Doxycycline Prophylaxis to Prevent Sexually Transmitted Infections in Women.

BACKGROUND: Doxycycline postexposure prophylaxis (PEP) has been shown to prevent sexually transmitted infections (STIs) among cisgender men and transgender women, but data from trials involving cisgender women are lacking. METHODS: We conducted a randomized, open-label trial comparing doxycycline PEP (doxycycline hyclate, 200 mg taken within 72 hours after condomless sex) with standard care among Kenyan women 18 to 30 years of age who were receiving preexposure prophylaxis against human immunodeficiency virus (HIV). The primary end point was any incident infection with Chlamydia trachomatis, Neisseria gonorrhoeae, or Treponema pallidum. Hair samples were collected quarterly for objective assessment of doxycycline use. RESULTS: A total of 449 participants underwent randomization; 224 were assigned to the doxycycline-PEP group and 225 to the standard-care group. Participants were followed quarterly over 12 months. A total of 109 incident STIs occurred (50 in the doxycycline-PEP group [25.1 per 100 person-years] and 59 in the standard-care group [29.0 per 100 person-years]), with no significant between-group difference in incidence (relative risk, 0.88; 95% confidence interval [CI], 0.60 to 1.29; P = 0.51). Among the 109 incident STIs, chlamydia accounted for 85 (78.0%) (35 in the doxycycline-PEP group and 50 in the standard-care group; relative risk, 0.73; 95% CI, 0.47 to 1.13). No serious adverse events were considered by the trial investigators to be related to doxycycline, and there were no incident HIV infections. Among 50 randomly selected participants in the doxycycline-PEP group, doxycycline was detected in 58 of 200 hair samples (29.0%). All N. gonorrhoeae-positive isolates were resistant to doxycycline. CONCLUSIONS: Among cisgender women, the incidence of STIs was not significantly lower with doxycycline PEP than with standard care. According to hair-sample analysis, the use of doxycycline PEP among those assigned to receive it was low. (Funded by the National Institutes of Health; dPEP ClinicalTrials.gov number, NCT04050540.).

Postexposure Doxycycline to Prevent Bacterial Sexually Transmitted Infections.

BACKGROUND: Interventions to reduce sexually transmitted infections (STIs) among men who have sex with men (MSM) are needed. METHODS: We conducted an open-label, randomized study involving MSM and transgender women who were taking preexposure prophylaxis (PrEP) against human immunodeficiency virus (HIV) infection (PrEP cohort) or living with HIV infection (persons living with HIV infection [PLWH] cohort) and who had had Neisseria gonorrhoeae (gonorrhea), Chlamydia trachomatis (chlamydia), or syphilis in the past year. Participants were randomly assigned in a 2:1 ratio to take 200 mg of doxycycline within 72 hours after condomless sex (doxycycline postexposure prophylaxis) or receive standard care without doxycycline. STI testing was performed quarterly. The primary end point was the incidence of at least one STI per follow-up quarter. RESULTS: Of 501 participants (327 in the PrEP cohort and 174 in the PLWH cohort), 67% were White, 7% Black, 11% Asian or Pacific Islander, and 30% Hispanic or Latino. In the PrEP cohort, an STI was diagnosed in 61 of 570 quarterly visits (10.7%) in the doxycycline group and 82 of 257 quarterly visits (31.9%) in the standard-care group, for an absolute difference of -21.2 percentage points and a relative risk of 0.34 (95% confidence interval [CI], 0.24 to 0.46; P<0.001). In the PLWH cohort, an STI was diagnosed in 36 of 305 quarterly visits (11.8%) in the doxycycline group and 39 of 128 quarterly visits (30.5%) in the standard-care group, for an absolute difference of -18.7 percentage points and a relative risk of 0.38 (95% CI, 0.24 to 0.60; P<0.001). The incidences of the three evaluated STIs were lower with doxycycline than with standard care; in the PrEP cohort, the relative risks were 0.45 (95% CI, 0.32 to 0.65) for gonorrhea, 0.12 (95% CI, 0.05 to 0.25) for chlamydia, and 0.13 (95% CI, 0.03 to 0.59) for syphilis, and in the PLWH cohort, the relative risks were 0.43 (95% CI, 0.26 to 0.71), 0.26 (95% CI, 0.12 to 0.57), and 0.23 (95% CI, 0.04 to 1.29), respectively. Five grade 3 adverse events and no serious adverse events were attributed to doxycycline. Of the participants with gonorrhea culture available, tetracycline-resistant gonorrhea occurred in 5 of 13 in the doxycycline groups and 2 of 16 in the standard-care groups. CONCLUSIONS: The combined incidence of gonorrhea, chlamydia, and syphilis was lower by two thirds with doxycycline postexposure prophylaxis than with standard care, a finding that supports its use among MSM with recent bacterial STIs. (Funded by the National Institutes of Health; DoxyPEP ClinicalTrials.gov number, NCT03980223.).

Dual species transcriptomics reveals conserved metabolic and immunologic processes in interactions between human neutrophils and Neisseria gonorrhoeae.

Neisseria gonorrhoeae (the gonococcus, Gc) causes the sexually transmitted infection gonorrhea. Gc is a prominent threat to human health by causing severe lifelong sequelae, including infertility and chronic pelvic pain, which is amplified by the emergence of "superbug" strains resistant to all current antibiotics. Gc is highly adapted to colonize human mucosal surfaces, where it survives despite initiating a robust inflammatory response and influx of polymorphonuclear leukocytes (PMNs, neutrophils) that typically clear bacteria. Here, dual-species RNA-sequencing was used to define Gc and PMN transcriptional profiles alone and after infection. Core host and bacterial responses were assessed for two strains of Gc and three human donors' PMNs. Comparative analysis of Gc transcripts revealed overlap between Gc responses to PMNs, iron, and hydrogen peroxide; 98 transcripts were differentially expressed across both Gc strains in response to PMN co-culture, including iron-responsive and oxidative stress response genes. We experimentally determined that the iron-dependent TbpB is suppressed by PMN co-culture, and iron-limited Gc have a survival advantage when cultured with PMNs. Analysis of PMN transcripts modulated by Gc infection revealed differential expression of genes driving cell adhesion, migration, inflammatory responses, and inflammation resolution pathways. Production of pro-inflammatory cytokines, including IL1B and IL8, the adhesion factor ICAM1, and prostaglandin PGE2 were induced in PMNs in response to Gc. Together, this study represents a comprehensive and experimentally validated dual-species transcriptomic analysis of two isolates of Gc and primary human PMNs that gives insight into how this bacterium survives innate immune onslaught to cause disease.

Experimental genital tract infection demonstrates Neisseria gonorrhoeae MtrCDE efflux pump is not required for in vivo human infection and identifies gonococcal colonization bottleneck.

The MtrCDE efflux pump of Neisseria gonorrhoeae exports a wide range of antimicrobial compounds that the gonococcus encounters at mucosal surfaces during colonization and infection and is a known gonococcal virulence factor. Here, we evaluate the role of this efflux pump system in strain FA1090 during in vivo human male urethral infection with N. gonorrhoeae using a controlled human infection model. With the strategy of competitive infections initiated with mixtures of wild-type FA1090 and an isogenic mutant FA1090 strain that does not contain a functional MtrCDE pump, we found that the presence of the efflux pump is not required for an infection to be established in the human male urethra. This finding contrasts with previous studies of in vivo infection in the lower genital tract of female mice, which demonstrated that mutant gonococci of a different strain (FA19) lacking a functional MtrCDE pump had a significantly reduced fitness compared to their wild-type parental FA19 strain. To determine if these conflicting results are due to strain or human vs. mouse differences, we conducted a series of systematic competitive infections in female mice with the same FA1090 strains as in humans, and with FA19 strains, including mutants that do not assemble a functional MtrCDE efflux pump. Our results indicate the fitness advantage provided by the MtrCDE efflux pump during infection of mice is strain dependent. Owing to the equal fitness of the two FA1090 strains in men, our experiments also demonstrated the presence of a colonization bottleneck of N. gonorrhoeae in the human male urethra, which may open a new area of inquiry into N. gonorrhoeae infection dynamics and control. TRIAL REGISTRATION. Clinicaltrials.gov NCT03840811.

Evolution, persistence, and host adaption of a gonococcal AMR plasmid that emerged in the pre-antibiotic era.

Plasmids are diverse extrachromosomal elements significantly that contribute to interspecies dissemination of antimicrobial resistance (AMR) genes. However, within clinically important bacteria, plasmids can exhibit unexpected narrow host ranges, a phenomenon that has scarcely been examined. Here we show that pConj is largely restricted to the human-specific pathogen, Neisseria gonorrhoeae. pConj can confer tetracycline resistance and is central to the dissemination of other AMR plasmids. We tracked pConj evolution from the pre-antibiotic era 80 years ago to the modern day and demonstrate that, aside from limited gene acquisition and loss events, pConj is remarkably conserved. Notably, pConj has remained prevalent in gonococcal populations despite cessation of tetracycline use, thereby demonstrating pConj adaptation to its host. Equally, pConj imposes no measurable fitness costs and is stably inherited by the gonococcus. Its maintenance depends on the co-operative activity of plasmid-encoded Toxin:Antitoxin (TA) and partitioning systems rather than host factors. An orphan VapD toxin encoded on pConj forms a split TA with antitoxins expressed from an ancestral co-resident plasmid or a horizontally-acquired chromosomal island, potentially explaining pConj's limited distribution. Finally, ciprofloxacin can induce loss of this highly stable plasmid, reflecting epidemiological evidence of transient reduction in pConj prevalence when fluoroquinolones were introduced to treat gonorrhoea.

Neisseria gonorrhoeae vaccines: a contemporary overview.

Neisseria gonorrhoeae infection is an important public health issue, with an annual global incidence of 87 million. N. gonorrhoeae infection causes significant morbidity and can have serious long-term impacts on reproductive and neonatal health and may rarely cause life-threatening disease. Global rates of N. gonorrhoeae infection have increased over the past 20 years. Importantly, rates of antimicrobial resistance to key antimicrobials also continue to increase, with the United States Centers for Disease Control and Prevention identifying drug-resistant N. gonorrhoeae as an urgent threat to public health. This review summarizes the current evidence for N. gonorrhoeae vaccines, including historical clinical trials, key N. gonorrhoeae vaccine preclinical studies, and studies of the impact of Neisseria meningitidis vaccines on N. gonorrhoeae infection. A comprehensive survey of potential vaccine antigens, including those identified through traditional vaccine immunogenicity approaches, as well as those identified using more contemporary reverse vaccinology approaches, are also described. Finally, the potential epidemiological impacts of a N. gonorrhoeae vaccine and research priorities for further vaccine development are described.

Novel lateral flow assay for point-of-care detection of Neisseria gonorrhoeae infection in syndromic management settings: a cross-sectional performance evaluation.

BACKGROUND: A rapid and affordable point-of-care test is a priority for Neisseria gonorrhoeae control. WHO and Foundation for Innovative New Diagnostics (FIND) have a target product profile for a non-molecular N gonorrhoeae rapid point-of-care test that requires a clinical sensitivity of greater than 80% and a specificity over 95% to be considered useful in syndromic management; test turnaround time should be 30 min or under, and the test should cost less than US$3. A novel lateral flow assay (LFA) was developed to achieve that profile. METHODS: In this cross-sectional study we evaluated the performance of the novel N gonorrhoeae lateral flow assay (NG-LFA) at the primary health-care level in South Africa. Male patients with urethral discharge syndrome and female patients with vaginal discharge syndrome were recruited from five primary health-care facilities in the Buffalo City Metropolitan Municipality health district of South Africa. First-void urine specimens and nurse-collected vaginal swabs were tested in-facility with the NG-LFA and Xpert CT/NG PCR assay. N gonorrhoeae multi-antigen sequence typing (NG-MAST) was performed on all LFA positive specimens. FINDINGS: Between March 7, and Sept 19, 2022, we enrolled 200 male patients with urethral discharge and 200 female patients with vaginal discharge. The median age of male patients was 24 years (IQR 21-31 years), and the median age of female patients was 25 years (IQR 21-32 years). In addition, 23 male patients and 12 female patients who presented at the facility with a partner notification slip were enrolled of whom one (4%) and five (42%) were symptomatic, respectively. NG-LFA and Xpert results were available for all participants. In urine specimens, NG-LFA sensitivity was 96·1% (Wilson 95% CI 91·2-98·3; 123 LFA-positive among 128 PCR-positive specimens) and 91·7% in vaginal swab specimens (78·2-97·1; 33 LFA-positive among 36 PCR-positive). The specificity was 97·2% in urine specimens (90·4-99·2; 70 LFA-negative among 72 PCR-negative) and 96·3% in vaginal specimens (92·2-98·3; 158 LFA-negative among 164 PCR-negative). In 156 LFA-positive specimens, NG-MAST showed 93 different sequence types. INTERPRETATION: The novel NG-LFA had excellent clinical sensitivity and specificity in symptomatic male and female patients. The test met the optimal requirement for sensitivity and the minimal requirement for specificity specified in the target product profile. NG-LFA could provide an important tool to optimise clinical management and reduce excess antibiotic use in settings without direct access to laboratory testing. FUNDING: Global Antimicrobial Resistance Innovation Fund (GAMRIF) via FIND and National Institutes of Health.

Functional characterization of the gonococcal polyphosphate pseudo-capsule.

Neisseria gonorrhoeae is an exclusively human pathogen able to evade the host immune system through multiple mechanisms. Gonococci accumulate a large portion of phosphate moieties as polyphosphate (polyP) on the exterior of the cell. Although its polyanionic nature has suggested that it may form a protective shield on the cell surface, its role remains controversial. Taking advantage of a recombinant His-tagged polyP-binding protein, the presence of a polyP pseudo-capsule in gonococcus was demonstrated. Interestingly, the polyP pseudo-capsule was found to be present in specific strains only. To investigate its putative role in host immune evasion mechanisms, such as resistance to serum bactericidal activity, antimicrobial peptides and phagocytosis, the enzymes involved in polyP metabolism were genetically deleted, generating mutants with altered polyP external content. The mutants with lower polyP content on their surface compared to the wild-type strains, became sensitive to complement-mediated killing in presence of normal human serum. Conversely, naturally serum sensitive strains that did not display a significant polyP pseudo-capsule became resistant to complement in the presence of exogenous polyP. The presence of polyP pseudo-capsule was also critical in the protection from antibacterial activity of cationic antimicrobial peptide, such as cathelicidin LL-37. Results showed that the minimum bactericidal concentration was lower in strains lacking polyP than in those harboring the pseudo-capsule. Data referring to phagocytic killing resistance, assessed by using neutrophil-like cells, showed a significant decrease in viability of mutants lacking polyP on their cell surface in comparison to the wild-type strain. The addition of exogenous polyP overturned the killing phenotype of sensitive strains suggesting that gonococcus could exploit environmental polyP to survive to complement-mediated, cathelicidin and intracellular killing. Taken together, data presented here indicate an essential role of the polyP pseudo-capsule in the gonococcal pathogenesis, opening new perspective on gonococcal biology and more effective treatments.

Neisseria gonorrhoeae co-opts C4b-binding protein to enhance complement-independent survival from neutrophils.

Neisseria gonorrhoeae (Gc) is a human-specific pathogen that causes the sexually transmitted infection gonorrhea. Gc survives in neutrophil-rich gonorrheal secretions, and recovered bacteria predominantly express phase-variable, surface-expressed opacity-associated (Opa) proteins (Opa+). However, expression of Opa proteins like OpaD decreases Gc survival when exposed to human neutrophils ex vivo. Here, we made the unexpected observation that incubation with normal human serum, which is found in inflamed mucosal secretions, enhances survival of Opa+ Gc from primary human neutrophils. We directly linked this phenomenon to a novel complement-independent function for C4b-binding protein (C4BP). When bound to the bacteria, C4BP was necessary and sufficient to suppress Gc-induced neutrophil reactive oxygen species production and prevent neutrophil phagocytosis of Opa+ Gc. This research identifies for the first time a complement-independent role for C4BP in enhancing the survival of a pathogenic bacterium from phagocytes, thereby revealing how Gc exploits inflammatory conditions to persist at human mucosal surfaces.

HIV, asymptomatic STI, and the rectal mucosal immune environment among young men who have sex with men.

Young men who have sex with men (YMSM) are disproportionately affected by HIV and bacterial sexually transmitted infections (STI) including gonorrhea, chlamydia, and syphilis; yet research into the immunologic effects of these infections is typically pursued in siloes. Here, we employed a syndemic approach to understand potential interactions of these infections on the rectal mucosal immune environment among YMSM. We enrolled YMSM aged 18-29 years with and without HIV and/or asymptomatic bacterial STI and collected blood, rectal secretions, and rectal tissue biopsies. YMSM with HIV were on suppressive antiretroviral therapy (ART) with preserved blood CD4 cell counts. We defined 7 innate and 19 adaptive immune cell subsets by flow cytometry, the rectal mucosal transcriptome by RNAseq, and the rectal mucosal microbiome by 16S rRNA sequencing and examined the effects of HIV and STI and their interactions. We measured tissue HIV RNA viral loads among YMSM with HIV and HIV replication in rectal explant challenge experiments among YMSM without HIV. HIV, but not asymptomatic STI, was associated with profound alterations in the cellular composition of the rectal mucosa. We did not detect a difference in the microbiome composition associated with HIV, but asymptomatic bacterial STI was associated with a higher probability of presence of potentially pathogenic taxa. When examining the rectal mucosal transcriptome, there was evidence of statistical interaction; asymptomatic bacterial STI was associated with upregulation of numerous inflammatory genes and enrichment for immune response pathways among YMSM with HIV, but not YMSM without HIV. Asymptomatic bacterial STI was not associated with differences in tissue HIV RNA viral loads or in HIV replication in explant challenge experiments. Our results suggest that asymptomatic bacterial STI may contribute to inflammation particularly among YMSM with HIV, and that future research should examine potential harms and interventions to reduce the health impact of these syndemic infections.

Urgent Need to Understand and Prevent Gonococcal Infection: From the Laboratory to Real-World Context.

Neisseria gonorrhoeae is widespread globally. Primary prevention is unsuccessful and antimicrobial resistance threatens optimal management. There is no specific vaccine and natural infection studies show that N gonorrhoeae can avoid and suppress immune responses. In addition to extensive variation in expression and specificity of many gonococcal surface antigens, it induces a robust inflammatory response through the Th17 pathway with a large influx of neutrophils and inflammatory cytokines but evades macrophages. The Th1- and Th2-mediated response is suppressed, resulting in low, short-lived antibody titers. Real-world evidence suggests that gonorrhea cases are reduced among recipients of Neisseria meningitidis group B vaccines containing outer membrane vesicles (OMVs). Although the first randomized trial of an OMV-containing MenB vaccine against N gonorrhoeae infection did not show statistically significant vaccine efficacy, ongoing trials might shed further light. Several candidate vaccine antigens for a gonococcal-specific vaccine are being evaluated preclinically but only one has reached clinical trials.

Development and Validation of Multiplex Assays for Mouse and Human IgG and IgA to Neisseria gonorrhoeae Antigens.

There is an urgent need for vaccines against Neisseria gonorrhoeae, the causative agent of gonorrhea. Vaccination with an outer membrane vesicle-based Neisseria meningitidis vaccine provides some protection from N. gonorrhoeae; however, the mechanisms underlying this cross-protection are unknown. To address this need, we developed multiplexed bead-based assays for the relative quantification of human and mouse IgG and IgA against N gonorrhoeae antigens. The assays were evaluated for analyte independence, dilutional linearity, specificity, sensitivity, intra- and interassay variability, and robustness to sample storage conditions. The assay was then used to test samples from mice and humans immunized with an N meningitidis outer membrane vesicle vaccine.

Changes in Inflammatory Cytokine Levels in Rectal Mucosa Associated With Neisseria gonorrheae and/or Chlamydia trachomatis Infection and Treatment Among Men Who Have Sex With Men in Lima, Peru.

BACKGROUND: Neisseria gonorrheae and Chlamydia trachomatis are associated with mucosal inflammation and human immunodeficiency virus 1 (HIV-1) transmission. We assessed levels of inflammatory cytokines in men who have sex with men (MSM) with and without rectal gonorrhea and/or chlamydia in Lima, Peru. METHODS: We screened 605 MSM reporting condomless receptive anal intercourse for rectal N. gonorrheae/C. trachomatis using nucleic acid testing. We identified 101 cases of gonorrhea and/or chlamydia and randomly selected 50 N. gonorrheae/C. trachomatis positive cases and matched 52 negative controls. We measured levels of IL-1ß, IL-6, IL-8, and TNF-α in rectal secretions. Tests for HIV-1, rectal N. gonorrheae/C. trachomatis, and mucosal cytokines were repeated after 3 and 6 months. Cytokine levels in cases and uninfected controls were compared using Wilcoxon rank-sum tests and linear regression. RESULTS: MSM with gonorrhea/chlamydia had elevated levels of all cytokines in rectal mucosa compared with matched controls (all P values <.001). Following antibiotic treatment there were no significant differences in cytokine levels at 3- or 6-month follow-up evaluations (all P values >.05). DISCUSSION: Rectal gonorrhea/chlamydia infection is associated with transient mucosal inflammation and cytokine recruitment. Our data provide proof of concept for rectal sexually transmitted infection screening as an HIV prevention strategy for MSM. Clinical Trials Registration. NCT03010020.

Prevalence of Sexually Transmitted Infections Among Transgender Women With and Without HIV in the Eastern and Southern United States.

BACKGROUND: Data on the epidemiology of sexually transmitted infections (STIs) among transgender women (TGW) with and without human immunodeficiency virus (HIV) are limited. METHODS: We analyzed baseline data collected from a cohort of adult TGW across 6 eastern and southern US cities between March 2018 and August 2020 (n = 1018). Participants completed oral HIV screening, provided self-collected rectal and urogenital specimens for chlamydia and gonorrhea testing, and provided sera specimens for syphilis testing. We assessed associations with ≥1 prevalent bacterial STI using modified Poisson regression. RESULTS: Bacterial STI prevalence was high and differed by HIV status: 32% among TGW with HIV and 11% among those without HIV (demographic-adjusted prevalence ratio = 1.91; 95% confidence interval = 1.39-2.62). Among TGW without HIV, bacterial STI prevalence differed by geographic region, race and ethnicity, and gender identity, and was positively associated with reporting >1 sexual partner, hazardous alcohol use, homelessness, having safety concerns regarding transit to health care, and no prior receipt of gender-affirming health services. Among TGW with HIV, older age was inversely associated with bacterial STI. CONCLUSIONS: TGW had a high prevalence of bacterial STIs. The prevalence and correlates of bacterial STI differed by HIV status, highlighting the unique needs and risks of TGW with and without HIV. Tailored interventions may reduce sexual health-related inequities.

Comparative analysis of the physical properties of murine and human S100A7: Insight into why zinc piracy is mediated by human but not murine S100A7.

S100 proteins are a subfamily of EF-hand calcium-binding proteins found primarily in vertebrate animals. They are distinguished by binding of transition metals and functioning in both the intracellular and extracellular milieu. S100A7 functions in the protection of the skin and mucous membranes and is a biomarker in inflammatory skin disease. A recent study of Neisseria gonorrhoeae infection revealed that human but not murine S100A7 could be used to evade host nutritional immunity. To understand the molecular basis for this difference, we carried out a comparative analysis of the physical and structural properties of human and murine S100A7. The X-ray crystal structure of Ca2+-loaded mouse S100A7 (mS100A7) was determined to 1.69 Å resolution, and Ca2+-induced conformational changes were assessed by NMR. Unlike human S100A7 (hS100A7), which exhibits conformational changes in response to binding of Ca2+, no significant changes in mS100A7 were detected. Dynamic light scattering, circular dichroism, and a competition chelator assay were used to compare the Zn2+ affinity and the effects of ion binding on mS100A7 versus hS100A7. Alignment of their sequences revealed a substantial difference in the C-terminal region, which is an important mediator of protein-protein interactions, suggesting a rationale for the specificity of N. gonorrhoeae for hS100A7. These data, along with more detailed analysis of S100A7 sequence conservation across different species, support the proposal that, although hS100A7 is highly conserved in many mammals, the murine protein is a distinct ortholog. Our results highlight the potential limitations of using mouse models for studying bacterial infections in humans.

Neisseria gonorrhoeae drives Chlamydia trachomatis into a persistence-like state during in vitro co-infection.

Chlamydia trachomatis and Neisseria gonorrhoeae are the most prevalent bacterial sexually transmitted infections (STIs) globally. Despite frequent co-infections in patients, few studies have investigated how mono-infections may differ from co-infections. We hypothesized that a symbiotic relationship between the pathogens could account for the high rates of clinical co-infection. During in vitro co-infection, we observed an unexpected phenotype where the C. trachomatis developmental cycle was impaired by N. gonorrhoeae. C. trachomatis is an obligate intracellular pathogen with a unique biphasic developmental cycle progressing from infectious elementary bodies (EB) to replicative reticulate bodies (RB), and back. After 12 hours of co-infection, we observed fewer EBs than in a mono-infection. Chlamydial genome copy number remained equivalent between mono- and co-infections. This is a hallmark of Chlamydial persistence. Chlamydial persistence alters inclusion morphology but varies depending on the stimulus/stress. We observed larger, but fewer, Chlamydia during co-infection. Tryptophan depletion can induce Chlamydial persistence, but tryptophan supplementation did not reverse the co-infection phenotype. Only viable and actively growing N. gonorrhoeae produced the inhibition phenotype in C. trachomatis. Piliated N. gonorrhoeae had the strongest effect on C. trachomatis, but hyperpiliated or non-piliated N. gonorrhoeae still produced the phenotype. EB development was modestly impaired when N. gonorrhoeae were grown in transwells above the infected monolayer. C. trachomatis serovar L2 was not impaired during co-infection. Chlamydial impairment could be due to cytoskeletal or osmotic stress caused by an as-yet-undefined mechanism. We conclude that N. gonorrhoeae induces a persistence-like state in C. trachomatis that is serovar dependent.

Investigating the importance of selected surface-exposed loops in HpuB for hemoglobin binding and utilization by Neisseria gonorrhoeae.

Neisseria gonorrhoeae is the etiological agent of the sexually transmitted infection gonorrhea. The pathogen is a global health challenge since no protective immunity results from infection, and far fewer treatment options are available with increasing antimicrobial resistance. With no efficacious vaccines, researchers are exploring new targets for vaccine development and innovative therapeutics. The outer membrane TonB-dependent transporters (TdTs) produced by N. gonorrhoeae are considered promising vaccine antigens as they are highly conserved and play crucial roles in overcoming nutritional immunity. One of these TdTs is part of the hemoglobin transport system comprised of HpuA and HpuB. This system allows N. gonorrhoeae to acquire iron from hemoglobin (hHb). In the current study, mutations in the hpuB gene were generated to better understand the structure-function relationships in HpuB. This study is one of the first to demonstrate that N. gonorrhoeae can bind to and utilize hemoglobin produced by animals other than humans. This study also determined that when HpuA is absent, mutations targeting extracellular loop 7 of HpuB led to defective hHb binding and utilization. However, when the lipoprotein HpuA is present, these loop 7 mutants recovered their ability to bind hHb, although the growth phenotype remained significantly impaired. Interestingly, loop 7 contains putative heme-binding motifs and a hypothetical α-helical region, both of which may be important for the use of hHb. Taken together, these results highlight the importance of loop 7 in the functionality of HpuB in binding hHb and extracting and internalizing iron.

Antimicrobial resistance and heterogeneity of Neisseria gonorrhoeae isolated from patients attending sexually transmitted infection clinics in Lusaka, Zambia.

BACKGROUND: Antimicrobial resistance (AMR) of Neisseria gonorrhoeae is a threat to public health as strains have developed resistance to antimicrobials available for the treatment of gonorrhea. Whole genome sequencing (WGS) can detect and predict antimicrobial resistance to enhance the control and prevention of gonorrhea. Data on the molecular epidemiology of N. gonorrhoeae is sparse in Zambia. This study aimed to determine the genetic diversity of N. gonorrhoeae isolated from patients attending sexually transmitted infection (STI) clinics in Lusaka, Zambia. METHODS: A cross-sectional study that sequenced 38 N. gonorrhoeae isolated from 122 patients with gonorrhea from 2019 to 2020 was conducted. The AMR profiles were determined by the E-test, and the DNA was extracted using the NucliSens easyMaG magnetic device. Whole genome sequencing was performed on the Illumina NextSeq550 platform. The Bacterial analysis pipeline (BAP) that is readily available at: https://cge.cbs.dtu.dk/services/CGEpipeline-1.1 was used for the identification of the species, assembling the genome, multi-locus sequence typing (MLST), detection of plasmids and AMR genes. Phylogeny by single nucleotide polymorphisms (SNPs) was determined with the CCphylo dataset. RESULTS: The most frequent STs with 18.4% of isolates each were ST7363, ST1921 and ST1582, followed by ST1583 (13%), novel ST17026 (7.9%), ST1588 (7.9%), ST1596 (5.3%), ST11181 (5.3%), ST11750 (2.6/%) and ST11241 (2.6%) among the 38 genotyped isolates. The blaTeM-1B and tetM (55%) was the most prevalent combination of AMR genes, followed by blaTeM-1B (18.4%), tetM (15.8%), and the combination of blaTeM-1B, ermT, and tetL was 2.6% of the isolates. The AMR phenotypes were predicted in ciprofloxacin, penicillin, tetracycline, azithromycin, and cefixime. The combination of mutations 23.7% was gryA (S91F), parC (E91G), ponA (L421) and rpsJ (V57M), followed by 18.4% in gyrA (S91F), ponA (L421P), rpsJ (V57M), and 18.4% in gyrA (D95G, S91F), ponA (L421P), and rpsJ (V57M). The combinations in gyrA (D95G, S91F) and rpsJ (V57M), and gyrA (D95G, S91F), parC (E91F), ponA (L421P) and rpsJ (V57M) were 13.2% each of the isolates. Plasmid TEM-1 (84.2%), tetM (15.8%), and gonococcal genetic island (GGI) was detected in all isolates. CONCLUSION: This study revealed remarkable heterogeneity of N. gonorrhoeae with blaTEM-1, tetM, ponA, gyrA, and parC genes associated with high resistance to penicillin, tetracycline, and ciprofloxacin demanding revision of the standard treatment guidelines and improved antimicrobial stewardship in Zambia.

Screening for Chlamydia and Gonorrhea in Youth Correctional Facilities, Utah, USA.

We reviewed data obtained in October 2021-May 2023 from youth who reported a history of sexual activity upon admission to 1 of 12 juvenile justice facilities in Utah, USA, that offered screening for Chlamydia trachomatis and Neisseria gonorrhoeae. Urinalysis revealed C. trachomatis positivity of 10.77%, N. gonorrhoeae positivity of 1.08%, and coinfection C. trachomatis N. gonorrhoeae) of 0.90%. Prevalence of infection was similar for youths in rural and urban facilities. A total of 12.01% of those identifying as male and 14.01% of those identifying as female tested positive for C. trachomatis, N. gonorrhoeae, or coinfection. Of young adults who tested positive, 74.65% received their results while incarcerated, all of whom accepted treatment. Our research underscores the feasibility of providing prompt C. trachomatis/N. gonorrhoeae screening and treatment in juvenile correctional facilities. The pervasiveness of infection emphasizes the urgent need for early identification and treatment for C. trachomatis and N. gonorrhoeae in incarcerated youth nationwide.

Rebound of Gonorrhea after Lifting of COVID-19 Preventive Measures, England.

After lifting of all COVID-19 preventive measures in England in July 2021, marked, widespread increases in gonorrhea diagnoses, but not testing numbers, were observed, particularly in persons 15-24 years of age. Continued close surveillance and public health messaging to young persons are needed to control and prevent gonorrhea transmission.