Frans Radebe

Human immunodeficiency virus infection and genital ulcer disease in South Africa: the herpetic connection.

Background and Objectives: While genital ulcers are a risk factor in HIV infection, the association of specific agents of genital ulcer disease (GUD) with HIV infection may vary. Goal: To determine the etiology of GUD in HIV‐infected and HIV‐uninfected men attending sexually transmitted disease (STD) clinics in Durban, Johannesburg, and Cape Town, South Africa, and the association of previous and current sexually transmitted infections with HIV infection in men with ulcerative and nonulcerative STDs. Study Design: A cross‐sectional study of 558 men with genital ulcers and 602 men with urethritis. Results: Patients with GUD were more likely to be infected with HIV than patients with urethritis (39.4% versus 21.4%, P ≤ 0.001). Herpes simplex virus 2 (HSV‐2) was the most common agent identified in ulcer specimens (35.9%), and was detected in a significantly higher proportion of ulcer specimens from HIV‐infected patients than in specimens from HIV‐uninfected patients (47.4% versus 28.2%, P ≤ 0.001). Patients infected with HIV‐1 were significantly more likely to have HSV‐2 infection, as measured by the presence of the antibody to glycoprotein G‐2, than patients not infected with HIV (63.1% versus 38.5%, P ≤ 0.001). Patients infected with HIV‐1 were also significantly more likely to have initial HSV‐2 infection than HIV‐uninfected patients with GUD (50.0% versus 31.6%, P = 0.007). Haemophilus ducreyi was detected in 31.7% of ulcer specimens; prevalence did not vary by HIV‐infection status. Treponema pallidum DNA was detected significantly less frequently in ulcer specimens from patients infected with HIV than in specimens from patients not infected with HIV (10.2% versus 26%, P ≤ 0.001); no association was found between HIV‐infection status and fluorescent treponemal antibody absorption test seroreactivity, even when men with M‐PCR‐positive syphilis lesions were excluded from the analyses. Conclusion: The authors found that HSV‐2 is a more common etiology of GUD than has been suggested by previous studies conducted in South Africa; serologic evidence of HSV‐2 infection and current cases of genital herpes are strongly associated with HIV infection among men who present to STD clinics with GUD or urethritis.

Whole-genome sequences of Chlamydia trachomatis directly from clinical samples without culture

The use of whole-genome sequencing as a tool for the study of infectious bacteria is of growing clinical interest. Chlamydia trachomatis is responsible for sexually transmitted infections and the blinding disease trachoma, which affect hundreds of millions of people worldwide. Recombination is widespread within the genome of C. trachomatis, thus whole-genome sequencing is necessary to understand the evolution, diversity, and epidemiology of this pathogen. Culture of C. trachomatis has, until now, been a prerequisite to obtain DNA for whole-genome sequencing; however, as C. trachomatis is an obligate intracellular pathogen, this procedure is technically demanding and time consuming. Discarded clinical samples represent a large resource for sequencing the genomes of pathogens, yet clinical swabs frequently contain very low levels of C. trachomatis DNA and large amounts of contaminating microbial and human DNA. To determine whether it is possible to obtain whole-genome sequences from bacteria without the need for culture, we have devised an approach that combines immunomagnetic separation (IMS) for targeted bacterial enrichment with multiple displacement amplification (MDA) for whole-genome amplification. Using IMS-MDA in conjunction with high-throughput multiplexed Illumina sequencing, we have produced the first whole bacterial genome sequences direct from clinical samples. We also show that this method can be used to generate genome data from nonviable archived samples. This method will prove a useful tool in answering questions relating to the biology of many difficult-to-culture or fastidious bacteria of clinical concern.

Etiology and STI/HIV coinfections among patients with urethral and vaginal discharge syndromes in South Africa.

Background: This study was undertaken to establish the etiology of the male urethral discharge (MUDS) and vaginal discharge (VDS) syndromes, to determine the prevalence of other sexually transmitted infections (STI) and human immunodeficiency virus (HIV) coinfections, and to examine associations between STIs and HIV serostatus among STI patients in South Africa. Methods: A total of 507 MUDS and 300 VDS patients were recruited in Cape Town (CPT) and Johannesburg (JHB). A multiplex polymerase chain reaction assay detected Neisseria gonorrhoeae, Chlamydia trachomatis, Trichomonas vaginalis, and Mycoplasma genitalium infections. Bacterial vaginosis and candidiasis were detected by microscopy. Sera were screened for syphilis, HSV-2, and HIV antibodies. Results: Etiological diagnoses were made for 92% of MUDS patients and 85% of VDS patients. Gonorrhoea accounted for 85% (CPT) and 71% (JHB) of MUDS presentations. Chlamydia was the second most frequently detected MUDS pathogen (CPT, 13%; JHB, 24%). Among VDS patients, bacterial vaginosis was the most common cause (CPT, 46%; JHB, 36%) and trichomoniasis the most frequently detected STI pathogen (CPT, 19%; JHB, 34%). Few patients (4%) had serological evidence of syphilis. The HSV-2 and HIV seroprevalence were higher in Johannesburg compared to Cape Town and among women compared to men. HIV infection was statistically significantly associated with HSV-2 seropositivity at both sites and with the presence of N. gonorrhoeae and absence of C. trachomatis in Cape Town MUDS patients. Conclusions: Gonorrhoea and bacterial vaginosis were confirmed as the most frequent causes of MUDS and VDS. The high HIV seroprevalence in STI patients emphasizes the need to address HIV testing among this population.

Comparison of a TaqMan-based real-time polymerase chain reaction with conventional tests for the detection of Trichomonas vaginalis

Objective: To compare a TaqMan-based real-time polymerase chain reaction (PCR) with conventional PCR, culture, and wet-mount microscopy for the diagnosis of trichomoniasis in women. Methods: Vaginal swabs from 119 women were tested for Trichomonas vaginalis by wet mount and culture. Paired vaginal lavage and urine specimens were tested by conventional and real-time PCR. Results: Using an expanded “gold standard”, defined as a positive culture result using vaginal swabs and/or a positive PCR test using TVK3/7 primers, the overall prevalence of T vaginalis in the study population was 65.5% (78/119). The detection rate of T vaginalis was 65.5% (78/119) and 36.9% (44/119) by conventional PCR using vaginal washings and urine specimens, respectively; 68.9% (82/119) by real-time PCR using vaginal washings and 61.3% (73/119) by real-time PCR using urine specimens. The sensitivities of conventional PCR using vaginal washings and urine and real-time PCR using vaginal washings and urine, compared with the gold standard were 100%, 56.4%, 100% and 76.7%, and the specificities of these tests were 100%, 97.6%, 82.9% and 97%, respectively. Conclusions: The real-time PCR test proved to be significantly more sensitive than culture and wet-mount microscopy, although its specificity was slightly lower than these tests. In addition, it was more sensitive, rapid and less time consuming than conventional PCR for the detection of T vaginalis.

Escalation in the relative prevalence of ciprofloxacin resistant gonorrhoea among men with urethral discharge in two South African cities - association with HIV Seropositivity

Objectives: The objectives of this study were to assess the prevalence of ciprofloxacin-resistant gonorrhoea in two South African cities and to investigate the association between the isolation of ciprofloxacin-resistant Neisseria gonorrhoeae and the HIV serostatus of patients. Methods: Gonococci were cultured from endourethral swabs taken from consecutive men with urethritis attending clinics in Johannesburg and Cape Town. Minimum inhibitory concentrations (MIC) for ciprofloxacin and ceftriaxone were determined with E-tests. Isolates with a ciprofloxacin MIC of 1 mg/l or greater were defined as resistant and isolates with a ceftriaxone MIC of 0.25 mg/l or less were defined as susceptible. Rapid tests were used to screen and confirm the presence of HIV antibodies. Survey data from 2004 were used as a baseline to assess trends in gonococcal resistance to ciprofloxacin. Results: In 2004, the prevalence of ciprofloxacin resistance was 7% in Cape Town and 11% in Johannesburg. In 2007, 37/139 (27%) Cape Town isolates and 47/149 (32%) Johannesburg isolates were resistant to ciprofloxacin; in comparison with 2004 data, this represents 2.9-fold and 1.9-fold increases, respectively. All isolates were fully susceptible to ceftriaxone. There was a significant association between HIV seropositivity and the presence of ciprofloxacin-resistant gonorrhoea among patients (p = 0.034). Conclusions: Johannesburg and Cape Town have witnessed significant rises in the prevalence of ciprofloxacin-resistant gonorrhoea among men with urethritis. The resistant phenotype is linked to HIV seropositivity. There is now an urgent need to change national first-line therapy for presumptive gonococcal infections within South Africa.

Prevalence and associations of genital ulcer and urethral pathogens in men presenting with genital ulcer syndrome to primary health care clinics in South Africa.

Background This study aimed to determine the prevalence of genital ulcer and urethral pathogens, as well as their association with clinical features, in men with genital ulcer disease (GUD) enrolled in a clinical trial. Methods Clinical data were collected by questionnaire. Ulcer swabs were tested for herpes simplex viruses (HSV-1/2), Treponema pallidum, Haemophilus ducreyi, and Chlamydia trachomatis L1-L3. First-pass urine was tested for urethral pathogens, namely Neisseria gonorrhoeae, C. trachomatis, Trichomonas vaginalis, and Mycoplasma genitalium. Pathogens were detected by real-time molecular assays. Blood was tested for HIV, HSV-2, and syphilis-associated antibodies. Pathogens and clinical associations were investigated using the &khgr;2 test. Results A total of 615 men with GUD were recruited. Herpes simplex virus (HSV-1, 4.2%; HSV-2, 98.2%) and bacterial pathogens were detected in 451 (73.6%) and 48 (7.8%) of genital ulcers, respectively. Human immunodeficiency virus, HSV-2, and treponemal antibodies were detected in 387 (62.9%), 434 (70.6%), and 141 (23.0%) men, respectively, whereas 54 men (8.8%) were rapid plasmin reagin (RPR) seropositive. A total of 223 urethral infections were diagnosed in 188 men (30.6%), including 69 (11.2%) M. genitalium, 64 (10.4%) T. vaginalis, 60 (9.8%) C. trachomatis, and 30 (4.9%) N. gonorrhoeae infections. Dysuria was reported by 170 men (27.6%), and 69 men (11.5%) had urethral discharge on examination. Urethral pathogens were detected in 102/409 (24.9%) men without these clinical features. Conclusions Herpes accounted for most GUD cases and urethral pathogen coinfections were common. Erythromycin, dispensed to treat infrequent chancroid and lymphogranuloma venereum cases, provided additional treatment of some asymptomatic urethral pathogens. Additional antibiotics would be required to treat asymptomatic trichomoniasis and gonorrhea.

Comprehensive global genome dynamics of Chlamydia trachomatis show ancient diversification followed by contemporary mixing and recent lineage expansion

Chlamydia trachomatis is the worlds 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 pathogens 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.

The detection of urethritis pathogens among patients with the male urethritis syndrome, genital ulcer syndrome and HIV voluntary counselling and testing clients: should South Africa’s syndromic management approach be revised?

Objectives: To determine the prevalence of urethritis pathogens amongst male symptomatic urethritis (MUS) patients, genital ulcer (GUS) patients without urethritis symptoms and men requesting HIV testing at a voluntary counselling and testing (VCT) clinic. Methods: A prospective study was conducted in Johannesburg, South Africa. Men from the three groups were screened for urethritis pathogens using molecular tests. Culture for Neisseria gonorrhoeae and, initially, trichomoniasis was performed. Antimicrobial susceptibility testing was undertaken for ciprofloxacin on all gonococcal isolates; ciprofloxacin resistant isolates were screened for ceftriaxone resistance. Results: 664 participants were recruited (438 MUS, 76 GUS and 158 VCT) over 2 years. Gonorrhoea was detected in 62.3% MUS, 15.8% GUS and 3.2% VCT participants. Chlamydial infection was detected in 19.3% MUS, 13.2% GUS and 8.2% VCT participants. Trichomoniasis was detected in 4.9% MUS, 19.7% GUS and 3.8% VCT participants. Mycoplasma genitalium infection was detected in 14.4% MUS, 13.2% GUS and 7.0% VCT participants. Ciprofloxacin resistance increased from 13.0% in the first year to 26.3% in the second year; all resistant isolates were susceptible to ceftriaxone. Conclusions: Urethritis pathogens, including Trichomonas vaginalis, should be covered in syndromic management treatment of genital ulcers in the absence of clinical urethritis. Consideration should be given to adding metronidazole to existing MUS treatment. Ciprofloxacin can no longer be relied upon to treat presumptive gonococcal infections in South Africa.

Trends and associations of Trichomonas vaginalis infection in men and women with genital discharge syndromes in Johannesburg, South Africa

Objectives To better understand the epidemiology of Trichomonas vaginalis infection, we investigated the association between T vaginalis and demographic, clinical, microbiological and behavioural characteristics of patients presenting with genital discharges to a primary healthcare clinic in Johannesburg, South Africa. Methods During six annual surveys (2007–2012), 1218 cases of male urethral discharge syndrome and 1232 cases of vaginal discharge syndrome were consecutively recruited. Diagnostic methods included nucleic acid amplification (Neisseria gonorrhoeae, Chlamydia trachomatis, T vaginalis and Mycoplasma genitalium), microscopy (bacterial vaginosis and Candida) and serology (Treponema pallidum, herpes simplex virus type 2 (HSV-2) and HIV). Logistic regression analyses and χ2 tests were used to identify predictors of T vaginalis infection. Results The prevalence of T vaginalis decreased from 2007 to 2012 (men from 13.4% to 4.8%; women from 33.8 to 23.1%). Overall, 74 (6.1%) men and 291 (23.6%) women were T vaginalis positive, with the highest prevalence in those aged ≥40 years (men 13.6%; women 30.9%). T vaginalis infection occurred more often in pregnant women (adjusted OR (aOR) 2.67; 95% CI 1.29 to 5.54) and in women with serological evidence of T pallidum (aOR 1.63; 95% CI 1.08 to 2.45) or HSV-2 infections (aOR 1.75; 95% CI 1.16 to 2.64). T vaginalis infection occurred less often in men with coexistent gonorrhoea (aOR 0.35; 95% CI 0.21 to 0.57) and in women with either bacterial vaginosis (aOR 0.60; 95% CI 0.44 to 0.82) or Candida morphotypes (OR 0.61; 95% CI 0.43 to 0.86). Conclusions Although the prevalence of T vaginalis infection has decreased over time, it remains an important cause of genital discharge in South Africa, particularly in older patients and pregnant women.

The burden of asymptomatic sexually transmitted infections among men in Carletonville, South Africa: implications for syndromic management

Objectives: To determine the prevalence of sexually transmitted infections (STI) and HIV among men attending an outreach STI/HIV screening service in informal settlements in South Africa over a 3-month period. Methods: A mobile clinic van was utilised to offer men: (1) urine screening for gonorrhoea, trichomoniasis, chlamydial and Mycoplasma genitalium infections; (2) serological screening for syphilis and herpes simplex virus type 2 (HSV-2); (3) on-site HIV voluntary counselling and testing. Urethritis pathogens were detected by molecular methods. HIV serostatus was determined using rapid tests. Demographic, sexual behaviour and clinical data were recorded on a nurse-administered questionnaire. Statistical analysis utilised the χ2 test. Results: 309 men attended the service; 304 (98%) requested serological screening for syphilis and HSV-2, 301 (97%) underwent urine-based screening for urethritis pathogens and 269 (87%) had an HIV test. Over 90% of men were asymptomatic for STI. Gonorrhoea was more prevalent in the symptomatic group (p<0.001); there were no significant differences in the prevalence of other urethritis pathogens between the groups. The total number of infections with each urethritis pathogen was highest in the asymptomatic group with twice as much gonorrhoea, 25 times as much chlamydial infection, six times as much trichomoniasis and nine times as much M genitalium infection compared with the symptomatic group. The overall HIV prevalence among clinic attendees was 29.7%. Conclusions: The uptake of both STI and HIV testing was high among men attending the service. The relatively high burden of both STI and HIV among the male clinic attendees has implications for the transmission of HIV.