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.

Prevalence of Four Sexually Transmitted Infection Pathogens Detected in Urine and Genital Tract Secretion in Hangzhou, China.

BACKGROUND: Sexually transmitted infections (STIs) increase gradually and have become a public health problem in the world. UU, CT, NG, and MG are four common STI pathogens. Our retrospective study analyzed the clinical situation and the laboratory data of patients infected with the four pathogens. The prevalence of the four pathogens, detected in urine and genital tract secretion, was studied in Hangzhou, China. METHODS: A total of 3,168 male and female patients were randomly selected from February 2023 to February 2024. Urine and genital secretions were collected, and four STI pathogens were controlled for detection. Data were collected from the hospital's electronic medical records, and SPSS 25.0 software was used to perform a statistical analysis. RESULTS: Among 3,168 patients, a total of 1,527 were detected as positive, and the positive rate was 48.20%. The age of patients ranged from 13 - 98 years, with an average age of 45.6. The total of patients consisted of 2,191 males and 977 females, which had a significant difference (p < 0.05). Specimens were mainly collected from the Department of Dermatovenerology, Urological Surgery, Obstetrics and Gynecology, and so on. The positive rate was statistically different between male and female patients (p < 0.05). Single infection performed a main role and accounted for 79.57% of all of the positive patients. In the ≤ 20 age group, the positive rate was the highest and was as high as 77.65%. In detail, single infection caused by UU dominated, especially in the 21 - 30 age group. Double infection caused by UU and CT and triple infection caused by UU, CT, and NG were the majority, both especially in the 21 - 30 age group. There were significant differences in the positive rates in the different age groups and in the four pathogens (p < 0.05). Quadruple infection was very rare and had only been detected in one patient. CONCLUSIONS: The prevalence of the four pathogens in Hangzhou was different from other regions. More male than female patients, more single than multiple infections, and more single and multiple infections occurring in young people were the features in Hangzhou. The study would provide reference for prevention, diagnosis, and treatment of STI.

Computational prediction of crucial genes involved in gonorrhea infection and neoplastic cell transformation: A multiomics approach.

Neisseria gonorrheae, the causative agent of genitourinary infections, has been associated with asymptomatic or recurrent infections and has the potential to form biofilms and induce inflammation and cell transformation. Herein, we aimed to use computational analysis to predict novel associations between chronic inflammation caused by gonorrhea infection and neoplastic transformation. Prioritization and gene enrichment strategies based on virulence and resistance genes utilizing essential genes from the DEG and PANTHER databases, respectively, were performed. Using the STRING database, protein‒protein interaction networks were constructed with 55 nodes of bacterial proteins and 72 nodes of proteins involved in the host immune response. MCODE and cytoHubba were used to identify 12 bacterial hub proteins (murA, murB, murC, murD, murE, purN, purL, thyA, uvrB, kdsB, lpxC, and ftsH) and 19 human hub proteins, of which TNF, STAT3 and AKT1 had high significance. The PPI networks are based on the connectivity degree (K), betweenness centrality (BC), and closeness centrality (CC) values. Hub genes are vital for cell survival and growth, and their significance as potential drug targets is discussed. This computational study provides a comprehensive understanding of inflammation and carcinogenesis pathways that are activated during gonorrhea infection.

Emergence of Extensively Drug-Resistant Neisseria gonorrhoeae, France, 2023.

Since 2022, Europe has had 4 cases of extensively drug-resistant Neisseria gonorrhoeae, sequence type 16406, that is resistant to ceftriaxone and highly resistant to azithromycin. We report 2 new cases from France in 2023 involving strains genetically related to the 4 cases from Europe as well as isolates from Cambodia.

Transcriptional responses of Neisseria gonorrhoeae to glucose and lactate: implications for resistance to oxidative damage and biofilm formation.

Understanding how bacteria adapt to different environmental conditions is crucial for advancing knowledge regarding pathogenic mechanisms that operate during infection as well as efforts to develop new therapeutic strategies to cure or prevent infections. Here, we investigated the transcriptional response of Neisseria gonorrhoeae, the causative agent of gonorrhea, to L-lactate and glucose, two important carbon sources found in the host environment. Our study revealed extensive transcriptional changes that gonococci make in response to L-lactate, with 37% of the gonococcal transcriptome being regulated, compared to only 9% by glucose. We found that L-lactate induces a transcriptional program that would negatively impact iron transport, potentially limiting the availability of labile iron, which would be important in the face of the multiple hydrogen peroxide attacks encountered by gonococci during its lifecycle. Furthermore, we found that L-lactate-mediated transcriptional response promoted aerobic respiration and dispersal of biofilms, contrasting with an anaerobic condition previously reported to favor biofilm formation. Our findings suggest an intricate interplay between carbon metabolism, iron homeostasis, biofilm formation, and stress response in N. gonorrhoeae, providing insights into its pathogenesis and identifying potential therapeutic targets.IMPORTANCEGonorrhea is a prevalent sexually transmitted infection caused by the human pathogen Neisseria gonorrhoeae, with ca. 82 million cases reported worldwide annually. The rise of antibiotic resistance in N. gonorrhoeae poses a significant public health threat, highlighting the urgent need for alternative treatment strategies. By elucidating how N. gonorrhoeae responds to host-derived carbon sources such as L-lactate and glucose, this study offers insights into the metabolic adaptations crucial for bacterial survival and virulence during infection. Understanding these adaptations provides a foundation for developing novel therapeutic approaches targeting bacterial metabolism, iron homeostasis, and virulence gene expression. Moreover, the findings reported herein regarding biofilm formation and L-lactate transport and metabolism contribute to our understanding of N. gonorrhoeae pathogenesis, offering potential avenues for preventing and treating gonorrhea infections.

Surge in Ceftriaxone-Resistant Neisseria gonorrhoeae FC428-Like Strains, Asia-Pacific Region, 2015-2022.

Ceftriaxone-resistant Neisseria gonorrhoeae FC428-like strains have disseminated across the Asia-Pacific region, with a continuous rise in prevalence during 2015-2022. To mitigate the effect of these strains, we advocate for enhanced molecular diagnostics, expanded surveillance networks, and a regionally coordinated effort to combat the global spread of FC428-like strains.

Combination of compound screening with an animal model identifies pentamidine to prevent Chlamydia trachomatis infection.

Chlamydia trachomatis (Ct) is the most common cause for bacterial sexually transmitted infections (STIs) worldwide with a tremendous impact on public health. With the aim to unravel novel targets of the chlamydia life cycle, we screen a compound library and identify 28 agents to significantly reduce Ct growth. The known anti-infective agent pentamidine-one of the top candidates of the screen-shows anti-chlamydia activity in low concentrations by changing the metabolism of host cells impairing chlamydia growth. Furthermore, it effectively decreases the Ct burden upon local or systemic application in mice. Pentamidine also inhibits the growth of Neisseria gonorrhea (Ng), which is a common co-infection of Ct. The conducted compound screen is powerful in exploring antimicrobial compounds against Ct in a medium-throughput format. Following thorough in vitro and in vivo assessments, pentamidine emerges as a promising agent for topical prophylaxis or treatment against Ct and possibly other bacterial STIs.

Data quality assessment of the Enhanced Gonococcal Antimicrobial Surveillance Programme (EGASP), Thailand, 2015-2021.

BACKGROUND: Quality assessments of gonococcal surveillance data are critical to improve data validity and to enhance the value of surveillance findings. Detecting data errors by systematic audits identifies areas for quality improvement. We designed and implemented an internal audit process to evaluate the accuracy and completeness of surveillance data for the Thailand Enhanced Gonococcal Antimicrobial Surveillance Programme (EGASP). METHODS: We conducted a data quality audit of source records by comparison with the data stored in the EGASP database for five audit cycles from 2015-2021. Ten percent of culture-confirmed cases of Neisseria gonorrhoeae were randomly sampled along with any cases identified with elevated antimicrobial susceptibility testing results and cases with repeat infections. Incorrect and incomplete data were investigated, and corrective action and preventive actions (CAPA) were implemented. Accuracy was defined as the percentage of identical data in both the source records and the database. Completeness was defined as the percentage of non-missing data from either the source document or the database. Statistical analyses were performed using the t-test and the Fisher's exact test. RESULTS: We sampled and reviewed 70, 162, 85, 68, and 46 EGASP records during the five audit cycles. Overall accuracy and completeness in the five audit cycles ranged from 93.6% to 99.4% and 95.0% to 99.9%, respectively. Overall, completeness was significantly higher than accuracy (p = 0.017). For each laboratory and clinical data element, concordance was >85% in all audit cycles except for two laboratory data elements in two audit cycles. These elements significantly improved following identification and CAPA implementation. DISCUSSION: We found a high level of data accuracy and completeness in the five audit cycles. The implementation of the audit process identified areas for improvement. Systematic quality assessments of laboratory and clinical data ensure high quality EGASP surveillance data to monitor for antimicrobial resistant Neisseria gonorrhoeae in Thailand.

Impact of mutations in the mtrR, rpdlVD and rrl genes on azithromycin resistance in Neisseria gonorrhoeae.

INTRODUCTION: Bacterial sexually transmitted infections (STIs) pose a major public health problem. The emergence of antibiotic-resistant strains of Neisseria gonorrhoeae represents a serious threat to successful treatment and epidemiological control. The first extensively drug-resistant (XDR) strains (ceftriaxone-resistant and high-level azithromycin-resistant [HLR AZY]) have been reported. AIMS: To identify molecular mechanisms implicated in azithromycin resistance in strains isolated from patients over a three-year period in a university hospital in Switzerland. MATERIAL AND METHODS: From January 2020 to December 2022, 34 isolates (one per patient) were recovered from samples analyzed at the University Hospital of Lausanne. Eight genes involved in azithromycin resistance were sequenced: mtrR repressor (mtrCDE operon repressor) and his promotor mtrR-pr, rplD gene (L4 ribosomal protein), rplV gene (L22 ribosomal protein) and the four alleles of the rrl gene (23S rRNA). RESULTS: With a cutoff value of 1 mg/L, 15 isolates were considered as being resistant to azithromycin, whereas the remaining 19 were susceptible. The C2597T mutation in 3 or 4 of the rrl allele confer a medium-level resistance to azithromycin (MIC = 16 mg/L, N = 2). The following mutations were significantly associated with MIC values ≥1 mg/L: the three mutations V125A, A147G, R157Q in the rplD gene (N = 10) and a substitution A->C in the mtrR promotor (N = 9). Specific mutations in the mtrR repressor and its promotor were observed in both susceptible and resistant isolates. CONCLUSIONS: Resistance to azithromycin was explained by the presence of mutations in many different copies of 23S RNA ribosomal genes and their regulatory genes. Other mutations, previously reported to be associated with azithromycin resistance, were documented in both susceptible and resistant isolates, suggesting they play little role, if any, in azithromycin resistance.

The effect of daily usage of Listerine Cool Mint mouthwash on the oropharyngeal microbiome: a substudy of the PReGo trial.

Introduction. ListerineÒ is a bactericidal mouthwash widely used to prevent oral health problems such as dental plaque and gingivitis. However, whether it promotes or undermines a healthy oral microbiome is unclear.Hypothesis/Gap Statement. We hypothesized that the daily use of Listerine Cool Mint would have a significant impact on the oropharyngeal microbiome.Aim. We aimed to assess if daily usage of Listerine Cool Mint influenced the composition of the pharyngeal microbiome.Methodology. The current microbiome substudy is part of the Preventing Resistance in Gonorrhoea trial. This was a double-blind single-centre, crossover, randomized controlled trial of antibacterial versus placebo mouthwash to reduce the incidence of gonorrhoea/chlamydia/syphilis in men who have sex with men (MSM) taking HIV pre-exposure prophylaxis (PrEP). Fifty-nine MSM taking HIV PrEP were enrolled. In this crossover trial, participants received 3 months of daily Listerine followed by 3 months of placebo mouthwash or vice versa. Oropharyngeal swabs were taken at baseline and after 3 months use of each mouthwash. DNA was extracted for shotgun metagenomic sequencing (Illumina Inc.). Non-host reads were taxonomically classified with MiniKraken and Bracken. The alpha and beta diversity indices were compared between baseline and after each mouthwash use. Differentially abundant bacterial taxa were identified using ANOVA-like differential expression analysis.Results. Streptococcus was the most abundant genus in most samples (n = 103, 61.7 %) with a median relative abundance of 31.5% (IQR 20.6-44.8), followed by Prevotella [13.5% (IQR 4.8-22.6)] and Veillonella [10.0% (IQR 4.0-16.8)]. Compared to baseline, the composition of the oral microbiome at the genus level (beta diversity) was significantly different after 3 months of Listerine (P = 0.006, pseudo-F = 2.29) or placebo (P = 0.003, pseudo-F = 2.49, permutational multivariate analysis of variance) use. Fusobacterium nucleatum and Streptococcus anginosus were significantly more abundant after Listerine use compared to baseline.Conclusion. Listerine use was associated with an increased abundance of common oral opportunistic bacteria previously reported to be enriched in periodontal diseases, oesophageal and colorectal cancer, and systemic diseases. These findings suggest that the regular use of Listerine mouthwash should be carefully considered.

Overcoming analytical and preanalytical challenges associated with extragenital home collected STI specimens.

Home sample collection for sexually transmitted infection (STI) screening options can improve access to sexual healthcare across communities. For Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG), genital infections have classically been the focus for remote collection options. However, infections may go undiagnosed if sampling is limited to urogenital sites because some individuals only participate in oral and/or anal intercourse. Here we evaluated samples for CT/NG detection after several pre-analytical collection challenges. A paired provider to self-collection validation was performed on rectal [n = 162; 22 + for CT and 9 + for NG by provider-collected (PC)] and throat (N = 158; 2 + for CT and 11 + for NG by provider-collected) swabs. The positive percent agreement for CT and NG ranged from 90.9% to 100%. The discrepancies were more often positive on self-collected (SC) (n = 9 SC+/PC-; n = 1 PC+/SC-; n = 1 PC+/SC Equiv.; n = 2 PC-/SC Equiv.). An empirical limit of detection (LoD) lower than the manufacturer's claim (0.031 vs 2.5 IFU/mL for CT and 0.063 vs 124.8 CFU/ml for NG, respectively) was used to challenge additional variables. Common hand contaminants, including soap, hand sanitizer, lotion, and sunscreen were added to known positive (3× empirical LoD) or negative samples and did not influence detection. Samples at 2× and 10× the empirical LoD were challenged with extreme temperature cycling and extended room temperature storage. Detection was not affected by these conditions. These results indicate that remote self-collection is an appropriate method of sample acquisition for detecting extragenital CT/NG infections. Additionally, they provide a foundation towards meeting the regulatory standards for commercial testing of home collected extragenital samples. IMPORTANCE: There is a clinical need for expanded extragenital bacterial sexually transmitted infection (STI) testing options, but the current regulatory landscape limits the wide-spread promotion and adoption of such services. Improved access, particularly for the LGBTQ+ community, can be achieved by validating testing for specimens that are self-collected at a remote location and arrive at the laboratory via a postal carrier or other intermediary route. Here we provide valuable data showing that self-collected samples for anal and oropharyngeal STI testing are equally or increasingly sensitive compared with those collected by a provider. We systematically consider the effects of storage time, exposure to temperature extremes, and the addition of common toiletries on results.

Isolation and Genomic Characterization of Schaalia turicensis from a Patient with Gonococcal Urethritis, Thailand.

Schaalia turicensis is facultative anaerobic Gram-positive bacillus that commonly inhabits the oropharynx, gastrointestinal, and genitourinary tract of healthy individuals. This organism has been co-isolated with Neisseria gonorrhoeae from 15-year-old Thai male patient with gonococcal urethritis in Bangkok, Thailand. In this study, we characterized the class 1 integron in S. turicensis isolate using whole-genome sequencing and bioinformatics analysis. Sequencing analysis confirmed the presence of an imperfect class 1 integron located on chromosome and a novel 24.5-kb-long composite transposon, named Tn7083. The transposon Tn7083 carried genes encoding chloramphenicol resistance (cmx), sulfonamide resistance (sul1), and aminoglycoside resistance [aph(6)-Id (strB), aph(3'')-Ib (strA), aph(3')-Ia].

Mini-Multilocus Sequence Typing Scheme for the Global Population of Neisseria gonorrhoeae.

The increasing problem of antimicrobial resistance in N. gonorrhoeae necessitates the development of molecular typing schemes that are suitable for rapid and mass screening. The objective of this study was to design and validate a mini-MLST scheme for N. gonorrhoeae based on global pathogen population data. Using sequences of seven housekeeping genes of 21,402 isolates with known MLSTs from the PubMLST database, we identified eighteen informative polymorphisms and obtained mini-MLST nucleotide profiles to predict MLSTs of isolates. We proposed a new MLST grouping system for N. gonorrhoeae based on mini-MLST profiles. Phylogenetic analysis revealed that MLST genogroups are a stable characteristic of the N. gonorrhoeae global population. The proposed grouping system has been shown to bring together isolates with similar antimicrobial susceptibility, as demonstrated by the characteristics of major genogroups. Established MLST prediction algorithms based on nucleotide profiles are now publicly available. The mini-MLST scheme was evaluated using a MLST detection/prediction method based on the original hydrogel DNA microarray. The results confirmed a high predictive ability up to the MLST genogroup. The proposed holistic approach to gonococcal population analysis can be used for the continuous surveillance of known and emerging resistant N. gonorrhoeae isolates.

Performance of Cepheid CT/NG Xpert rapid PCR test for detection of Chlamydia trachomatis and Neisseria gonorrhoeae at a tertiary care centre in North India.

INTRODUCTION: Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) are prevalent causes of sexually transmitted infections (STIs) globally, leading to substantial morbidity and transmission risks. METHODS: This study evaluates the diagnostic efficacy of Xpert CT/NG compared to conventional PCR and culture methods in 121 patients at a tertiary care centre in North India. RESULTS: Xpert CT/NG demonstrated high sensitivity (85.8%) and specificity (96.3%) outperforming conventional PCR. Xpert CT/NG's rapidity and accuracy underscore its utility in timely diagnosis and control of STIs. CONCLUSION: As sexually transmitted infections pose a serious health concern implementation of such rapid diagnostic methods/point of care testing methods are to be implemented for early diagnosis.

The novel 2024 WHO Neisseria gonorrhoeae reference strains for global quality assurance of laboratory investigations and superseded WHO N. gonorrhoeae reference strains-phenotypic, genetic and reference genome characterization.

OBJECTIVES: MDR and XDR Neisseria gonorrhoeae strains remain major public health concerns internationally, and quality-assured global gonococcal antimicrobial resistance (AMR) surveillance is imperative. The WHO global Gonococcal Antimicrobial Surveillance Programme (GASP) and WHO Enhanced GASP (EGASP), including metadata and WGS, are expanding internationally. We present the phenotypic, genetic and reference genome characteristics of the 2024 WHO gonococcal reference strains (n = 15) for quality assurance worldwide. All superseded WHO gonococcal reference strains (n = 14) were identically characterized. MATERIAL AND METHODS: The 2024 WHO reference strains include 11 of the 2016 WHO reference strains, which were further characterized, and four novel strains. The superseded WHO reference strains include 11 WHO reference strains previously unpublished. All strains were characterized phenotypically and genomically (single-molecule PacBio or Oxford Nanopore and Illumina sequencing). RESULTS: The 2024 WHO reference strains represent all available susceptible and resistant phenotypes and genotypes for antimicrobials currently and previously used (n = 22), or considered for future use (n = 3) in gonorrhoea treatment. The novel WHO strains include internationally spreading ceftriaxone resistance, ceftriaxone resistance due to new penA mutations, ceftriaxone plus high-level azithromycin resistance and azithromycin resistance due to mosaic MtrRCDE efflux pump. AMR, serogroup, prolyliminopeptidase, genetic AMR determinants, plasmid types, molecular epidemiological types and reference genome characteristics are presented for all strains. CONCLUSIONS: The 2024 WHO gonococcal reference strains are recommended for internal and external quality assurance in laboratory examinations, especially in the WHO GASP, EGASP and other GASPs, but also in phenotypic and molecular diagnostics, AMR prediction, pharmacodynamics, epidemiology, research and as complete reference genomes in WGS analysis.

Gonococcal mastitis. / Gonokokkmastitt.

A young woman experienced pain and swelling in a non-lactating breast. The culture test result showed an unusual microbe, which is increasingly prevalent in Norway and internationally.