General dynamics of the URT microbiome and microbial signs of recovery in COVID-19 patients.

COVID-19 is caused by an airborne virus, SARS-CoV-2. The upper respiratory tract (URT) is, therefore, the first system to endure the attack. Inhabited by an assemblage of microbial communities, a healthy URT wards off the invasion. However, once invaded, it becomes destabilised, which could be crucial to the establishment and progression of the infection. We examined 696 URT samples collected from 285 COVID-19 patients at three time-points throughout their hospital stay and 100 URT samples from 100 healthy controls. We used 16S ribosomal RNA sequencing to evaluate the abundance of various bacterial taxa, α-diversity, and ß-diversity of the URT microbiome. Ordinary least squares regression was used to establish associations between the variables, with age, sex, and antibiotics as covariates. The URT microbiome in the COVID-19 patients was distinctively different from that of healthy controls. In COVID-19 patients, the abundance of 16 genera was significantly reduced. A total of 47 genera were specific to patients, whereas only 2 were unique to controls. The URT samples collected at admission differed more from the control than from the samples collected at later stages of treatment. The following four genera originally depleted in the patients grew significantly by the end of treatment: Fusobacterium, Haemophilus, Neisseria, and Stenotrophomonas. Our findings strongly suggest that SARS-CoV-2 caused significant changes in the URT microbiome, including the emergence of numerous atypical taxa. These findings may indicate increased instability of the URT microbiome in COVID-19 patients. In the course of the treatment, the microbial composition of the URT of COVID-19 patients tended toward that of controls. These microbial changes may be interpreted as markers of recovery.

The use of 4-Hexylresorcinol as antibiotic adjuvant.

Ever decreasing efficiency of antibiotic treatment due to growing antibiotic resistance of pathogenic bacteria is a critical issue in clinical practice. The two generally accepted major approaches to this problem are the search for new antibiotics and the development of antibiotic adjuvants to enhance the antimicrobial activity of known compounds. It was therefore the aim of the present study to test whether alkylresorcinols, a class of phenolic lipids, can be used as adjuvants to potentiate the effect of various classes of antibiotics. Alkylresorcinols were combined with 12 clinically used antibiotics. Growth-inhibiting activity against a broad range of pro- and eukaryotic microorganisms was determined. Test organisms did comprise 10 bacterial and 2 fungal collection strains, including E. coli and S. aureus, and clinical isolates of K. pneumoniae. The highest adjuvant activity was observed in the case of 4-hexylresorcinol (4-HR), a natural compound found in plants with antimicrobial activity. 50% of the minimal inhibitory concentration (MIC) of 4-HR caused an up to 50-fold decrease in the MIC of antibiotics of various classes. Application of 4-HR as an adjuvant revealed its efficiency against germination of bacterial dormant forms (spores) and prevented formation of antibiotic-tolerant persister cells. Using an in vivo mouse model of K. pneumoniae-induced sepsis, we could demonstrate that the combination of 4-HR and polymyxin was highly effective. 75% of animals were free of infection after treatment as compared to none of the animals receiving the antibiotic alone. We conclude that alkylresorcinols such as 4-HR can be used as an adjuvant to increase the efficiency of several known antibiotics. We suggest that by this approach the risk for development of genetically determined antibiotic resistance can be minimized due to the multimodal mode of action of 4-HR.

Molecular Mechanisms of Ceftaroline Susceptibility Reduction in Methicillin-Resistant Staphylococcus aureus.

Ceftaroline is a unique cephalosporin with activity against methicillin resistant Staphylococcus aureus (MRSA). It was approved for clinical use in the USA, Europe and Russian Federation since 2010 for the treatment of the skin and soft tissue infection and community-acquired pneumoniae. In the present study there was used molecular typing of 24 isolates of MRSA with reduced susceptibility to ceftaroline. For 8 isolates belonging to different genetic lines (ST8, ST239 and ST228) and requiring MICs there were determined antibiotic concentrations preventing formation of resistant mutants (mutant prevention concentration) and the ranges of the mutant selection window (MSW). The last majority of the isolates with reduced susceptibility to ceftaroline (MIC of 2 mcg/ml) belonged to the clonal line ST228. The whole genome sequencing of two isolates of ST228 showed that they belonged to the epidemic South Germany genetic line and were characterized by the presence of mutations in PBP2a (N146K) and PBP2 (C197Y) responsible for reduced susceptibility. The highest rates of MPC (32 mcg/ml) and MSW (2-16 mcg/ml) were observed in the clinical isolates belonging to the genetic line ST8. The isolates of ST239 and ST228 had the selection window within 2-4 mcg/ml. No dependence of the MIC and MPC/MSW levels was detected.

SNP Polymorphism in Genomes of CC320 Isolates of Streptococcuspneumoniae Resistant to Beta-Lactams.

Bioinformatic analysis of the data on the genome sequencing of the isolates of the Streptococcuspneumoniae clonal complex SS320 from the Russian Federation, as well as the data on SS320 isolates from public sources in the penicillin resistant isolates resulted in detection of 139 missense mutations in 45 genes. In addition to the mutations in the genes of the main penicillin-binding proteins (PSB - PBP1A, PBP2B and PBP2X) there was detected high frequency of mutations in the genes of the (division and cell wall) dcw-cluster, as well as in RegR protein belonging to the transcription regulators of the LacI/GaIR family. Development of resistance to beta-lactams in S.pneumoniae is defined not only by modification of the PSB, but also by adaptive changes in the metabolic pathways involved in the bacterial cell growth and division.

[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.

[Antibiotic Resistance and Serotype Pattern of Streptococcus pneumoniae Isolated from Children in St. Petersburg in 2010-2013].

The surveillance of the serotype pattern and antibiotic resistance of S. pneumoniae in various geographical regions is required for the validity of rational etiotrophic therapy of pneumococcal infections and the choice of the optimal vaccines for their prophylaxis. 250 S. pneumoniae isolates from children with acute otitis or pneumonia and healthy carriers in St. Petersburg in 2010-2013 were investigated. The analysis of the serotype pattern of the pneumococci showed that 13-valent conjugate vaccine was the most active (86.1% of pneumococci causing pneumonia and 86.4% of pneumococci causing acute otitis). The isolates were higly resistant to beta-lactams and macrolides. By the EUCAST criteria, the decrease in the susceptibility to penicillin, cefotaxime, erythromycin and ceftarolin was observed in 32.4%, 14%, 33.2 and 6% of the isolates respectively. 22.4% of the isolates showed associate resistance to penicillin and erythromycin.. No resistance to moxifloxacin was detected. The frequency of resistance to tetracycline, co-trimoxasole and chloramphenicol in various patients ranged within 30-50%. The prevalence of the antibiotic resistance was mainly characteristic of the isolates serotypes 19A, 19F, 14 and serogroup 6.

[Antibiotic Resistance of Coagulase-Negative Staphylococci Isolated at Hospitals of St. Petersburg and Moscow].

Antibiotic susceptibility of 119 coagulase-negative staphylococci isolated at hospitals of St. Petersburg and Moscow was investigated and estimated at the local laboratories as oxacillin resistant. The following species were identified: Staphylococcus epidermidis, S. haemolyticus, S. hominis, S.capitis, S. simulans, S. pettenkoferi, S. lentus, S. carnosus and S. warneri. The oxacillin resistance was confirmed in 79.8% of the isolates. The frequency of the associated resistance to non-beta-lactams was much higher in the oxacillin resistant isolates vs. the oxacillin susceptible ones. When the CLSI and EUCAST susceptibility criteria were used, 1-3% difference in the resistance levels was recorded. Among the oxacillin resistant isolates the frequency of resistance to gentamicin, ciprofloxacin, erythromycin, moxifloxacin, tetracycline and clindamycin equaled 90, 88, 88, 63, 43 and 26% respectively. Two linezolid resistant isolates of S. epidermidis with lower susceptibility to tedizolid were isolated. Eight isolates of S. epidermidis showed lower resistance to mupirocin. The MIC of ceftarolin for oxacillin resistant coagulase-negative staphylococci varied from 0.5 to 2.0 mcg/ml, while for the oxacillin susceptible ones it was lower than 0.25 mcg/ml. No resistance to tigecyclin and vancomycin was observed.

[Structural Polymorphism of Genome Islands Encoding Resistance to Beta-Lactams in Coagulase-Negative Staphylococci Isolated at Hospitals of the Russian Federation].

Coagulase-negative staphylococci (CoNS) are considered as a reservoir of mobile genetic elements and first of all of the staphylococcal cassette chromosome mec (SCCmec), defining staphylococci resistance to beta-lactams. Types II, IV, IVa, V, VII and VIII SCCmec were detected among 95 staphylococcal strains isolated in different regions of the Russian Federation. Subtypes C1a, C1b, C1c and C1 SCCmec were also identified (class B mec complex and two complexes of ccr1 and ccr2 genes recombinases). Some other cassette types carrying A, C1 and C2 classes of the mec complexes in combination with various recombinase genes were detected. The S.epidermidis isolates mainly formed cassettes carrying mec complex B, while the S. haemolyticus isolates had cassettes carrying classes C1 and C2 mec complex. Out of 9 isolates of S. hominis 5 isolates carried a new type cassette: class A mec complex in combination with the complex of the recombinase ccr1 genes. SCCmec was not identified in S. capitis and S. pasteuri. Their representatives carried either mec complex (1 isolate of S. pasteuri) or the recombinase complexes (2 isolates of S. capitis). The detected SCCmec variants in CoNS could be a source of emergence of new genetic lines of MRSA.

[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.

[Staphylococcal cassette chromosome mec, evolution and genetic lines of methicillin resistant Staphylococcus aureus].

Methicillin resistant Staphylococcus aureus (MRSA) is the main pathogen of hospital- and community-associated infections. Methicillin resistance is due to mecA gene located in a mobile complex element, staphylococcal cassette chromosome mec (SCCmec). The structure of the staphylococcal cassettes is diverse. At present eleven types of the cassettes are described. Types I-IV SCCmec are always associated with epidemiologically significant genetic lines of Staphylococcus. Thus, the pandemic hospital-associated MRSA (HA MRSA) belonging to CC5 and CC8 are of the types I-III SCCmec. The prevalence of virulent community-associated MRSA (CA MRSA) in many regions of the world is first of all connected with the characteristics of the type IV SCCmec structure and the presence of a recently described arginine catabolic mobile element (ACME) increasing the colonization activity of Staphylococcus. The review presents the up-to-date data on the origin, genetic structure and classification of SCCmec. Global genetic lines of MRSA are described and the problem of CA MRSA is discussed.

[Diaphragmatic lesions in closed injury]. / Povredeniia diafragmy pri zakrytoi travme

Bases on the analysis of 10 clinical observations, it was found that closed injuries of the diaphragm occur mainly in the use of great mechanic force (compression, a fall from great height, an impact of quick-acting object). In the acute period these are frequently not accompanied by diaphragmatic hernia and remain unrecognized. It is the author's opinion that in early diagnosis of acute diaphramatic hernia clinical symptoms play no leading role, but chest roentgenography is of primary importance. The terms of surgical treatment and the choice of the operative access are determined by general patient's state and the character of associated lesions.