[Phenotypic and molecular genetic properties of Escherichia coli clinical strains isolated from patients with urological diseases].

OBJECTIVE: Microbiological and molecular genetic characterization resistance profiles of Escherichia coli strains isolated in a pilot single-center clinical study from patients of the urological department in Yaroslavl in 2016-2017. MATERIALS AND METHODS: Clinical strains of E. coli (n=18) were isolated from the urine of women aged 23-84 years. The mobility of bacteria, colicinogenicity, and sensitivity to lactobacilli antagonism, biofilm formation, and susceptibility to antimicrobials were evaluated. The antibiotic resistance genes were identified. RESULTS: The E. coli strains had a wide heterogeneity in mobility, colicinogenicity, and biofilm formation. They were sensitive to Lactobacillus acidophilus antagonism, as well as to nitrofurantoin, meropenem, fosfomycin and the main functional classes of disinfectants and antiseptics, but are resistant to beta-lactams, fluoroquinolones and aminoglycosides. The mcr-1 gene providing resistance to colistin was identified in two strains. CONCLUSIONS: Analysis of genetic antibiotic resistance determinants revealed the genetic diversity of clinical E. coli strains. The obtained data on the strain sensitivity to antibacterials and disinfectants can be used by clinicians in choosing the optimal antibiotic therapy and treatment of abiotic surfaces in urological departments.

[Trials of the domestically produced nutrient medium «Agar Muller-Hinton II - Obolensk¼.]

The results of the comparative tests of the «Agar Muller-Hinton II - Obolensk¼ nutrient medium developed in SRCAMB, Obolensk, and the control nutrient medium imported «Mueller Hinton II Agar¼ are presented in the study. The susceptibility of bacterial clinical strains to antimicrobial agents (AMP) was determined by the disc diffusion method and the method of gradient diffusion (E-test). The carbapenemase activity of the strains carrying the carbapenemase genes was determined by CIM-test. Total 173 characterized bacterial strains of species Klebsiella pneumoniae, Pseudomonas aeruginosa, Acinetobacter baumannii, Proteus mirabilis, Serratia marcescens, Enterobacter aerogenes, Escherichia coli; Photorhabdus spp., Staphylococcus aureus, Enterococcus spp. were used in the study, including producers of OXA- and NDM-types carbapenemases for gram negative bacteria. A high degree of coincidence of the results obtained on both nutrient media was shown. The consistency index of the strain sensitivity categories to AMPs (S, I, and R) was 98.2% for the disc diffusion method, and 94.4-100% - for E-test and CIM-test methods. Thus, within the framework of the Import Substitution Program, the domestic nutrient medium «MHA II-Obolensk¼ has been successfully developed. The nutrient medium meets the requirements of GOST R ISO 20776-2-2010 «Clinical laboratory testing and in vitro diagnostic test systems - Susceptibility testing of infectious agents and evaluation of performance of antimicrobial susceptibility test devices¼.

[Clinical trials of salmonella enrichment medium.]

Salmonella infections continue to be a serious problem in modern medicine. Being intestinal infections-associated pathogens, representatives of the genus Salmonella manifest themselves as pathogenic bacteria, especially in developing nosocomial infections. Given the polymorphism of clinical symptoms of salmonellosis, laboratory studies using bacteriological and serological methods are an important link in the diagnosis. In addition, the general prevention of salmonellosis includes measures to identify bacteria carriers, to ensure control over the incidence in farm animals and birds, food safety, etc. The list of nutrient media to isolate and identify Salmonella is lengthy and steadily extending, and the choice of specific media is largely relies on the nature of the material under study as well as on the idea of the potential availability of Salmonella bacteria in it, with research, diagnosis or epidemic situation being taken into account. The SRCAMB (Rospotrebnadzor) has designed two nutrient media allowing the enrichment and isolation Salmonella from various clinical samples. These are "Nutrient Medium for Enrichment of Salmonella, Dry (Magnesium medium) and "Nutrient Agar with Brilliant Green, Phenolic Red, Lactose and Sucrose, Dry (BPLS-FMH agar)." Growth and inhibitory properties of the new culture media produced by the SRCAMB and commercial domestic and foreign counterparts have been compared by using clinical material. Domestic nutrient media such as Magnesium medium and BPLS-FMH agar were proved to correspond to their commercial analogues when being used for enrichment, isolating and counting Salmonella bacteria in clinical specimens to have bacteriological control real data.

[The characteristic of diarrheagenic Escherichia separated from children aged under 5 years old in Yaroslavl.]

The diarrheagenic bacteria coli take a significant place among agents of acute intestinal infections in children aged under 5 years. The main danger among these pathogens is represented by both enterotoxigenic E. coli causing enteritis and enterocolitis accompanied by acute dehydration diarrhea and Escherichia producing shiga-toxin being agents of hemorrhagic colitis and hemolytic uremic syndrome. The fast and proper identification of agents of these two groups of pathogens is an important task of bacteriologists to be resolved for successful treatment of patient because tactics of therapy of enterotoxigenic diarrhea and hemorrhagic colitis and hemolytic uremic syndrome significantly differ. The high capacity of Escherichia coli to form populations resistant to anti-microbial medications, including pan-resistant ones, also is a serious problem for science and public health. The object of study was a collection of isolates of E. coli (n = 112), separated from 112 children aged under 5 years with clinical manifestations of acute intestinal infection, food toxic infection hemocolitis and diarrhea of obscure etiology in Yaroslavl in 2015-2017. Initially, the strains of E. coli were tested using diagnostic agglutinating coli-serums and then using reagents' kit «AmpliSens®Escherichiosis-FL¼ for detection and differentiation DNAof diarrheagenic bacteria coli and also with specific oligonucleotide primers to genes of virulence and O-serum group belonging. The obtained data permitted to determine belonging of analyzed strains of E. coli to four sub-groups: ЕНЕС (n = 9), EPEC (n = 17), ETEC (n = 1) и EAgEС (n = 1). All of them were agents of genes of pathogenicity specific for every pathogroup. The most numerous group EPEC was represented by strains of five serogroups with dominating among them serogroup O26 (9 strains). Therefore, studying collection of strains of diarrheagenic Escherichia isolated during 2015-1017 in Yaroslavl from children aged under 5 years with acute intestinal infections permitted to demonstrate efficiency of application of molecular genetic methods of analysis for characterizing E. coli i.e. establishment their serogroups, detection of genes of virulence and attributing to pathogroups.

[Antibiotic Resistance of MRSA in the Russian Federation].

The results of the multicentre trial on estimation of MRSA antibiotic susceptibility to 17 antibiotics are presented. 474 nonrepeting isolates of MRSA (mecA+), collected in 2011-2014 in 10 cities of the Russian Federation were used in the trial. The antibiotic susceptibility was determined by the method of serial microdilutions in broth with estimation of the MICs in accordance with the international standards CLSI 2014 and EUCAST 2014. The highest levels of the MRSA resistance were stated against ciprofloxacin--92%(MIC50 32 mcg/ml), gentamicin--85% (MIC50 128 mcg/ml), erythromycin--54% (MIC50 32-mcg/ml) and clindainycin - 45% (MIC50 0.03 mcg/ml), as well as against rifampicin--38% (MIC50 0.06 mcg/ml). The frequency of MRSA isolated at the vancomycin dose of 2 mcg/ml equaled 26%. No correlation of the decrease in susceptibility to vancomycin and rifampicin was observed. In 5% of MRSA isolated from infected surgical wounds in patients with bone infection or sepsis, there was observed a decrease in the susceptibility to ceftarolin (MIC 2-4 mcg/ml). Co-trimoxasole, fusidic acid (MIC50 0.06 mcg/ml) and mupirocin (MIC50 0.5 mcg/ml) showed high antibacterial activity, 93-98% of the isolates being susceptible to the drugs. No resistance to linezolid and tigecycline was detected. By the associate resistance spectrum, most of the MRSA isolates were characterized by resistance to drugs of 3-7 groups (56%). The phenotypes with simultaneous resistance to drugs of 8-10 groups amounted to 6%. As a whole, 70 variants of associate resistance combinations were detected.

[Estimation of MRSA susceptibility to oxacillin, cefoxitine, vancomycin and daptomycin].

Prevalence and therapy of infections due to MRSA remain one of the most serious problems in the world. Therefore, correct laboratory identification of the MRSA phenotype based on the use of the marker antibiotic cefoxitine, as a more susceptibile one vs. oxacillin, is of great importance. There is lately being observed a tendency towards emergence of strains with lower susceptibility to the last reserve drugs protecting from MRSA, i. e. vancomycin and daptomycin. Susceptibility of MSRA to these drugs was not investigated in Russia and there are no data on the prevalence of the VISA and hVISA phenotypes. The results of our study on estimation of susceptibility of 316 MRSA isolates from several regions of Russia to oxacillin, cefoxitine, vancomycin and daptomycin are presented herein. It was shown that the ranges of the oxacillin MIC were extremely wide, i. e. 0.5 to 512 mcg/ml, while 2.2 +/- 1% of the isolates was susceptible by the phenotype to oxacillin, in spite of the mecA gene presence. As for cefoxitine, the MRSA isolates were rather resistant to it at the MIC > 16 mcg/ml. The tests with serial microdilutions revealed that 30.7 +/- 7% of the isolates had a critical level of susceptibility to vancomycin at the MIC 2 mcg/ml. The E-tests revealed 1.3 +/- 1% of the isolates which were susceptible at the MIC 2-4 mcg/ml. The MRSA isolates were highly susceptible to daptomycin, while high levels of the MIC (2 mcg/ml) were characteristic of 2.8 +/- 1% of the isolates. Cross reduction of the susceptibility to vancomycin and daptomycin was observed.