Overlapping Streptococcus pyogenes and Streptococcus dysgalactiae subspecies equisimilis household transmission and mobile genetic element exchange.

Streptococcus dysgalactiae subspecies equisimilis (SDSE) and Streptococcus pyogenes share skin and throat niches with extensive genomic homology and horizontal gene transfer (HGT) possibly underlying shared disease phenotypes. It is unknown if cross-species transmission interaction occurs. Here, we conduct a genomic analysis of a longitudinal household survey in remote Australian First Nations communities for patterns of cross-species transmission interaction and HGT. Collected from 4547 person-consultations, we analyse 294 SDSE and 315 S. pyogenes genomes. We find SDSE and S. pyogenes transmission intersects extensively among households and show that patterns of co-occurrence and transmission links are consistent with independent transmission without inter-species interference. We identify at least one of three near-identical cross-species mobile genetic elements (MGEs) carrying antimicrobial resistance or streptodornase virulence genes in 55 (19%) SDSE and 23 (7%) S. pyogenes isolates. These findings demonstrate co-circulation of both pathogens and HGT in communities with a high burden of streptococcal disease, supporting a need to integrate SDSE and S. pyogenes surveillance and control efforts.

An outbreak of acute rheumatic fever in a remote Aboriginal community.

OBJECTIVES: We describe the public health response to an outbreak of acute rheumatic fever (ARF) in a remote Aboriginal community. METHODS: In August 2021, the Northern Territory Rheumatic Heart Disease Control Program identified an outbreak of acute rheumatic fever in a remote Aboriginal community. A public health response was developed using a modified acute poststreptococcal glomerulonephritis protocol and the National Acute Rheumatic Fever Guideline for Public Health Units. RESULTS: 12 cases were diagnosed during the outbreak; six-times the average number of cases in the same period in the five years prior (n=1.8). Half (n=6) of the outbreak cases were classified as recurrent episodes with overdue secondary prophylaxis. Contact tracing and screening of 11 households identified 86 close contacts. CONCLUSIONS: This outbreak represented an increase in both first episodes and recurrences of acute rheumatic fever and highlights the critical need for strengthened delivery of acute rheumatic fever secondary prophylaxis, and for improvements to the social determinants of health in the region. IMPLICATIONS FOR PUBLIC HEALTH: Outbreaks of acute rheumatic fever are rare despite continuing high rates of acute rheumatic fever experienced by remote Aboriginal communities. Nevertheless, there can be improvements in the current national public health guidance relating to acute rheumatic fever cluster and outbreak management.

Evaluating the role of asymptomatic throat carriage of Streptococcus pyogenes in impetigo transmission in remote Aboriginal communities in Northern Territory, Australia: a retrospective genomic analysis.

BACKGROUND: Streptococcus pyogenes, or group A Streptococcus (GAS), infections contribute to a high burden of disease in Aboriginal Australians, causing skin infections and immune sequelae such as rheumatic heart disease. Controlling skin infections in these populations has proven difficult, with transmission dynamics being poorly understood. We aimed to identify the relative contributions of impetigo and asymptomatic throat carriage to GAS transmission. METHODS: In this genomic analysis, we retrospectively applied whole genome sequencing to GAS isolates that were collected as part of an impetigo surveillance longitudinal household survey conducted in three remote Aboriginal communities in the Northern Territory of Australia between Aug 6, 2003, and June 22, 2005. We included GAS isolates from all throats and impetigo lesions of people living in two of the previously studied communities. We classified isolates into genomic lineages based on pairwise shared core genomes of more than 99% with five or fewer single nucleotide polymorphisms. We used a household network analysis of epidemiologically and genomically linked lineages to quantify the transmission of GAS within and between households. FINDINGS: We included 320 GAS isolates in our analysis: 203 (63%) from asymptomatic throat swabs and 117 (37%) from impetigo lesions. Among 64 genomic lineages (encompassing 39 emm types) we identified 264 transmission links (involving 93% of isolates), for which the probable source was asymptomatic throat carriage in 166 (63%) and impetigo lesions in 98 (37%). Links originating from impetigo cases were more frequent between households than within households. Households were infected with GAS for a mean of 57 days (SD 39 days), and once cleared, reinfected 62 days (SD 40 days) later. Increased household size and community presence of GAS and scabies were associated with slower clearance of GAS. INTERPRETATION: In communities with high prevalence of endemic GAS-associated skin infection, asymptomatic throat carriage is a GAS reservoir. Public health interventions such as vaccination or community infection control programmes aimed at interrupting transmission of GAS might need to include consideration of asymptomatic throat carriage. FUNDING: Australian National Health and Medical Research Council.

minSNPs: an R package for the derivation of resolution-optimised SNP sets from microbial genomic data.

Here, we present the R package, minSNPs. This is a re-development of a previously described Java application named Minimum SNPs. MinSNPs assembles resolution-optimised sets of single nucleotide polymorphisms (SNPs) from sequence alignments such as genome-wide orthologous SNP matrices. MinSNPs can derive sets of SNPs optimised for discriminating any user-defined combination of sequences from all others. Alternatively, SNP sets may be optimised to determine all sequences from all other sequences, i.e., to maximise diversity. MinSNPs encompasses functions that facilitate rapid and flexible SNP mining, and clear and comprehensive presentation of the results. The minSNPs' running time scales in a linear fashion with input data volume and the numbers of SNPs and SNPs sets specified in the output. MinSNPs was tested using a previously reported orthologous SNP matrix of Staphylococcus aureus and an orthologous SNP matrix of 3,279 genomes with 164,335 SNPs assembled from four S. aureus short read genomic data sets. MinSNPs was shown to be effective for deriving discriminatory SNP sets for potential surveillance targets and in identifying SNP sets optimised to discriminate isolates from different clonal complexes. MinSNPs was also tested with a large Plasmodium vivax orthologous SNP matrix. A set of five SNPs was derived that reliably indicated the country of origin within three south-east Asian countries. In summary, we report the capacity to assemble comprehensive SNP matrices that effectively capture microbial genomic diversity, and to rapidly and flexibly mine these entities for optimised marker sets.

First Description of the Composition and the Functional Capabilities of the Skin Microbial Community Accompanying Severe Scabies Infestation in Humans.

Epidemiological studies link Sarcoptes scabiei infection and impetigo. Scabies mites can promote Streptococcus pyogenes (Group A Streptococcus) and Staphylococcus aureus infections by breaching the skin barrier and excreting molecules that inhibit host innate immune responses. However, little is known about the composition and the function of the scabies-associated microbiota. Here, high-throughput whole-metagenome sequencing was used to explore the scabies-associated microbiome. Scabies mites including their immediate microenvironments were isolated from two patients with severe scabies in Northern Australia. Two ~45-50 million paired-end reads Illumina libraries were generated of which ~2 (5.1%) and 0.7 million (1.3%) microbial reads were filtered out by mapping to human (hg19) and mite draft genomes. Taxonomic profiling revealed a microbial community dominated by the phylum Firmicutes (A: 79% and B: 59%) and genera that comprise Streptococcus, Staphylococcus, Acinetobacter, and Corynebacterium. Assembly of the metagenome reads resulted in genome bins representing reference genomes of Acinetobacter baumannii, Streptococcus dysgalactiae (Group C/G), Proteus mirablis and Staphylococcus aureus. The contigs contained genes relevant to pathogenicity and antibiotics resistance. Confocal microscopy of a patient skin sample confirmed A. baumannii, Streptococci and S. aureus in scabies mite gut and faeces and the surrounding skin. The study provides fundamental evidence for the association of opportunistic pathogens with scabies infection.

Longitudinal whole-genome based comparison of carriage and infection associated Staphylococcus aureus in northern Australian dialysis clinics.

BACKGROUND: The study objective was to reveal reservoirs potentially leading to Staphylococcus aureus infections in haemodialysis clinic clients in the tropical north of the Australian Northern Territory (NT). This client population are primarily Aboriginal Australians who have a greater burden of ill health than other Australians. Reservoir identification will enhance infection control in this client group, including informing potential S. aureus decolonisation strategies. METHODS AND FINDINGS: The study participants were 83 clients of four haemodialysis clinics in the Darwin region of the NT, and 46 clinical staff and researchers who had contact with the clinic clients. The study design was longitudinal, encompassing swabbing of anatomical sites at two month intervals to yield carriage isolates, and also progressive collection of infection isolates. Swab sampling was performed for all participants, and infection isolates collected for dialysis clients only. Analysis was based on the comparison of 139 carriage isolates and 27 infection isolates using whole genome sequencing. Genome comparisons were based on of 20,651 genome-wide orthologous SNPs, presence/absence of the mecA and pvl genes, and inferred multilocus sequence type and clonal complex. Pairs of genomes meeting the definition of "not discriminated" were classed as defining potential transmission events. The primary outcome was instances of potential transmission between a carriage site other than a skin lesion and an infection site, in the same individual. Three such instances were identified. Two involved ST762 (CC1) PVL- MRSA, and one instance ST121 PVL+ MSSA. Three additional instances were identified where the carriage strains were derived from skin lesions. Also identified were six instances of potential transmission of a carriage strains between participants, including transmission of strains between dialysis clients and staff/researchers, and one potential transmission of a clinical strain between participants. There were frequent occurrences of longitudinal persistence of carriage strains in individual participants, and two examples of the same strain causing infection in the same participants at different times. Strains associated with infections and skin lesions were enriched for PVL and mecA in comparison to strains associated with long term carriage. CONCLUSIONS: This study indicated that strains differ with respect to propensity to stably colonise sites such as the nose, and cause skin infections. PVL+ strains were associated with infection and skin lesions and were almost absent from the carriage sites. PVL- MRSA (mainly CC1) strains were associated with infection and also with potential transmission events involving carriage sites, while PVL- MSSA were frequently observed to stably colonise individuals without causing infection, and to be rarely transmitted. Current clinical guidelines for dialysis patients suggest MRSA decolonisation. Implementation in this client group may impact infections by PVL- MRSA, but may have little effect on infection by PVL+ strains. In this study, the PVL+ strains were predominant causes of infection but rarely colonised typical carriage sites such as the nose, and in the case of ST121, were MSSA. The important reservoirs for infection by PVL+ strains appeared to be prior infections.

Molecular diagnosis of scabies using a novel probe-based polymerase chain reaction assay targeting high-copy number repetitive sequences in the Sarcoptes scabiei genome.

BACKGROUND: The suboptimal sensitivity and specificity of available diagnostic methods for scabies hampers clinical management, trials of new therapies and epidemiologic studies. Additionally, parasitologic diagnosis by microscopic examination of skin scrapings requires sample collection with a sharp scalpel blade, causing discomfort to patients and difficulty in children. Polymerase chain reaction (PCR)-based diagnostic assays, combined with non-invasive sampling methods, represent an attractive approach. In this study, we aimed to develop a real-time probe-based PCR test for scabies, test a non-invasive sampling method and evaluate its diagnostic performance in two clinical settings. METHODOLOGY/PRINCIPAL FINDINGS: High copy-number repetitive DNA elements were identified in draft Sarcoptes scabiei genome sequences and used as assay targets for diagnostic PCR. Two suitable repetitive DNA sequences, a 375 base pair microsatellite (SSR5) and a 606 base pair long tandem repeat (SSR6), were identified. Diagnostic sensitivity and specificity were tested using relevant positive and negative control materials and compared to a published assay targeting the mitochondrial cox1 gene. Both assays were positive at a 1:100 dilution of DNA from a single mite; no amplification was observed in DNA from samples from 19 patients with other skin conditions nor from house dust, sheep or dog mites, head and body lice or from six common skin bacterial and fungal species. Moderate sensitivity of the assays was achieved in a pilot study, detecting 5/7 (71.4% [95% CI: 29.0% - 96.3%]) of clinically diagnosed untreated scabies patients). Greater sensitivity was observed in samples collected by FLOQ swabs compared to skin scrapings. CONCLUSIONS/SIGNIFICANCE: This newly developed qPCR assay, combined with the use of an alternative non-invasive swab sampling technique offers the possibility of enhanced diagnosis of scabies. Further studies will be required to better define the diagnostic performance of these tests.

Clinical and Molecular Epidemiology of an Emerging Panton-Valentine Leukocidin-Positive ST5 Methicillin-Resistant Staphylococcus aureus Clone in Northern Australia.

Recently, we identified a Staphylococcus aureus sequence type 5 (ST5) clone in northern Australia with discrepant trimethoprim-sulfamethoxazole (SXT) susceptibility results. We aimed to identify isolates of this clone using Vitek 2 SXT resistance as a proxy and to compare its epidemiology with those of other circulating S. aureus strains. We collated Vitek 2 susceptibility data for S. aureus isolates collected through our laboratory and conducted a prospective, case-control study comparing clinical, microbiological, epidemiological, and genomic data for subsets of isolates reported as SXT resistant (cases) and SXT susceptible (controls) by Vitek 2. While overall SXT resistance rates remained relatively stable from 2011 to 2018 among 27,721 S. aureus isolates, non-multidrug-resistant methicillin-resistant S. aureus (MRSA) strains almost completely replaced multidrug-resistant MRSA strains as the predominant SXT-resistant MRSA phenotype. Demographic and clinical features of 51 case-control pairs were similar, but genotyping revealed stark differences: clonal complex 5 (CC5) MRSA predominated among SXT-resistant cases (34/51 [67%]), while CC93 MRSA predominated among susceptible controls (26/51 [51%]). All CC5 isolates were an ST5 clonal lineage that possessed the trimethoprim resistance gene dfrG within SCCmec IVo; all were SXT susceptible by Etest. The replacement of Vitek 2 reported SXT-resistant multidrug-resistant MRSA by non-multidrug-resistant MRSA appears related to the emergence of an ST5-MRSA-SCCmec IVo clone that is SXT susceptible by Etest and causes clinical disease similar to that caused by ST93-MRSA-SCCmec IVa. Reliance on Vitek 2 SXT reporting may lead to unnecessary restriction of effective oral treatment options for S. aureus infections. Whether the presence of dfrG within SCCmec IVo provides a selective advantage at the population level is currently unclear.IMPORTANCEStaphylococcus aureus is an important human pathogen that causes a wide range of clinical infections. In the past 2 decades, an epidemic of community-associated skin and soft tissue infections has been driven by S. aureus strains with specific virulence factors and resistance to beta-lactam antibiotics. Recently, an S. aureus strain with discrepant antimicrobial susceptibility testing results has emerged in northern Australia. This ST5-MRSA-SCCmec IVo clone is reported as resistant to trimethoprim-sulfamethoxazole by Vitek 2 but susceptible by phenotypic methods. ST5-MRSA-SCCmec IVo is now the second most common community-associated MRSA clone in parts of Australia and causes a spectrum of clinical disease similar to that caused by the virulent ST93-MRSA lineage. Whole-genome sequence analysis demonstrates that ST5-MRSA-SCCmecIVo is causing a clonal outbreak across a large geographical region. Although phenotypic testing suggests in vitro susceptibility to trimethoprim-sulfamethoxazole, it is unclear at this stage whether the presence of dfrG within SCCmec IVo provides a selective advantage at the population level.

High-quality nuclear genome for Sarcoptes scabiei-A critical resource for a neglected parasite.

The parasitic mite Sarcoptes scabiei is an economically highly significant parasite of the skin of humans and animals worldwide. In humans, this mite causes a neglected tropical disease (NTD), called scabies. This disease results in major morbidity, disability, stigma and poverty globally and is often associated with secondary bacterial infections. Currently, anti-scabies treatments are not sufficiently effective, resistance to them is emerging and no vaccine is available. Here, we report the first high-quality genome and transcriptomic data for S. scabiei. The genome is 56.6 Mb in size, has a a repeat content of 10.6% and codes for 9,174 proteins. We explored key molecules involved in development, reproduction, host-parasite interactions, immunity and disease. The enhanced 'omic data sets for S. scabiei represent comprehensive and critical resources for genetic, functional genomic, metabolomic, phylogenetic, ecological and/or epidemiological investigations, and will underpin the design and development of new treatments, vaccines and/or diagnostic tests.

Identification and Discrimination of Chlamydia trachomatis Ocular and Urogenital Strains and Major Phylogenetic Lineages by CtGEM Typing, A Double-Locus Genotyping Method.

CtGEM typing was developed to subdivide the bacterial species Chlamydia trachomatis on the basis of genome phylogeny and anatomical tropism. The rationale was facilitation of surveillance for ocular strains, although the method is applicable to essentially any C. trachomatis surveillance application that does not require high resolution. CtGEM is a double-locus genotyping method. The loci included in the assay were identified by computerized analysis of 65 complete genomes for resolution optimized sets of single nucleotide polymorphisms (SNPs). From this, two PCR amplifiable fragments were defined. One, rg1, is within a hypothetical gene annotated as Jali-1891 within the C. trachomatis B_Jali20 genome. The other, ofr, is within the ompA gene which encodes the major outer membrane protein. Variation in rg1 is conferred by two SNPs defining four haplotypes that exhibit concordance with genome phylogeny. Variation within ofr is more complex and allows for inference of ompA genotype, either to the level of single genotype, or group of closely related genotypes. Two CtGEM formats were developed. One is based on interrogation of the two loci by high resolution melting analysis (HRMA), and the other based on analysis of the loci by Sanger sequencing. The genotypes defined identify known ocular genotypes, discriminate known ocular genotypes from each other, discriminate the major phylogenetic lineages of the species, and discriminate all ompA genotypes with the exception of closely related variants within the genotypes H, I, J cluster. The Sanger sequencing format provides slightly more resolution that the HRMA format with respect to ompA genotype. An unusual aspect of this method is that all possible combinations of rg1 haplotype, and inferred ompA genotype(s) have been given CtGEM typing numbers. This includes types that at this time have not been shown to exist.

Correction to: Nevus of Ota - an intraoral presentation: a case report.

In the publication of this article [1], the figures were accidentally omitted due to an error. This has now been updated in the original article within the Case presentation section.

Concerns for efficacy of a 30-valent M-protein-based Streptococcus pyogenes vaccine in regions with high rates of rheumatic heart disease.

The prevalence of rheumatic heart disease (RHD) in the Aboriginal population of the Australian Northern Territory is high, and Streptococcus pyogenes skin infections likely contribute to this. A promising candidate S. pyogenes "30mer" vaccine is composed of 30 pharyngitis associated type-specific antigens from the S. pyogenes M protein. Cross opsonisation experiments suggest that 30mer vaccine protection may extend to non-cognate emm types. A new "emm cluster" scheme for classifying M protein is based on the full-length coding sequence, and correlates with functional and immunological properties, and anatomical tropism. Twenty-seven years of research in the Northern Territory has yielded 1810 S. pyogenes isolates with clinical and emm type data. The primary aim was to analyse these data with reference to the emm cluster scheme and cross opsonisation information, to inform estimation of 30mer vaccine efficacy in the Northern Territory. The isolates encompass 101 emm types. Variants of cluster A-C were enriched in throat isolates, and variants of emm cluster D enriched in skin isolates. Throat isolates were enriched for 30mer vaccine cognate emm types in comparison with skin isolates of which only 25% were vaccine emm types. While cross opsonisation data indicates potential for enhancing 30mer vaccine coverage, more than one third of skin isolates were within 38 emm types untested for cross opsonisation. Emm cluster D variants, in particular emm cluster D4, were not only all non-cognate with the vaccine, but were abundant and diverse, and less likely to be cross-opsonisation positive than other emm clusters. Long term persistence of many emm types in the study area was revealed. It was concluded that the 30mer vaccine efficacy in the Northern Territory will likely require both cross protection, and additional measures to elicit immunity against variants of emm cluster D.

Nevus of Ota - an intraoral presentation: a case report.

BACKGROUND: Nevus of Ota or "oculodermal melanocytosis" is a rare congenital hamartoma of dermal melanocytes causing a blue-gray hyperpigmentation of the eye and surrounding structures. The condition, originally described by Ota and Tanino in 1939, mainly affects the ophthalmic and maxillary divisions of the trigeminal nerve. We describe the first reported case of unilateral oculodermal melanocytosis in a Caucasian woman with oral buccal mucosal involvement. Oral involvement of nevus of Ota is very rare. CASE PRESENTATION: A 48-year-old Caucasian woman was referred by the dermatology division to the oral medicine department at the University of Liverpool School of Dentistry with new-onset oral pigmentation to the left buccal mucosa. The patient had a previous diagnosis of oculodermal nevus. CONCLUSION: An incisional biopsy of the left buccal mucosa was completed. The report stated that histological and immunohistochemical features were in keeping with a blue nevus, but within the context of the preexisting occulodermal pigmentation, a diagnosis of oculodermal melanocytosis, also known as "nevus of Ota," was made. The patient will be kept under review in the oral medicine department because the progression of the lesion on the left buccal mucosa requires active monitoring owing to the potential for malignant change. The patient also requires regular review in the dermatology and ophthalmology divisions.

A cluster of acute rheumatic fever cases among Aboriginal Australians in a remote community with high baseline incidence.

OBJECTIVES: We report a cluster of acute rheumatic fever (ARF) cases and the public health response in a high-burden Australian setting. METHODS: The public health unit was notified of an increase in ARF cases in a remote Australian Aboriginal community. A multi-disciplinary group coordinated the response. Household contacts were screened for ARF or group A Streptococcus (GAS) infection by questionnaire and swab collection, offered an echocardiogram if aged 5-20 years, and intramuscular benzathine benzylpenicillin if aged over one year or if less than one year with impetigo. RESULTS: Fifteen definite and seven probable ARF cases were diagnosed in the community in July-December 2014 (all-age incidence of definite ARF: 1,473/100,000). The public health response identified two additional cases of ARF. A total of 81 contacts were screened; GAS was detected in 3/76 (4%) throat swabs and 11/24 (46%) skin swabs. Molecular typing revealed high GAS strain diversity. CONCLUSIONS: The incidence of ARF during this cluster was very high. Carriage and infection with GAS was observed, but no outbreak strain identified. Implications for public health: A national public health guideline has since been developed that includes advice on the investigation of an ARF outbreak/cluster. Sustained efforts with strong community engagement are required to tackle high ARF rates.

Potential for Molecular Testing for Group A Streptococcus to Improve Diagnosis and Management in a High-Risk Population: A Prospective Study.

BACKGROUND: In high-burden settings, guidelines recommend antibiotic treatment for all suspected group A Streptococcus (GAS) infections to prevent rheumatic fever and poststreptococcal glomerulonephritis. Highly sensitive rapid GAS tests could reduce unnecessary antibiotic use in these settings. METHODS: This was a prospective study of the Xpert Xpress Strep A (Cepheid) molecular test compared with culture of throat swab samples collected at a referral hospital in northern Australia. Demographic and clinical data and results of streptococcal serology and culture were collected. RESULTS: Of 164 throat swab samples, 145 (88%) were eligible for inclusion; 49 (34%) were molecular test positive and 24 (17%) were culture positive for GAS. The sensitivity, specificity, and positive and negative predictive values for the molecular test versus culture were 100.0%, 79.3%, 48.8%, and 100.0%, respectively. Among 25 samples testing positive with the molecular test and negative with culture, group C or G streptococci were cultured in 2, and a plausible clinical explanation, such as pharyngotonsillitis, or rheumatic fever with positive results of streptococcal serology, was apparent in 19 instances. In 25 patients with rheumatic fever or poststreptococcal glomerulonephritis diagnoses, molecular testing nearly trebled the detection of GAS in throat swab samples, from 3 (12%) detected with culture to 8 (32%) detected with molecular testing. Reasons for "false-positive" molecular test results could include the presence of GAS below the threshold of culture detection or persistence of nonviable organisms after infection. CONCLUSION: Implementation of molecular testing could improve antibiotic use in this high-burden setting. The incremental yield in poststreptococcal syndromes, by which time cultures are negative, has high potential in the diagnostic workup of autoimmune poststreptococcal syndromes and warrants further investigation.

Global Scale Dissemination of ST93: A Divergent Staphylococcus aureus Epidemic Lineage That Has Recently Emerged From Remote Northern Australia.

Background: In Australia, community-associated methicillin-resistant Staphylococcus aureus (MRSA) lineage sequence type (ST) 93 has rapidly risen to dominance since being described in the early 1990s. We examined 459 ST93 genome sequences from Australia, New Zealand, Samoa, and Europe to investigate the evolutionary history of ST93, its emergence in Australia and subsequent spread overseas. Results: Comparisons with other S. aureus genomes indicate that ST93 is an early diverging and recombinant lineage, comprising of segments from the ST59/ST121 lineage and from a divergent but currently unsampled Staphylococcal population. However, within extant ST93 strains limited genetic diversity was apparent with the most recent common ancestor dated to 1977 (95% highest posterior density 1973-1981). An epidemic ST93 population arose from a methicillin-susceptible progenitor in remote Northern Australia, which has a proportionally large Indigenous population, with documented overcrowded housing and a high burden of skin infection. Methicillin-resistance was acquired three times in these regions, with a clade harboring a staphylococcal cassette chromosome mec (SCCmec) IVa expanding and spreading to Australia's east coast by 2000. We observed sporadic and non-sustained introductions of ST93-MRSA-IVa to the United Kingdom. In contrast, in New Zealand, ST93-MRSA-IVa was sustainably transmitted with clonal expansion within the Pacific Islander population, who experience similar disadvantages as Australian Indigenous populations. Conclusion: ST93 has a highly recombinant genome including portions derived from an early diverging S. aureus population. Our findings highlight the need to understand host population factors in the emergence and spread of antimicrobial resistant community pathogens.

Phylogenetic relationships, stage-specific expression and localisation of a unique family of inactive cysteine proteases in Sarcoptes scabiei.

BACKGROUND: Scabies is worldwide one of the most common, yet neglected, parasitic skin infections, affecting a wide range of mammals including humans. Limited treatment options and evidence of emerging mite resistance against the currently used drugs drive our research to explore new therapeutic candidates. Previously, we discovered a multicopy family of genes encoding cysteine proteases with their catalytic sites inactivated by mutation (SMIPP-Cs). This protein family is unique in parasitic scabies mites and is absent in related non-burrowing mites. We postulated that the SMIPP-Cs have evolved as an adaptation to the parasitic lifestyle of the scabies mite. To formulate testable hypotheses for their functions and to propose possible strategies for translational research we investigated whether the SMIPP-Cs are common to all scabies mite varieties and where within the mite body as well as when throughout the parasitic life-cycle they are expressed. RESULTS: SMIPP-C sequences from human, pig and dog mites were analysed bioinformatically and the phylogenetic relationships between the SMIPP-C multi-copy gene families of human, pig and dog mites were established. Results suggest that amplification of the SMIPP-C genes occurred in a common ancestor and individual genes evolved independently in the different mite varieties. Recombinant human mite SMIPP-C proteins were produced and used for murine polyclonal antibody production. Immunohistology on skin sections from human patients localised the SMIPP-Cs in the mite gut and in mite faeces within in the epidermal skin burrows. SMIPP-C transcription into mRNA in different life stages was assessed in human and pig mites by reverse transcription followed by droplet digital PCR (ddPCR). High transcription levels of SMIPP-C genes were detected in the adult female life stage in comparison to all other life stages. CONCLUSIONS: The fact that the SMIPP-Cs are unique to three Sarcoptes varieties, present in all burrowing life stages and highly expressed in the digestive system of the infective adult female life stage may highlight an essential role in parasitism. As they are excreted from the gut in scybala they presumably are able to interact or interfere with host proteins present in the epidermis.

CtGEM typing: Discrimination of Chlamydia trachomatis ocular and urogenital strains and major evolutionary lineages by high resolution melting analysis of two amplified DNA fragments.

Chlamydia trachomatis infects the urogenital tract (UGT) and eyes. Anatomical tropism is correlated with variation in the major outer membrane protein encoded by ompA. Strains possessing the ocular ompA variants A, B, Ba and C are typically found within the phylogenetically coherent "classical ocular lineage". However, variants B, Ba and C have also been found within three distinct strains in Australia, all associated with ocular disease in children and outside the classical ocular lineage. CtGEM genotyping is a method for detecting and discriminating ocular strains and also the major phylogenetic lineages. The rationale was facilitation of surveillance to inform responses to C. trachomatis detection in UGT specimens from young children. CtGEM typing is based on high resolution melting analysis (HRMA) of two PCR amplified fragments with high combinatorial resolving power, as defined by computerised comparison of 65 whole genomes. One fragment is from the hypothetical gene defined by Jali-1891 in the C. trachomatis B_Jali20 genome, while the other is from ompA. Twenty combinatorial CtGEM types have been shown to exist, and these encompass unique genotypes for all known ocular strains, and also delineate the TI and T2 major phylogenetic lineages, identify LGV strains and provide additional resolution beyond this. CtGEM typing and Sanger sequencing were compared with 42 C. trachomatis positive clinical specimens, and there were no disjunctions. CtGEM typing is a highly efficient method designed and tested using large scale comparative genomics. It divides C. trachomatis into clinically and biologically meaningful groups, and may have broad application in surveillance.

Contaminated fingers: a potential cause of Chlamydia trachomatis-positive urine specimens.

OBJECTIVES: The detection of an STI agent in a urogenital tract (UGT) specimen from a young child is regarded as being indicative of sexual abuse. However, the probabilities of contamination events that could conceivably lead to STI positive specimens in the absence of sexual contact are unclear. The objective was to estimate the potential for fingers that have come in contact with Chlamydia trachomatis-positive urine to detectably contaminate C. trachomatis-negative urine. METHODS: The study design was based on self-experimentation. Dilutions of C. trachomatis elementary bodies (EBs) were prepared. A participant contacted an EB dilution then a urine surrogate specimen. The experiment was performed by three participants using three C. trachomatis isolates, of genotype E, F and B. Two surrogate urine contact methods were used to mimic contamination of a carer assisting with a child's urine collection. All EB dilutions and urine surrogate specimens were subjected to C. trachomatis assay and quantification in a real-time PCR-based diagnostic system. RESULTS: The amplimer crossing point (Cq) for EB dilutions was 10.0±1.6 less than for corresponding finger contacted urine specimens, which corresponds to ~10 µL of EB suspension transferred. This was largely independent of participant identity, C. trachomatis strain or EB dilution. Hand decontamination led to large reductions in EBs transferred, but transfer remained consistently detectable. Recent Cq data from C. trachomatis-positive clinical urine specimens were collated, and 20% clearly contained sufficient C. trachomatis to detectably contaminate another specimen by finger-mediated transfer, as in this experiment. CONCLUSIONS: This study directly demonstrated the potential for urine contaminated fingers to convert a C. trachomatis-negative urine specimen to C. trachomatis positive as a result of contact. Accordingly, procedures for urine specimen collection, particularly from children, need to be designed to prevent contamination.