Objective. To study the prevalence and mechanisms of antibiotic resistance, including carbapenemase production, in clinical isolates of Klebsiella pneumoniae and Escherichia coli isolated in different regions of Russia as part of the sentinel multicenter surveillance study in 2020–2021, and to explore the population structure of K. pneumoniae and the impact of “high-risk clones” on antibiotic resistance. Materials and Methods. Consecutive, non-duplicate isolates of K. pneumoniae (n = 2503) and E. coli (n = 2055) isolated from various specimens (blood, cerebrospinal fluid, respiratory samples, urine, wound secretions, etc.) of hospitalized patients with clinical signs of infection in 55 hospitals of 29 cities of Russia were studied. Species identification of isolates was performed by matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry (MALDI-TOF MS). Antibiotic susceptibilities were determined by serial broth microdilution or, in the case of fosfomycin, by agar dilution method, and results were interpreted according to EUCAST v13 MIC breakpoints. Carbapenemase production was determined phenotypically by carbapenem inactivation method (CIM), the presence of genes of the most common serine carbapenemases (KPC, OXA-48) and metallo-β-lactamases (VIM, IMP, NDM) was determined by real-time PCR. K. pneumoniae clinical isolates were genotyped and assigned to the known clonal complexes (CC) and sequence types (ST) using SNP typing and multilocus sequencing typing (MLST) methods. K- and O-serotypes, acquired resistance and virulence genes, and plasmids carrying these genes were characterized using whole-genome sequencing of selected isolates (n = 215). Results. The resistance rates of nosocomial/community-acquired isolates of K. pneumoniae were as follows: amoxicillin-clavulanate – 88.63⁄57.99%, piperacillin-tazobactam – 82.92⁄45.49%, cefotaxime – 87.74⁄56.97%, ceftazidime – 84.76⁄53.07%, cefepime – 81.43⁄49.18%, aztreonam – 1.63⁄53.28%, ceftazidime-avibactam – 30, 88⁄9.22%, ceftolozan-tazobactam – 70.06/31.35%, ertapenem – 72.10⁄28.69%, meropenem – 49.60⁄15.16%, imipenem – 44.54⁄13.73%, gentamicin – 60.82⁄30.33%, amikacin – 42.06/17.21%, ciprofloxacin – 85.10⁄49.39%; trimethoprimsulfamethoxazole – 74.38⁄48.16%, colistin – 5.96⁄2.25%. The resistance of nosocomial/outpatient isolates of E. coli were: ampicillin – 84.93⁄67.67%, amoxicillin-clavulanate – 57.37⁄39.73%, piperacillin-tazobactam – 19.48⁄8.70%, cefotaxime – 63.83⁄34.19%, ceftazidime – 45.32⁄20.34%, cefepime – 35.95⁄16.61%, aztreonam – 51.78⁄26.11%, ceftazidime-avibactam – 5.71⁄0.80%, ceftolozane-tazobactam – 11, 95⁄2.22%, ertapenem – 8.18⁄1.42%, meropenem – 5.17⁄0.53%, imipenem – 4.95⁄0.36%, gentamicin – 24.54⁄13.68%, amikacin – 5.49⁄1.42%, ciprofloxacin – 54, 14⁄32.50%, trimethoprim-sulfamethoxazole – 52.21⁄38.54%, fosfomycin – 2.48⁄1.43%, colistin – 1.60⁄1.07%, tigecycline – 6.35⁄3.11%. The frequency of carbapenemase production among K. pneumoniae nosocomial isolates was 65.32% (OXA-48 – 40.75%, NDM – 30.28%, KPC – 8.74%, OXA-48 + NDM – 10.62%, OXA-48 + KPC – 2.98%, NDM + KPC – 0.45%, OXA-48 + NDM + KPC – 0.20%). More than 70% of nosocomial isolates of K. pneumoniae belonged to only 7 major genetic lineages known as “high-risk international clones”: CC395 – 37.40%, CC23 – 9.59%, CC307 – 8.64%, CC147 – 7.61%, CC15 – 2.95%, CC258 – 2.92%, and CC11 – 2.41%. The population of community-acquired K. pneumoniae was characterized by significantly greater genetic diversity (Simpson diversity index: D = 0.919; 95% CI: 0.904 to 0.933) compared with the population of nosocomial strains (Simpson diversity index: D = 0.815; 95% CI: 0.802 to 0.827). Strains of the “hypervirulent” genetic lineage of K. pneumoniae CC23 were more common in community-acquired infections. Conclusions. The extremely high frequency of resistance to cephalosporins in K. pneumoniae (> 80%) and E. coli (> 60%), as well as the high frequency of combined resistance to aminoglycosides and fluoroquinolones precludes their empirical use for the treatment of serious nosocomial infections caused by these pathogens. K. pneumoniae shows a rapid increase in resistance to carbapenems, mainly due to the spread of carbapenemases of three major groups: OXA-48, NDM and KPC. The overall increase in the frequency of carbapenemase production is accompanied by the growing diversity of carbapenemases, the increasing prevalence of strains producing NDM and KPC enzymes and those co-producing multiple carbapenemases simultaneously. In community-acquired infections, the high prevalence of resistance to cephalosporins in E. coli (> 30%) and K. pneumoniae (> 50%) remains the most important problem.

Objective. To assess the prevalence and antimicrobial susceptibility of carbapenemase-producing Enterobacterales (CPE). Materials and Methods. A total of 5539 Enterobacterales isolates recovered from hospitalized patients in 52 medical institutions in 27 cities in Russia in 2014-2016 were tested. Antibiotic susceptibility testing (AST) was performed using broth microdilution method according to ISO 20776-1:2006. Avibactam was tested in combinations with beta-lactams at fixed concentration of 4 mg/l. AST results were interpreted according to EUCAST v8.

Objective. To assess the in vitro activity of ceftazidime-avibactam against clinical Enterobacterales and Pseudomonas aeruginosa isolates in various regions of Russia based on results of local susceptibility testing by disk diffusion method. Materials and Methods. Overall, 160 laboratories located in 61 Russian cities participated in this surveillance during 2018-2020. All consecutive clinical isolates of Enterobacterales and Pseudomonas aeruginosa in each participating laboratory were included in the study. Ceftazidime-avibactam susceptibility testing was done by disc-diffusion method in accordance with current EUCAST recommendations. Susceptibility data for carbapenems and III-IV generation cephalosporins, as well as results of carbapenemases detection, were also reported, if available. All the data were recorded in electronic case report form developed on the OpenClinica online platform (www.openclinica.com). Data analysis and reporting were done using AMRcloud online platform (https://amrcloud.net/). Results. In total, we received information on antimicrobial susceptibility of 22,121 isolates, including 17,456 (78.9%) Enterobacterales and 4,665 (21.1%) P. aeruginosa. Less than 9% of Enterobacterales isolates were resistant to ceftazidime-avibactam. At the same time rates of resistance to ceftazidime, cefotaxime, cefepime, ertapenem, imipenem, and meropenem were 54.1%, 58.9%, 59.4%, 41.4%, 23.9%, and 21.3%. Among Enterobacterales the highest level of resistance to ceftazidime-avibactam was detected in K. pneumoniae (16.5%), lowest – in E. coli (2.1%). Some increase of resistance to ceftazidimeavibactam was noted during the study – from 7.8% in 2018-2019 to 9.6% in 2020 (p = 0.0001). Rate of resistance to ceftazidime-avibactam in P. aeruginosa was 33.1%. At the same time rates of resistance to ceftazidime, cefepime, imipenem, and meropenem were 51.1%, 54.5%, 50%, and 47.3%. During the study there was statistically significant decrease in resistance to ceftazidime-avibactam in P. aeruginosa (p = 0.0001). Resistance rates for all beta-lactams for both Enterobacterales and P. aeruginosa were higher in nosocomial isolates than in community-acquired isolates. Conclusions. Ceftazidime-avibactam demonstrated significantly higher in vitro activity against Enterobacterales and P. aeruginosa Russian clinical isolates comparing with commonly used carbapenems and extended spectrum cephalosporins. Access for all study data available at the AMRcloud online platform (https://amrcloud.net/ru/project/cazavi-1-2/).

Objective. To analyze the current taxonomic structure of the genus Stenotrophomonas based on genomic data, taking into account limitations associated with nomenclatural validation processes. Materials and Methods. A reference dataset was constructed using taxonomic information from LPSN (List of Prokaryotic names with Standing in Nomenclature) and genome assemblies retrieved from the NCBI database. Pairwise average nucleotide identity (ANI) and average amino acid identity (AAI) values were calculated. Species delineation was performed using the commonly accepted threshold of ANI ≥ 95%. Hierarchical clustering was conducted using the average linkage method based on distance matrices. Results. Based on taxonomic information provided by LPSN, an initial reference dataset comprising 36 genomes corresponding to validly named species of the genus Stenotrophomonas was established. Pairwise ANI values confirmed clear separation of these genomes at the level of species. Analysis of pairwise AAI values revealed several genomically distinct clusters consistent with a recently proposed taxonomic framework suggesting subdivision of the genus Stenotrophomonas into multiple genera. Analysis of 2,332 Stenotrophomonas genomes from the NCBI database revealed that 15.7% had ANI values below the 95% threshold relative to reference genomes of LPSN-listed taxa, precluding assignment to any recognized species. Clustering at ANI ≥95% yielded 102 genomic clusters, some lacking genomes from LPSN-listed species. To enable classification of such genomes, an expanded reference dataset was constructed by including representative genomes of the corresponding ANI clusters. This expanded reference dataset enabled assignment of all analyzed genomes to either LPSN-listed taxa or to distinct genomic clusters lacking validated nomenclatural status. Conclusions. Even with abundant genomic data and quantitative delineation criteria, taxonomic interpretation of genomic collections remains constrained by the lag between genome-based identification of putative new taxa and their formal description with type strain designation. Our proposed analytical approach systematically accounts for such genomes and offers a reproducible framework for analyzing the taxonomic structure of the nomen genus Stenotrophomonas amid ongoing revisions.

Objective. To determine the population structure of clinical P. aeruginosa isolates recovered from hospitals in different regions of Russia and to assess the role of high-risk clones in the dissemination of carbapenemases and carbapenem resistance. Materials and Methods. The study included consecutive, non-duplicate clinical isolates of P. aeruginosa (N = 1379) obtained from 55 hospitals across 30 Russian cities over a 24-month period. Species identification was performed using MALDI-TOF mass-spectrometry. Antimicrobial susceptibility testing was carried out by broth microdilution. The presence of acquired carbapenemase genes (VIM, IMP, NDM, GES-2/-5) was determined by real-time PCR. Subspecies typing and assignment to known sequence types (STs) and clonal complexes (CCs) were performed by identifying single nucleotide polymorphisms (SNPs) in seven chromosomal loci used in the multilocus sequence typing (MLST) scheme. Whole-genome sequencing (WGS) and resistance gene annotation were performed for selected isolates representing the main high-risk clones. Results. All P. aeruginosa isolates were assigned to 264 genetic lineages, among which four clonal complexes corresponding to internationally recognized high-risk clones were predominant: CC235 (13.05%), CC244 (7.11%), CC654 (5.51%), and CC357 (4.21%). Three of these lineages – CC235, CC357, and CC654 – showed significantly higher levels of resistance to most antibiotics, including carbapenems, and were more frequently associated with nosocomial than communityacquired infections. Carbapenemase production was detected in 14.5% of isolates (VIM, 11.02%; GES-5, 2.61%; IMP, 0.44%; NDM, 0.44%), with the majority of carbapenemase-producing strains belonging to CC235 (52.0%) and CC654 (35.0%). WGS data revealed that CC235 isolates collected from different regions of Russia exhibited the greatest diversity of resistance determinants, including carbapenemase genes VIM-2, GES-5, and IMP-1. In contrast, all CC654 isolates carried the VIM-2 metallo-β-lactamase gene and shared similar genotypic and phenotypic resistance profiles to various antibiotic classes. Compared with previous surveillance data from the Russia, a decreasing trend was observed in the proportion of CC235 isolates, including carbapenemase producers, while the prevalence of other high-risk clones within the P. aeruginosa population remained stable. Conclusions. Despite a decline in carbapenemase production frequency and overall carbapenem resistance, the data show that high-risk clones continue to serve as the main reservoirs and drivers of resistance dissemination, underscoring the importance of maintaining systematic molecular surveillance and control of their circulation.