Development of a genomics-based PCR assay for detection of Mycoplasma pneumoniae in a large outbreak in New York State.

A genomics-based PCR method was developed and used to test specimens from patients involved in a large outbreak of Mycoplasma pneumoniae in a closed religious community in New York State. New P1 adhesin gene primers were designed to bind to 9 of 10 target sequences in the repetitive-element sequences obtained from the whole genome sequence of M. pneumoniae. This PCR method had a sensitivity of 0.006 CFU and a specificity of 100% for M. pneumoniae. The PCR was validated by testing a subset of patient samples by culture and comparing the results to those obtained by PCR. Of the initial 280 samples tested, 73 were positive by PCR and 22 were positive by culture. All samples positive by culture were also positive by PCR. Follow-up testing of selected patients 3 to 6 weeks after antibiotic treatment revealed that eight samples remained positive by PCR and that three samples remained positive by culture. Additionally, no nonspecific PCR inhibition was detected as a result of the specimen type, transport medium, or sample preparation methodology. The study demonstrates that the PCR described here is a rapid, sensitive, and specific method for the identification of M. pneumoniae and was helpful for the detection and monitoring of the outbreak.

The emergence of Neisseria gonorrhoeae with decreased susceptibility to ciprofloxacin in Cleveland, Ohio: epidemiology and risk factors.

BACKGROUND: Until 1992, almost all strains of Neisseria gonorrhoeae that had been tested in the United States were susceptible to fluoroquinolones, including ciprofloxacin. However, among men with urethral gonococcal infections who attended one sexually transmitted disease clinic in Cleveland, Ohio, the prevalence of gonococci with decreased susceptibility to ciprofloxacin increased from 2% in 1991 to 16% in 1994. OBJECTIVE: To describe the emergence of and risk factors for gonococcal urethritis caused by gonococci with decreased susceptibility to ciprofloxacin. Resistance to ciprofloxacin was considered to be decreased if the mean inhibitory concentration was at least 0.12 microgram/mL, and was less than or equal to 0.25 microgram/mL; this definition did not equate with the definition of clinical resistance. DESIGN: Case-control study. SETTING: An urban sexually transmitted disease clinic. PARTICIPANTS: 51 case-patients and 106 controls. MEASUREMENTS: Pulsed-field gel electrophoresis was used to identify individual genotypes of ciprofloxacin-resistant and ciprofloxacin-susceptible isolates. RESULTS: 55 of the 746 isolates of N. gonorrhoeae that were tested (7.4%) had decreased susceptibility to ciprofloxacin, and the prevalence of N. gonorrhoeae with decreased susceptibility significantly increased during the study period. Case-patients were significantly less likely to have gram-negative diplococci seen on microscopic examination of urethral discharge (P < or = 0.01) and were less likely to be treated for gonococcal urethritis than were controls (P < or = 0.001). Molecular typing suggested the spread of a single genotype of N. gonorrhoeae. CONCLUSIONS: Strains of gonococci with decreased susceptibility to ciprofloxacin appear to have become endemic in Cleveland, Ohio. The clinical significance of these isolates is not clear, but the potential for the emergence of clinically important resistance may preclude the use of fluoroquinolones as an alternative treatment for uncomplicated gonorrhea.

Activities of three investigational fluoroquinolones (BAY y 3118, DU-6859a, and clinafloxacin) against Neisseria gonorrhoeae isolates with diminished susceptibilities to ciprofloxacin and ofloxacin.

Between 1 January 1992 and 31 December 1993, our laboratory, as part of the Gonococcal Isolate Surveillance Project, found that 39 of 673 isolates of Neisseria gonorrhoeae from one local sexually transmitted diseases clinic demonstrated decreased susceptibilities to both ciprofloxacin and ofloxacin. The MICs of BAY y 3118, DU-6859a, and clinafloxacin at which 90% of the gonococci with decreased susceptibility to ciprofloxacin and ofloxacin were inhibited were all 0.016 microgram/ml, which was eightfold higher than those for ciprofloxacin-susceptible gonococci. Our report substantiates prior observations that diminished susceptibility to one quinolone is often associated with diminished susceptibility to other quinolones.