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ObjectivesThe 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.MethodsThe 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.ResultsThe 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.ConclusionsThis 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.

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.

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., Author summary The bacterium Streptococcus pyogenes causes throat and skin infections. A danger from such infections is an immune response that attacks human heart tissue, leading to rheumatic heart disease, which is difficult to treat and potentially deadly. Disadvantaged populations such as the Indigenous people in remote tropical northern Australia have high burdens of S. pyogenes skin infection, and rheumatic heart disease. An effective vaccine would be a benefit, but none is approved for clinical use. We analysed data from 1810 S. pyogenes isolates from north Australia obtained over 28 years, to determine the potential of a previously described S. pyogenes vaccine candidate to be effective in this region. Only one quarter of the isolates from skin infections had a surface antigen corresponding to any one of the 30 antigen variants in the candidate vaccine. Previous work in animals indicates potential cross-protection from the vaccine against strains with mismatched antigens. However, even if this occurs in humans, protection against skin infection strains would likely remain compromised, unless there were additional components in the vaccine. Further studies on cross-protection are critical to defining the potential of this type of vaccine in populations burdened with S. pyogenes skin infections and rheumatic heart disease.

Group A Streptococcus (GAS; Streptococcus pyogenes) is a bacterial pathogen for which a commercial vaccine for humans is not available. Employing the advantages of high-throughput DNA sequencing technology to vaccine design, we have analyzed 2,083 globally sampled GAS genomes. The global GAS population structure reveals extensive genomic heterogeneity driven by homologous recombination and overlaid with high levels of accessory gene plasticity. We identified the existence of more than 290 clinically associated genomic phylogroups across 22 countries, highlighting challenges in designing vaccines of global utility. To determine vaccine candidate coverage, we investigated all of the previously described GAS candidate antigens for gene carriage and gene sequence heterogeneity. Only 15 of 28 vaccine antigen candidates were found to have both low naturally occurring sequence variation and high (>99%) coverage across this diverse GAS population. This technological platform for vaccine coverage determination is equally applicable to prospective GAS vaccine antigens identified in future studies., info:eu-repo/semantics/published

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.

Chlamydia trachomatis causes sexually transmitted infections and the blinding disease trachoma. Current data on C. trachomatis phylogeny show that there is only a single trachoma-causing clade, which is distinct from the lineages causing urogenital tract (UGT) and lymphogranuloma venerum diseases. Here we report the whole-genome sequences of ocular C. trachomatis isolates obtained from young children with clinical signs of trachoma in a trachoma endemic region of northern Australia. The isolates form two lineages that fall outside the classical trachoma lineage, instead being placed within UGT clades of the C. trachomatis phylogenetic tree. The Australian trachoma isolates appear to be recombinants with UGT C. trachomatis genome backbones, in which loci that encode immunodominant surface proteins (ompA and pmpEFGH) have been replaced by those characteristic of classical ocular isolates. This suggests that ocular tropism and association with trachoma are functionally associated with some sequence variants of ompA and pmpEFGH., Chlamydia trachomatis isolates causing a blinding disease (trachoma) form a single lineage that is different from the lineages causing urogenital infections. Here, Andersson et al. show however that trachoma isolates from Australia are more closely related to urogenital strains than to other trachoma isolates.

Two meticillin-resistant Staphylococcus aureus (MRSA) clones, sequence type (ST) 22 and ST239, have successfully spread globally. Across Australia, ST22 has supplanted ST239 as the main healthcare-associated MRSA. To understand the reasons underlying this shift, the epidemiology and clinical features of infections due to ST22 and ST239 MRSA isolates from a tertiary hospital in Melbourne, Australia were compared.Over six months, consecutive MRSA isolates with clinical data were collected from specimens referred to Alfred Health Pathology (AHP). Isolates were genotyped by a multi-locus-sequence-typing-based high-resolution melting method.Three hundred and twenty-eight of 1079 (30%) S. aureus isolated by AHP were MRSA. Of these, 313 were genotyped; 78 (25%) were clonal complex (CC) 22 (representing ST22) and 142 (45%) were CC239 (representing ST239). Common clinical syndromes included skin or soft tissue, respiratory tract and osteo-articular infections. On multi-variate logistic regression, compared with CC239, CC22 was associated with older patients [adjusted odds ratio (aOR) 1.04 for each year increase, 95% confidence interval (CI) 1.02-1.07)], and patients from subacute hospitals (aOR 2.7, 95% CI 1.2-5.8) or long-term care facilities (LTCFs; aOR 5.5, 95% CI 2.0-14.5). Median time from patient admission to MRSA isolation was nine days for CC239 and one day for CC22 (P 0.01). MRSA strain epidemiology varied according to hospital unit.CC22 and CC239 MRSA have differing ecological niches. CC22 is associated with elderly patients in LTCFs, and CC239 is associated with nosocomial acquisition. Infection control strategies involving LTCFs and their residents will likely be required to achieve continued MRSA control.

SUMMARYImpetigo is common in remote Indigenous children of northern Australia, with the primary driver in this context beingStreptococcus pyogenes[or group AStreptococcus(GAS)]. To reduce the high burden of impetigo, the transmission dynamics of GAS must be more clearly elucidated. We performed whole genome sequencing on 31 GAS isolates collected in a single community from children in 11 households with ⩾2 GAS-infected children. We aimed to determine whether transmission was occurring principally within households or across the community. The 31 isolates were represented by nine multilocus sequence types and isolates within each sequence type differed from one another by only 0–3 single nucleotide polymorphisms. There was evidence of extensive transmission both within households and across the community. Our findings suggest that strategies to reduce the burden of impetigo in this setting will need to extend beyond individual households, and incorporate multi-faceted, community-wide approaches.

Chlamydia trachomatis is the world's most prevalent bacterial sexually transmitted infection and leading infectious cause of blindness, yet it is one of the least understood human pathogens, in part due to the difficulties of in vitro culturing and the lack of available tools for genetic manipulation. Genome sequencing has reinvigorated this field, shedding light on the contemporary history of this pathogen. Here, we analyze 563 full genomes, 455 of which are novel, to show that the history of the species comprises two phases, and conclude that the currently circulating lineages are the result of evolution in different genomic ecotypes. Temporal analysis indicates these lineages have recently expanded in the space of thousands of years, rather than the millions of years as previously thought, a finding that dramatically changes our understanding of this pathogen's history. Finally, at a time when almost every pathogen is becoming increasingly resistant to antimicrobials, we show that there is no evidence of circulating genomic resistance in C. trachomatis., Funding agencies:Sanger Institute through the Wellcome Trust 098051 Australian National Health and Medical Research Council 1060768

We report two cases of severe pneumonia due to clone ST93 methicillin-resistant Staphylococcus aureus (MRSA) presenting from a remote Australian Indigenous community within a 2-week period, and the utilization of whole genome sequences to determine whether these were part of an outbreak. S. aureus was isolated from 12 of 92 nasal swabs collected from 25 community households (including the two index households); one isolate was ST93. Three of five skin lesion S. aureus isolates obtained at the community were ST93. Whole genome sequencing of the ST93 isolates from this study and a further 20 ST93 isolates from the same region suggested that recent transmission and progression to disease had not taken place. The proximity in time and space of the two severe pneumonia cases is probably a reflection of the high burden of disease due to ST93 MRSA in this population where skin infections and household crowding are common.

Chlamydia trachomatis is the world's most prevalent bacterial sexually transmitted infection and leading infectious cause of blindness, yet it is one of the least understood human pathogens, in part due to the difficulties of in vitro culturing and the lack of available tools for genetic manipulation. Genome sequencing has reinvigorated this field, shedding light on the contemporary history of this pathogen. Here, we analyze 563 full genomes, 455 of which are novel, to show that the history of the species comprises two phases, and conclude that the currently circulating lineages are the result of evolution in different genomic ecotypes. Temporal analysis indicates these lineages have recently expanded in the space of thousands of years, rather than the millions of years as previously thought, a finding that dramatically changes our understanding of this pathogen's history. Finally, at a time when almost every pathogen is becoming increasingly resistant to antimicrobials, we show that there is no evidence of circulating genomic resistance in C. trachomatis.

Objectives The objective was to determine the frequency of trachoma genotypes of Chlamydia trachomatis- positive urogenital tract (UGT) specimens from remote areas of the Australian Northern Territory (NT). Setting The setting was analysis of remnants of C. trachomatis positive primarily UGT specimens obtained in the course of clinical practice. The specimens were obtained from two pathology service providers. Participants From 3356 C. trachomatis specimens collected during May 2012–April 2013, 439 were selected for genotyping, with a focus on specimens from postpubescent patients, in remote Aboriginal communities where ocular trachoma is potentially present. Primary and secondary outcome measures The primary outcome measure was the proportion of successfully genotyped UGT specimens that were trachoma genotypes. The secondary outcome measures were the distribution of genotypes, and the frequencies of different classes of specimens able to be genotyped. Results Zero of 217 successfully genotyped UGT specimens yielded trachoma genotypes (95% CI for frequency=0–0.017). For UGT specimens, the genotypes were E (41%), F (22%), D (21%) and K (7%), with J, H and G and mixed genotypes each at 1–4%. Four of the five genotyped eye swabs yielded trachoma genotype Ba, and the other genotype J. Two hundred twenty-two specimens (50.6%) were successfully genotyped. Urine specimens were less likely to be typable than vaginal swabs (p Conclusions Unlike in some other studies, in the remote NT, trachoma genotypes of C. trachomatis were not found circulating in UGT specimens from 2012 to 2013. Therefore, C. trachomatis genotypes in UGT specimens from young children can be informative as to whether the organism has been acquired through sexual contact. We suggest inclusion of C. trachomatis genotyping in guidelines examining the source of sexually transmitted infections in young children in areas where trachoma genotypes may continue to circulate, and continued surveillance of UGT C. trachomatis genotypes.

The notion that sexually transmitted infections (STIs) can be acquired through non-sexual means is a well-established meme in the public consciousness. The public toilet seat is the most commonly blamed vector. This is despite the lack of evidence for acquisition of STIs from toilet seats, and the ready availability of reputable advice to this effect.1 While the ‘toilet seat hypothesis’ is not currently regarded as supported by data, other modes of non-sexual acquisition of STIs, though rare, have been reported. Examples include pharyngeal gonorrhoea by sharing of confectionery,2 transmission of urogenital (UGT) gonorrhoea by sharing of a sex doll,3 and transmission of Trichomonas vaginalis via the fingers of a traditional healer in west Africa.4 This provides credibility to the notion that STI detection in UGT specimens from young children is not always a consequence of ‘conventional’ sexual contact or, for very young children, vertical transmission. Estreich and Forster5 reviewed relevant evidence and concluded that inferring the cause of an STI in a young child is a ‘complex subject’, with vertical transmission, child sexual abuse and accidental/fomite transmission all conceivable mechanisms. This begs the question that is critical to service providers: How strongly does the presence of an STI in a young child indicate sexual abuse? This dilemma remains a challenge.6 In individual cases, potential modes of non-sexual transmission may be invoked, and the plausibility of these can impact child protection and/or justice system responses. Incorrect interpretation has the potential for a child remaining in an unsafe environment, or conversely leading to a false conclusion that a crime has taken place, resulting in initiation of an investigation. The strong imperatives to avoid either of these errors can create instability in institutional responses, and inappropriate management …

Chlamydia trachomatis causes both urogenital tract (UGT) and ocular infections. One class of ocular infection is trachoma. Trachoma has a distinctive pathology, encompassing repeated rounds of conjunctivitis, which can lead to chronic inflammation, corneal scarring and blindness. It has long been that while strains that cause UGT infections are essentially all able to cause conjunctivitis, the strains that cause trachoma are rarely associated with UGT infection. Previous genetic studies have shown that the trachoma strains form a single evolutionary lineage within the C. trachomatis species. This presentation will encompass recent advances in the understanding of the evolutionary structure and dynamics of C. trachomatis . Much of the work described was directed towards understanding better how service providers should respond to instances in which UGT specimens from young children are found to be C. trachomatis positive. While this is regarded as indicative of sexual abuse, a conceivable mechanism by which this can arise is autoinoculation from an ocular C. trachomatis infection. This is particularly relevant to remote northern and central Australia, where trachoma still exists, and where the extent of child sexual abuse is contentious and politically loaded. Understanding the strains associated with trachoma in Australia could potentially underpin procedures for determining whether or not C. trachomatis in a paediatric UGT specimen is derived from an ocular infection. Genome sequences of historic trachoma associated C. trachoma isolates from Northern Australia will be presented. The results do not support the established model of a single trachoma lineage. While the isolates possessed alleles of the ompA gene previously associated with trachoma strains, these appeared to be recombined into multiple non-trachoma-associated lineages. Furthermore, there was evidence for acquisition of of pmpEFGH alleles similar to those found in trachoma lineages. It was concluded that ompA and pmpEFGH may be functionally associated with trachoma pathology. Disclosure of interest statement Much of the work presented was funded by the NHMRC project grants 1004123 and 1060768. Phil Giffard declares no other interests.