing.

BACKGROUND: A novel Panton-Valentine leukocidin (PVL)-positive meticillin-resistant Staphylococcus aureus (MRSA) clonal complex (CC)5-MRSA-IVc ('Sri Lankan' clone) was recently described from Sri Lanka. Similar isolates caused a recent Irish hospital outbreak. AIM: To investigate the international dissemination and diversity of PVL-positive CC5-MRSA-IVc isolates from hospital and community settings using whole-genome sequencing (WGS). METHODS: Core-genome single nucleotide polymorphism (cgSNP) analysis, core-genome multi-locus sequence typing (cgMLST) and microarray-based detection of antimicrobial-resistance and virulence genes were used to investigate PVL-positive CC5-MRSA-IVc (N = 214 including 46 'Sri Lankan' clone) from hospital and community settings in 12 countries over 17 years. Comparators included 29 PVL-positive and 23 PVL-negative CC5/ST5-MRSA-I/II/IVa/IVc/IVg/V. RESULTS: Maximum-likelihood cgSNP analysis grouped 209/214 (97.7%) CC5-MRSA-IVc into Clade I; average of 110 cgSNPs between isolates. Clade III contained the five remaining CC5-MRSA-IVc; average of 92 cgSNPs between isolates. Clade II contained seven PVL-positive CC5-MRSA-IVa comparators, whereas the remaining 45 comparators formed an outlier group. Minimum-spanning cgMLST analysis revealed a comparably low average of 57 allelic differences between all CC5/ST5-MRSA-IVc. All 214 CC5/ST5-MRSA-IVc were identified as 'Sri Lankan' clone, predominantly spa type t002 (186/214) with low population diversity and harboured a similar range of virulence genes and variable antimicrobial-resistance genes. All 214 Sri Lankan clone isolates and Clade II comparators harboured a 9616-bp chromosomal PVL-encoding phage remnant, suggesting both arose from a PVL-positive meticillin-susceptible ancestor. Over half of Sri Lankan clone isolates were from infections (142/214), and where detailed metadata were available (168/214), most were community associated (85/168). CONCLUSIONS: Stable chromosomal retention of pvl may facilitate Sri-Lankan clone dissemination.

Effective cleaning and decontamination of the internal air and water channels, heads and head-gears of multiple contra-angle dental handpieces using an enzymatic detergent and automated washer-disinfection in a dental hospital setting.

BACKGROUND: Dental handpieces (DHPs) are reusable invasive medical devices that must be cleaned, decontaminated, lubricated and steam sterilized after use. DHPs have a complex internal design including narrow channels, contamination of which can compromise sterilization. DHPs are not designed for routine disassembly, making cleaning/decontamination efficacy difficult to monitor. Washer-disinfection is the preferred method of decontaminating DHPs, but few studies have investigated its direct effectiveness at reducing microbial contamination internally. AIMS: To use contra-angle DHPs as a model system to investigate the effectiveness of washer-disinfection at reducing microbial contamination of internal components of multiple DHPs. METHODS: The air and water channels and heads of 10 disassembled contra-angle DHPs (BienAir, Biel/Bienne, Switzerland) were inoculated separately with 108 colony forming units (cfu) of Pseudomanas aeruginosa, Staphylococcus aureus, Enterococcus hirae or Candida albicans in the presence of 0.3% bovine serum albumin (BSA) (clean conditions), 3.0% BSA or 10% artificial test soil (dirty conditions). After reassembly, all 10 DHPs underwent washer-disinfection simultaneously in a Míele (Míele Ireland Ltd., Dublin, Ireland) PG8528 washer-disinfector and were tested for reductions in micro-organisms and protein. Additional experiments were undertaken with three lubricated DHPs inoculated with S. aureus and 10% test soil. All experiments were repeated in triplicate. FINDINGS: On average, an approximate 5 log or greater reduction in microbial cfu and a >93% reduction in protein from DHP heads and channels was consistently recorded following washer-disinfection for all DHPs under all conditions tested. CONCLUSIONS: The internal components of multiple DHPs can be effectively cleaned and decontaminated by washer-disinfection.

Meticillin-susceptible Staphylococcus aureus transmission among healthcare workers, patients and the environment in a large acute hospital under non-outbreak conditions investigated using whole-genome sequencing.

BACKGROUND: The role of meticillin-susceptible Staphylococcus aureus (MSSA) colonization of healthcare workers (HCWs), patients and the hospital environment in MSSA transmission events (TEs) is poorly understood. AIMS: The role of meticillin-resistant Staphylococcus aureus (MRSA) was investigated recently under non-outbreak conditions in a large hospital with a history of endemic MRSA over 2 years using whole-genome sequencing (WGS). Numerous potential MRSA TEs were identified. The present study investigated MSSA TEs from the same sources during the same 2-year hospital study. METHODS: HCW (N=326) and patient (N=388) volunteers on nine wards were tested for nasal and oral MSSA colonization over 2 years. Near-patient environment (N=1164), high-frequency touch sites (N=810) and air (N=445) samples were screened for MSSA. Representative MSSA and clinical isolates were sequenced and analysed by core genome multi-locus sequence typing. Closely related isolates (≤24 allelic differences) were segregated into related isolate groups (RIGs). Potential TEs involving MSSA in RIGs from HCWs, patients and patient infections were identified in combination with epidemiological data. FINDINGS: In total, 635 MSSA were recovered: clinical isolates (N=82), HCWs (N=170), patients (N=120), and environmental isolates (N=263). Twenty-four clonal complexes (CCs) were identified among 406/635 MSSA sequenced, of which 183/406 segregated into 59 RIGs. Numerous potential HCW-to-patient, HCW-to-HCW and patient-to-patient TEs were identified, predominantly among CC5-MSSA, CC30-MSSA and CC45-MSSA. HCW, patient, clinical and environmental isolates were identified in 33, 24, six and 32 RIGs, respectively, with 19/32 of these containing MSSA related to HCW and/or patient isolates. CONCLUSIONS: WGS detected numerous potential hospital MSSA TEs involving HCWs, patients and environmental contamination under non-outbreak conditions.

Meticillin-resistant Staphylococcus aureus transmission among healthcare workers, patients and the environment in a large acute hospital under non-outbreak conditions investigated using whole-genome sequencing.

BACKGROUND: The role of meticillin-resistant Staphylococcus aureus (MRSA) colonization of healthcare workers (HCWs), patients and the hospital environment in MRSA transmission in non-outbreak settings is poorly understood. AIMS: To investigate transmission events (TEs) involving HCWs, patients and the environment under non-outbreak conditions in a hospital with a history of endemic MRSA using whole-genome sequencing (WGS). METHODS: HCW (N = 326) and patient (N = 388) volunteers on nine wards were tested for nasal and oral MRSA colonization over two years. Near-patient environment (N = 1164), high-frequency touch sites (N = 810) and air (N = 445) samples were screened for MRSA. Representative MRSA and clinical isolates were analysed by WGS and core-genome multi-locus sequence typing (cgMLST). Closely related isolates (≤24 allelic differences) were segregated into related isolated groups (RIGs). FINDINGS: In total, 155 MRSA were recovered: clinical isolates (N = 41), HCWs (N = 22), patients (N = 37), environmental isolates (N = 55). Nine clonal complexes (CCs) were identified among 110/155 MRSA sequenced with 77/110 assigned to CC22. Seventy-nine MRSA segregated into 17 RIGs. Numerous potential TEs were associated with CC22-MRSA (RIGs 1-15), CC45-MRSA (RIG-16) and CC8-MRSA (RIG-17). RIG-1, (the largest RIG) contained 24 ST22-MRSA-IVh from six HCWs, six patients, four clinical and eight environmental samples recovered over 17 months involving 7/9 wards. TEs involving HCW-to-patient, HCW-to-HCW, patient-to-patient and environmental contamination by HCW/patient isolates were evident. HCW, patient, clinical and environmental isolates were identified in four, nine, seven and 13 RIGs, respectively, with 12/13 of these containing isolates closely related to HCW and/or patient isolates. CONCLUSIONS: WGS detected numerous potential hospital MRSA TEs involving HCWs, patients and the environment under non-outbreak conditions.

Effects of a randomized controlled trial of a brief, student-nurse led, parent-based sexual health intervention on parental protective factors and HPV vaccination uptake.

BACKGROUND: Parents play a pivotal role in adolescent sexual health and Human Papillomavirus (HPV) vaccination. Nurses are on the frontlines of healthcare and play a critical role in promoting HPV vaccination and parent-child sexual health communication. We enhanced the Families Talking Together (FTT) parent-based sexual health curriculum to include adolescent vaccinations herein, FTT + HPV, and trained student nurses to provide a strong HPV vaccination and parent-child sexual health communication endorsement. METHODS: Using a randomized attention-controlled trial design, we examined the efficacy of FTT + HPV among 519 parents and their 11-14 year old youth recruited from medically underserved communities between 2015 and 2018. Participants were recruited from 22 after-school programs (e.g., Boys and Girls Clubs) and 19 charter schools. For parents, we examined protective factors including parent-child sexual health communication and parental involvement. For youth, we examined sexual health knowledge, parent-child sexual health communication, and parent-child connectedness. To assess HPV vaccination initiation and completion, we searched IMMTRAC immunization registry records for 85% of youth and used parental report for youth without registry records. Group differences were calculated using the estimated mean difference at one- and six months post-intervention with significance set at the p < 0.05 level. RESULTS: Baseline rates of HPV vaccination were low at 55.7%. No significant difference between the groups was seen in vaccination initiation or completion rates by one-month post-intervention. However, by six-months post intervention, there was a significant difference between the groups with 70.3% of the intervention group initiating the HPV vaccination series vs. 60.6% for the control group (p = 0.02). No difference between the groups was found for HPV series completion at six-months. There were significant differences in condom knowledge (p = 0.04), parent-child connectedness (p = 0.04), and communication frequency (p = 0.001) with greater improvement in the intervention vs. the control group. Rates of sexual activity remained low in both groups throughout the six-month follow-up period. CONCLUSION: A brief parent-based adolescent sexual health and HPV vaccination intervention delivered by student nurses can improve sexual health outcomes including protective parental factors, adolescent sexual health knowledge, and HPV vaccination initiation rates. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02600884 . Prospectively registered September 1, 2015.

Multiple distinct outbreaks of Panton-Valentine leucocidin-positive community-associated meticillin-resistant Staphylococcus aureus in Ireland investigated by whole-genome sequencing.

BACKGROUND: Panton-Valentine leucocidin (PVL)-positive community-associated meticillin-resistant Staphylococcus aureus (CA-MRSA) is increasingly associated with infection outbreaks. AIM: To investigate multiple suspected PVL-positive CA-MRSA outbreaks using whole-genome sequencing (WGS). METHODS: Forty-six suspected outbreak-associated isolates from 36 individuals at three separate Irish hospitals (H1-H3) and from separate incidents involving separate families associated with H2 were investigated by whole-genome multi-locus sequence typing (wgMLST). FINDINGS: Two clusters (CH1 and CH2) consisting of 8/10 and 6/6 PVL-positive t008 ST8-MRSA-IVa isolates from H1 and H2, respectively, were identified. Within each cluster, neighbouring isolates were separated by ≤5 allelic differences; however, ≥73 allelic differences were identified between the clusters, indicating two independent outbreaks. Isolates from the H3 maternity unit formed two clusters (CH3-SCI and CH3-SCII) composed of four PVL-negative t4667 ST5-MRSA-V and 14 PVL-positive t002 ST5-MRSA-IVc isolates, respectively. Within clusters, neighbouring isolates were separated by ≤24 allelic differences, whereas both clusters were separated by 1822 allelic differences, indicating two distinct H3 outbreaks. Eight PVL-positive t127 ST1-MRSA-V+fus and three PVL-negative t267 ST97-MRSA-V+fus isolates from two distinct H2-associated families FC1 (N = 4) and FC2 (N = 7) formed three separate clusters (FC1 (t127), FC2 (t127) and FC2 (t267)). Neighbouring isolates within clusters were closely related and exhibited ≤7 allelic differences. Intrafamilial transmission was apparent, but the detection of ≥48 allelic differences between clusters indicated no interfamilial transmission. CONCLUSION: The frequent importation of PVL-positive CA-MRSA into healthcare settings, transmission and association with outbreaks is a serious ongoing concern. WGS is a highly discriminatory, informative method for deciphering such outbreaks conclusively.

Hospital outbreak of linezolid-resistant and vancomycin-resistant ST80 Enterococcus faecium harbouring an optrA-encoding conjugative plasmid investigated by whole-genome sequencing.

BACKGROUND: Linezolid is an antibiotic used to treat infections caused by multi-drug-resistant Gram-positive bacteria. Linezolid resistance in enterococci has been reported with increasing frequency, with a recent rise in resistance encoded by optrA, poxtA or cfr. AIM: To investigate a hospital outbreak of linezolid- and vancomycin-resistant Enterococcus faecium (LVREfm) using whole-genome sequencing (WGS). METHODS: Thirty-nine VREfm from patient screening (19 isolates, 17 patients) and environmental sites (20 isolates) recovered in October 2019 were investigated. Isolates were screened using polymerase chain reaction for optrA, poxtA and cfr, and underwent Illumina MiSeq WGS. Isolate relatedness was assessed using E. faecium core genome multi-locus sequence typing (cgMLST). One LVREfm underwent MinION long-read WGS (Oxford Nanopore Technologies) and hybrid assembly with MiSeq short-read sequences to resolve an optrA-encoding plasmid. FINDINGS: Twenty isolates (51.3%) were LVREfm and optrA-positive, including the LVREfm from the index patient. A closely related cluster of 28 sequence type (ST) 80 isolates was identified by cgMLST, including all 20 LVREfm and eight linezolid-susceptible VREfm, with an average allelic difference of two (range 0-10), indicating an outbreak. Nineteen (95%) LVREfm harboured a 56,684-bp conjugative plasmid (pEfmO_03). The remaining LVREfm exhibited 44.1% sequence coverage to pEfmO_03. The presence of pEfmO_03 in LVREfm and the close relatedness of the outbreak cluster isolates indicated the spread of a single strain. The outbreak was terminated by enhanced infection prevention and control (IPC) and environmental cleaning measures, ceasing ward admissions and ward-dedicated staff. CONCLUSION: WGS was central in investigating an outbreak of ST80 LVREfm. The rapid implementation of enhanced IPC measures terminated the outbreak.

The oral cavity revealed as a significant reservoir of Staphylococcus aureus in an acute hospital by extensive patient, healthcare worker and environmental sampling.

BACKGROUND: Methicillin-susceptible Staphylococcus aureus (MSSA) bloodstream infection rates have risen steadily in recent years, with a marked decline in the corresponding rates due to methicillin-resistant S. aureus (MRSA). Screening for MSSA carriage is not routinely undertaken and MRSA screening is not universal, so the extent of S. aureus colonisation pressure in nosocomial settings is unknown. METHODS: We conducted a prospective, observational study of patients and healthcare workers (HCWs) across nine inpatient wards in a tertiary referral hospital over a two-year period. Participants were screened for MSSA and MRSA using nasal swabs and oral rinses. Environmental surfaces and air were also tested for S. aureus using contact plates and active air sampling. FINDINGS: We enrolled 388 patients and 326 HCWs; and took 758 contact plate samples from surfaces and 428 air samples. MSSA was recovered from 24% of patients, 31.3% of HCWs, 16% of air samples and 7.9% of surface samples. MRSA was recovered from 6.4% of patients, 3.7% of HCWs, 2.5% of air samples and 2.2% of surface samples. Inclusion of the oral cavity in addition to the anterior nares in the sampling regimen identified 30 patients and 36 HCWs who exhibited exclusive oral colonisation. CONCLUSIONS: The oral cavity comprises a significant nosocomial reservoir for S. aureus that is currently under-appreciated. Oral screening should be considered both in terms of the colonisation pressure in a healthcare facility, and on an individual patient level, especially in patients where decolonisation attempts have repeatedly failed and those undergoing high risk procedures.

Whole-genome sequencing identifies highly related Pseudomonas aeruginosa strains in multiple washbasin U-bends at several locations in one hospital: evidence for trafficking of potential pathogens via wastewater pipes.

BACKGROUND: Hand washbasin U-bends have increasingly been associated with nosocomial outbreaks by Gram-negative bacteria, including Pseudomonas aeruginosa which is virtually ubiquitous in U-bends. Wastewater networks servicing U-bends are potential highways for trafficking pathogenic bacteria. AIM: To use P. aeruginosa to investigate trafficking of bacteria between hospital washbasin U-bends. METHODS: Twenty-five washbasin U-bends in five locations in Dublin Dental University Hospital (DDUH) were investigated for trafficking of P. aeruginosa: 10 in Clinic 2 (C2), 10 in the Accident & Emergency Department (A&E) and five in three other locations. In addition, washbasin tap samples (N=80) and mains and tap water samples (N=72) were cultured for P. aeruginosa. Selected P. aeruginosa isolates recovered over 29 months underwent whole-genome sequencing, and relatedness was interpreted using whole-genome multi-locus sequence typing and pairwise single nucleotide polymorphism (SNP) analysis. FINDINGS: P. aeruginosa was recovered from all U-bends but not from taps or water. Eighty-three U-bend isolates yielded 10 sequence types (STs), with ST560 and ST179 from A&E, C2 and two other locations predominating (70%). ST560 was also recovered from a common downstream pipe. Isolates within ST560 and ST179 were highly related regardless of source. ST560 was divided into Cluster I (N=25) and Cluster II (N=2) with average allelic differences and SNPs of three and zero, and two and five, respectively. The 31 ST179 isolates exhibited an average allelic difference and SNPs of three and 12, respectively. CONCLUSION: Highly related P. aeruginosa strains were identified in multiple U-bends in several DDUH locations, indicating trafficking via the wastewater network.

Contribution of whole-genome sequencing to understanding of the epidemiology and control of meticillin-resistant Staphylococcus aureus.

In recent years, approaches to tracking the spread of meticillin-resistant Staphylococcus aureus (MRSA) as part of outbreak management have used conventional DNA-based methods, including pulsed-field gel electrophoresis and spa typing. However, when a predominant clone is present, these methods may be insufficiently discriminatory. A literature search was undertaken to highlight how whole-genome sequencing (WGS) has revolutionized the investigation of outbreaks of MRSA, including intrahospital spread and MRSA in the community, and to review its future potential. WGS provides enhanced isolate discrimination, as it permits the entire genomic DNA sequence of isolates to be determined and compared rapidly. Software packages used for the analysis of WGS data are becoming increasingly available. To date, WGS has been more sensitive in confirming outbreaks, often persisting for prolonged periods, previously undetected by conventional molecular typing. The evolving dynamic of spread from the community to hospitals, within and between hospitals, and from hospitals to the community is only becoming clear with WGS studies, and is more complex and convoluted than widely appreciated. Also, WGS can exclude cross-transmission, when isolates are different. The challenges now are to make WGS technology more amenable for routine use, and to develop an evidence-based consensus for sequence difference thresholds for isolates that are deemed to be part of the same outbreak, including protracted outbreaks. Using such data in a timely way will provide increased sensitivity in detecting cross-transmission events at an earlier stage, with the potential to prevent outbreaks, and have a positive impact on infection prevention and control.

Minimizing microbial contamination risk simultaneously from multiple hospital washbasins by automated cleaning and disinfection of U-bends with electrochemically activated solutions.

BACKGROUND: Outbreaks of infection associated with microbial biofilm in hospital hand washbasin U-bends are being reported increasingly. In a previous study, the efficacy of a prototype automated U-bend decontamination method was demonstrated for a single non-hospital pattern washbasin. It used two electrochemically activated solutions (ECA) generated from brine: catholyte with detergent properties and anolyte with disinfectant properties. AIM: To develop and test a large-scale automated ECA treatment system to decontaminate 10 hospital pattern washbasin U-bends simultaneously in a busy hospital clinic. METHODS: A programmable system was developed whereby the washbasin drain outlets, U-bends and proximal wastewater pipework automatically underwent 10-min treatments with catholyte followed by anolyte, three times weekly, over five months. Six untreated washbasins served as controls. Quantitative bacterial counts from U-bends were determined on Columbia blood agar, Reasoner's 2A agar and Pseudomonas aeruginosa selective agar following treatment and 24 h later. FINDINGS: The average bacterial densities in colony-forming units/swab from treated U-bends showed a >3 log reduction compared with controls, and reductions were highly significant (P<0.0001) on all media. There was no significant increase in average bacterial counts from treated U-bends 24 h later on all media (P>0.1). P. aeruginosa was the most prevalent organism recovered throughout the study. Internal examination of untreated U-bends using electron microscopy showed dense biofilm extending to the washbasin drain outlet junction, whereas treated U-bends were free from biofilm. CONCLUSION: Simultaneous automated treatment of multiple hospital washbasin U-bends with ECA consistently minimizes microbial contamination and thus the associated risk of infection.

Reduced pro-inflammatory responses to Staphylococcus aureus bloodstream infection and low prevalence of enterotoxin genes in isolates from patients on haemodialysis.

Patients with end-stage renal failure undergo regular haemodialysis (HD) and often develop episodes of Staphylococcus aureus bloodstream infection (BSI), which can re-occur. However, clinically, patients on HD, with S. aureus BSI, respond well to treatment, rarely developing overt signs of sepsis. We investigated the contributions of bacterial virulence and cytokine responses to the clinical course of S. aureus BSI in HD and non-HD patients. Seventy patients were recruited, including 27 (38.6 %) patients on HD. Isolates were spa-typed and virulence and antimicrobial resistance gene carriage was investigated using DNA microarray analysis. Four inflammatory cytokines, IL-6, RANTES, GROγ and leptin, were measured in patient plasma on the day of diagnosis and after 7 days. There was no significant difference in the prevalence of genotypes or antimicrobial resistance genes in S. aureus isolates from HD compared to non-HD patients. The enterotoxin gene cluster (containing staphylococcal enterotoxins seg, sei, sem, sen, seo and seu) was significantly less prevalent among BSI isolates from HD patients compared to non-HD patients. Comparing inflammatory cytokine response to S. aureus BSI in HD patients to non-HD patients, IL-6 and GROγ were significantly lower (p = 0.021 and p = 0.001, respectively) in HD patients compared to other patients on the day of diagnosis and RANTES levels were significantly lower (p = 0.025) in HD patients on day 7 following diagnosis. Lowered cytokine responses in HD patients and a reduced potential for super-antigen production by infecting isolates may partly explain the favourable clinical responses to episodes of S. aureus BSI in HD patients that we noted clinically.

Elimination of biofilm and microbial contamination reservoirs in hospital washbasin U-bends by automated cleaning and disinfection with electrochemically activated solutions.

BACKGROUND: Washbasin U-bends are reservoirs of microbial contamination in healthcare environments. U-Bends are constantly full of water and harbour microbial biofilm. AIM: To develop an effective automated cleaning and disinfection system for U-bends using two solutions generated by electrochemical activation of brine including the disinfectant anolyte (predominantly hypochlorous acid) and catholyte (predominantly sodium hydroxide) with detergent properties. METHODS: Initially three washbasin U-bends were manually filled with catholyte followed by anolyte for 5min each once weekly for five weeks. A programmable system was then developed with one washbasin that automated this process. This U-bend had three cycles of 5min catholyte followed by 5min anolyte treatment per week for three months. Quantitative bacterial counts from treated and control U-bends were determined on blood agar (CBA), R2A, PAS, and PA agars following automated treatment and on CBA and R2A following manual treatment. FINDINGS: The average bacterial density from untreated U-bends throughout the study was >1×10(5) cfu/swab on all media with Pseudomonas aeruginosa accounting for ∼50% of counts. Manual U-bend electrochemically activated (ECA) solution treatment reduced counts significantly (<100cfu/swab) (P<0.01 for CBA; P<0.005 for R2A). Similarly, counts from the automated ECA-treatment U-bend were significantly reduced with average counts for 35 cycles on CBA, R2A, PAS, and PA of 2.1±4.5 (P<0.0001), 13.1±30.1 (P<0.05), 0.7±2.8 (P<0.001), and 0 (P<0.05) cfu/swab, respectively. P. aeruginosa was eliminated from all treated U-bends. CONCLUSION: Automated ECA treatment of washbasin U-bends consistently minimizes microbial contamination.

Evaluation of commercial chromogenic media for the detection of meticillin-resistant Staphylococcus aureus.

BACKGROUND: Selective chromogenic media allowing one-step meticillin-resistant Staphylococcus aureus (MRSA) isolation and identification are widely used. However, the changing epidemiology of MRSA means that the suitability of these chromogenic media requires investigation. AIM: To evaluate the following chromogenic media - Colorex MRSA, MRSA Select II, ChromID MRSA, and MRSA Brilliance 2 - for the detection of divergent strain types. METHODS: We used a diverse collection of S. aureus, including strains harbouring the mecC gene, strains expressing varying levels of meticillin resistance, and isolates recovered from patient samples. FINDINGS: MRSA Select II, Colorex MRSA, and ChromID each grew at a density of 1.5 × 10(1)cfu/mL for each SCCmec type investigated. Brilliance 2 demonstrated growth at 1.5 × 10(1)cfu/mL for mecC MRSA but at a higher density (1.5 × 10(4)cfu/mL) for the three mecA MRSA strains. All four media demonstrated excellent sensitivity for MRSA detection (≥99%), but reduced levels of specificity (85-73%) when challenged with a range of meticillin-susceptible S. aureus (MSSA) isolates. High levels of false positives (∼50%) were also obtained with all chromogenic media when tested with mec-negative borderline oxacillin-resistant S. aureus (BORSA) isolates. CONCLUSION: Although false positives may be obtained with some strains of MSSA and BORSA, the high sensitivity of these media and their ability to recover almost all MRSA tested (including oxacillin-susceptible and mecC-positive strains) confirm the value of chromogenic agar in MRSA detection.

Overcoming the problem of residual microbial contamination in dental suction units left by conventional disinfection using novel single component suction handpieces in combination with automated flood disinfection.

OBJECTIVES: Decontaminating dental chair unit (DCU) suction systems in a convenient, safe and effective manner is problematic. This study aimed to identify and quantify the extent of the problems using 25 DCUs, methodically eliminate these problems and develop an efficient approach for reliable, effective, automated disinfection. METHODS: DCU suction system residual contamination by environmental and human-derived bacteria was evaluated by microbiological culture following standard aspiration disinfection with a quaternary ammonium disinfectant or alternatively, a novel flooding approach to disinfection. Disinfection of multicomponent suction handpieces, assembled and disassembled, was also studied. A prototype manual and a novel automated Suction Tube Cleaning System (STCS) were developed and tested, as were novel single component suction handpieces. RESULTS: Standard aspiration disinfection consistently failed to decontaminate DCU suction systems effectively. Semi-confluent bacterial growth (101-500 colony forming units (CFU) per culture plate) was recovered from up to 60% of suction filter housings and from up to 19% of high and 37% of low volume suction hoses. Manual and automated flood disinfection of DCU suction systems reduced this dramatically (ranges for filter cage and high and low volume hoses of 0-22, 0-16 and 0-14CFU/plate, respectively) (P<0.0001). Multicomponent suction handpieces could not be adequately disinfected without prior removal and disassembly. Novel single component handpieces, allowed their effective disinfection in situ using the STCS, which virtually eliminated contamination from the entire suction system. CONCLUSION: Flood disinfection of DCU suction systems and single component handpieces radically improves disinfection efficacy and considerably reduces potential cross-infection and cross-contamination risks. CLINICAL SIGNIFICANCE: DCU suction systems become heavily contaminated during use. Conventional disinfection does not adequately control this. Furthermore, multicomponent suction handpieces cannot be adequately disinfected without disassembly, which is costly in time, staff and resources. The automated STCS DCU suction disinfection system used with single component handpieces provides an effective solution.

A longitudinal study of Staphylococcus aureus colonization in pigs in Ireland.

The emergence of methicillin-resistant Staphylococcus aureus (MRSA) in livestock has refocused attention on S. aureus colonization and transmission in pigs. This study investigated the effect of the S. aureus colonization status of a sow on the colonization status of her piglets, and whether pigs carry the same strain of S. aureus throughout production. Nasal swabs were collected from the piglets of six healthy sows two days after birth and two days before and two days after they were moved into each production stage. The average prevalence of S. aureus colonization varied between 26% and 73%. The odds of being S. aureus positive were almost 12 times higher for piglets born to nasal-positive sows than for those born to nasal-negative sows, and three times higher again for piglets born to sows that were both nasal- and vaginal-positive. Isolates recovered from piglets immediately after birth were indistinguishable from those of the dam as determined by phenotypic and molecular typing, including microarray analysis and optical mapping. All isolates belonged to clonal complex 9 and the majority exhibited a novel spa type, t10449. The findings show that the S. aureus colonization status of the sow influences the colonization status of her piglets in the early production stages but strains carried by pigs change over time. Multiresistant S. aureus was detected, in particular post-weaning. Results suggest that sow status and management practices, including mixing of pigs and antimicrobial usage at weaning, should be considered when implementing control measures for S. aureus on a farm.

Air and surface contamination patterns of meticillin-resistant Staphylococcus aureus on eight acute hospital wards.

BACKGROUND: Meticillin-resistant Staphylococcus aureus (MRSA) can be recovered from hospital air and from environmental surfaces. This poses a potential risk of transmission to patients. AIM: To investigate associations between MRSA isolates recovered from air and environmental surfaces with those from patients when undertaking extensive patient and environmental sampling. METHODS: This was a prospective observational study of patients and their environment in eight wards of a 700-bed tertiary care hospital during 2010 and 2011. Sampling of patients, air and surfaces was carried out on all ward bays, with more extended environmental sampling in ward high-dependency bays and at particular times of the day. The genetic relatedness of isolates was determined by DNA microarray profiling and spa typing. FINDINGS: MRSA was recovered from 30/706 (4.3%) patients and from 19/132 (14.4%) air samples. On 9/132 (6.8%) occasions both patient and air samples yielded MRSA. In 32 high-dependency bays, MRSA was recovered from 12/161 (7.4%) patients, 8/32 (25%) air samples, and 21/644 (3.3%) environmental surface samples. On 10/132 (7.6%) occasions, MRSA was isolated from air in the absence of MRSA-positive patients. Patient demographic data combined with spa typing and DNA microarray profiling revealed four likely transmission clusters, where patient and environmental isolates were deemed to be very closely related. CONCLUSION: Air sampling yielded MRSA on frequent occasions, especially in high-dependency bays. Environmental and air sampling combined with patient demographic data, spa typing and DNA microarray profiling indicated the presence of clusters that were not otherwise apparent.