Inter-species gene flow drives ongoing evolution of Streptococcus pyogenes and Streptococcus dysgalactiae subsp. equisimilis.

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

Development and pre-clinical evaluation of a Zika virus diagnostic for low resource settings.

Introduction: Zika virus (ZIKV) is a re-emerging flavivirus that poses a significant public health threat. ZIKV exhibits a wide array of non-vector borne human transmission routes, such as sexual transmission, transplacental transmission and blood transfusion. Detection and surveillance of ZIKV is considered paramount in prevention of major outbreaks. With the majority of cases reported in low-resource locations, simple, low-cost detection methods are considered highly desirable. Materials and Methods: Here we have developed a sensitive and specific ZIKV diagnostic using reverse transcription recombinase-aided amplification (RT-RAA) coupled with lateral flow detection (LFD) targeting a highly conserved region of the ZIKV NS1 gene. Results: We show our rapid, isothermal-ZIKV-diagnostic (Iso-ZIKV-Dx) can detect 500 copies of synthetic ZIKV RNA/µL in under 30 min at a constant 39°C. Using simulated urine samples, we observed that Iso-ZIKV-Dx also detects as low as 34.28 RNA copies/reaction of ZIKV (MR766 strain). Specificity testing confirmed that our test does not detect any co-circulating flaviviruses (dengue, West Nile, Japanese encephalitis, Murray Valley encephalitis and yellow fever viruses) or chikungunya virus. Sample processing results show complete inactivation of ZIKV (MR766 strain) in 5 min at room temperature using our novel viral RNA sample preparation reagent. Furthermore, lateral flow strips testing demonstrates positive diagnoses in as little as 5 min in running buffer. Discussion: Contrary to conventional RT-qPCR, our Iso-ZIKV-Dx does not require expensive machinery, specialised laboratory settings or extensively trained personnel. Pre-clinical evaluation demonstrates that our test exhibits robust, in-field capabilities without compromising sensitivity or specificity. When compared to the gold-standard RT-qPCR, our Iso-ZIKV-Dx test offers an array of applications that extend beyond diagnostics alone, including potential for surveillance and monitoring of ZIKV vector competency.

Characterization of an experimental model to determine streptococcal M protein-induced autoimmune cardiac and neurobehavioral abnormalities.

Group A streptococcal (GAS) infection is associated with a spectrum of autoimmune diseases including acute rheumatic fever/rheumatic heart disease (ARF/RHD) and neurobehavioral abnormalities. Antibodies against GAS M proteins cross-react with host tissue proteins in the heart and brain leading to the symptomatology observed in ARF/RHD. As throat carriage of Streptococcus dysgalactiae subspecies equisimilis (SDSE) has been reported to be relatively high in some ARF/RHD endemic regions compared with GAS, and both SDSE and GAS express coiled-coil surface protein called M protein, we hypothesized that streptococci other than GAS can also associated with ARF/RHD and neurobehavioral abnormalities. Neurobehavioral assessments and electrocardiography were performed on Lewis rats before and after exposure to recombinant GAS and SDSE M proteins. Histological assessments were performed to confirm inflammatory changes in cardiac and neuronal tissues. ELISA and Western blot analysis were performed to determine the cross-reactivity of antibodies with host connective, cardiac and neuronal tissue proteins. Lewis rats injected with M proteins either from GAS or SDSE developed significant cardiac functional and neurobehavioral abnormalities in comparison to control rats injected with phosphate-buffered saline. Antibodies against GAS and SDSE M proteins cross-reacted with cardiac, connective and neuronal proteins. Serum from rats injected with streptococcal antigens showed higher immunoglobulin G binding to the striatum and cortex of the brain. Cardiac and neurobehavioral abnormalities observed in our experimental model were comparable to the cardinal symptoms observed in patients with ARF/RHD. Here for the first time, we demonstrate in an experimental model that M proteins from different streptococcal species could initiate and drive the autoimmune-mediated cardiac tissue damage and neurobehavioral abnormalities.

Group A streptococcal antigen exposed rat model to investigate neurobehavioral and cardiac complications associated with post-streptococcal autoimmune sequelae.

Background: The neuropsychiatric disorders due to post-streptococcal autoimmune complications such as Sydenham's chorea (SC) are associated with acute rheumatic fever and rheumatic heart disease (ARF/RHD). An animal model that exhibits characteristics of both cardiac and neurobehavioral defects in ARF/RHD would be an important adjunct for future studies. Since age, gender, strain differences, and genotypes impact on the development of autoimmunity, we investigated the behavior of male and female Wistar and Lewis rat strains in two age cohorts (<6 weeks and >12 weeks) under normal husbandry conditions and following exposure to group A streptococcus (GAS). Methods: Standard behavioral assessments were performed to determine the impairments in fine motor control (food manipulation test), gait and balance (beam walking test), and obsessive-compulsive behavior (grooming and marble burying tests). Furthermore, electrocardiography, histology, and behavioral assessments were performed on male and female Lewis rats injected with GAS antigens. Results: For control Lewis rats there were no significant age and gender dependent differences in marble burying, food manipulation, beam walking and grooming behaviors. In contrast significant age-dependent differences were observed in Wistar rats in all the behavioral tests except for food manipulation. Therefore, Lewis rats were selected for further experiments to determine the effect of GAS. After exposure to GAS, Lewis rats demonstrated neurobehavioral abnormalities and cardiac pathology akin to SC and ARF/RHD, respectively. Conclusion: We have characterised a new model that provides longitudinal stability of age-dependent behavior, to simultaneously investigate both neurobehavioral and cardiac abnormalities associated with post-streptococcal complications.

Requirements for a Robust Animal Model to Investigate the Disease Mechanism of Autoimmune Complications Associated With ARF/RHD.

The pathogenesis of Acute Rheumatic Fever/Rheumatic Heart Disease (ARF/RHD) and associated neurobehavioral complications including Sydenham's chorea (SC) is complex. Disease complications triggered by Group A streptococcal (GAS) infection are confined to human and determining the early events leading to pathology requires a robust animal model that reflects the hallmark features of the disease. However, modeling these conditions in a laboratory animal, of a uniquely human disease is challenging. Animal models including cattle, sheep, pig, dog, cat, guinea pigs rats and mice have been used extensively to dissect molecular mechanisms of the autoimmune inflammatory responses in ARF/RHD. Despite the characteristic limitations of some animal models, several rodent models have significantly contributed to better understanding of the fundamental mechanisms underpinning features of ARF/RHD. In the Lewis rat autoimmune valvulitis model the development of myocarditis and valvulitis with the infiltration of mononuclear cells along with generation of antibodies that cross-react with cardiac tissue proteins following exposure to GAS antigens were found to be similar to ARF/RHD. We have recently shown that Lewis rats injected with recombinant GAS antigens simultaneously developed cardiac and neurobehavioral changes. Since ARF/RHD is multifactorial in origin, an animal model which exhibit the characteristics of several of the cardinal diagnostic criteria observed in ARF/RHD, would be advantageous to determine the early immune responses to facilitate biomarker discovery as well as provide a suitable model to evaluate treatment options, safety and efficacy of vaccine candidates. This review focuses on some of the common small animals and their advantages and limitations.

In Search of the Holy Grail: A Specific Diagnostic Test for Rheumatic Fever.

Current diagnosis of Acute Rheumatic Fever and Rheumatic Heart Disease (ARF/RHD) relies on a battery of clinical observations aided by technologically advanced diagnostic tools and non-specific laboratory tests. The laboratory-based assays fall into two categories: those that (1) detect "evidence of preceding streptococcal infections" (ASOT, anti-DNAse B, isolation of the Group A Streptococcus from a throat swab) and (2) those that detect an ongoing inflammatory process (ESR and CRP). These laboratory tests are positive during any streptococcal infection and are non-specific for the diagnosis of ARF/RHD. Over the last few decades, we have accumulated considerable knowledge about streptococcal biology and the immunopathological mechanisms that contribute to the development, progression and exacerbation of ARF/RHD. Although our knowledge is incomplete and many more years will be devoted to understanding the exact molecular and cellular mechanisms involved in the spectrum of clinical manifestations of ARF/RHD, in this commentary we contend that there is sufficient understanding of the disease process that using currently available technologies it is possible to identify pathogen associated peptides and develop a specific test for ARF/RHD. It is our view that with collaboration and sharing of well-characterised serial blood samples from patients with ARF/RHD from different regions, antibody array technology and/or T-cell tetramers could be used to identify streptococcal peptides specific to ARF/RHD. The availability of an appropriate animal model for this uniquely human disease can further facilitate the determination as to whether these peptides are pathognomonic. Identification of such peptides will also facilitate testing of potential anti-streptococcal vaccines for safety and avoid potential candidates that may pre-dispose potential vaccine recipients to adverse outcomes. Such peptides can also be readily incorporated into a universally affordable point of care device for both primary and tertiary care.

In silico characterisation of stand-alone response regulators of Streptococcus pyogenes.

Bacterial "stand-alone" response regulators (RRs) are pivotal to the control of gene transcription in response to changing cytosolic and extracellular microenvironments during infection. The genome of group A Streptococcus (GAS) encodes more than 30 stand-alone RRs that orchestrate the expression of virulence factors involved in infecting multiple tissues, so causing an array of potentially lethal human diseases. Here, we analysed the molecular epidemiology and biological associations in the coding sequences (CDSs) and upstream intergenic regions (IGRs) of 35 stand-alone RRs from a collection of global GAS genomes. Of the 944 genomes analysed, 97% encoded 32 or more of the 35 tested RRs. The length of RR CDSs ranged from 297 to 1587 nucleotides with an average nucleotide diversity (π) of 0.012, while the IGRs ranged from 51 to 666 nucleotides with average π of 0.017. We present new evidence of recombination in multiple RRs including mga, leading to mga-2 switching, emm-switching and emm-like gene chimerization, and the first instance of an isolate that encodes both mga-1 and mga-2. Recombination was also evident in rofA/nra and msmR loci with 15 emm-types represented in multiple FCT (fibronectin-binding, collagen-binding, T-antigen)-types, including novel emm-type/FCT-type pairings. Strong associations were observed between concatenated RR allele types, and emm-type, MLST-type, core genome phylogroup, and country of sampling. No strong associations were observed between individual loci and disease outcome. We propose that 11 RRs may form part of future refinement of GAS typing systems that reflect core genome evolutionary associations. This subgenomic analysis revealed allelic traits that were informative to the biological function, GAS strain definition, and regional outbreak detection.

Potent Antibacterial Prenylated Acetophenones from the Australian Endemic Plant Acronychia crassipetala.

Acronychia crassipetala is an endemic plant species in Australia. Its phytochemistry and therapeutic properties are underexplored. The hexane extract of the fruit A. crassipetala T. G. Hartley was found to inhibit the growth of the Gram-positive bacteria Staphylococcus aureus. Following bio-activity guided fractionation, two prenylated acetophenones, crassipetalonol A (1) and crassipetalone A (2), were isolated. Their structures were determined mainly by NMR and MS spectroscopic analyses. This is the first record of the isolation and structural characterisation of secondary metabolites from the species A. crassipetala. Their antibacterial and cytotoxic assessments indicated that the known compound (2) had more potent antibacterial activity than the antibiotic chloramphenicol, while the new compound (1) showed moderate cytotoxicity.

Antibacterial 5α-Spirostane Saponins from the Fruit of Cordyline manners-suttoniae.

Antibacterial-activity-guided fractionation of a dichloromethane extract from the fruit of Cordyline manners-suttoniae and subsequent structure-activity investigations resulted in the identification of 10 new (1-10) and one known (11) 5α-spirostane saponin. The structures of the new compounds were established by 1D and 2D NMR analyses. The absolute configurations of the isolated compounds were determined by X-ray diffraction analysis or chemical derivatizations. The most active compound, suttonigenin F (6), inhibited the Gram-positive bacteria Staphylococcus aureus with MIC75 values that were comparable to those of the antibiotic chloramphenicol. Structure-activity relationships were also obtained from the assessment of antibacterial and cytotoxic activities of the isolated saponins.

Skin colonization at peripheral intravenous catheter insertion sites increases the risk of catheter colonization and infection.

BACKGROUND: Peripheral intravenous catheters (PIVCs) break the skin barrier, and preinsertion antiseptic disinfection and sterile dressings are used to reduce risk of catheter-related bloodstream infection (CRBSI). In this study, the impact of PIVC skin site colonization on tip colonization and the development of CRBSI was investigated. METHODS: A total of 137 patients' PIVC skin site swabs and paired PIVC tips were collected at catheter removal, cultured, and bacterial species and clonality were identified. RESULTS: Of 137 patients, 45 (33%) had colonized skin sites and/or PIVC tips. Of 16 patients with paired colonization of both the skin site and PIVC tips, 11 (69%) were colonized with the same bacterial species. Of these, 77% were clonally related, including 1 identical clone of Pseudomonas aeruginosa in a patient with systemic infection and the same organism identified in blood culture. CONCLUSIONS: The results demonstrate that opportunistic pathogen colonization at the skin site poses a significant risk for PIVC colonization and CRBSI. Further research is needed to improve current preinsertion antiseptic disinfection of PIVC skin site and the sterile insertion procedure to potentially reduce PIVC colonization and infection risk.

Dressings and securements for the prevention of peripheral intravenous catheter failure in adults (SAVE): a pragmatic, randomised controlled, superiority trial.

BACKGROUND: Two billion peripheral intravenous catheters (PIVCs) are used globally each year, but optimal dressing and securement methods are not well established. We aimed to compare the efficacy and costs of three alternative approaches to standard non-bordered polyurethane dressings. METHODS: We did a pragmatic, randomised controlled, parallel-group superiority trial at two hospitals in Queensland, Australia. Eligible patients were aged 18 years or older and required PIVC insertion for clinical treatment, which was expected to be required for longer than 24 h. Patients were randomly assigned (1:1:1:1) via a centralised web-based randomisation service using random block sizes, stratified by hospital, to receive tissue adhesive with polyurethane dressing, bordered polyurethane dressing, a securement device with polyurethane dressing, or polyurethane dressing (control). Randomisation was concealed before allocation. Patients, clinicians, and research staff were not masked because of the nature of the intervention, but infections were adjudicated by a physician who was masked to treatment allocation. The primary outcome was all-cause PIVC failure (as a composite of complete dislodgement, occlusion, phlebitis, and infection [primary bloodstream infection or local infection]). Analysis was by modified intention to treat. This trial is registered with the Australian New Zealand Clinical Trials Registry, number ACTRN12611000769987. FINDINGS: Between March 18, 2013, and Sept 9, 2014, we randomly assigned 1807 patients to receive tissue adhesive with polyurethane (n=446), bordered polyurethane (n=454), securement device with polyurethane (n=453), or polyurethane (n=454); 1697 patients comprised the modified intention-to-treat population. 163 (38%) of 427 patients in the tissue adhesive with polyurethane group (absolute risk difference -4·5% [95% CI -11·1 to 2·1%], p=0·19), 169 (40%) of 423 of patients in the bordered polyurethane group (-2·7% [-9·3 to 3·9%] p=0·44), 176 (41%) of 425 patients in the securement device with poplyurethane group (-1·2% [-7·9% to 5·4%], p=0·73), and 180 (43%) of 422 patients in the polyurethane group had PIVC failure. 17 patients in the tissue adhesive with polyurethane group, two patients in the bordered polyurethane group, eight patients in the securement device with polyurethane group, and seven patients in the polyurethane group had skin adverse events. Total costs of the trial interventions did not differ significantly between groups. INTERPRETATION: Current dressing and securement methods are commonly associated with PIVC failure and poor durability, with simultaneous use of multiple products commonly required. Cost is currently the main factor that determines product choice. Innovations to achieve effective, durable dressings and securements, and randomised controlled trials assessing their effectiveness are urgently needed. FUNDING: Australian National Health and Medical Research Council.

In silico characterisation of the two-component system regulators of Streptococcus pyogenes.

Bacteria respond to environmental changes through the co-ordinated regulation of gene expression, often mediated by two-component regulatory systems (TCS). Group A Streptococcus (GAS), a bacterium which infects multiple human body sites and causes multiple diseases, possesses up to 14 TCS. In this study we examined genetic variation in the coding sequences and non-coding DNA upstream of these TCS as a method for evaluating relationships between different GAS emm-types, and potential associations with GAS disease. Twelve of the 14 TCS were present in 90% of the genomes examined. The length of the intergenic regions (IGRs) upstream of TCS coding regions varied from 39 to 345 nucleotides, with an average nucleotide diversity of 0.0064. Overall, IGR allelic variation was generally conserved with an emm-type. Subsequent phylogenetic analysis of concatenated sequences based on all TCS IGR sequences grouped genomes of the same emm-type together. However grouping with emm-pattern and emm-cluster-types was much weaker, suggesting epidemiological and functional properties associated with the latter are not due to evolutionary relatedness of emm-types. All emm5, emm6 and most of the emm18 genomes, all historically considered rheumatogenic emm-types clustered together, suggesting a shared evolutionary history. However emm1, emm3 and several emm18 genomes did not cluster within this group. These latter emm18 isolates were epidemiologically distinct from other emm18 genomes in study, providing evidence for local variation. emm-types associated with invasive disease or nephritogenicity also did not cluster together. Considering the TCS coding sequences (cds), correlation with emm-type was weaker than for the IGRs, and no strong correlation with disease was observed. Deletion of the malate transporter, maeP, was identified that serves as a putative marker for the emm89.0 subtype, which has been implicated in invasive outbreaks. A recombination-related, subclade-forming DNA motif was identified in the putative receiver domain of the Spy1556 response regulator that correlated with throat-associated emm-pattern-type A-C strains.

Extracorporeal membrane oxygenation line-associated complications: in vitro testing of cyanoacrylate tissue adhesive and securement devices to prevent infection and dislodgement.

BACKGROUND: Extracorporeal membrane oxygenation (ECMO) delivers cardiac and/or respiratory support to critically ill patients who have failed conventional medical therapies. If the large-bore cannulas used to deliver ECMO become infected or dislodged, the patient consequences can be catastrophic. ECMO cannula-related infection has been reported to be double the rate of other vascular devices (7.1 vs 3.4 episodes/1000 ECMO days respectively). The aim of this study was to assess the ability of cyanoacrylate tissue adhesive (TA) to inhibit bacterial growth at the ECMO cannulation site, and the effectiveness of TA and securement devices in securing ECMO cannulas and tubing. METHODS: This in vitro study tested the (1) antimicrobial qualities of TA against standard transparent dressing with ECMO cannula; (2) chemical compatibility between cannula, TA and removal agent; (3) pull-out strength of transparent dressing and TA at the cannula insertion site; and (4) pull-out strength of adhesive bandage and commercial sutureless securement devices (SSDs) on circuit tubing. Fisher's exact test was used to evaluate differences in bacterial growth observed between the transparent dressing and TA groups. Data from mechanical testing were analysed using one-way ANOVA, followed by Tukey's multiple comparison test or t test as appropriate. Statistical significance was defined as p < 0.05. RESULTS: No bacterial growth occurred under TA-covered cannulas compared with transparent dressing-covered cannulas (p = 0.002). Compared to plates lacking TA or transparent dressing, growth was observed at the insertion point and under the dressing in the transparent dressing group; however, no growth was observed in the TA group (p = 0.019). TA did not weaken the cannulas; however, the TA removal agent did after 60 min of exposure, compared with control (p < 0.01). Compared with transparent dressing, TA increased the pull-out force required for cannula dislodgement from the insertion point (p < 0.0001). SSDs significantly increased the force required to remove the tubing from the fixation points compared with adhesive bandage (p < 0.01). CONCLUSIONS: Our findings suggest that the combined use of TA at the cannula insertion site with a commercial device for tubing securement could provide an effective bedside strategy to prevent or minimise infection and line dislodgement.

Does a Dedicated Lumen for Parenteral Nutrition Administration Reduce the Risk of Catheter-Related Bloodstream Infections? A Systematic Literature Review.

Guidelines recommend using single-lumen central vascular access devices (CVADs) for the administration of parenteral nutrition (PN) or lipid-based solutions, or a dedicated lumen on a multilumen CVAD. Publications reviewed by the authors reported comparative rates of catheter-related bloodstream infection (CR-BSI) in patients with CVADs who received PN through a dedicated lumen compared with those who had PN administered through multilumen CVADs. Two studies included 650 patients with 1349 CVADs. CR-BSIs were equally distributed between the 2 groups. Both studies were poorly reported and had significant risk of bias. These results should be interpreted with caution.

Group G Streptococcus Induces an Autoimmune Carditis Mediated by Interleukin 17A and Interferon γ in the Lewis Rat Model of Rheumatic Heart Disease.

Acute rheumatic fever and rheumatic heart disease (ARF/RHD) have long been described as autoimmune sequelae of Streptococcus pyogenes or group A streptococcal (GAS) infection. Both antibody and T-cell responses against immunodominant GAS virulence factors, including M protein, cross-react with host tissue proteins, triggering an inflammatory response leading to permanent heart damage. However, in some ARF/RHD-endemic regions, throat carriage of GAS is low. Because Streptococcus dysgalactiae subspecies equisimilis organisms, also known as ß-hemolytic group C streptococci and group G streptococci (GGS), also express M protein, we postulated that streptococci other than GAS may have the potential to initiate or exacerbate ARF/RHD. Using a model initially developed to investigate the uniquely human disease of ARF/RHD, we have discovered that GGS causes interleukin 17A/interferon γ-induced myocarditis and valvulitis, hallmarks of ARF/RHD. Remarkably the histological, immunological, and functional changes in the hearts of rats exposed to GGS are identical to those exposed to GAS. Furthermore, antibody cross-reactivity to cardiac myosin was comparable in both GGS- and GAS-exposed animals, providing additional evidence that GGS can induce and/or exacerbate ARF/RHD.

Peripherally InSerted CEntral catheter dressing and securement in patients with cancer: the PISCES trial. Protocol for a 2x2 factorial, superiority randomised controlled trial.

INTRODUCTION: Around 30% of peripherally inserted central catheters (PICCs) fail from vascular, infectious or mechanical complications. Patients with cancer are at highest risk, and this increases morbidity, mortality and costs. Effective PICC dressing and securement may prevent PICC failure; however, no large randomised controlled trial (RCT) has compared alternative approaches. We designed this RCT to assess the clinical and cost-effectiveness of dressing and securements to prevent PICC failure. METHODS AND ANALYSIS: Pragmatic, multicentre, 2×2 factorial, superiority RCT of (1) dressings (chlorhexidine gluconate disc (CHG) vs no disc) and (2) securements (integrated securement dressing (ISD) vs securement device (SED)). A qualitative evaluation using a knowledge translation framework is included. Recruitment of 1240 patients will occur over 3 years with allocation concealment until randomisation by a centralised service. For the dressing hypothesis, we hypothesise CHG discs will reduce catheter-associated bloodstream infection (CABSI) compared with no CHG disc. For the securement hypothesis, we hypothesise that ISD will reduce composite PICC failure (infection (CABSI/local infection), occlusion, dislodgement or thrombosis), compared with SED. SECONDARY OUTCOMES: types of PICC failure; safety; costs; dressing/securement failure; dwell time; microbial colonisation; reversible PICC complications and consumer acceptability. Relative incidence rates of CABSI and PICC failure/100 devices and/1000 PICC days (with 95% CIs) will summarise treatment impact. Kaplan-Meier survival curves (and log rank Mantel-Haenszel test) will compare outcomes over time. Secondary end points will be compared between groups using parametric/non-parametric techniques; p values <0.05 will be considered to be statistically significant. ETHICS AND DISSEMINATION: Ethical approval from Queensland Health (HREC/15/QRCH/241) and Griffith University (Ref. No. 2016/063). Results will be published. TRIAL REGISTRATION: Trial registration number is: ACTRN12616000315415.

Culture-independent detection of chlorhexidine resistance genes qacA/B and smr in bacterial DNA recovered from body sites treated with chlorhexidine-containing dressings.

PURPOSE: Dressings containing chlorhexidine gluconate (CHG) are increasingly used in clinical environments for prevention of infection at central venous catheter insertion sites. Increased tolerance to this biocide in staphylococci is primarily associated with the presence of qacA/B and smr genes. METHODOLOGY: We used a culture-independent method to assess the prevalence of these genes in 78 DNA specimens recovered from the skin of 43 patients at catheter insertion sites in the arm that were covered with CHG dressings. RESULTS: Of the 78 DNA specimens analysed, 52 (67 %) possessed qacA/B and 14 (18 %) possessed smr; all samples positive for smr were also positive for qacA/B. These prevalence rates were not statistically greater than those observed in a subsample of specimens taken from non-CHG treated contralateral arms and non-CHG-dressing exposed arms. A statistically greater proportion of specimens with greater than 72 h exposure to CHG dressings were qac-positive (P=0.04), suggesting that the patients were contaminated with bacteria or DNA containing qacA/B during their hospital stay. The presence of qac genes was not positively associated with the presence of DNA specific for Staphylococcusepidermidis and Staphylococcusaureus in these specimens. CONCLUSION: Our results show that CHG genes are highly prevalent on hospital patients' skin, even in the absence of viable bacteria.

Draft Genome Sequence of Roseomonas mucosa Strain AU37, Isolated from a Peripheral Intravenous Catheter.

Roseomonas mucosa is an opportunistic pathogen that causes infections in humans and is often associated with vascular catheter-related bacteremia. Here, we report the draft genome sequence of Roseomonas mucosa strain AU37, isolated from a peripheral intravenous catheter tip.

Predicted Coverage and Immuno-Safety of a Recombinant C-Repeat Region Based Streptococcus pyogenes Vaccine Candidate.

The C-terminal region of the M-protein of Streptococcus pyogenes is a major target for vaccine development. The major feature is the C-repeat region, consisting of 35-42 amino acid repeat units that display high but not perfect identity. SV1 is a S. pyogenes vaccine candidate that incorporates five 14mer amino acid sequences (called J14i variants) from differing C-repeat units in a single recombinant construct. Here we show that the J14i variants chosen for inclusion in SV1 are the most common variants in a dataset of 176 unique M-proteins. Murine antibodies raised against SV1 were shown to bind to each of the J14i variants present in SV1, as well as variants not present in the vaccine. Antibodies raised to the individual J14i variants were also shown to bind to multiple but different combinations of J14i variants, supporting the underlying rationale for the design of SV1. A Lewis Rat Model of valvulitis was then used to assess the capacity of SV1 to induce deleterious immune response associated with rheumatic heart disease. In this model, both SV1 and the M5 positive control protein were immunogenic. Neither of these antibodies were cross-reactive with cardiac myosin or collagen. Splenic T cells from SV1/CFA and SV1/alum immunized rats did not proliferate in response to cardiac myosin or collagen. Subsequent histological examination of heart tissue showed that 4 of 5 mice from the M5/CFA group had valvulitis and inflammatory cell infiltration into valvular tissue, whereas mice immunised with SV1/CFA, SV1/alum showed no sign of valvulitis. These results suggest that SV1 is a safe vaccine candidate that will elicit antibodies that recognise the vast majority of circulating GAS M-types.