Nicoletta Banzola

Evaluation of the New Test VERSANT CT/GC DNA 1.0 Assay for the Detection of Chlamydia trachomatis and Neisseria gonorrhoeae in Urine Specimens

Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (GC) are the two most common sexually transmitted bacterial infections in developed countries. The purpose of the present study was evaluating a new system for CT/GC detection in urine specimens. A total of 700 urine specimens were obtained from patients attending the STD Outpatients Clinic of St. Orsola University Hospital, Bologna, Italy. Samples were tested by VERSANT® CT/GC DNA 1.0 Assay (Siemens Healthcare Diagnostics Inc., Tarrytown, NY), a multiplex Real‐Time PCR assay, for simultaneous CT/GC detection. Results obtained by VERSANT assay were compared with those obtained by culturing genital secretions of the same patients. Moreover, urine specimens testing positive in VERSANT assay were retested by in‐house PCR assays, used as confirmatory tests. VERSANT® CT/GC DNA 1.0 Assay performed with 99.4% and 99.2% of specificity for GC and CT detection, respectively, whereas sensitivity was 100% both for CT and GC. Culture methods were 100% specific, but far less sensitive than VERSANT assay. VERSANT® CT/GC DNA 1.0 Assay demonstrated to be a highly sensitive and specific technique for CT/GC detection.

Use of Dermasilk briefs in recurrent vulvovaginal candidosis: safety and effectiveness

Despite the generally excellent results achieved with fluconazole 150 mg weekly in recurrent vulvovaginal candidosis (RVVC), some patients with a long history of disease do not achieve complete resolution of symptoms following antimycotic treatment. It is thought that use of tight synthetic fabric underwear could be a significant factor in causing recurrence. We decided to compare underwear made of Dermasilk®, a pure fibroin fabric impregnated with a permanent antimicrobial protection, with a cotton placebo to see whether it could be a useful adjunctive tool in the management of RVVC. We recruited 96 women who had a long‐term history of RVVC and had not responded to oral antimycotics with complete satisfaction. The patients were randomly divided into two groups and instructed to use either white cotton placebo briefs or Dermasilk® briefs. Both groups were treated with fluconazole 150 mg once weekly for 6 months. After 6 months, the Dermasilk group showed a statistically significant greater decrease of itching, burning, erythema and a smaller number of recurrences than the cotton group. Our work suggests that Dermasilk® briefs could be a useful adjunctive tool in addition to antimycotic treatment to help relieve the discomfort of recurrent vulvovaginitis.

Carbon dioxide laser: first-line therapy in vulvar inflammatory linear verrucous epidermal nevus

Inflammatory linear verrucous epidermal nevus (ILVEN) is normally associated with the failure of topical and systemic treatments and with recurrences on interruption of therapy. Many physical approaches have been used, but they generally resulted in varying rates of recurrence and unacceptable scarring. We reported a case of ILVEN treated with a single session CO2 laser treatment. In our experience, CO2 laser was quick, easy, effective, and safe; we therefore believe that this approach should be considered as a first‐line surgical option in the treatment of genital ILVEN, particularly in cases of mucosal involvement

A Case of Reactive Arthritis Associated With Lymphogranuloma Venereum Infection in a Woman.

We report the first case of reactive arthritis associated with lymphogranuloma venereum (LGV) in an Italian human immunodeficiency virus-negative woman with urogenital and rectal Chlamydia trachomatis L2 serovar infection. The LGV-associated arthritis has to be considered even when classic symptoms of arthritis are missing and in case of asymptomatic or cryptic LGV localizations.

AIDS-related Kaposi’s sarcoma involving the genital and inguinal regions

with leukocytoclasis throughout the dermis (Fig. 2). Interestingly, the polymerase chain reaction (PCR) assay for HSV type 2 DNA was positive in the same specimen using Seeplex HSV2 ACE Detection (Seegene, Seoul, Korea). Cutaneous vasculitis is a common diagnosis in routine dermatology practice. Underlying autoimmune diseases, malignancy, drugs, and systemic vasculitis are often found to be etiological factors. Infections also play a major role, including streptococcus, staphylococcus, mycobacterium, and hepatitis B or C. But, with better techniques such as PCR and various immunodiagnostic techniques, looking at the characteristic antibodies for certain infections now offers us the ability to sort out and determine the true incidence of vasculitis related to the myriad infectious agents. As far as we know, leukocytoclastic vasculitis associated with HSV has not been previously reported. Although our patient did not have any history of herpetic infection, an unapparent HSV infection was implicated as the cause associated with vasculitis. This relationship of HSV and vasculitis is supported by an animal study involving ICR strain mice. The mice were inoculated with an HSV-1 solution, and 29.8% of 258 mice showed Behcet’s diseaselike symptoms that revealed vasculitis. Moreover, the unique plantar location of cutaneous vasculitis supports this etiological association, because the plantar area is known as a predilection site of erythema multiforme, in which hematogenous spread of HSV DNA from the infected lesion to the plantar area has already been proven, and the benign course but frequent recurrences in the presented case seem to be consistent with the natural course of mucosal HSV infections. Here we report a case of plantar leukocytoclastic vasculitis associated with HSV-2 infection proven by PCR assay. Although the specificity of a PCR assay is not high enough, HSV seems to be one of various agents that can cause leukocytoclastic vasculitis. We think the causality link has been overlooked by both patients and physicians due to the asymptomatic nature of HSV infection. More sophisticated laboratory techniques and subsequent reports are required to confirm the etiology of plantar leukocytoclastic vasculitis.

P3-S1.07 Chlamydia trachomatis serovar distribution and other sexually transmitted coinfections in subjects attending a STD outpatients clinic in Italy

Background Chlamydia trachomatis is the leading cause of bacterial sexually transmitted diseases (STDs) in industrialised countries. omp1 (ompA), the gene encoding the major outer membrane protein (MOMP), has been widely used for molecular epidemiology, because it contains four spaced variable domains. Methods A total of 1625 patients attending the STD Outpatients Clinic of St. Orsola University Hospital of Bologna, Italy were enrolled for this study. Each patient was clinically visited, bled in order to perform serological tests, than three urethral or endocervical swabs were obtained. Two swabs were cultured for the detection of C trachomatis and Neisseria gonorrhoeae, whereas the third was stored at −80°C. When a positive result was obtained by C trachomatis culture, the corresponding frozen sample was withdrawn, its DNA was extracted by VERSANT kPCR SP Module (Siemens Healthcare Diagnostics Inc.) and used as a template for omp1 gene fragment amplification. PCR products were purified and both strands were sequenced. Nucleotide sequences were compared to omp1 sequences using the BLAST search tool at the National Center for Biotechnology Information. The sequences were manually aligned using BioEdit (version 7.0.0) software. χ2 Test was used and a p value of <0.05 was considered statistically significant. Results C trachomatis was detected in 103 out of 1625 (6.3%) swabs by culture. Prevalence was significantly higher in men (p<0.01), with 60 positives out of 525 tested (11.4%), than in women (43/1100; 3.9%), as well as presence of clinical symptoms: 81.7% (49/60) of infected men and 44.2% of infected women (19/43) were symptomatic. Also prevalence of STD coinfections was significantly higher (p<0.01) in men (35/60; 58.3%) than in women (8/43; 18.6%). In our population the most common serovar was E, with a prevalence of 38.8%, followed by G (23.3%), F (13.5%), D/Da (11.6%), and J (4.8%). Statistically significant differences (p=0.042) in serovar prevalence between men and women were detected. Finally, significant differences (p=0.035) were detected when serovar distribution among patients with or without coinfection was studied: patients with an infection due to D/Da had the highest coinfection rate (75.0%), whereas coinfection rates among patients with serovars F, E, and G were 57.1%, 37.5%, and 29.2%, respectively see Abstract P3-S1.07 table 1. Abstract P3-S1.07 Table 1 Primary demographic, epidemiological, and clinical data and rates of infection with C trachomatis serovars for male and female patients Sex No (%) of patients Males Female p Value (χ2 test) Place of birth  Italy 42 (70.0) 22 (51.2) 0.052  Other 18 (30.0) 21 (48.8) Symptoms  Yes 49 (81.7) 19 (44.2) 0.000*  No 11 (18.3) 24 (55.8) N gonorrhoeae coinfection  Yes 27 (45.0) 3 (7.0) 0.000*  No 33 (55.0) 40 (93.0) T pallidum coinfection  Yes 5 (8.3) 2 (4.7) 0.696  No 55 (91.7) 41 (95.3) Human papillomavirus coinfection  Yes 6 (10.0) 3 (7.0) 0.592  No 54 (90.0) 40 (93.0) HIV coinfection  Yes 5 (8.3) 0 (0,0) 0.073  No 55 (91.7) 43 (100) C trachomatis serovar  B 0 (0,0) 2 (4.7) 0.042*  D/Da 10 (16.7) 2 (4.7)  E 24 (40.0) 16 (37.2)  F 11 (18.3) 3 (7.0)  G 10 (16.7) 14 (32.6)  H 0 (0.0) 2 (4.7)  I/Ia 2 (3.3) 0 (0.0)  J 2 (3.3) 3 (7.0)  K 1 (1.7) 1 (2.3) * Statistically significant (p<0.01). Conclusions The present study contributed to increase the knowledge on serovar distribution of C trachomatis in Italy.

Insights Into Vaginal Bacterial Communities and Metabolic Profiles of Chlamydia trachomatis Infection: Positioning Between Eubiosis and Dysbiosis

The vaginal microbiota plays a crucial role in maintaining the health and functioning of the female genital tract, preventing the colonization of urogenital pathogens and sexually transmitted infections. In this study, we characterized the vaginal bacterial communities and the metabolome associated to Chlamydia trachomatis infection (CT: 20 women), compared to healthy condition (H: 22 women) and bacterial vaginosis (BV: 19 women). A microarray-based tool (VaginArray), implemented with a real-time PCR for Gardnerella vaginalis, was used to determine the vaginal bacterial composition, whereas the metabolic profiles were assessed by a proton-based nuclear magnetic resonance (1H-NMR) spectroscopy. CT infection was characterized by bacterial and metabolic signatures similar to healthy condition, even though higher amounts of Lactobacillus iners, as well as depletion of some amino acids, biogenic amines, and succinate marked CT infection. Moreover, the frequency of Lactobacillus crispatus was higher in asymptomatic CT-positive patients than in women with CT-correlated symptoms. We also confirmed the marked differences in the microbiome and metabolome between healthy and BV-affected women. In conclusion, we highlighted microbial and metabolic peculiarities of the vaginal ecosystem in the case of CT infection, even though further studies are needed to understand if the observed alterations precede the infection onset or if the pathogen itself perturbs the vaginal environment.

P1.14 Evaluation of the aptima assays for the detection of bacterial sexually transmitted infections in a selected population of women

Introduction: Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG) and Mycoplasma genitalium (MG) represent the most common agents of bacterial sexually transmitted infections (STIs), worldwide. In women, uro-genital infections caused by these microorganisms are often asymptomatic and, left untreated, can lead to several sequelae. Nucleic acid amplification techniques (NAATs) have become the reference methods for the diagnosis, thanks to the suitability for different specimens and the outstanding sensitivity and specificity. The aim of this study was to assess the performance of Aptima Assays for CT, NG and MG detection in a group of selected women, by a head-to-head comparison with other NAATs. Moreover, an evaluation about the suitability for the Aptima assays with one of the most used swab collection device (E-Swab; Copan) was carried out. Methods Routinely, all the women attending the STI Outpatients Clinic of Sant’Orsola-Malpighi Hospital of Bologna (Italy) complaining of genital STI-related symptoms or reporting unsafe intercourse, are managed as follows. After a clinical visit, a sample of first-void urines and a vaginal swab collected in E-swab, are obtained for CT, NG and MG detection. A duplex real-time PCR (Versant CT/GC DNA 1.0 assay; Siemens) is used for CT and NG detection, while, MG presence is investigated by a home-made PCR, starting from the remaining eluate of Versant PCR plate. From January 2016, a total of 100 patients were selected and their samples were also tested with Aptima assays. Previously frozen samples were thawed and transferred to the suitable collection devices for Aptima assays: in particular, 2 ml of urines and 100 µl of vaginal E-swab were used. All the specimens were processed by Aptima Combo2® for CT and NG detection and by the Aptima® Mycoplasma genitalium assay for MG infection diagnosis. These assays were run on Panther system (Hologic). A comparison between the different molecular methods, stratified by type of sample and microorganism, was conducted. Results In the group of 100 women selected, 25 patients were positive for CT, 4 for NG and 6 for MG. One case of CT-NG and two cases of CT-MG co-infections were found. Interestingly, more than 50% of CT-positive women were completely asymptomatic. By the routine tests, all positive cases were simultaneously found both on the urine sample and on the vaginal swab, except for 3 CT, 1 NG and 1 MG infections, detected only on the vaginal swab. A complete concordance with Aptima assays, both for the type of sample and microorganism was found, with only one discordant result (a CT case detected by Versant on urines and vaginal swab, found by Aptima only on urines). Any interference due to the different liquid components of E-Swab was excluded. Conclusion Given the outstanding performance, Aptima assays can represent an excellent choice for CT, NG and MG molecular detection. Moreover, it is noteworthy that Aptima assays allow testing of specimens collected by E-Swab, enabling the possibility to use the same sample for both NG molecular detection and culture.

P1.13 Vaginal microbiome signatures in chlamydia trachomatis infected women

Introduction In healthy women, lactobacilli play a crucial role in maintaining the microbial homeostasis of the vaginal niche. In case of bacterial vaginosis (BV), a condition characterised by a depletion of lactobacilli and an increasing number of anaerobes, a higher risk of urogenital and sexually transmitted infections (STIs) is reported. The vaginal environment of healthy and BV-positive women have been extensively studied, leading to the identification of the microbial species dominating these opposite conditions and to the description of specific metabolic profiles. Besides that, less is known about the vaginal microbiome in case of STIs, as Chlamydia trachomatis (CT) infections. The aim of this study was to analyse the composition of the endogenous microbiota and the metabolic signatures of the vaginal niche in 3 different conditions: healthy, BV and CT infections. Methods From July 2016, all the pre-menopausal women attending the STI Outpatients Clinic of Sant’Orsola-Malpighi Hospital in Bologna (Italy) and meeting one of the following criteria were enrolled: presence of vaginal symptoms or presence of risk factors for CT infection. Patients with vaginal candidiasis were excluded. For all the patients, a vaginal swab was collected for molecular CT detection (Versant CT/GC DNA 1.0 Assay; Siemens), whereas Amsel criteria were used for BV assessment. Moreover, for each woman, an additional vaginal swab stored in saline was collected and centrifuged. Cell pellets were examined with a DNA-microarray platform including 17 probe sets specific for the most representative vaginal bacterial groups and with a quantitative real-time PCR targeting 16s rRNA gene of Gardnerella vaginalis (GV). Cell-free supernatants were used for metabolomic analysis by means of 1H-NMR spectroscopy. NMR spectra were recorded with an AVANCE spectrometer (Bruker). Similarities among microbial and metabolic profiles of samples were investigated by means of a principal component analysis (PCA). Differences in GV DNA loads and metabolites concentrations were analysed by ANOVA test. The study was approved by the Hospital Ethical Committee. Results Among all the women enrolled, 25 were considered healthy, 18 received a diagnosis of BV and 22 were positive for CT. PCA revealed that the vaginal microbiome of healthy and BV-subjects were clearly distinct and that CT-positive women were more similar to healthy women rather than to BV-positives, both for microbial composition and for metabolic profile. The mean GV DNA load was significantly different between the groups (p=0.03): healthy and CT positive women showed similar and lower mean loads compared to BV group. At a metabolic level, significantly higher concentrations of formate, ethanolamine and methylamine were found in BV-patients, while tryptophan and lactate were more present in healthy and CT-positive women. Conclusion Specific microbial and metabolic signatures characterise different clinical conditions of the vaginal tract. In this context, CT-positive women are definitely more similar to healthy than BV-subjects.