Cervical cancer prevention in transgender men: a review.

There is increased awareness of transgender physical and mental health widely and in academic research. A significant proportion of transgender men will retain their cervix with an increased risk of cervical cancer. In this review of cervical cancer screening among transgender men, we try to estimate how many transgender men still have a cervix, understand to identify challenges and barriers to cervical screening and propose possible solutions. Organised cervical screening programmes need to consider the needs of this population, in particular the provision of HPV self-sampling. TWEETABLE ABSTRACT: Transgender men need access to cervical screening.

Sex is associated with the persistence of non-optimal vaginal microbiota following treatment for bacterial vaginosis: a prospective cohort study.

OBJECTIVE: Determine the associations between factors and sexual practices and the composition of the vaginal microbiome (VM) of women treated for bacterial vaginosis (BV). DESIGN: Prospective cohort study. SETTING: The Melbourne Sexual Health Centre, Melbourne, Australia. POPULATION: Seventy-five reproductive-age women diagnosed with clinical BV, treated with first-line antibiotics and followed for up to 6 months. METHODS: Women self-collected vaginal swabs and completed questionnaires at enrolment, the day following antibiotics and monthly for up to 6months until BV recurrence or no BV recurrence (n = 430 specimens). Bacterial composition was determined using 16S rRNA gene amplicon sequencing. The effects of ongoing factors on VM composition (utilising 291 monthly specimens) were assessed using generalised estimating equations population-averaged models, which accounted for repeated measures within individuals. MAIN OUTCOME MEASURES: The relative abundance of vaginal bacterial taxa. RESULTS: Women who reported ongoing sex with a regular sexual partner (RSP) had a VM comprised of increased relative abundance of non-optimal BV-associated bacteria (Adjusted co-efficient [Adjusted co-eff] = 11.91, 95% CI 3.39to20.43, P = 0.006) and a decreased relative abundance of optimal, Lactobacillus species (Adjusted co-eff = -12.76, 95% CI -23.03 to -2.49, P = 0.015). A history of BV was also associated with a decreased relative abundance of Lactobacillus spp. (Adjusted co-eff = -12.35, 95% CI -22.68, P = 0.019). The relative abundance of Gardnerella, Atopobium and Sneathia spp. increased following sex with an RSP. CONCLUSIONS: Sex with an untreated RSP after BV treatment was associated with a VM comprised of non-optimal BV-associated bacteria. BV treatment approaches may need to include partner treatment if they are to achieve a sustained optimal VM associated with improved health outcomes. TWEETABLE ABSTRACT: Sex drives a return to a 'non-optimal' vaginal microbiota after antibiotics for bacterial vaginosis.

Evaluation of the Roche MagNA Pure 96 nucleic acid extraction platform for the Seegene Anyplex II HPV28 detection assay.

AIM: Validate the Roche, MagNAPure96 (MP96) nucleic acid extraction platform for Seegene Anyplex II HPV28 (Anyplex28) detection of Human Papillomavirus. METHODS AND RESULTS: Comparisons were made for Anyplex28 genotyping from 115 cervical samples extracted on the Hamilton, STARlet and the MP96. Two DNA concentrations were used for the MP96, one matched for sample input to the STARlet and another 5× concentration (laboratory standard). Agreement of HPV detection was 89·8% (κ = 0·798; P = 0·007), with HPV detected in 10 more samples for the MP96. There was a high concordance of detection for any oncogenic HPV genotype (κ = 0·77; P = 0·007) and for any low-risk HPV genotype (κ = 0·85; P = 0·008). DNA extracted at laboratory standard had a lower overall agreement 85·2% (κ = 0·708; P < 0·001), with 17/115 discordant positive samples that tested negative after STARlet extraction. Of the discordant genotypes, 72·7% were detected in the lowest signal range for Anyplex28 ('+'). CONCLUSIONS: MP96 performed with high concordance to STARlet, although produced DNA with a higher analytical sensitivity on the Anyplex28. SIGNIFICANCE AND IMPACT OF THE STUDY: This analysis supports the use of samples extracted on the MP96 for HPV genotyping using the Anyplex28. Furthermore, an increase in DNA concentration increased analytical sensitivity of the Anyplex28, particularly appropriate for prevalence studies.

The impact of sample storage on molecular-based detection of Mycoplasma genitalium.

AIMS: Mycoplasma genitalium causes a common, sexually transmitted bacterial infection. This study assessed the detection of M. genitalium in stored urine samples to understand the impact of sample storage on M. genitalium detection. METHODS: Aliquots of M. genitalium-positive urine (n = 20 patients) were stored at either room temperature (22°C) or 4°C, without a preservative. At weekly intervals, samples were tested using the commercial test ResistancePlus MG® (SpeeDx® , Australia). We report the analysis at 1 week, an acceptable collection-to-test turnaround time, with further analysis over 5 weeks to illustrate degradation trends. RESULTS: After storing at 4°C, the proportion of specimens that remained positive for M. genitalium was 100% after 1 week and 95% after 4 weeks. Storage at 22°C led to more rapid decline in detection in the first 4 weeks, with 95% detected after 1 week and 85% at 2 weeks onwards. At 5 weeks, samples stored at both temperatures had an 85% M. genitalium detection rate, with increase in crossing points (Cq) of 0·72 (95% confidence interval (CI) 0·01-1·43; P-trend = 0·027) at 4°C, and 1·75 ((95% CI 0·79-2·71), P-trend <0·001) at 22°C. CONCLUSIONS: Urine samples stored without preservative, and unfrozen, retained high M. genitalium detection levels over the short term (up to 5 weeks). To minimize degradation, storing at 4°C is recommended. SIGNIFICANCE AND IMPACT OF THE STUDY: There is little known about the stability of clinical samples for M. genitalium detection. This study found that a high proportion (85-100%) of samples are still suitable for M. genitalium detection after storage for up to 5 weeks.

Efficacy, Immunogenicity, and Safety of a 9-Valent Human Papillomavirus Vaccine: Subgroup Analysis of Participants From Asian Countries.

Background: A 9-valent human papillomavirus-6/11/16/18/31/33/45/52/58 (9vHPV) vaccine extends coverage to 5 next most common oncogenic types (31/33/45/52/58) in cervical cancer versus quadrivalent HPV (qHPV) vaccine. We describe efficacy, immunogenicity, and safety in Asian participants (India, Hong Kong, South Korea, Japan, Taiwan, and Thailand) from 2 international studies: a randomized, double-blinded, qHPV vaccine-controlled efficacy study (young women aged 16-26 years; NCT00543543; Study 001); and an immunogenicity study (girls and boys aged 9-15 years; NCT00943722; Study 002). Methods: Participants (N = 2519) were vaccinated at day 1 and months 2 and 6. Gynecological samples (Study 001 only) and serum were collected for HPV DNA and antibody assessments, respectively. Injection-site and systemic adverse events (AEs) were monitored. Data were analyzed by country and vaccination group. Results: 9vHPV vaccine prevented HPV-31/33/45/52/58-related persistent infection with 90.4%-100% efficacy across included countries. At month 7, ≥97.9% of participants seroconverted for each HPV type. Injection-site AEs occurred in 77.7%-83.1% and 81.9%-87.5% of qHPV and 9vHPV vaccine recipients in Study 001, respectively, and 62.4%-85.7% of girls/boys in Study 002; most were mild to moderate. Conclusions: The 9vHPV vaccine is efficacious, immunogenic, and well tolerated in Asian participants. Data support 9vHPV vaccination programs in Asia. Clinical Trials Registration: NCT00543543; NCT00943722.

Prospective Evaluation of ResistancePlus MG, a New Multiplex Quantitative PCR Assay for Detection of Mycoplasma genitalium and Macrolide Resistance.

Mycoplasma genitalium is a significant pathogen for which first-line treatment is becoming less effective due to increased resistance to macrolides. As conventional culture and antimicrobial susceptibility testing is not feasible for routine detection of this pathogen, molecular markers such as detection of mutations in the 23S rRNA gene have been described to predict resistance. Recently, a novel multiplex quantitative PCR (qPCR) assay, ResistancePlus MG, has been described for the simultaneous detection of Mycoplasma genitalium and macrolide resistance. In the current study, the clinical performance of the assay was evaluated on 1,089 consecutive urine and anogenital swab samples in symptomatic and asymptomatic male and female patients. Overall, 6.0% were positive for M. genitalium, with 63.1% having macrolide resistance-associated mutations. Compared to the laboratory-validated qPCR method targeting the 16S rRNA gene and Sanger sequencing to determine 23S rRNA mutations, the sensitivity and specificity of M. genitalium detection were 98.5% and 100% and for detection of macrolide resistance mutations were 100.0% and 96.2%, respectively. This assay offers a considerable advantage in clinical settings for M. genitalium testing by making the results of macrolide resistance and mutation analyses simultaneously available, which is increasingly important with escalating macrolide resistance.

Determinants of bone mineral density in young Australian women; results from the Safe-D study.

The study aimed to explore determinants of bone parameters in young women. Most bone parameters were associated with height and lean mass. Bone parameters were not associated with vitamin D status. Future research should address whether interventions aimed at improving lean mass are beneficial to bone health in young women. INTRODUCTION: The implementation of prevention strategies during young adulthood may be crucial for osteoporosis prevention in later life, yet literature examining the determinants of bone health in premenopausal women is limited. We aimed to assess determinants of bone health, including serum 25-hydroxyvitamin D (25OHD), in females aged 16-25 years, living in Victoria, Australia, recruited through Facebook advertising. METHODS: Serum 25OHD was measured by liquid chromatography-tandem mass spectrometry and bone health was measured using dual-energy X-ray absorptiometry (DXA) and peripheral quantitative computed tomography (pQCT) in 326 participants. RESULTS: Mean (± standard deviation) serum 25OHD was 69 ± 28 nmol/L and the prevalence of vitamin D deficiency (serum 25OHD <50 nmol/L) was 26%. Seven percent of participants (n = 23) reported taking a vitamin D supplement. Two percent of participants had low lumbar spine bone mineral density (Z-score <-2.0), 5% at the hip and 7% at the femoral neck. Serum 25OHD levels were not associated with DXA bone parameters, nor with pQCT bone parameters. Most bone parameters were positively associated with height and lean mass. CONCLUSION: Vitamin D status was not associated with bone health in young women in the current study. Our findings suggest that targeting other modifiable factors, such as lean body mass, is likely to be beneficial to bone health in young women. Longitudinal studies examining the association between vitamin D status and bone health in young women are necessary to confirm our findings. In addition, whether raising 25OHD levels is advantageous for young women's bone health is yet to be determined.

A snapshot of Chlamydia trachomatis genetic diversity using multilocus sequence type analysis in an Australian metropolitan setting.

High-resolution screening methodologies which enable the differentiation of Chlamydia trachomatis at the strain level, directly from clinical samples, can provide the detailed information required for epidemiological questions such as the dynamics of treatment failure. In addition, they give a detailed snapshot of circulating C. trachomatis genetic variation, data which are currently lacking for the Australian population. In the context of two Australian clinical trials, we assessed the genetic diversity of C. trachomatis and compared these to strains circulating globally. We used high-resolution multilocus sequence typing (MLST) of five highly variable genetic regions of C. trachomatis to examine variation in Australia. Samples with established genovars were drawn from a pool of 880 C. trachomatis-positive samples from two clinical studies, whereby 76 sample pairs which remained C. trachomatis-positive for the same genovar after treatment underwent MLST analysis to distinguish between treatment failure and reinfection. MLST analysis revealed a total of 25 sequence types (STs), six new allele variants and seven new STs not described anywhere else in the world, when compared to those in the international C. trachomatis MLST database. Of the eight most common global STs, seven were found in Australia (four derived from men who have sex with men (MSM) and three from heterosexuals). Newly identified STs were predominantly found in samples from the MSM population. In conclusion, MLST provided a diverse C. trachomatis strain profile, with novel circulating STs, and could be used to identify local sexual networks to focus on interventions such as testing and partner notification to prevent reinfection.

Anyplex II HPV28 detection and Anyplex II HPV HR detection assays are highly concordant with other commercial assays for detection of high-risk HPV genotypes in women with high grade cervical abnormalities.

PURPOSE: to evaluate the performance of Anyplex II HPV28 and HPV HR Detection assays against the EuroArray HPV, Cobas 4800 HPV (Cobas), HPV Amplicor (Amp), Linear Array HPV (LA) and Hybrid Capture 2 (HC2) in detection of high-risk HPV (HR-HPV) from liquid-based cervical cytology samples. METHODS: cervical specimens from 404 women undergoing management of high-grade cytological abnormality were evaluated by Anyplex II HPV28 and HPV HR Detection assays for detection of HR-HPV genotypes and prediction of histologically-confirmed cervical intraepithelial neoplasia grade 2 or higher (≥CIN2). The results were compared to EuroArray, HC2, Cobas, Amp, and LA. RESULTS: specimens were evaluated from 404 women with an average age of 30 years, including 336 with a histological diagnosis of ≥ CIN2 and 68 with ≤ CIN1. Concordance of HR-HPV detection between Anyplex II HPV28 and other genotyping assays was 94.79 % (κ = 0.84; EuroArray) and 97.27 % (κ = 0.91; LA); and between Anyplex II HPV HR and other HR-HPV detection assays was 86.35 % (κ = 0.62; HC2), 96.03 % (κ = 0.87; Cobas) and 96.77 % (κ = 0.89; Amp). Using HR-HPV detection for prediction of ≥ CIN2 by Anyplex II HPV28 and HPV HR, sensitivity (90.18, 95 % CI 86.48-93.14; 90.77, 95 % CI 87.16-93.65) and specificity (both 67.16, 95 % CI 54.60-78.15) were not significantly different to the other HPV assays tested, with one exception. Both Anyplex assays had significantly higher sensitivity than HC2 (p < 0.0001), with a specificity of 96 % (p > 0.05) of HC2 in this high-risk population. CONCLUSIONS: both Anyplex II HPV detection assays were concordant with other commercial assays for HR-HPV detection, with comparable sensitivity and specificity for ≥ CIN2 detection.

Evaluation of the Hologic Panther Transcription-Mediated Amplification Assay for Detection of Mycoplasma genitalium.

The detection of Mycoplasma genitalium was evaluated on 1,080 urine samples by the use of a Panther instrument. Overall sensitivity, specificity, positive predictive values, and negative predictive values were 100%, 99.4%, 93.6%, and 100%, respectively. Detection of M. genitalium by the use of the Panther transcription-mediated amplification assay offers a simple, accurate, and sensitive platform for diagnostic laboratories.

Comparison of the Roche Cobas® 4800 HPV assay to Roche Amplicor for detection of high-risk human papillomavirus.

UNLABELLED: Roche Amplicor HPV (AMP) had previously been used for detection of high-risk human papillomavirus (HR-HPV) in epidemiological and clinical studies. As this assay is no longer available, we compared its performance using PreservCyt samples from women aged of 18-24 years attending for routine cervical cytology screening to Roche Cobas® 4800 (Cobas) to determine if subsequent studies could continue using the Cobas assay. Overall 507 samples were tested on Cobas and compared to previous AMP results, with discrepant samples tested on Roche Linear Array. RESULTS: Overall, agreement between the Cobas and AMP for the presence of HR HPV types was very high (κ = 0.81) (95 % CI: 0.76 - 0.87) with percentage agreement of 91.57 %. Cobas is comparable to AMP for the detection of HR-HPV types in a community recruited cohort of healthy women.

EUROarray human papillomavirus (HPV) assay is highly concordant with other commercial assays for detection of high-risk HPV genotypes in women with high grade cervical abnormalities.

The purpose of this study was to evaluate the performance of the EUROIMMUN EUROArray HPV genotyping assay against the Roche Cobas 4800, Roche HPV Amplicor, Roche Linear Array and Qiagen Hybrid Capture 2 assays in the detection of high-risk HPV (HR-HPV) from liquid based cervical cytology samples collected from women undergoing follow-up for abnormal cervical cytology results. Cervical specimens from 404 women undergoing management of high-grade cytological abnormality were evaluated by EUROarray HPV for detection of HR-HPV genotypes and prediction of histologically-confirmed cervical intraepithelial neoplasia grade 2 or higher (≥CIN2). The results were compared to Hybrid Capture 2, Cobas 4800 HPV, Amplicor and Linear Array HPV. Positivity for 14 HR-HPV types was 80.0 % for EUROarray (95 % CI; 75.7-83.8 %). Agreement (κ, 95 % CI) between the EUROarray and other HPV tests for detection of HR-HPV was good to very good [Hybrid Capture κ = 0.62 (0.54-0.71); Cobas κ = 0.81 (0.74-0.88); Amplicor κ = 0.68 (0.60-0.77); Linear Array κ = 0.77 (0.70-0.85)]. For detection of HR-HPV, agreement with EUROarray was 87.90 % (Hybrid Capture), 93.58 % (Cobas), 92.84 % (Amplicor) and 92.59 % (Linear Array). Detection of HR-HPV was not significantly different between EUROarray and any other test (p < 0.001). EUROarray was concordant with other assays evaluated for detection of high-risk HPV and showed sensitivity and specificity for detection of ≥ CIN2 of 86 % and 71 %, respectively.

The acceptability and cost of a home-based chlamydia retesting strategy: findings from the REACT randomised controlled trial.

BACKGROUND: Chlamydia retesting three months after treatment is recommended to detect reinfections, but retesting rates are typically low. The REACT (retest after Chlamydia trachomatis) randomised trial demonstrated that home-based retesting using postal home-collection kits and SMS reminders, resulted in substantial improvements in retesting rates in women, heterosexual men and men who have sex with men (MSM), with detection of more repeat positive tests compared with SMS reminder alone. In the context of this trial, the acceptability of the home-based strategy was evaluated and the costs of the two strategies were compared. METHODS: REACT participants (200 women, 200 heterosexual men, 200 MSM) were asked to complete an online survey that included home-testing acceptability and preferred methods of retesting. The demographics, sexual behaviour and acceptability of home collection were compared between those preferring home-testing versus clinic-based retesting or no preference, using a chi-square test. The costs to the health system of the clinic-based and home retesting strategies and the cost per infection for each were also compared. RESULTS: Overall 445/600 (74 %) participants completed the survey; 236/445 from the home-testing arm, and 141 of these (60 %) retested at home. The majority of home arm retesters were comfortable having the kit posted to their home (86 %); found it easy to follow the instructions and collect the specimens (96 %); were confident they had collected the specimens correctly (90 %); and reported no problems (70 %). Most (65 %) preferred home retesting, 21 % had no preference and 14 % preferred clinic retesting. Comparing those with a preference for home testing to those who didn't, there were significant differences in being comfortable having a kit sent to their home (p = 0.045); not having been diagnosed with chlamydia previously (p = 0.030); and living with friends (p = 0.034). The overall cost for the home retest pathway was $154 (AUD), compared to $169 for the clinic-based retesting pathway and the cost per repeat infection detected was $1409 vs $3133. CONCLUSIONS: Among individuals initially diagnosed with chlamydia in a sexual health clinic setting, home-based retesting was shown to be highly acceptable, preferred by most participants, and cost-efficient. However some clients preferred clinic-based testing, often due to confidentiality concerns in their home environment. Both options should be provided to maximise retesting rates. TRIAL REGISTRATION: The trial was registered with the Australia New Zealand Clinical Trials Registry on September 9, 2011: ACTRN12611000968976.

The effects of oral garlic on vaginal candida colony counts: a randomised placebo controlled double-blind trial.

OBJECTIVE: Garlic is effective against Candida species in vitro, and along with other alternative therapies, is used by women with vulvovaginal candidiasis. The objective of this study was to ascertain whether oral garlic reduced vaginal candida counts during the second half of the menstrual cycle in asymptomatic women colonised with Candida species. DESIGN: A simple randomised double-blinded controlled trial. SETTING: Melbourne, Australia. SAMPLE: Sixty-three asymptomatic women who were culture-positive for Candida species at screening. METHODS: Participants were randomised to three garlic tablets or placebo orally, twice daily, for 14 days. MAIN OUTCOME MEASURES: The primary outcome was the proportion of women with colony counts of candida >100 colony-forming units per ml in any given day during the last 7 days before menstruation, defined as a 'case'. Secondary outcomes included the mean quantitative colony counts of candida over 14 days prior to menses. RESULTS: There was no evidence of a difference between the proportion of cases in the garlic and placebo groups (76 versus 90%; relative risk, RR 0.85; 95% confidence interval, 95% CI 0.67-1.08), in the mean colony counts in both groups (ratio of geometric means of candidal colony counts 0.63; 95% CI 0.39-10.03; P = 0.74), or difference in the number of women reporting abnormal vaginal symptoms during the 2 weeks before menstruation (RR 1.03; 95% CI 0.67-1.58; P = 0.91). The garlic group reported more adverse effects (83% compared 43% in the placebo group; difference in proportions 39%; 95% CI 17-%; P < 0.01). CONCLUSIONS: This study provided data for sample size calculations in future studies on the antifungal effect of garlic, but provided no evidence to inform clinical practice regarding the use of garlic in vaginal candidiasis. Further studies might investigate longer courses or topical formulations.

Chlamydia trachomatis serovars in community-based HIV-positive and HIV-negative men who have sex with men in Sydney, Australia.

OBJECTIVES: There are few data on the distribution of specific Chlamydia trachomatis serovars among men who have sex with men (MSM) outside clinical settings. To investigate these patterns, serovar determination was performed on chlamydia-positive samples from two community-based cohort studies of HIV-positive and HIV-negative MSM in Sydney, Australia. METHODS: From January 2005 to June 2007 all positive C trachomatis pharyngeal, urine and anal samples were evaluated. The serovar of each C trachomatis infection was determined by omp1 gene sequencing with confirmatory quantitative PCR screening. Symptom data were routinely reported by study participants at the time of testing. RESULTS: Serovar determination was possible for 54 samples among 52 participants. Seven samples were not able to be typed. Site-specific symptoms were reported by fewer than 10% of participants diagnosed with pharyngeal and anogenital chlamydia. The most commonly identified serovars were serovar D (n=32, 59.3%, 95% CI 45.0 to 72.4), followed by serovar G (n=11, 20.4%, 95% CI 10.6 to 33.5) and serovar J (n=5, 9.3%, 95% CI 3.1 to 20.3). Only one lymphogranuloma venereum serovar was identified (L2b). CONCLUSIONS: This community-based study found a similar distribution of chlamydia serovars to that observed among Australian community-based MSM several years ago, and serovar distribution recently observed among predominantly symptomatic MSM at a Sydney clinic. These findings suggest little change in C trachomatis serovar distribution in Australian MSM over the past decade and a lack of correlation of specific chlamydia serovars with anogenital symptoms among MSM.

Pilot study of the utility and acceptability of tampon sampling for the diagnosis of Neisseria gonorrhoeae and Chlamydia trachomatis infections by duplex realtime polymerase chain reaction in United Kingdom sex workers.

We aimed to evaluate the acceptability of self-collected tampon samples for the screening of female sex workers for sexually transmitted infections. We recruited 65 sex workers, and 63 agreed to provide tampon samples. The tampon samples were processed by realtime polymerase chain reaction (PCR) targeting Neisseria gonorrhoeae and Chlamydia trachomatis. Urethral and endocervical swabs were also obtained from 61 of 63 participants and tested using culture (N. gonorrhoeae) and the BD ProbeTec strand displacement amplification (SDA) (C. trachomatis) assay. Tampon sampling was preferred by 95% of the women and all favoured being tested away from genitourinary medicine clinics; the most common reasons cited were avoidance of embarrassment (40%) and convenience (30%). Besides near-universal acceptability of tampon sampling, the tampon sampling-PCR approach described in this study appeared to have enhanced sensitivity compared with conventional testing, suggesting the possibility of a residual hidden burden of N. gonorrhoeae and/or C. trachomatis genital infections in UK female sex workers.

A custom amplicon sequencing approach to detect resistance associated mutations and sequence types in Mycoplasma genitalium.

BACKGROUND: Mycoplasma genitalium resistance to antibiotic treatments is increasing, with very limited treatment alternatives on the horizon. Surveillance via sequencing of multiple M. genitalium loci would allow: monitoring of known antibiotic resistance mutations, associations between resistance/treatment failure and specific mutations, and strain typing for epidemiological purposes. In this study we assessed the performance of a custom amplicon sequencing approach, which negates the cost of library preparation for next generation sequencing. METHODS: Fifty-two M. genitalium positive samples (cervical, vaginal, anal and rectal swabs, and urine) were used. Three regions associated with M. genitalium antibiotic resistance (23S rRNA, parC and gyrA genes) were targeted, in conjunction with a locus used for differentiation of sequence types in the mgpB gene, and findings compared to Sanger sequencing. RESULTS: Amplicon sequencing provided adequate sequence read coverage (>30×) for the majority of samples for 23S rRNA gene (96%) and mgpB (97%), parC (78%) and gyrA (75%). Single nucleotide polymorphisms (SNPs) were characterised in samples for 23S rRNA gene (94%), parC (56%) and gyrA (4%). Unlike Sanger sequencing, mixed mutations could be identified by the amplicon sequencing method, and ratios of mutation types determined. All results, with one exception, were concordant to Sanger sequence results. Sequence diversity in the mgpB region was represented by 15 sequence types, 4 being observed in multiple samples. CONCLUSIONS: We have demonstrated the utility of this custom amplicon sequencing approach for generating highly informative datasets with the capacity to identify and determine ratios of mixed sequences. The use of this customisable amplicon sequencing method enables cost effective, scalable amplicon sequencing of multiple target regions of interest in M. genitalium.

Prevalence and associations of larger burden of intra-anal high-grade squamous intraepithelial lesions at baseline in an Australian cohort of gay and bisexual men: The Study of the Prevention of Anal Cancer.

OBJECTIVES: To investigate factors associated with larger burden of intra-anal high-grade squamous intraepithelial lesions (HSIL) in a natural history study of HSIL. METHODS: 617 gay and bisexual men (GBM) attended a baseline visit. High-resolution anoscopy-directed biopsy was performed of suspected HSIL. GBM with biopsy-confirmed HSIL (bHSIL) affecting a single-octant were compared with those who had bHSIL affecting a larger area. RESULTS: Of 196 men with bHSIL at baseline, 73 (37.2 %) had larger bHSIL burden. Larger burden was independently associated with anal HPV16 detection (aOR 2.06, 95 % CI 1.09-3.89, p = 0.026) and infection with a greater number of high-risk HPV types (aOR per type increase 1.25, 95 % CI 1.05-1.49, p-trend = 0.010). CONCLUSION: The observation that men with a larger burden of HSIL also had more risk factors for anal cancer suggests this group may warrant closer observation to ensure earlier detection, and thus improved prognosis, of individuals whose HSIL may progress to anal cancer.