Global phylogeny of Treponema pallidum lineages reveals recent expansion and spread of contemporary syphilis.

Syphilis, which is caused by the sexually transmitted bacterium Treponema pallidum subsp. pallidum, has an estimated 6.3 million cases worldwide per annum. In the past ten years, the incidence of syphilis has increased by more than 150% in some high-income countries, but the evolution and epidemiology of the epidemic are poorly understood. To characterize the global population structure of T. pallidum, we assembled a geographically and temporally diverse collection of 726 genomes from 626 clinical and 100 laboratory samples collected in 23 countries. We applied phylogenetic analyses and clustering, and found that the global syphilis population comprises just two deeply branching lineages, Nichols and SS14. Both lineages are currently circulating in 12 of the 23 countries sampled. We subdivided T. p. pallidum into 17 distinct sublineages to provide further phylodynamic resolution. Importantly, two Nichols sublineages have expanded clonally across 9 countries contemporaneously with SS14. Moreover, pairwise genome analyses revealed examples of isolates collected within the last 20 years from 14 different countries that had genetically identical core genomes, which might indicate frequent exchange through international transmission. It is striking that most samples collected before 1983 are phylogenetically distinct from more recently isolated sublineages. Using Bayesian temporal analysis, we detected a population bottleneck occurring during the late 1990s, followed by rapid population expansion in the 2000s that was driven by the dominant T. pallidum sublineages circulating today. This expansion may be linked to changing epidemiology, immune evasion or fitness under antimicrobial selection pressure, since many of the contemporary syphilis lineages we have characterized are resistant to macrolides.

Blastomycosis in Africa and the Middle East: A Comprehensive Review of Reported Cases and Reanalysis of Historical Isolates Based on Molecular Data.

BACKGROUND: Blastomycosis has been reported from countries in Africa and the Middle East, but a decades-long debate has persisted regarding whether this is the same disease known in North America and caused by Blastomyces dermatitidis and Blastomyces gilchristii. METHODS: We reviewed published cases of human and veterinary blastomycosis from Africa and the Middle East. We abstracted epidemiological and clinical features of cases, including sites of disease, diagnosis, management, outcomes, and, where available, genetic and antigenic typing of case isolates. In addition, we sequenced nucleic acids from 9 clinical isolates from Africa deposited in global collections as B. dermatitidis; for 5, we sequenced the internal transcribed spacer regions, and for the other 4 we sequenced the whole genomes. RESULTS: We identified 172 unique human patients with blastomycosis, including 159 patients from 25 African countries and 12 patients from 5 Middle Eastern countries, and also identified 7 reports of veterinary blastomycosis. In humans, cutaneous disease predominated (n = 100/137, 73%), followed by pulmonary (n = 73/129, 57%) and osteoarticular involvement (n = 61/128, 48%). Unusual direct microscopy/histopathological presentations included short hyphal fragments in tissues (n = 23/129, 18%). There were 34 genotyped case isolates that comprised 4 species: Blastomyces percursus (n = 22, 65%), from 8 countries throughout all regions; Blastomyces emzantsi (n = 9, 26%), from South Africa; B. dermatitidis (n = 1, 3%), from the Democratic Republic of Congo; and B. gilchristii (n = 2, 6%), from South Africa and Zimbabwe. CONCLUSIONS: Blastomycosis occurs throughout Africa and the Middle East and is caused predominantly by B. percursus and, at least in South Africa, B. emzantsi, resulting in distinct clinical and pathological patterns of disease.

Role of IgM testing in the diagnosis and post-treatment follow-up of syphilis: a prospective cohort study.

OBJECTIVES: The diagnosis of repeat syphilis and its follow-up remains challenging. We aimed to investigate if IgM testing may assist in the diagnosis of syphilis reinfection/relapse and its treatment follow-up. METHODS: This substudy was conducted in the context of a syphilis biomarker discovery study (ClinicalTrials.gov Nr: NCT02059525). Sera were collected from 120 individuals with a new diagnosis of syphilis (72 with repeat infections) and 30 syphilis negative controls during a cohort study investigating syphilis biomarkers conducted at a sexually transmitted infection/HIV clinic in Antwerp, Belgium. Syphilis was diagnosed based on a simultaneous positive treponemal and non-treponemal assay result and/or positive serum PCR targeting polA. Specimens collected at visit of diagnosis, and 3 and 6 months post-treatment were tested by two enzyme immunoassays (EIAs), recomWell (Mikrogen; MI) and Euroimmun (EU), to detect anti-treponemal IgM. Baseline specimens were also tested for anti-treponemal IgM using a line immunoassay (LIA) recomLine (MI). Quantitative kinetic decay curves were constructed from the longitudinal quantitative EIA results. RESULTS: An overall sensitivity for the diagnosis of syphilis of 59.8% (95% CI: 50.3%-68.7%), 75.0% (95% CI: 66.1%-82.3%) and 63.3% (95% CI: 54.8%-72.6%) was obtained for the EU, MI EIAs and MI LIA, respectively. When only considering repeat syphilis, the diagnostic sensitivity decreased to 45.7% (95% CI: 33.9%-58.0%), 63.9% (95% CI: 51.7%-74.6%) and 47.2% (95% CI: 35.5%-59.3%), respectively. IgM seroreverted in most cases 6 months after treatment. Post-treatment IgM concentrations decreased almost 30% faster for initial syphilis compared with repeat infection. The IgM EIAs and IgM LIA agreed from fairly to moderately (Cohen's kappa (κ): 0.36 (EU EIA); κ: 0.53 (MI EIA); κ: 0.40 (MI LIA)) with the diagnosis of syphilis. CONCLUSIONS: IgM detection was not a sensitive method to diagnose syphilis and was even poorer in the diagnosis of syphilis repeat infections.

HIV prevalence correlated with circumcision prevalence and high-risk sexual behavior in India's states: an ecological study.

Background: HIV prevalence varies between 0% and 1.6% in India's states. The factors underpinning this variation are poorly defined. Methods: We evaluated the relationship between HIV prevalence by state and a range of risk factors in the Indian 2015 National Family Health Survey. Pearson's correlation was used to assess the relationship between HIV prevalence and each variable. The prevalence of each risk factor was compared between five high-HIV-prevalence states (>1% prevalence) and a large low-HIV-prevalence state (Uttar Pradesh; HIV prevalence, 0.06%). Results: There was an association between HIV prevalence and men's mean lifetime number of partners (r = 0.55; P = 0.001) and men reporting sex with a non-married, non-cohabiting partner (r = 0.40; P = 0.014). In general, men in high-prevalence states were less likely to be circumcised and (with the exception of Chandigarh) use condoms at last sex. In two high prevalence states (Mizoram and Nagaland), men reported a higher number of lifetime partners and a higher prevalence of multiple partners and high-risk sex in the past year. Conclusions: Variation in circumcision prevalence and sexual behavior may contribute to the large variations in HIV prevalence by state in India.

Superimposing incident sexually transmitted infections on HIV phylogram to investigate possible misclassification of men who have sex with men as heterosexuals in a cohort in Antwerp, Belgium.

In this study, we assessed if the superimposition of incident sexually transmitted infections (STIs) on HIV phylogenetic analyses could reveal possible sexual behaviour misclassifications in our HIV-infected population. HIV-1 sequences collected between 1997 and 2014 from 1169 individuals attending a HIV clinic in Antwerp, Belgium were analysed to infer a partial HIV transmission network. Individual demographic, clinical and laboratory data collected during routine HIV follow-up were used to compare clustered and non-clustered individuals using logistic regression analyses. In total, 438 (37.5%) individuals were identified in 136 clusters, including 76 transmission pairs and 60 clusters consisting of three or more individuals. Individuals in a cluster were more likely to have a history of syphilis, Chlamydia and/or gonorrhoea (P < 0.05); however, when analyses were stratified by HIV transmission risk groups (heterosexual and men who have sex with men [MSM]), this association only remained significant for heterosexuals with syphilis (P = 0.001). Under closer scrutiny, this association was driven by six heterosexual men who were located in six almost exclusively MSM clusters. A parsimonious conclusion is that these six individuals were potentially misclassified as heterosexual. Improving the accuracy of sexual behaviour reporting could improve care.

Differential sexual network connectivity offers a parsimonious explanation for population-level variations in the prevalence of bacterial vaginosis: a data-driven, model-supported hypothesis.

BACKGROUND: The prevalence of bacterial vaginosis (BV) and vaginal microbiota types varies dramatically between different populations around the world. Understanding what underpins these differences is important, as high-diversity microbiotas associated with BV are implicated in adverse pregnancy outcomes and enhanced susceptibility to and transmission of sexually transmitted infections. MAIN TEXT: We hypothesize that these variations in the vaginal microbiota can, in part, be explained by variations in the connectivity of sexual networks. We argue: 1) Couple-level data suggest that BV-associated bacteria can be sexually transmitted and hence high sexual network connectivity would be expected to promote the spread of BV-associated bacteria. Epidemiological studies have found positive associations between indicators of network connectivity and the prevalence of BV; 2) The relationship between BV prevalence and STI incidence/prevalence can be parsimoniously explained by differential network connectivity; 3) Studies from other mammals are generally supportive of the association between network connectivity and high-diversity vaginal microbiota. CONCLUSION: To test this hypothesis, we propose a combination of empirical and simulation-based study designs.

Needle lost in the haystack: multiple reaction monitoring fails to detect Treponema pallidum candidate protein biomarkers in plasma and urine samples from individuals with syphilis.

Background: Current syphilis diagnostic strategies are lacking a sensitive manner of directly detecting Treponema pallidum antigens. A diagnostic test that could directly detect T. pallidum antigens in individuals with syphilis would be of considerable clinical utility, especially for the diagnosis of reinfections and for post-treatment serological follow-up. Methods: In this study, 11 candidate T. pallidum biomarker proteins were chosen according to their physiochemical characteristics, T. pallidum specificity and predicted abundance. Thirty isotopically labelled proteotypic surrogate peptides (hPTPs) were synthesized and incorporated into a scheduled multiple reaction monitoring assay. Protein extracts from undepleted/unenriched plasma (N = 18) and urine (N = 4) samples from 18 individuals with syphilis in various clinical stages were tryptically digested, spiked with the hPTP mixture and analysed with a triple quadruple mass spectrometer. Results: No endogenous PTPs corresponding to the eleven candidate biomarkers were detected in any samples analysed. To estimate the Limit of Detection (LOD) of a comparably sensitive mass spectrometer (LTQ-Orbitrap), two dilution series of rabbit cultured purified T. pallidum were prepared in PBS. Polyclonal anti- T. pallidum antibodies coupled to magnetic Dynabeads were used to enrich one sample series; no LOD improvement was found compared to the unenriched series. The estimated LOD of MS instruments is 300 T. pallidum/ml in PBS. Conclusions: Biomarker protein detection likely failed due to the low (femtomoles/liter) predicted concentration of T. pallidum proteins. Alternative sample preparation strategies may improve the detectability of T. pallidum proteins in biofluids.

Could differences in implicit attitudes to sexual concurrency play a role in generalized HIV epidemics?

Background: Sexual partner concurrency has been implicated in the genesis of generalized HIV epidemic in South Africa. Most South Africans, however, disapprove of concurrency in surveys. These surveys test individuals' explicit attitudes which are susceptible to a number of important biases such as the social desirability bias. Assessment of implicit cognitions have been found to be better predictors of behaviour in socially sensitive domains. We hypothesized that South Africans may have implicit attitudes more tolerant of concurrency than lower concurrency prevalence populations. Methods: To test this hypothesis, we developed a concurrency-implicit association test (C-IAT) and compared the C-IATs of samples of South African and Belgian university students. Results: We found a large and statistically significant difference in the C-IAT between the South Africans (D600-score = -0.009, indicating absence of preference for concurrency or monogamy) and Belgians (D600-score = 0.783, indicating a strong preference for monogamy; t-test = 13.3; P < 0.0001). The effect size measure, Cohen's d, was found to be 0.88, which is considered a large effect size in this field. Conclusions: Our results are compatible with the thesis that differences in implicit attitudes to concurrency play a role in the genesis of generalised HIV epidemics.

Candidate Treponema pallidum biomarkers uncovered in urine from individuals with syphilis using mass spectrometry.

AIM: A diagnostic test that could detect Treponema pallidum antigens in urine would facilitate the prompt diagnosis of syphilis. MATERIALS & METHODS: Urine from 54 individuals with various clinical stages of syphilis and 6 controls were pooled according to disease stage and interrogated with complementary mass spectrometry techniques to uncover potential syphilis biomarkers. RESULTS & CONCLUSION: In total, 26 unique peptides were uncovered corresponding to four unique T. pallidum proteins that have low genetic sequence similarity to other prokaryotes and human proteins. This is the first account of direct T. pallidum protein detection in human clinical samples using mass spectrometry. The implications of these findings for future diagnostic test development is discussed. Data are available via ProteomeXchange with identifier PXD009707.

Does gonorrhoea screening intensity play a role in the early selection of antimicrobial resistance in men who have sex with men (MSM)? A comparative study of Belgium and the United Kingdom.

Background: It is unclear why antimicrobial resistance in Neisseriagonorrhoeae in the United Kingdom (UK) and the United States has tended to first appear in men who have sex with men (MSM). We hypothesize that increased exposure to antimicrobials from intensive STI screening programmes plays a role. Methods: We assess if there is a difference in the distribution of azithromycin, cefixime and ceftriaxone minimum inhibitory concentrations (MICs) between MSM and women in the United Kingdom (UK) where 70% of MSM report STI screening in the past year vs. Belgium where 9% report STI screening in the past year. Our hypothesis is that MICs of the MSM should be higher than those of the women in the UK but not Belgium. Data for the MICs were taken from the Gonococcal Resistance to Antimicrobials Surveillance Programme (GRASP) in the UK in 2010/2011 and 2014 and a similar national surveillance programme in Belgium in 2013/2014 (the first most complete available data). We used the Mann-Whitney test to compare the MIC distributions between MSM and women within each country Results: In the UK the MICs for all three antimicrobials were significantly higher in MSM than women at both time points (P all <0.0005). In Belgium only the MIC distribution for azithromycin was higher in MSM (P<0.0005). Conclusion: The findings for cefixime and ceftriaxone, but not azithromycin are compatible with our hypothesis that screening-intensity could contribute to the emergence of AMR. Numerous other interpretations of our results are discussed.

Effects of Sexual Network Connectivity and Antimicrobial Drug Use on Antimicrobial Resistance in Neisseria gonorrhoeae.

Contemporary strategies to curtail the emergence of antimicrobial resistance in Neisseria gonorrhoeae include screening for and treating asymptomatic infections in high-prevalence populations in whom antimicrobial drug-resistant infections have typically emerged. We argue that antimicrobial resistance in these groups is driven by a combination of dense sexual network connectivity and antimicrobial drug exposure (for example, through screen-and-treat strategies for asymptomatic N. gonorrhoeae infection). Sexual network connectivity sustains a high-equilibrium prevalence of N. gonorrhoeae and increases likelihood of reinfection, whereas antimicrobial drug exposure results in selection pressure for reinfecting N. gonorrhoeae strains to acquire antimicrobial resistance genes from commensal pharyngeal or rectal flora. We propose study designs to test this hypothesis.

Implicit attitudes to sexual partner concurrency vary by sexual orientation but not by gender-A cross sectional study of Belgian students.

High rates of sexual partner concurrency have been shown to facilitate the spread of various sexually transmitted infections. Assessments of explicit attitudes to concurrency have however found little difference between populations. Implicit attitudes to concurrency may vary between populations and play a role in generating differences in the prevalence of concurrency. We developed a concurrency implicit associations test (C-IAT) to assess if implicit attitudes towards concurrency may vary between individuals and populations and what the correlates of these variations are. A sample of 869 Belgian students (mean age 23, SD 5.1) completed an online version of the C-IAT together with a questionnaire concerning sexual behavior and explicit attitudes to concurrency. The study participants C-IATs demonstrated a strong preference for monogamy (-0.78, SD = 0.41). 93.2% of participants had a pro-monogamy C-IAT. There was no difference in this implicit preference for monogamy between heterosexual men and women. Men who have sex with men and women who have sex with women were more likely to exhibit implicit but not explicit preferences for concurrency compared to heterosexual men and women. Correlates of the C-IAT varied between men and women.

HIV prevalence by ethnic group covaries with prevalence of herpes simplex virus-2 and high-risk sex in Uganda: An ecological study.

BACKGROUND: HIV prevalence varies from 1.7% to 14.8% between ethnic groups in Uganda. Understanding the factors responsible for this heterogeneity in HIV spread may guide prevention efforts. METHODS: We evaluated the relationship between HIV prevalence by ethnic group and a range of risk factors as well as the prevalence of herpes simplex virus-2 (HSV-2), syphilis and symptomatic STIs in the 2004/2005 Uganda HIV/AIDS Sero-Behavioural Survey-a two stage, nationally representative, population based survey of 15-59-year-olds. Spearman's correlation was used to assess the relationship between HIV prevalence and each variable. RESULTS: There was a positive association between HIV prevalence and HSV-2, symptomatic STIs and high-risk sex (sex with a non-cohabiting, non-marital partner) for women. Non-significant positive associations were present between HIV and high-risk sex for men and lifetime number of partners for men and women. CONCLUSION: Variation in sexual behavior may contribute to the variations in HIV, HSV-2 and other STI prevalence by ethnic group in Uganda. Further work is necessary to delineate which combinations of risk factors determine differential STI spread in Uganda.

What Is the Role of Paired Rapid Plasma Reagin Testing (Simultaneous Testing of Acute and Convalescent Samples) in the Diagnosis of Repeat Syphilis and the Follow-up of Syphilis?

BACKGROUND: Repeat syphilis is playing an increasing role in syphilis transmission in several populations. The assessment of repeat syphilis and response to treatment depends on accurately measuring intraindividual changes in non-treponemal tests. For a 0- to 6-month delta rapid plasma reagin (RPR) to be determined by routine individual RPR testing, samples are tested 6 months apart with differences in reagent batches, environmental conditions, and observers all leading to measurement errors. We hypothesized that conducting paired RPR testing (simultaneous testing of acute and convalescent samples) would enable a more accurate determination of delta RPR compared with individual testing. METHODS: A total of 120 study participants with a new diagnosis of syphilis were followed up at 0, 3, 6, 9, 12, 18, and 24 months, with RPR testing performed via individual testing at each study visit and at any suspected repeat syphilis. Rapid plasma reagin paired testing was performed on samples from 0 and 6 months and at any suspected repeat syphilis. RESULTS: The quantitative agreement ±1 dilution among paired and individual testing was 97.2%. There was no difference in the proportion with serofast status at 6 months: 21 (19.4%) and 19 (17.6%) according to paired and individual testing, respectively (P = 0.726). There was no statistically significant difference between 0- and 6-month delta RPR as determined by paired and individual testing in predicting seroresponse at 12 months (86.1% and 91.6% agreement with 12-month serofast/nonserofast classification, respectively; P = 0.262). CONCLUSIONS: In our setting, individual testing performed equally well compared with paired testing. Follow-up of syphilis will remain onerous for the patient and the health care provider until new tests that can more accurately assess the response to therapy and repeat syphilis/treatment failure are developed.

Association between intensity of STI screening and development of antimicrobial resistance in N. gonorrhoeae in 12 cities in the USA: An ecological study.

In this study, we assessed if there was a city-level association between sexually transmitted infection (STI) screening intensity in men who have sex with men and antimicrobial sensitivity in Neisseria gonorrhoeae in the United States, 2007 to 2013.  We found positive associations between STI screening intensity and increases in minimum inhibitory concentrations for cefixime and azithromycin, but not ceftriaxone when using change in city geometric mean N. gonorrhoeae MIC between 2005 and 2013.

Association between bacterial vaginosis and partner concurrency: a longitudinal study.

OBJECTIVES: The study aimed to test if there was an association between prevalent bacterial vaginosis (BV) and women reporting that their partner had other partners at the same time (partner concurrency). This association has not been assessed in a longitudinal cohort. METHODS: The Longitudinal Study of Vaginal Flora recruited a cohort of 3620 non-pregnant women aged 15-44 years who presented for routine primary healthcare at 12 clinics in Birmingham, Alabama. Behavioural questionnaires and vaginal smears were obtained quarterly for a year and BV was defined by a Nugent score 7 or higher as well as Amsel criteria. Mixed effects logistic regression was used to assess the relationship between prevalent BV and reporting that one's partner had other partners in the preceding 3-6 months time interval. RESULTS: Nugent score prevalent BV was associated with both reporting that one's partner definitely (adjusted OR (aOR) 1.5; 95% CI 1.2 to 1.8) and possibly (aOR 1.5; 95% CI 1.2 to 1.8) engaged in partner concurrency in the preceding 3-6 months time period. Prevalent BV diagnosed by Amsel criteria was similar. CONCLUSIONS: A diagnosis of prevalent BV was associated with reporting that one's partner possibly or definitely engaged in partner concurrency.

Strong association between higher-risk sex and HIV prevalence at the regional level: an ecological study of 27 sub-Saharan African countries.

Background: It is unclear why HIV prevalence varies by nearly two orders of magnitude between regions within countries in sub-Saharan Africa. In this ecological study, we assess if HIV prevalence by region is associated with any of four markers of higher risk sexual behavior: lifetime number of partners, multiple partners in past year, higher risk sex (defined as sex with non-cohabiting, non-marital partners) and age at debut. Methods: We performed Pearson's correlation between the 4 behavioral risk factors and HIV prevalence by region in 47 nationally representative surveys from 27 sub-Saharan African countries, separately by gender. In addition, principal components analysis was used to reduce the eight risk factors (four for each gender) to two principal components (PCs). Mixed effects linear regression was used to assess the relationship between the resulting two PCs and HIV prevalence after controlling for the prevalence of male circumcision. Results: HIV prevalence varied by a median 3.7 fold (IQR 2.9-7.9) between regions within countries. HIV prevalence was strongly associated with higher risk sex and, to a lesser extent, the other risk factors evaluated. Both PCs were strongly associated with HIV prevalence when assessed via linear regression. Conclusions: Differences in sexual behavior may underpin the large differences in HIV-prevalence between subpopulation within sub-Saharan African countries.

It's the network, stupid: a population's sexual network connectivity determines its STI prevalence.

There is little consensus as to why sexually transmitted infections (STIs), including HIV and bacterial vaginosis (BV) are more prevalent in some populations than others. Using a broad definition of sexual network connectivity that includes both structural and conductivity-related factors, we argue that the available evidence suggests that high prevalence of traditional STIs, HIV and BV can be parsimoniously explained by these populations having more connected sexual networks. Positive feedback, whereby BV and various STIs enhance the spread of other STIs, then further accentuates the spread of BV, HIV and other STIs. We review evidence that support this hypothesis and end by suggesting study designs that could further evaluate the hypothesis, as well as implications of this hypothesis for the prevention and management of STIs.

Incident Trichomonas vaginalis Is Associated With Partnership Concurrency: A Longitudinal Cohort Study.

BACKGROUND: Sexual partner concurrency (PC) has been shown to be a risk factor for a number of sexually transmitted infections but it is unknown if it is a risk factor for Trichomonas vaginalis (TV). OBJECTIVE: We assess if there is an association between PC and incident TV infection. STUDY DESIGN: We used mixed effects logistic regression to assess the association between PC and incident TV in the Longitudinal Study of Vaginal Flora, a cohort study of 3620 women followed quarterly for 5 visits. RESULTS: Trichomonas vaginalis was more common in those reporting definite/possible/unknown PC (15.6%/15.0%/18.3%) than those reporting no PC (5.2%; P < 0.001 for all 3 comparisons). After controlling for a range of confounders, incident TV remained associated with reporting that one's partner definitely (adjusted odds ratio, 5.4; 95% confidence interval, 3.7-8.0) and possibly (adjusted odds ratio, 3.4; 95% confidence interval, 2.2-5.1) engaged in PC in the preceding period. CONCLUSIONS: Partner concurrency was associated with incident TV infection.