The present study investigates the suitability of direct bacterial profiling as a tool for the identification and subtyping of pathogenic Neisseria. The genus Neisseria includes two human pathogens , Neisseria meningitidis and Neisseria gonorrhoeae, as well as several nonpathogenic Neisseria species. Here , a matrix-assisted laser desorption/ionization time-of-flight mass spectrometry profiling protocol was optimized using a laboratory strain of E. coli DH5␣ to guarantee high quality and reproducible results. Subsequently , mass spectra for both laboratory and clinical strains of N. gonorrhoeae, N. meningitidis, and several nonpathogenic Neisseria species were collected. Significant interspecies differences but little intraspecies diversity were revealed by means of a visual inspection and bioinformatics examination using the MALDI BioTyper software. Cluster analysis successfully separated mass spectra collected from three groups that corresponded to N. gonorrhoeae , N. meningitidis , and nonpathogenic Neisseria isolates. Requiring only one bacterial colony for testing and using a fast and easy measuring protocol , this approach represents a powerful tool for the rapid identification of pathogenic Neisseria and can be adopted for other microorganisms.

The present study documents the first case of fluoroquinolone-resistant N. gonorrhoeae in Russia.

The main goal of this work is to clarify the predictive value of known genetic markers of Neisseria gonorrhoeae resistance to penicillin, tetracycline, and fluoroquinolones. The correlation between the presence of certain genetic markers and susceptibility of N. gonorrhoeae isolates to penicillin, tetracycline, and fluoroquinolones has been analyzed by means of statistical methods. Susceptibility testing with penicillin, tetracycline, and fluoroquinolones was performed by the agar dilution method. N. gonorrhoeae genomic DNA was isolated. The presence of bla TEM-1 and tet(M) genes was analyzed by PCR. A novel method of polymorphism discovery based on a minisequencing reaction followed by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry was applied for the analysis of chromosomal N. gonorrhoeae genes involved in antimicrobial resistance development. Clinical N. gonorrhoeae isolates (n ‫؍‬ 464) were collected. Susceptibility levels to penicillin, tetracycline, and fluoroquinolones were found to be 25.9%, 35.9%, and 54.1%, respectively. Among the 19 N. gonorrhoeae isolates with penicillin MICs of >4 g/ml, the bla TEM-1 gene was detected in 12. The Tet(M) determinant was found in 4 of 12 N. gonorrhoeae isolates with tetracycline MICs of >16 g/ml. The chromosomal genetic markers of penicillin and tetracycline resistance were detected especially in isolates with penicillin MICs of 0.25 to 2.0 g/ml and tetracycline MICs of 0.5 to 4 g/ml. Mutations in GyrA and ParC were found in 208 of 211 quinoloneresistant N. gonorrhoeae isolates. This work is the first representative molecular research of the N. gonorrhoeae population in Russia. Information about the prevalence of antibiotic resistance mechanisms and the positive predictive value of certain genetic determinants is given. The positive predictive values of the analyzed genetic markers were found to be different for fluoroquinolones (90.3%), penicillin (91.1%), and tetracycline (81.9%).

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Genetic polymorphism of Russian population of N. gonorrhoeae was detected and a system for genotyping of its clinical strains was introduced into practice. Comparative analysis of the prevalence of N. gonorrhoeae genotypes in Russia and abroad was carried out. For adaptation of the methods of molecular typing of N. gonorrhoeae strains and its approbation on clinical strains isolated in Russia 41 clinical strains of N. gonorrhoeae were typed. The predominance of PIB serovar (83%) was demonstrated.