Rhein inhibits Chlamydia trachomatis infection by regulating pathogen-host cell.

The global incidence of genital Chlamydia trachomatis infection increased rapidly as the primary available treatment of C. trachomatis infection being the use of antibiotics. However, the development of antibiotics resistant stain and other treatment failures are often observed in patients. Consequently, novel therapeutics are urgently required. Rhein is a monomer derivative of anthraquinone compounds with an anti-infection activity. This study investigated the effects of rhein on treating C. trachomatis infection. Rhein showed significant inhibitory effects on the growth of C. trachomatis in multiple serovars of C. trachomatis, including D, E, F and L1, and in various host cells, including HeLa, McCoy and Vero. Rhein could not directly inactivate C. trachomatis but could inhibit the growth of C. trachomatis by regulating pathogen-host cell interactions. Combined with azithromycin, the inhibitory effect of rehin was synergistic both in vitro and in vivo. Together these findings suggest that rhein could be developed for the treatment of C. trachomatis infections.

Evaluation of the Predictive Value of Urine Leukocyte Esterase Test in Chlamydia trachomatis and Neisseria gonorrhoeae Infection Among Males Attending HIV/STI Clinics in Guangdong Province, China.

Leukocyte esterase test (LET) detection is a simple and inexpensive test performed by urinalysis. This study investigated the predictive value of LET for Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) infection among men attending HIV and sexually transmitted infection (HIV/STI) clinics in Guangdong Province, China. A total of 5,509 urine samples were collected from HIV and sexually transmitted infection clinics in Guangdong Province between 2017 and 2019. Specimens from 5,464 males were tested by both LET and nucleic acid amplification test (NAAT). Of 5,464 males, 497 (9.1%) tested positive for CT or NG by NAAT, with respective prevalence rates of 6.4% (95% confidence interval [95% CI]: 5.8-7.1%) and 3.8% (95% CI: 3.3-4.3%), including 1.2% (95% CI: 0.9-1.4%) co-infected. Compared to the HIV-negative individuals, individuals living with HIV tend to have a higher prevalence of CT, NG and co-infection with CT and NG. The LET sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) for CT were 46.4% (95% CI: 41.2-51.7%), 92.0% (95% CI: 91.2-92.7%), 28.4% (95% CI: 24.8-32.1%), and 96.1% (95% CI: 95.6-96.7%), respectively. The LET sensitivity, specificity, PPV, and NPV for NG were 68.4% (95% CI: 62.1-74.7%), 91.8% (95% CI: 91.1-92.6%), 25.0% (95% CI: 21.4-28.5%), and 98.7% (95% CI: 98.3-99%), respectively. Compared to the HIV-negative individuals, higher sensitivity and specificity were observed for HIV-positive individuals, but there was no statistical difference. The incremental cost-effectiveness ratio (ICER) using economic costs per additional person CT positive and NG positive was -$238.74 and -$145.60 compared with LET positive, respectively. LET is a cost-effective test and will be valuable for predicting CT and NG infection, which is highly prevalent in low- and middle-income countries.

Development and application of Cas13a-based diagnostic assay for Neisseria gonorrhoeae detection and azithromycin resistance identification.

BACKGROUND: Gonorrhoea, caused by Neisseria gonorrhoeae, has spread worldwide. Strains resistant to most antibiotics, including ceftriaxone and azithromycin, have emerged to an alarming level. Rapid testing for N. gonorrhoeae and its antimicrobial resistance will therefore contribute to clinical decision making for early diagnosis and rational drug use. METHODS: A Cas13a-based assay (specific high-sensitivity enzymatic reporter unlocking; SHERLOCK) was developed for N. gonorrhoeae detection (porA gene) and azithromycin resistance identification (A2059G, C2611T). Assays were evaluated for sensitivity with purified dsDNA and specificity with 17 non-gonococcal strains. Performance of SHERLOCK (porA) was compared with Roche Cobas 4800 using 43 urine samples. Identification of azithromycin resistance mutations (A2059G, C2611T) was evaluated using a total of 84 clinical isolates and 18 urine samples. Lateral flow was tested for this assay as a readout tool. Moreover, we directly assayed 27 urethral swabs from patients with urethritis to evaluate their status in terms of N. gonorrhoeae infection and azithromycin resistance. RESULTS: The SHERLOCK assay was successfully developed with a sensitivity of 10 copies/reaction, except 100 copies/reaction for A2059G, and no cross-reaction with other species. Comparison of the SHERLOCK assay with the Cobas 4800 revealed 100% concordance within 18 positive and 25 negative urine samples. Of the 84 isolates, 21 strains with azithromycin resistance mutations were distinguished and further verified by sequencing and MIC determination. In addition, 62.96% (17/27) strains from swab samples were detected with no mutant strains confirmed by sequencing. CONCLUSIONS: The SHERLOCK assay for rapid N. gonorrhoeae detection combined with azithromycin resistance testing is a promising method for application in clinical practice.

Inhibition of the Extracellular Signal-Regulated Kinase/Ribosomal S6 Kinase Cascade Limits Chlamydia trachomatis Infection.

Chlamydiatrachomatis is the cause of the most common bacterial sexually transmitted infection worldwide. Azithromycin is effective in treating chlamydial infection; however, resistance to this antibiotic is increasing, and it is important that new therapeutic strategies are developed. In this study, we demonstrated that inhibitors targeting each kinase in the extracellular signal-regulated kinase/ribosomal S6 kinase cascade significantly decreased the size and number of inclusions as well as the number of infectious progeny. The suppressive effects of the inhibitors were observed across the Chlamydia serotypes D, E, F, and L1 and across HeLa, McCoy, and Vero host cells. When combined with azithromycin, all the inhibitors exerted a synergistic suppressive effect on chlamydial infection. Knockdown experiments using small interfering RNA demonstrated that extracellular signal-regulated kinase 1/2 and ribosomal S6 kinase 1 were crucial for chlamydial infection. Moreover, BVD-523, a first-in-class extracellular signal-regulated kinase 1/2 inhibitor currently undergoing a phase II clinical trial, suppressed chlamydial infection both in cell culture and in a mouse model. These observations demonstrated not only that the extracellular signal-regulated kinase/ribosomal S6 kinase pathway plays a critical role in chlamydial infection but also that these kinases have potential as targets for host-directed therapy against C. trachomatis.

Prevalence and Genotype Distribution of Chlamydia trachomatis in Urine among Men Attending Sexually Transmitted Disease Clinics in Guangdong Province, China, in 2016.

Studies have rarely assessed the genotype distribution of Chlamydia trachomatis (CT) in urine among men attending sexually transmitted disease clinics (MSCs) in China. This study was aimed at investigating the prevalence and molecular epidemiology of CT infection by examining urine samples among MSCs from different geographic areas of Guangdong Province, China. A cross-sectional study was conducted among MSCs from 10 human immunodeficiency virus sentinel surveillance sites in Guangdong Province. CT DNA was extracted from male urine samples and analyzed using a Roche cobas 4800 CT/NG. The ompA genes were amplified by nested PCR and sequenced. The leukocyte esterase test was performed by routine urine analysis at local clinics. Of the 1,903 samples, 163 (8.6%, 95% confidence interval [CI] 3.8-16.3%) tested positive for CT. The highest prevalence (10.5%) of CT infection was observed among participants aged between 21 and 30 years. A total of 130 CT-positive samples (79.8%, 130/163) were successfully genotyped by nested PCR, resulting in 8 genotypes. The most prevalent genotypes were D, E, F, and J, with proportions of 20.8%, 20.0%, 17.7%, and 16.9%, respectively. There were no significant correlations between the geographical areas, leukocyte esterase test results and genotype distribution. Promotion of detection and molecular epidemiology research is needed for effective and comprehensive prevention and control programs.

An in vitro model of azithromycin-induced persistent Chlamydia trachomatis infection.

Single-dose azithromycin is recommended for treating Chlamydia trachomatis infections. Here, we established an in vitro cell model of azithromycin-induced persistent infection. Azithromycin inhibited the replication of C. trachomatis in a dose-time-dependent manner. Electron microscopy indicated that small inclusions in the induced model contained enlarged, aberrant and non-infectious reticulate bodies. RT-PCR showed that C. trachomatis still has the ability to express the unprocessed 16S rRNA gene in the model and that C. trachomatis recovered after the removal of azithromycin with a peak recovery time of 24 h. The mutations in 23S rRNA, L4 and L22 genes were not found in persistent infection, and qRT-PCR analysis showed that the relative expression level of euo in azithromycin treated infection was upregulated while omcB was downregulated. In summary, this study provides a novel in vitro cell model to examine the characteristics of azithromycin-induced persistent infection and contribute to the development of treatments for C. trachomatis infection.