Ulcer Specimens Research Articles

Common Pathogens Isolated from Infected Diabetic Foot Ulcers at King Abdulaziz University Hospital, Saudi Arabia: A Retrospective Study

Objectives: To determine the common pathogens isolated from DFI to administer appropriate antibiotic treatment, followed by surgical interventions. Methods: A retrospective study was performed on 260 patients who presented with diabetic foot at King Abdulaziz University Hospital, Jeddah, Saudi Arabia, from October 2014 to September 2020. All patients underwent swabs and tissue culture for microbiological evaluation. Patient medical records were reviewed to collect demographic and clinical data, including Glycated Hemoglobin (HgA1C), Diabetes Mellitus (DM) type, duration of diabetes, swab, tissue culture, type of surgery, and type of isolated organism. Results: Escherichia coli was the most common organism isolated from the feet of diabetic patients, followed by Staphylococcus aureus, Klebsiella pneumoniae, Streptococcus agalactiae, and Pseudomonas aeruginosa. Amputations were significantly higher in patients who did not have Staphylococcus aureus or Pseudomonas aeruginosa infection, with no significant relationship between amputation and any other isolated organisms. A significant negative correlation between patient age and HgA1C level was discovered, as well as a significant positive correlation between HgA1C and the number of minor amputations. Patients with major and minor amputations had a significantly higher percentage of non-Multidrug Resistance (MDR), whereas patients with MDR had a significantly higher rate of infection with Klebsiella pneumoniae, Escherichia coli, Pseudomonas aeruginosa, or Staphylococcus aureus. Conclusion: DM is a metabolic syndrome that affects all the body systems and impacts both morbidity and mortality. The most common organism isolated from the feet of diabetic patients was Escherichia coli, followed by Staphylococcus aureus. Ulcer specimens should be collected for culture and identification of causative organisms. Preventive measures such as good glycemic control, appropriate foot care, targeted antibiotic therapy, and patient education can reduce the incidence of amputation.

Inhibition of USP7 suppresses advanced glycation end-induced cell cycle arrest and senescence of human umbilical vein endothelial cells through ubiquitination of p53.

Diabetes mellitus is a n arising public health concern, and diabetic foot is one of the most common complications of diabetes. Current management for diabetic foot cannot reach optimal remission. In this study, we aim to explore the mechanism underlying the pathogenesis of diabetic foot and provide novel strategies for the treatment of diabetic foot. A total of 10 normal skin tissues and 20 diabetic foot ulcer specimens are collected. Cell proliferation is determined by CCK-8 assay. Cell cycle is determined by flow cytometry, and cell senescence is evaluated by β-galactosidase staining. Co-immunoprecipitation assay is used to explore the interaction between USP7 and p53. Advanced glycation end products (AGEs) are used to establish diabetic cell model, and streptozotocin (STZ) is used to establish diabetic rat model. Our results showed that USP7 expression is increased in diabetic foot ulcer and in human umbilical vein endothelial cells (HUVECs) after treatment with AGEs. Inhibition of USP7 can reduce cell cycle arrest and cell senescence in HUVECs. Moreover, USP7 can interact with p53 and promote its expression through mediating its deubiquitination. Knockdown of p53 can reverse USP7-mediated cell cycle arrest and cell senescence in HUVECs. In diabetic rats, HBX 41108, the specific inhibitor of USP7, can significantly accelerate wound healing. Our study reveals that the inhibition of USP7 can suppress AGEs-induced cell cycle arrest and cell senescence of HUVECs through promoting p53 ubiquitination. USP7 is a potential target for the treatment of diabetic foot ulcers.

The First Case of Monkeypox in the Republic of Korea

A rapid outbreak of monkeypox is ongoing in non-endemic countries since May 2022. We report the first case of monkeypox in the Republic of Korea. This occurred in a 34-year-old male patient who traveled to Europe in June 2022. On the day of his return to the Republic of Korea (June 21, 2022), the patient presented with a genital lesion. The results of the monkeypox real-time polymerase chain reaction tests were positive in the penile ulcer, oropharyngeal and nasopharyngeal specimens. The patient subsequently developed fever and skin rash after hospital admission. Careful history taking along physical examination should be conducted in the patients who have epidemiologic risk factors for monkeypox. Moreover, appropriate specimens should be obtained from lesions and tested for the monkeypox virus.

Aktivitas Antibakteri Ekstrak dan Fraksi dari Ekstrak Metanol Daun Manggis (Garcinia mangostana) pada Bakteri Penyebab Ulkus Diabetik

Diabetes mellitus (DM) is a degenerative disease due to metabolic disorders with acute to chronic hyperglycemic conditions. The complication of this disease is peripheral neuropathy exacerbated by bacterial infection, causing ulcers (sores) on the feet, hands, knees, back, and other parts of the body. The mangosteen plant (Garcinia mangostana) is empirically believed to have antidiabetic and cure diabetic ulcers. This study aims to compare the antibacterial activity of extracts and fractions of methanol extract of mangosteen leaves at concentrations of 20%, 40%, 60%, 80%, and 100% against the bacteria causing diabetic ulcers in vitro. The research began with the maceration of mangosteen leaf powder extraction using methanol as a solvent. The fractions were obtained by the vacuum liquid chromatography (KCV) method. The bacteria that cause diabetic ulcers are obtained from ulcer specimens of diabetic ulcer patients. Antibacterial activity of extracts and fractions at 5 concentration series was tested using the disc diffusion method with 10% DMSO negative control and positive controls, namely Amoxicillin, Ampicillin, Ciprofloxacin, Cefotaxime, Gentamycin, Erythromycin, and Doxycycline. Antibacterial activity was expressed by the large area of the inhibition zone on the media inoculated with ulcer bacteria and incubated for 24 hours at 370 C. The results showed that there were 5 dominant bacteria found in the diabetic ulcer specimens, namely Escherichia spp, Klebsiella spp, Staphylococcus spp, salmonella spp, and Shigella spp. The results of the antibacterial sensitivity test showed that ciprofloxacin had the largest area of inhibition zone compared to Amoxicillin, Ampicillin, Cefotaxime, Gentamycin, Erythromycin, and Doxycycline so that Ciprofloxacin could be used as a positive control. The results of the antibacterial activity test for the methanol extract of mangosteen leaves with a concentration of 100% produced the largest inhibition zone, namely 27.10 mm ± 0.1 compared to methanol extract, fractions A, B, and C in all concentration series. However, the inhibition zone of the 100% concentration of mangosteen leaves methanol extract was smaller than the positive control of ciprofloxacin 5 μg / disk, which was 35.01 mm ± 0.1.

The changing epidemiology of genital ulcer disease in South Africa: has donovanosis been eliminated?

ObjectivesWe used an in-house molecular assay for the detection of Klebsiella granulomatis in ulcer specimens collected over a 12-year surveillance period in order to determine whether a diagnosis of donovanosis...

Successful isolation of Treponema pallidum strains from patients' cryopreserved ulcer exudate using the rabbit model.

Clinical isolates of Treponema pallidum subspecies pallidum (T. pallidum) would facilitate study of prevalent strains. We describe the first successful rabbit propagation of T. pallidum from cryopreserved ulcer specimens. Fresh ulcer exudates were collected and cryopreserved with consent from syphilis-diagnosed patients (N = 8). Each of eight age-matched adult male rabbits were later inoculated with a thawed specimen, with two rabbits receiving 1.3 ml intratesticularly (IT), and six receiving 0.6 ml intravenously (IV) and IT. Monitoring of serology, blood PCR and orchitis showed that T. pallidum grew in 2/8 rabbits that were inoculated IV and IT with either a penile primary lesion specimen (CDC-SF003) or a perianal secondary lesion specimen (CDC-SF007). Rabbit CDC-SF003 was seroreactive by T. pallidum Particle Agglutination (TP-PA) and Rapid Plasma Reagin (RPR) testing, PCR+, and showed orchitis by week 6. Euthanasia was performed in week 7, with treponemal growth in the testes confirmed and quantified by qPCR and darkfield microscopy (DF). Serial passage of the extract in a second age-matched rabbit also yielded treponemes. Similarly, rabbit CDC-SF007 showed negligible orchitis, but was seroreactive and PCR+ by week 4 and euthanized in week 6 to yield T. pallidum, which was further propagated by second passage. Using the 4-component molecular typing system for syphilis, 3 propagated strains (CDC-SF003, CDC-SF007, CDC-SF008) were typed as 14d9f, 14d9g, and 14d10c, respectively. All 3 isolates including strain CDC-SF011, which was not successfully propagated, had the A2058G mutation associated with azithromycin resistance. Our results show that immediate cryopreservation of syphilitic ulcer exudate can maintain T. pallidum viability for rabbit propagation.

Trends in the relative prevalence of genital ulcer disease pathogens and association with HIV infection in Johannesburg, South Africa, 2007-2015.

BackgroundIn South Africa, treatment of genital ulcer disease (GUD) occurs in the context of syndromic management. GUD aetiological studies have been conducted in Johannesburg since 2007. We report on GUD pathogen prevalence, sero-prevalence of STI co-infections and aetiological trends among GUD patients presenting to a community-based primary healthcare facility in Johannesburg over a 9-year period.Methods and findingsGUD surveys were conducted from January to April each year. Consecutive genital ulcers were sampled from consenting adults. Swab-extracted DNA was tested by multiplex real-time PCR assays for herpes simplex virus (HSV), Treponema pallidum (TP), Haemophilus ducreyi (HD) and Chlamydia trachomatis (CT). HSV-positive DNA extracts were further subtyped into HSV-1 and HSV-2 using a commercial PCR assay; CT-positive extracts were tested with an in-house PCR assay specific for serovars L1-L3 (lymphogranuloma venereum). Sera were tested for HIV, HSV-2, and syphilis co-infections. Giemsa-stained ulcer smears were screened for Klebsiella granulomatis by microscopy. Data were analysed with STATATM version 14. Of 771 GUD specimens, 503 (65.2%) had a detectable pathogen: HSV 468 (60.7%); TP 30 (3.9%); CT L1-3 7 (0.9%); HD 4 (0.5%). No aetiological agents were detected in 270 (34.8%) ulcer specimens. Seroprevalence rates were as follows: HIV 61.7%; HSV-2 80.2% and syphilis 5.8%. There was a strong association between GUD pathogen detection and HIV seropositivity (p < 0.001); 68% of cases caused by HSV were co-infected with HIV. There was a significant decline in the relative prevalence of ulcer-derived HSV over time, predominantly from 2013–2015 (p-value for trend = 0.023); and a trend towards a decrease in the HIV seropositivity rate (p-value for trend = 0.209).ConclusionsHSV remains the leading cause of pathogen-detectable GUD in South Africa. The prevalence of HIV co-infection among GUD patients is high, underlining the importance of linkage to universal HIV testing and treatment in primary healthcare settings.

The Etiology of Genital Ulcer Disease and Coinfections With Chlamydia trachomatis and Neisseria gonorrhoeae in Zimbabwe: Results From the Zimbabwe STI Etiology Study

In many countries, sexually transmitted infections (STIs) are treated syndromically. Thus, patients diagnosed as having genital ulcer disease (GUD) in Zimbabwe receive a combination of antimicrobials to treat syphilis, chancroid, lymphogranuloma venereum (LGV), and genital herpes. Periodic studies are necessary to assess the current etiology of GUD and assure the appropriateness of current treatment guidelines. We selected 6 geographically diverse clinics in Zimbabwe serving high numbers of STI cases to enroll men and women with STI syndromes, including GUD. Sexually transmitted infection history and risk behavioral data were collected by questionnaire and uploaded to a Web-based database. Ulcer specimens were obtained for testing using a validated multiplex polymerase chain reaction (M-PCR) assay for Treponema pallidum (TP; primary syphilis), Haemophilus ducreyi (chancroid), LGV-associated strains of Chlamydia trachomatis, and herpes simplex virus (HSV) types 1 and 2. Blood samples were collected for testing with HIV, treponemal, and nontreponemal serologic assays. Among 200 GUD patients, 77 (38.5%) were positive for HSV, 32 (16%) were positive for TP, and 2 (1%) were positive for LGV-associated strains of C trachomatis. No H ducreyi infections were detected. No organism was found in 98 (49.5%) of participants. The overall HIV positivity rate was 52.2% for all GUD patients, with higher rates among women compared with men (59.8% vs 45.2%, P < 0.05) and among patients with HSV (68.6% vs 41.8%, P < 0.0001). Among patients with GUD, 54 (27.3%) had gonorrhea and/or chlamydia infection. However, in this latter group, 66.7% of women and 70.0% of men did not have abnormal vaginal or urethral discharge on examination. Herpes simplex virus is the most common cause of GUD in our survey, followed by T. pallidum. No cases of chancroid were detected. The association of HIV infections with HSV suggests high risk for cotransmission; however, some HSV ulcerations may be due to HSV reactivation among immunocompromised patients. The overall prevalence of gonorrhea and chlamydia was high among patients with GUD and most of them did not meet the criteria for concomitant syndromic management covering these infections.

Microbial Diversity of Genital Ulcers of HSV-2 Seropositive Women

We measured the microbial community structure of genital ulcers in women. Swabs from clinically detected ulcers were tested for HSV-2 and Treponema pallidum by polymerase chain reaction (PCR). HSV-2 and T. pallidum were detected by serum antibody testing. Microbial community structure was characterized by high-throughput 16 s rRNA gene amplicon sequencing. Multiple group testing and Elastic net and Lasso regressions identified taxa associated with differences in factors of interest. Among 49 ulcer specimens from 49 HSV-2 seropositive women, by PCR HSV-2 was recovered from 28 (57%) specimens and T. pallidum from none; one woman showed serologic evidence of syphilis. Overall, 63% of women were HIV-positive and 49% had an uncircumcised male sex partner. By both multiple group testing and regression, Porphyromonas (FDR p-value = 0.02), Prevotella (FDR p-value = 0.03), Anaerococcus (FDR p-value = 0.07), and Dialister (FDR p-value = 0.09) were detected at higher relative abundance in HSV-2 PCR-positive than negative ulcers. The presence of HSV-2 in a lesion was associated with presumed bacterial agents of Bacterial vaginosis. Differences in bacterial communities may contribute to HSV-2 ulcer pathogenesis, severity, or prolonged healing. If these results are confirmed, future studies may consider the influence of BV treatment on women’s GUD and HSV-2 incidence and recurrence.

A Comparative Analysis of Polymerase Chain Reaction and Direct Fluorescent Antibody Test for Diagnosis of Genital Herpes.

Objective:To compare laboratory tests that can simultaneously detect and type herpes simplex virus (HSV) directly from the genital ulcer specimens in clinically suspected cases of genital herpes.Materials and Methods:A study was conducted over 10 months and 44 adult male and female patients clinically suspected with genital herpes were recruited. Genital ulcer swab specimens were subjected to glycoprotein-G gene-based conventional polymerase chain reaction (PCR) and commercially available direct fluorescent antibody (DFA) test and the results were compared.Results:PCR for HSV was positive in 82% (36/44) cases. DFA was positive in 68.2% (30/44) cases. There was 100% agreement between HSV types detected by DFA and PCR. The strength of agreement between the results was better in primary genital herpes than recurrent cases.Conclusion:PCR was found to be better in the detection of HSV in recurrent genital herpes patients. It is a better modality, especially when genital herpes clinically presents with ulcerative or crusted lesions, and is also a cheaper alternative as compared to DFA.

Performance of the HSV OligoGen kit for the diagnosis of herpes simplex virus type 1 and 2

PCR methods are nowadays between the most rapid and sensitive methods for screening and diagnosing herpes simplex virus (HSV) type 1 and 2. The aim of this study was to analyze the reliability, accuracy, and usefulness of the new assay HSV OligoGen kit in comparison with the Roche LightCycler HSV ½ Qual Kit assay for the detection of HSV in clinical samples. For this analysis, a prospective study was designed for detection of HSV-1 and HSV-2 including 110 ulcer specimens, 48 urine, 48 endocervical, 43 cerebral spinal fluids, 4 urethral and 3 pharyngeal swabs that were sent from a regional STI clinic or an Intensive Clinical Unit, both in Seville, Spain. In comparison to the Roche LightCycler HSV ½ Qual Kit assay, sensitivity, specificity, positive and negative predicative values, and kappa value for HSV detection using the HSV OligoGen kit were 96.2%, 100%, 100%, 98.3%, and 0.97 for HSV-1, respectively. For HSV-2, the corresponding values were 98.3%, 100%, 100%, 99.5%, and 0.98, respectively. Statistical data obtained in this study confirms the usefulness and reliable results of this new assay.

Syphilis epidemiology in 1994-2013, molecular epidemiological strain typing and determination of macrolide resistance in Treponema pallidum in 2013-2014 in Tuva Republic, Russia.

The incidence of syphilis in the Tuva Republic (geographical centre of Asia), Russia has been exceedingly high historically. No detailed examinations and no molecular investigations of Treponema pallidum strains transmitted in the Tuva Republic, or in general, in Russia, were published internationally. We examined the syphilis epidemiology in 1994-2013, and the molecular epidemiology and macrolide resistance in T. pallidum strains in 2013-2014 in the Tuva Republic. Among 95 mainly primary or secondary syphilis patients, the arp, tpr, tp0548 and 23S rRNA genes in 85 polA gene-positive genital ulcer specimens were characterized. The syphilis incidence in Tuva Republic peaked in 1998 (1562), however declined to 177 in 2013. Among the 70 (82%) completely genotyped specimens, six molecular strain types were found. Strain type 14d/f accounted for 91%, but also 14c/f, 14d/g, 14b/f, 14i/f, 9d/f, and 4d/f were identified. Two (2.4%) specimens contained the 23S rRNA A2058G macrolide resistance mutation. This is the first internationally published typing study regarding T. pallidum in Russia, performed in the Tuva Republic with the highest syphilis incidence in Russia. The two molecular strain types 4d/f and 9d/f have previously been described only in Eastern and Northern China and for the first time, macrolide-resistant syphilis was described in Russia.

P05.03 Performance of two enzyme and one strip immune assay for the detection of igm anti-treponema pallidumantibodies

Background The detection of anti- Treponema pallidum (Tp) IgM may be useful in the diagnosis of very early syphilis, re-infection and in the assessment of the newborn. We aimed to evaluate two enzyme and one strip immune assay for the detection of anti-Tp IgM in blood specimens. Methods A total of 33 blood specimens collected from 16 patients presenting with a syphilitic ulcer were analysed. The specimens were tested employing: Rapid Plasma Reagin; Treponema Pallidum Particle Agglutination (TPPA); recomLine Treponema IgM (Mikrogen Diagnostik); recomWell Treponema IgM (Mikrogen Diagnostik); Anti- Treponema pallidum ELISA -IgM (Euroimmun). Ulcer specimens were tested by qPCR. A specimen was considered positive for anti-Tp IgM when both TPPA and IgM, obtained with any of the assays, were positive. Specimens containing antibodies against Epstein Barr Virus, Leptospira sp, Borrelia sp, Plasmodium sp, Herpes Simplex Virus 1 and 2 and specimens from pregnant women were tested with the three IgM assays to determine cross reactivity. Results Anti-Tp IgM was found in 22/33 specimens. The recomLine assay detected 17/22 (77%), the recomWell 15/22 (68%) and the Euroimmun 14/22 (64%) positive specimens. None of the three assays provided false positive results. Borderline results were obtained with the recomLine (N = 6) and the recomWell (N = 2) assay. Considering the borderline results as positive increased the detection rate to 95% for the recomLine and to 72% for the recomWell. However, two and one false positive result was then obtained with the recomLine and recomWell assay, respectively. Tp DNA was detected in 13/16 (81%) ulcer specimens. The Euroimmun assay cross reacted with malaria antibodies in one sample. Conclusions None of the three assays showed to be highly sensitive. Surprisingly, the highest sensitivity was obtained with the recomLine assay. The sensitivity improved by defining the borderline results as positive but decreased the specificity. Disclosure of interest The IgM assays were kindly provided by the two companies, Euroimmun and Mikrogen Diagnostik. No other funding was received for this work.

P09.24 The aetiology of genital ulcer disease and association with hiv infection in zimbabwe

Background In many countries, sexually transmitted infections (STI) are treated syndromically. Thus, patients diagnosed with genital ulcer disease (GUD) in Zimbabwe receive antimicrobials that cover infections with Treponema pallidum (TP: benzathine penicillin), Haemophilus ducreyi (HD: erythromycin) and herpes simplex virus (HSV: acyclovir). However, periodic surveys into the aetiology of GUD are necessary to inform treatment guidelines. Methods For this study, we recently completed enrollment of 200 patients with GUD at 6 clinics in Zimbabwe. To date, test procedures have been completed for patients enrolled at Harare clinics (N = 70). Ulcer specimens were obtained for testing using a validated multiplex polymerase chain reaction assay (M-PCR, National Institute of Communicable Diseases, Johannesburg) for TP (primary syphilis), HD (chancroid), HSV (genital herpes) and Chlamydia trachomatis strains associated with lymphogranuloma venereum (CT-LGV). Blood samples were collected for HIV testing by a standard rapid HIV test algorithm (First Response™ followed by Alere HIV 1/2™) and considered positive when reactive on both. Results To date, M-PCR testing is complete for all 70 patients with GUD recruited from the Harare clinics (38 men and 32 women). Of these, 17 (24.2%) were positive for HSV, 8 (11.4%) were positive for TP, and 1 was positive for CT-LGV. No cases of chancroid were detected. The overall HIV positivity rate was 43.1%. HIV rates were higher among patients with HSV (62.5%, p = 0.07, Chi Square Test) and TP (87.5%, p Conclusions Genital herpes was the most common cause of GUD in our survey, followed by primary syphilis. The association of HIV infection with HSV and TP suggests high risk for co-transmission; however some HSV ulcerations among HIV-infected patients may be the result of HSV reactivation among immunocompromised patients. Our study methods and data should be relevant for other countries using a syndromic management to STI control.

Distribution and antimicrobial susceptibility profile of extended-spectrum β-lactamase-producing Proteus mirabilis strains recently isolated in Japan

Here we report on the prevalence of extended-spectrum β-lactamase (ESBL)-producing Proteus mirabilis from a nationwide antimicrobial resistance survey in different geographical regions of Japan. A total of 799 P. mirabilis isolates recovered between July 2009 and June 2010 from 314 healthcare facilities were characterised according to ESBL production, source, location and antimicrobial susceptibility pattern. ESBL production was found in 364 (45.6%) of the isolates, among which 354 (97.3%) produced CTX-M-2 group β-lactamases. Of the 349 ESBL-producing isolates in which the inpatient or outpatient status of the source was known, 324 (92.8%) were from inpatients and 25 (7.2%) were from outpatients (P<0.05). Results of pulsed-field gel electrophoresis (PFGE) analysis performed on 66 of the ESBL-producers generated a distribution of PFGE patterns into 21 groups. Genetic relatedness was seen among isolates within a region, which is consistent with horizontal transmission. With respect to the frequency of ESBL-producers by specimen source, 12/14 (85.7%) central venous catheter specimens yielded ESBL-producing P. mirabilis compared with 159/405 (39.3%), 119/209 (56.9%), 42/77 (54.5%) and 20/49 (40.8%), respectively, for isolates from urine, sputum, decubitus ulcer and wound specimens. Among the ESBL-producers, non-susceptibility to ciprofloxacin was found in 74.2% of the ESBL-producing isolates compared with 17.7% of the ESBL-non-producing isolates. These results show that approximately one-half of the P. mirabilis isolates from clinical specimens in Japan are ESBL-producers and that the potential for concomitant fluoroquinolone resistance must also be considered.

P5.068 Role of Type Specific Herpes Simplex Virus Serology in Diagnosis of Primary and Recurrent Genital Herpes- a Study in the Indian Population

Background Type specific serologic tests for herpes simplex virus (HSV)-1&2 are used for sero-epidemiological studies. Its early application has also been shown to be of benefit in testing for primary & recurrent infections in diagnosis and patient management. The present study was conducted to evaluate the role of type specific HSV-1&2 antibody detection as a diagnostic modality in patients with clinically suspected genital herpes. Methods Sera obtained from 44 patients with clinically suspected genital herpes attending the STD clinic of our institution over last 7 months were tested for type specific HSV-1&2 IgG antibody by glycoprotein G based ELISA (HerpeSelect 1&2, Focus Diagnostics, USA). Direct Fluorescent Antibody testing (DFA) (BIO-RAD laboratories, USA) was used for identification and typing of HSV-1& 2 in genital ulcer specimens in all cases. Results There were 21 cases of primary & 23 cases of recurrent genital ulceration. DFA was positive in 68% samples (43% only HSV-2, 9% only HSV-1, 16% both HSV-1&2). On comparison with DFA, the sensitivity, specificity, positive and negative predictive value of HSV-2 IgG ELISA in primary herpetic ulcer was 8%, 75%, 33%, 33%, while it was 79%, 89%, 92%, 73% in recurrent genital ulceration. The sensitivity, specificity, positive predictive and negative predictive values of HSV-1 IgG ELISA in primary herpetic ulcer was 50%, 67%, 37.5%, 77%, while it was 80%, 39%, 27%, and 87.5% in recurrent genital ulceration. Conclusion Our results highlight the importance of HSV-2 IgG detection in strengthening the diagnosis of recurrent HSV-2 disease, while absence of HSV-2 IgG antibody helps in excluding genital herpes as a likely cause of recurrent genital ulceration. The identification of HSV-1 IgG antibody may not be useful for diagnosis in patients of genital ulcer disease highlighting the need for typing HSV-1 strains from the genital lesion.

Genetic Characterization of Herpes Simplex Virus Type 2 UL23 Thymidine Kinase in Johannesburg, South Africa

Herpes simplex virus type 2 (HSV-2) is currently the leading cause of Genital Ulcer Disease (GUD) both globally and within South Africa. HSV-2 infections are most often treated with acyclovir (ACV), a guanosine nucleoside analogue that requires phosphorylation by virus-encoded thymidine kinase (TK). Resistance to ACV is mainly due to mutations in the viral UL23 gene that codes for TK. ACV was added as part of the first-line syndromic management treatment algorithm for GUD in South Africa in late 2008. In order to assess the prevalence of TK-associated ACV resistance among HSV-2 virions detected in genital ulcer specimens, pre- and post-introduction of ACV, we amplified and fully sequenced the UL23 gene of 254 HSV-2 positive specimens obtained from participants in GUD aetiological surveys conducted between 2007 and 2011 in Johannesburg, South Africa. We identified 63 nucleotide mutations in the UL23 genes analysed, that resulted in 30 silent mutations and 32 amino acid changes. A large proportion (41%) of these amino acid changes were due to previously described natural polymorphisms that occur in both sensitive and resistant HSV strains. In addition, we identified 19 unknown amino acid changes in 30 samples that have not been described before. All mutations detected were outside the recognised TK conserved domains where ACV resistance mutations typically occur. No frameshift mutations or mutations causing stop codons were identified in those UL23 genes analysed. Importantly, no evidence was found of known ACV resistance mutations in HSV-2 following the addition of ACV as first-line therapy for GUD.

Macrolide resistance testing and molecular subtyping of Treponema pallidum strains from southern Africa

ObjectivesTo determine whether the 23S ribosomal RNA (rRNA) A2058G and A2059G mutations that confer macrolide resistance are present among southern African strains of Treponema pallidum and to determine their subtype...

P1-S5.10 Acute genital ulceration in South African Men: importance of age and sexual risk behaviours

BackgroundHerpes simplex virus (HSV) is the leading cause of genital ulcer disease (GUD) in Africa. Although some studies have examined the demographics and sexual behaviours of men with GUD, less...

P4-S3.01 Macrolide-resistance testing and molecular subtyping of Treponema pallidum strains from South Africa

ObjectivesTo determine whether the main 23S rRNA mutation (A2058G) that confers macrolide resistance inTreponema pallidumis present among DNA obtained from syphilitic ulcers in South Africa and to determine the strain subtype distribution using molecular methods.MethodsDNA was re-extracted from sixty ulcer swabs, previously shown to containT pallidumDNA by in-house real-time multiplex PCR assay, using a MagNA Pure Compact DNA extraction system (Roche, Germany). The genital ulcer specimens were collected in South Africa between 2005 and 2009 during either national microbiological surveillance activities or as part of large episodic acyclovir treatment trial (HSV4294). The re-extracted DNA was screened for the A2058G point mutation in the peptidyltransferase region of the 23S rRNA subunit using a rapid PCR-based restriction digest assay. Subtyping ofT pallidum, using a previously-published technique, was based on the molecular characterisation of two variable treponemal repeat genes, arp and tpr.ResultsMacrolide resistance profiles and the subtype distribution for all 60 DNA samples were obtained. The mean age of all participants was 27.4 years (range 18–43; SD 5.24) of which 55 (92%) were male and 5 (8%) were female. None of the samples analysed contained the 23S rRNA gene point mutation that confers macrolide resistance. A total of eight arp repeat sizes, 8 RFLP patterns and a combined total of 17 subtypes were identified in this study population. The most common subtypes were 14d (43%), followed by 17d (13%), 14b (7%), 22b (5%) and 23b (5%).ConclusionsThis is the first South African study to examine both macrolide-resistance profiles and the subtype distribution of syphilis strains using molecular techniques. Macrolide-resistantT pallidumstrains appear to be uncommon in South Africa compared to more developed countries. Our results indicated subtype 14d to be the predominant circulatingT pallidumstrain in South Africa and there seem to be a high degree of genetic heterogeneity within this population.