Search

Men having sex with men (MSM) represent a key population, in which sexually transmitted rectal infections (STIs) caused by Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG) and high-risk HPV (HR-HPV) are very common and linked to significant morbidity. Investigating the anorectal microbiome associated with rectal STIs holds potential for deeper insights into the pathogenesis of these infections and the development of innovative control strategies. In this study, we explored the interplay at the rectal site between C. trachomatis, N. gonorrhoeae, HR-HPV infection, and the anorectal microbiome in a cohort of 92 MSM (47 infected by CT and/or NG vs 45 controls). Moreover, we assessed the presence of Torquetenovirus (TTV), a non-pathogenic endogenous virus, considered as a possible predictor of immune system activation. We found a high prevalence of HR-HPV rectal infections (61%), especially in subjects with a concurrent CT/NG rectal infection (70.2%) and in people living with HIV (84%). In addition, we observed that TTV was more prevalent in subjects with CT/NG rectal infections than in non-infected ones (70.2% vs 46.7%, respectively). The anorectal microbiome of patients infected by CT and/or NG exhibited a reduction in Escherichia, while the presence of TTV was significantly associated with higher levels of Bacteroides. We observed a positive correlation of HR-HPV types with Escherichia and Corynebacterium, and a negative correlation with the Firmicutes phylum, and with Prevotella, Oscillospira, Sutterella. Our findings shed light on some of the dynamics occurring within the rectal environment involving chlamydial/gonococcal infections, HPV, TTV, and the anorectal microbiome. These data could open new perspectives for the control and prevention of STIs in MSM., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Ceccarani et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: Chlamydia trachomatis (CT) is the agent of the most common bacterial sexually transmitted infection worldwide. Until now, little information is available about the microbial composition of urine samples during CT urethritis. Therefore, in this study, we characterized the microbiome and metabolome profiles of first-void urines in a cohort of women with CT urethral infection attending an STI clinic., Methods: Based on CT positivity by nucleic acid amplification techniques on urine samples, the enrolled women were divided into two groups, i.e., "CT-negative" ( n = 21) and "CT-positive" ( n = 11). Urine samples were employed for (i) the microbiome profile analysis by means of 16s rRNA gene sequencing and (ii) the metabolome analysis by 1 H-NMR., Results: Irrespective of CT infection, the microbiome of first-void urines was mainly dominated by Lactobacillus , L. iners and L. crispatus being the most represented species. CT-positive samples were characterized by reduced microbial biodiversity compared to the controls. Moreover, a significant reduction of the Mycoplasmataceae family-in particular, of the Ureaplasma parvum species-was observed during CT infection. The Chlamydia genus was positively correlated with urine hippurate and lactulose., Conclusions: These data can help elucidate the pathogenesis of chlamydial urogenital infections, as well as to set up innovative diagnostic and therapeutic approaches.

Chlamydia persistence is a viable but non-replicative stage, induced by several sub-lethal stressor agents, including beta-lactam antibiotics. So far, no data about the connection between doxycycline and chlamydial persistence has been described in literature. We investigated the ability of doxycycline to induce C. trachomatis (CT) persistence in an in vitro model of epithelial cell infection (HeLa cells), comparing the results with the well-established model of penicillin-induced persistence. The effect of doxycycline was explored on 10 different CT strains by analysing (i) the presence of aberrant inclusions, (ii) chlamydial recovery, (iii) the expression of different chlamydial genes (omcB, euo, Ct110, Ct604, Ct755, HtrA) and (iv) the effects on epithelial cell viability. For each strain, the presence of foreign genomic islands responsible of tetracycline resistance was excluded. We found that low doses of doxycycline can induce a condition of CT persistence. For concentrations of doxycycline equal to 0.03-0.015 mg/L, CT inclusions are smaller and aberrant and CT cycle is characterized by the presence of viable but non-dividing RBs with the complete abolishment of chlamydial cytotoxic effect. Infectious EBs can be recovered after removal of the drug. During doxycycline-induced persistence, the expression of the late gene omcB is decreased, indicating the blocking of RB-to-EB conversion. Conversely, as for penicillin G, a significant up-regulation of the stress response HtrA gene is found in doxycycline-treated cells. This study provides a novel in vitro cell model to examine the characteristics of doxycycline-induced persistent CT infection., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Chlamydia trachomatis and Neisseria gonorrhoeae are the most common agents of bacterial sexually transmitted infections (STIs) worldwide. Here, we evaluated genital and extra-genital C. trachomatis and N. gonorrhoeae infection prevalence in a cohort of young women attending an STI Outpatients Clinic in Italy. From May 2019 to December 2019, 134 women aged 18-26 years were enrolled. A vaginal, a pharyngeal and a rectal swab for the molecular detection of C. trachomatis and N. gonorrhoeae were collected from each patient. Chlamydia-positive samples underwent a molecular genotyping based on pmpH gene. Total prevalence of C. trachomatis and N. gonorrhoeae infections was 17.9% and 11.2%, respectively. Chlamydial infections were prevalent in the urogenital (16.4%) and rectal (13.4%) sites, whereas N. gonorrhoeae predominated in the genital (7.4%) and pharyngeal (6%) mucosa. Overall, 5.2% of cases would have been missed if extra-genital sites had not been tested. Notably, 60% of women with a rectal infection did not report anal sex. A history of sexual contacts with a positive partner (P=0.03) and a history of ≥3 partners in the last 6 months (P=0.0075) were highly predictive of a bacterial STI. No cases of lymphogranuloma venereum were found. These data could help set up effective strategies to prevent the spread of STIs.

Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) represent the most common agents of sexually transmitted rectal infections among men having sex with other men (MSM). In this study, we assessed the bacterial composition of the rectal microbiota associated with CT and/or NG infections in a cohort of men reporting unsafe rectal intercourse. A total of 125 rectal swabs were collected and four groups were compared: non-infected subjects ( n = 53), patients with CT ( n = 37), or NG rectal infection ( n = 17) and patients with contemporary positivity for CT/NG ( n = 18). CT and NG infections were detected by a real-time commercial test and the rectal microbiota composition was analyzed from rectal swabs through sequencing of the hypervariable V3-V4 regions of the 16S rRNA gene. The rectal microbiota of all subgroups was dominated by Prevotellaceae, Enterobacteriaceae , and Ruminococcaceae families. Irrespective of the analyzed subgroup, we found that the rectal environment of all the enrolled MSM was rich in Prevotella and Escherichia genera. Moreover, a shift in the bacterial composition between patients with sexually transmitted rectal infections and controls was noticed: infected patients were characterized by a depletion of Escherichia species, associated with an increase of anaerobic genera, including Peptoniphilus, Peptostreptococcus , and Parvimonas . Overall, the presence of rectal symptoms did not significantly modify the rectal microbiota profiles among the four groups of analyzed patients. We confirmed that HIV-positive patients are characterized by a lower bacterial richness than HIV-negative subjects. However, we found that the presence of HIV has a different impact on bacterial rectal communities compared to CT and NG infections, modifying the relative abundance of several genera, including Gardnerella, Lactobacillus, Corynebacterium , and Sutterella . Information about the rectal microbiota composition in CT and NG infections could shed light on the pathogenesis of these conditions and could contribute to the onset of new strategies for their control., (Copyright © 2019 Ceccarani, Marangoni, Severgnini, Camboni, Laghi, Gaspari, D'Antuono, Foschi, Re and Consolandi.)

We characterized the vaginal ecosystem during common infections of the female genital tract, as vulvovaginal candidiasis (VVC, n = 18) and Chlamydia trachomatis infection (CT, n = 20), recruiting healthy (HC, n = 21) and bacterial vaginosis-affected (BV, n = 20) women as references of eubiosis and dysbiosis. The profiles of the vaginal microbiome and metabolome were studied in 79 reproductive-aged women, by means of next generation sequencing and proton based-nuclear magnetic resonance spectroscopy. Lactobacillus genus was profoundly depleted in all the genital infections herein considered, and species-level analysis revealed that healthy vaginal microbiome was dominated by L. crispatus. In the shift from HC to CT, VVC, and BV, L. crispatus was progressively replaced by L. iners. CT infection and VVC, as well as BV condition, were mainly characterised by anaerobe genera, e.g. Gardnerella, Prevotella, Megasphaera, Roseburia and Atopobium. The changes in the bacterial communities occurring during the genital infections resulted in significant alterations in the vaginal metabolites composition, being the decrease of lactate a common marker of all the pathological conditions. In conclusion, according to the taxonomic and metabolomics analysis, we found that each of the four conditions is characterized by a peculiar vaginal microbiome/metabolome fingerprint.

The sexually transmitted pathogen Chlamydia trachomatis (CT) is able to replicate and survive in human intestinal epithelial cells, being the gastro-intestinal tract a suitable site of residence for this microorganism. In this context, no detailed information about the mechanisms of cell death in intestinal cell lines after a chlamydial infection is available. The aim of this study was to compare the effect of two different CT serovars (D and L2) on the survival/death of different intestinal cell lines (Caco-2 and COLO-205), using endocervical cells (HeLa) as a reference model of genital infection. Seventy two hours after chlamydial infection at different multiplicity of infection (MOI) levels, the viability of HeLa, Caco-2 and COLO 205 cells was evaluated through dose-response experiments by means of a MTS-based assay. To get deeper insights in the mechanisms of cell death induced by CT, cell viability was assessed in presence of different inhibitors (i.e. pan-caspase inhibitor Z-VAD, necroptosis inhibitor Necrostatin-1, hydrogen peroxide scavenger catalase, caspase-1 inhibitor Ac-YVAD-cmk). Moreover, the activation of effector caspases and the presence of cellular apoptotic/necrotic changes were evaluated at different time points after CT infection. Our results demonstrated that, for both chlamydial serovars, intestinal cell lines are more resistant to CT-induced cell death compared to HeLa, thus representing a suitable 'niche' for chlamydial residence and replication. In literature, apoptosis has been widely described to be the main cell death mechanism elicited by chlamydia infection. However, our data demonstrate that necroptosis plays a relevant role, proceeding in parallel with apoptosis. The protective effect of catalase suggests the involvement of oxidative stress in triggering both cell death pathways. Moreover, we demonstrated that caspase-1 is involved in CT-induced cell death, potentially contributing to host inflammatory response and tissue damage. Cells infected by L2 serovar displayed a higher activation of effector caspases compared to cells infected with serovar D, suggesting a serovar-specific activation of apoptotic pathways and potentially explaining the greater virulence of L serovars. Finally, we found that Chlamydia elicits the early externalization of phosphatidylserine on the external leaflet of plasma membrane independently of caspase activation., Competing Interests: The authors have declared that no competing interests exist.

Purpose: We assessed the prevalence and predictors of Chlamydia trachomatis, Neisseriagonorrhoeae and Mycoplasmagenitalium rectal infections in a population of 'men having sex with men' (MSM)., Methodology: From January to November 2017, 165 MSM attending a STI outpatients clinic in Bologna (Italy) and reporting unsafe anal intercourses were enrolled. An ano-rectal swab was collected from each patient: chlamydial and gonococcal infections were diagnosed by a commercial NAAT, whereas an in-house quantitative PCR was used for M. genitalium detection. In addition, 131 urine samples and 84 pharyngeal swabs underwent testing for C. trachomatis and N. gonorrhoeae. A molecular C. trachomatis typing, a serological screening for anti-Chlamydia IgG and IgA, as well as the assessment of HIV, HCV and syphilis infections, were performed.Results/Key findings. The prevalence of C. trachomatis, N. gonorrhoeae and M. genitalium rectal infections was 27.2, 25.4 and 4.8 %, respectively. Globally, 63.1 % of cases were asymptomatic and up to 80 % of chlamydial and gonococcal infections would have been missed if the rectal site had not been tested. All the patients with rectal M. genitalium carriage were asymptomatic and characterized by low bacterial loads (<2500 DNA copies/reaction). Lymphogranuloma venereum (LGV) prevalence was 12.1 % with a considerable proportion of asymptomatic infections (35 %). The presence of symptoms, age >30, HIV-positivity and elevated levels of anti-Chlamydia antibodies were the most significant predictors of LGV., Conclusions: Sexually transmitted rectal infections are frequent and often asymptomatic among MSM. LGV prevalence is high in our country and there is increasing evidence of symptomless cases.

The aim of this study was to characterize the urine metabolome of women with Chlamydia trachomatis (CT) uro-genital infection (n = 21), comparing it with a group of CT-negative subjects (n = 98). By means of a proton-based nuclear magnetic resonance (1H-NMR) spectroscopy, we detected and quantified the urine metabolites of a cohort of 119 pre-menopausal Caucasian women, attending a STI Outpatients Clinic in Italy. In case of a CT positive result, CT molecular genotyping was performed by omp1 gene semi-nested PCR followed by RFLP analysis. We were able to identify several metabolites whose concentrations were significantly higher in the urine samples of CT-positive subjects, including sucrose, mannitol, pyruvate and lactate. In contrast, higher urinary levels of acetone represented the main feature of CT-negative women. These results were not influenced by the age of patients nor by the CT serovars (D, E, F, G, K) responsible of the urethral infections. Since the presence of sugars can increase the stability of chlamydial proteins, higher levels of sucrose and mannitol in the urethral lumen, related to a higher sugar consumption, could have favoured CT infection acquisition or could have been of aid for the bacterial viability. Peculiar dietary habits of the subjects enrolled, in term of type and amount of food consumed, could probably explain these findings. Lactate and pyruvate could result from CT-induced immunopathology, as a product of the inflammatory microenvironment. Further studies are needed to understand the potential role of these metabolites in the pathogenesis of CT infection, as well as their diagnostic/prognostic use.

The aim of this study was to assess Chlamydia trachomatis (CT) infection prevalence and serovar distribution in a high-density urban area in the north of Italy, by comparing different groups of subjects divided on the basis of the type of care provider they referred to (STI Clinic, gynaecologists or general practitioners). From January 2011 to May 2014, all the specimens submitted to the Microbiology Laboratory of St Orsola Hospital in Bologna for CT detection were tested by PCR assay. For positive specimens, molecular genotyping based on RFLP analysis was performed. Total prevalence of CT infection was 8.1 %, with significant differences between subgroups (P<0.01) but stable during the study period. The STI Clinic was mainly responsible for CT diagnosis, whereas the lowest infection prevalence was detected in gynaecological clinics, despite the high number of tests performed. Extra-genital samples were almost exclusively collected from males at the STI Clinic. Interestingly, 13.3 % of patients providing extra-genital specimens were positive for CT on rectal and/or pharyngeal swabs, and 4.4 % of cases would have been missed if extra-genital sites had not been tested. The most common serovar was E, and serovar distribution was influenced by gender (P<0.01), age (P<0.01), care provider (P=0.01) and anatomical site (P<0.01). The L2 serovar was detected only in extra-genital samples from males at the STI Clinic. Knowledge about care providers' contributions in CT testing and diagnosis is essential for infection control. CT typing is crucial for appropriate management of specific infections, such as lymphogranuloma venereum in extra-genital samples of high-risk populations.

Objective: Chlamydia pneumoniae has been linked to atherosclerosis, strictly associated with hyperlipidemia. The liver plays a central role in the regulation of lipid metabolism. Since in animal models C. pneumoniae can be found at hepatic level, this study aims to elucidate whether C. pneumoniae infection accelerates atherosclerosis by affecting lipid metabolism., Methods: Thirty Balb/c mice were challenged intra-peritoneally with C. pneumoniae elementary bodies and thirty with Chlamydia trachomatis, serovar D. Thirty mice were injected with sucrose-phosphate-glutamate buffer, as negative controls. Seven days after infection, liver samples were examined both for presence of chlamydia and expression of genes involved in inflammation and lipid metabolism., Results: C. pneumoniae was isolated from 26 liver homogenates, whereas C. trachomatis was never re-cultivated (P < 0.001). C. pneumoniae infected mice showed significantly increased serum cholesterol and triglycerides levels compared both with negative controls (P < 0.001 and P = 0.0197, respectively) and C. trachomatis infected mice (P < 0.001). Liver bile acids were significantly reduced in C. pneumoniae compared to controls and C. trachomatis infected mice. In C. pneumoniae infected livers, cholesterol 7α-hydroxylase (Cyp7a1) and low-density lipoprotein receptor (Ldlr) mRNA levels were reduced, while inducible degrader of the low-density lipoprotein receptor (Idol) expression was increased. Hypertriglyceridemia was associated to reduced expression of hepatic carnitine palmitoyltransferase-1a (Cpt1a) and medium chain acyl-Coenzyme A dehydrogenase (Acadm). Pro-inflammatory cytokines gene expression was increased compared to negative controls. Conversely, in C. trachomatis infected animals, normal serum lipid levels were associated with elevated pro-inflammatory cytokines gene expression, linked to only a mild disturbance of lipid regulatory genes., Conclusion: Our results indicate that C. pneumoniae mouse liver infection induces dyslipidemic effects with significant modifications of genes involved in lipid metabolism., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Screening for extra-genital Chlamydia trachomatis and Neisseria gonorrhoeae infections is a crucial component for sexually transmitted diseases management, even if at present days no commercial methods have been approved for use on pharyngeal and rectal specimens by the US FDA or have received the conformity CE marking. Here we report the analytical sensitivities of the Versant CT/GC 1.0 assay (Siemens Healthcare Diagnostics, Tarrytown, NY, USA) on rectal and pharyngeal swabs, and an evaluation about the suitability for this assay with two widely used swab collection devices (E-Swab and eNAT, Copan, Brescia, Italy). The limits of detection for rectal and pharyngeal specimens with the Versant assay were 10 copies/ml and 1.0 copies/ml, for C. trachomatis and N. gonorrhoeae, respectively. False positive results due to the presence of non-gonococcal Neisseria species were excluded when clinical rectal and pharyngeal samples containing organisms identified as N. meningitidis, N. sicca, N. flavescens and N. subflava were tested. Due to its sensitivity and specificity, the Versant assay represents a good choice for the diagnosis of chlamydial and/or gonococcal infections not only in genito-urinary samples, but also on rectal and pharyngeal swabs.

An Italian HIV-positive man having sex with men (MSM) attended the STIs Outpatients Clinic of Sant'Orsola Hospital in Bologna complaining of anal pain and constipation. According to patient's sexual history and repertoires, NAAT testing for Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (GC) was performed. Pharyngeal and anal swabs resulted positive for CT DNA detection and the following molecular genotyping identified a L2 serovar, coming to the final diagnosis of pharyngeal and rectal lymphogranuloma venereum (LGV) infection. After an antibiotic therapy with doxycycline 100 mg twice a day for 3 weeks, the patient completely recovered and the test of cure was negative for LGV infection.

Purpose: The aim of this study is to explore the feasibility of 11C-Choline PET in the assessment of the degree of inflammation in the Chlamydia muridarum genital infection model., Procedures: Forty female Balb/c mice received 2.5 mg of medroxyprogesterone acetate i.m. 9 and 2 days prior to the infection: 21 mice were infected by C. muridarum into the vaginal vault, 12 mice were treated with inactivated chlamydiae, and 7 mice were SPG buffer-treated as negative controls. Three healthy control mice were not treated with progesterone. Mice in each category were randomly subdivided in two groups: (1) sacrificed at 5, 10, 15, and 20 days for histological analysis and (2) undergoing 11C-Choline PET at days 5, 10, and 20 post-infection (20 MBq of 11C-Choline, uptake time of 10 min, acquisition through a small-animal PET tomograph for 15 min)., Results: Infected animals showed a significantly higher standardized uptake value than both controls and animals inoculated with heat-inactivated chlamydiae in each PET scan (P<0.05). All organs of the infected animals had scores of inflammation ranging between 2 and 3 at day 5, decreasing to 1-2 at day 20., Conclusions: This preliminary result demonstrated that 11C-Choline PET can highlight a specific proliferation mechanism of inflammatory cells induced by C. muridarum, thanks to a very high sensitivity in detecting very small amounts of tracer in inflammatory cells.

We studied the prevalence of Chlamydia trachomatis (CT) urogenital infection and the distribution of different genotypes in a non-selected STD population of 1625 patients, evaluating presence of coinfections with other sexually transmitted diseases. Each patient was bled to perform serological tests for syphilis and HIV, then urethral or endocervical swabs were obtained for the detection of CT and Neisseria gonorrhoeae by culture. DNA extracted from remnant positive swabs was amplified by omp1 Nested PCR and products were sequenced. Total prevalence of CT infection was 6.3% (103/1625), with strong differences between men and women (11.4% vs 3.9%, P<0.01). Clinical symptoms and coinfections were much more frequent in men than in women (P<0.01). The most common serovar was E (prevalence of 38.8%), followed by G (23.3%), F (13.5%) D/Da (11.6%) and J (4.8%). Serovars distribution was statistically different between men and women (P=0.042) and among patients with or without coinfection (P=0.035); patients infected by serovar D/Da showed the highest coinfection rate. This study can be considered a contribution in increasing knowledge on CT serovar distribution in Italy. Further studies are needed to better define molecular epidemiology of CT infection and to investigate its correlation with other STDs.