Vakcinaciya yavlyaetsya effektivnym sredstvom preduprezhdeniya zabolevaniya koklyushnoj infekciej. Cel'yu raboty bylo usovershenstvovanie vakcinnyh preparatov v RF, v tom chisle aktualizaciya proizvodstvennyh vakcinnyh shtammov. Izuchali shtammy B. pertussis, vydelennye v g. Moskve, Voronezhskoj, Novosibirskoj, Ul'yanovskoj i CHelyabinskoj oblastyah, i vosem' proizvodstvennyh shtammov dlya adsorbirovannoj koklyushno-difterijno-stolbnyachnoj (AKDS) vakciny. Dlya genotipirovaniya ispol'zovali mul'tilokusnoe antigennoe sikvens-tipirovanie (MAST) i polnogenomnoe mul'tilokusnoe sikvens-tipirovanie (wgMLST). Izucheny kul'tural'no-morfologicheskie, fermentantivnye, serologicheskie i genotipicheskie svojstva shtammov B. pertussis — kandidatov v proizvodstvennye vakcinnye shtammy i proveden sravnitel'nyj analiz genotipicheskih svojstv etih shtammov i proizvodstvennyh vakcinnyh shtammov B. pertussis dlya vakciny AKDS. Kandidatnye shtammy yavlyayutsya predstavitelyami chetyrekh genotipov — ptxA1/ptxV2/ptxS2/ptxR3/fim2-2/fim3-2/prn2, ptxA1/ptxV2/ptxS2/ptxR3/fim2-2/fim3-2/prn9, ptxA1/ptxV2/ptxS2/ptxR3/fim2-1/fim3-1/prn1 i ptxA1/ptxV2/ptxS2/ptxR3/fim2-2/fim3-1/prn2. Proizvodstvennye vakcinnye shtammy prinadlezhali k drugim shesti genotipam: ptxA2/ptxV1/ptxS1/ptxR1/ fim2-1/fim3-1/prn1, ptxA2/ptxV2/ptxS1/ptxR2/fim2-1/fim3-1/prn1, ptxA4/ptxV1/ptxS1/ptxR2/fim2-1/fim3-1/prn1, ptxA2/ptxV2/ptxS1/ptxR1/fim2-1/fim3-1/prn1, ptxA4/ptxV2/ptxS1/ptxR2/fim2-1/fim3-1/prn1 i ptxA1/ptxV2/ptxS1/ptxR1/fim2-1/fim3-1/prn1. S pomoshch'yu wgMLST ustanovlena prinadlezhnost' vsekh kandidatnyh shtammov B. pertussis k ST2.

Vaccination is an effective means of preventing pertussis infection. The purpose of this work was to improve vaccines available in the Russian Federation in general and actualize vaccine strains used for the production thereof in particular. We studied B. pertussis strains isolated in Moscow, Voronezh, Novosibirsk, Ulyanovsk and Chelyabinsk regions, and eight production strains part of the adsorbed diphtheria-pertussis-tetanus (DPT) vaccine. Multilocus antigenic sequence typing (MAST) and whole genome multilocus sequence typing (wgMLST) were used for genotyping. We studied cultural morphological, enzymatic, serological, and genotypic properties of the candidate B. pertussis strains, and compared their genotypic properties to those of B. pertussis vaccine strains from the current composition of the DPT vaccine. Candidate strains belong to four genotypes: ptxА1/ptxВ2/ptxС2/ptxР3/fim2-2/fim3-2/prn2, ptxА1/ptxВ2/ptxС2/ptxР3/fim2-2/fim3-2/prn9, ptxА1/ptxВ2/ptxС2/ptxР3/fim2-1/fim3-1/prn1 and ptxА1/ptxВ2/ptxС2/ptxР3/fim2-2/fim3-1/prn2. Current vaccine strains were from other six genotypes: ptxА2/ptxВ1/ptxС1/ptxР1/fim2-1/fim3-1/prn1, ptxА2/ptxВ2/ptxС1/ptxР2/fim2-1/fim3-1/prn1, ptxА4/ptxВ1/ptxС1/ptxР2/fim2-1/fim3-1/prn1, ptxА2/ptxВ2/ptxС1/ptxР1/fim2-1/fim3-1/prn1, ptxА4/ptxВ2/ptxС1/ptxР2/fim2-1/fim3-1/prn1 and ptxА1/ptxВ2/ptxС1/ptxР1/fim2-1/fim3-1/prn1. With the help of wgMLST, we established affiliation of all candidate strains of B. pertussis to ST2.

Objective: to assess anti-pertussis immunity in schoolchildren aged 7–17 who complained of a prolonged cough during the 11-year follow-up period. Materials and methods. The study included 1046 patients aged 7 to 17 years who applied to the Consultative and Diagnostic Center of the G.N. Gabrichevsky Research Institute of Epidemiology and Microbiology with complaints of prolonged cough in the period from 2010 to 2020. Blood serums were examined in ELISA with the determination of IgM, IgG, IgA antibodies using RIDASCREEN test system (Germany). Results. An active infection with the detection of IgM and/ or IgA, IgG antibodies above threshold levels was detected in 51,3% of children with prolonged cough, while annually in a fairly high percentage throughout the follow-up period. Active pertussis infection, established based on the detection of IgM, IgG, IgA antibodies above thresholds in blood serum samples, prevailed in children 12–15 years old, accounting for more than 60% in children with prolonged cough. Antipertussis immunity as a result of childhood vaccination or previous disease was detected in 16.1-20.2% of people in the period 2010–2014 and in 12,8-20,9% in 2015–2020. Conclusion. The results obtained by us on the study of anti-pertussis immunity in schoolchildren confirm the presence of active latent circulation of the pathogen whooping cough among children of this age cohort and, therefore, the presence of unaccounted for cases of the disease. This confirms the importance of timely diagnosis of pertussis, isolation of children for the period of active infection and justifies the need for the widespread introduction of a second revaccination against pertussis.

Bakterii roda Aerococcus chasto associirovany s infekciyami mochevyvodyashchih putej i krovotoka u cheloveka. SHtamm Aerococcus sp. 1KP-2016, vydelennyj iz lejkocitarnogo sloya krovi, obladal posledovatel'nost'yu 16S rRNK, sovpadayushchej na 98,7% i menee s ranee opisannymi predstavitelyami dannogo roda. Cel'yu raboty bylo provesti polnogenomnoe sekvenirovanie Aerococcus 1KP-2016 s posleduyushchej filogeneticheskoj rekonstrukciej. Pokazano, chto Aerococcus 1KP-2016 yavlyaetsya predstavitelem novogo vida roda Aerococcus, naibolee blizkogo k Aerococcus viridans i Aerococcus urinaeequi. Genomnaya posledovatel'nost', imeyushchaya dlinu 2,042 mln p.n. i GC-sostav 38,5%, deponirovana v DBJ/EMBL/GenBank pod identifikatorom NEEY00000000.

Aerococcus genus bacteria are often associated with human urinary tract and bloodstream infections. The Aerococcus sp. 1KP-2016 strain isolated from the buffy coat had the 16S rRNA sequence that was a 98.7% (and less) match with the previously described members of this genus. The purpose of this study was to perform whole genome sequencing of Aerococcus 1KP-2016 followed by phylogenetic reconstruction. We have shown that Aerococcus 1KP-2016 belongs to the new species of the Aerococcus genus that is closest to Aerococcus viridans and Aerococcus urinaeequi. The genomic sequence, which consists of 2.042 million bps with GC content at 38.5%, was deposited in the DBJ/EMBL/GenBank under identifier NEEY00000000.

Community-acquired bloodstream infections (CBSIs) occur in the out-of-hospital setting (44%) and increase the overall mortality from bloodstream infections (BSIs) by 7.2% per year. The development of CBSIs depends on both comorbid and polymorbid diseases and the patients' age. The causes of CBSIs are: respiratory, hepatobiliary gastrointestinal and urogenital tracts and dental interventions. The etiology of CBSIs is characterized by the isolation of coagulase-negative staphylococci (CNS) (32%), E. coli (27%). To investigate community-acquired bloodstream infection in therapeutic patients. The study included out-of-hospital patients (n=382). 4.5 ml of blood were taken intravenously into a closed vacuum system in order to obtain a buffy coat of blood, which was put on glasses for microscopy and Petri dishes with blood agar for cultivating under aerobic and anaerobic conditions. Microorganisms were identified by mass spectrometry. Microscopy of blood smears was used for rapid diagnosis of infection in the bloodstream. BSI was diagnosed in 183 (48.0%) out of 382 out-of-hospital patients. The etiology of CBSIs was studied on 297 isolated strains of microorganisms. CBSIs rather often complicated the underlying disease in women and young people. The spectrum of CBSI pathogens included aerobic and anaerobic bacteria and fungi. Gram-positive cocci with the leadership of S.epidermidis (25.7%) were more often isolated among bacteria. 70% of all isolated pathogens grew under anaerobic conditions. CBSIs were characterized by polymicrobiality (33.5%) of two to four different microorganisms in one blood culture; the species of associates of polymicrobial blood cultures are shown. Microscopic examination of blood smears revealed microorganisms in 97.1% of cases, including associations of bacteria with fungi (66.9%). CBSIs occurred after contour plastic, in diseases of the respiratory system, genitourinary system, oral cavity, skin and subcutaneous tissue. Microbiological examination of the buffy coat is an alternative microbiological method of CBSIs diagnosis, which includes microscopy and blood cultivating and has a high diagnostic efficiency (97.1% and 48% respectively). It can become an option for replacing imported blood culture automated systems.

The aim of the study was to characterize toxigenic strains of Corynebacterium diphtheriae by examining 12 toxigenic strains of C.diphtheriae isolated in Russia between January, 2017 to June, 2019. The morphological, toxigenic and biochemical properties of C.diphtheriae was studied. Genotyping of C.diphtheriae strains was performed by using MLST and dtxR gene sequencing with subsequent phylogenetic analysis. Results. Toxigenic strains of C.diphtheriae were isolated in the Novosibirsk, Samara and Chelyabinsk regions, the Khanty-Mansi autonomous okrug - Yugra as well as the republic Northern Ossetia - Alania. Among these strains, 5 of them were isolated from diphtheria patients (moderate disease found in one case, mild course – remaining patients) and 7 strains were isolated from bacterial carriers. In two cases C.diphtheriae from diphtheria patients were identified as ST25 sequence type, gravis variant; in one case – ST8 type, gravis variant; two cases – ST67 sequence type, mitis variant. In asymptomatic carriers of tox-positive C.diphtheriae strains they belonged to ST25 sequence type, gravis variant – in two cases, ST67 type, mitis variant – in four cases. A sequencing type was not identified in one case. All sequence types were widespread globally being presented by a large number of isolates in the PubMLST and characterized by a substantial amount of derivative sequence types. At the same time, they belonged to different clonal complexes and differed markedly from each other contributing to their reliable difference as assessed by MLST. Study of gene dtxR sequence diversity showed that all allelic variants were typical for the representatives of these a sequence types. New alleles of gene dtxR were not revealed in strains examined. It was shown that non-synonymous substitution C440T leading to A147V amino acid substitution was found solely in one allele distributed in ST8, ST185, ST195 and ST451 types suggesting at late mutation. In contrast, the polymorphism C640A resulting in the amino acid substitution L214I was found not only in the same allele, but also in the basal tree branches indicating that isoleucine was in the ancestral sequence of the protein.

The aim of the study was to characterize toxigenic strains of Corynebacterium diphtheriae by examining 12 toxigenic strains of C. diphtheriae isolated in Russia between January, 2017 to June, 2019. The morphological, toxigenic and biochemical properties of C. diphtheriae was studied. Genotyping of C. diphtheriae strains was performed using MLST and dtxR gene sequencing with subsequent phylogenetic analysis. Results. Toxigenic strains of C. diphtheriae were isolated in the Novosibirsk, Samara and Chelyabinsk Regions, the Khanty-Mansi Autonomous Okrug - Yugra as well as the Republic of Northern Ossetia - Alania. Among these strains, 5 were isolated from diphtheria patients (moderate disease found in one case, mild course - remaining patients) and 7 strains were isolated from bacterial carriers. In two cases C. diphtheriae from diphtheria patients were identified as ST25 sequence type, gravis variant; in one case - ST8 type, gravis variant; two cases - ST67 sequence type, mitis variant. In asymptomatic carriers of tox-positive C. diphtheriae strains they belonged to ST25 sequence type, gravis variant - in two cases, ST67 type, mitis variant - in four cases. A sequencing type was not identified in one case. All sequence types were widespread globally being presented by a large number of isolates in the PubMLST and characterized by a substantial amount of derivative sequence types. At the same time, they belonged to different clonal complexes and differed markedly from each other contributing to their reliable difference as assessed by MLST. Study of gene dtxR sequence diversity showed that all allelic variants were typical for the representatives of these sequence types. New alleles of gene dtxR were not revealed in strains examined. It was shown that non-synonymous substitution C440T leading to A147V amino acid substitution was found solely in one allele distributed in ST8, ST185, ST195 and ST451 types suggesting at late mutation. In contrast, the polymorphism C640A resulting in the amino acid substitution L214I was found not only in the same allele, but also in the basal tree branches indicating that isoleucine was in the ancestral sequence of the protein.

The main objective of laboratory diagnosis of a diphtheria is identification of the causative agent by means of the minimum quantity of diagnostic tests for obtaining the authentic answer in the most short time. One of the major stages is capture and delivery of pathological material on which the efficiency of carrying out and timeliness of issue of the final answer depends. Considering emergence in the market of commercial liquid transport mediums, assessment of their efficiency for diagnosis of diphtheria is advisable. In the real work the pilot studies allowing to predict efficiency of use of the commercial transport liquid medium ∑-Transwab® with the liquid medium of Ames in two systems - with the standard applicator (system 1) and with the thin extended tampon for sampling from narrow cavities - urethral and nazofarengialny are conducted (system 2). In a research used a control toxigenic strain of Corynebacterium diphtheriae of a biovar of gravis No. 665. In an experiment "imitated" operating conditions of the medical organizations for storage of tampons with pathological material on diphtheria before their transportation in laboratory - on a table at the room temperature (6 and 20 hours), in the refrigerator (6 and 20 hours), in the thermostat (6 and 20 hours). After an incubation all tampons sowed the environment for primary crops of pathological material on a blood tellurite agar.. Accounting of results was carried out in 24 and 48 hours of growth. It is established that the commercial transport liquid medium of Ames can be used for capture of pathological material on diphtheria in the second half of the working day at storage in the conditions of the refrigerator. At the same time, it is necessary to consider a tampon form as the best results on a identification of the causative agent of diphtheria have been received when using a universal tampon.

The pathogenic mechanism used by Corynebacterium diphtheriae is attributed to the ability of the diphtheria toxin to disrupt protein synthesis in human cells. Diphtheria toxin production is regulated by the DtxR protein. The latter is involved in the iron-mediated repression of the toxin gene and coordinates activities of other genes essential for the survival of C. diphtheriae. The DtxR-encoding gene occurs in both toxigenic and non-toxigenic strains; therefore it can be used to analyze the population structure of the species. In our work we have studied 45 strains of C. diphtheriae isolated in the Russian Federation in 2010-2015. These strains were analyzed to reveal that gene dtxR is a highly conservative region of.. diphtheriae genome that can be found in all members of the studied species. The majority of the discovered polymorphisms were synonymous (16 of 18 single nucleotide polymorphisms identified). In spite of the low phylogenetic signal, the allelic variant of dtxR was associated with the strain's phenotype (biovar, toxigenicity). The obtained data indicate the presence of aggressive negative selection aimed to maintain the existing protein sequence in the population. Based on the results, we recommend dtxR polymerase chain reaction as an additional technique for pathogen identification, which is especially relevant considering the increasing prevalence of the disease associated with non-toxigenic C. diphtheriae strains.

Считается, что патогенез Corynebacterium diphtheriae основан на воздействии дифтерийного токсина на синтез белка в клетках человека. Регуляция синтеза токсина находится под контролем белка DtxR. Данный белок осуществляет железоопосредованную репрессию гена дифтерийного токсина, а также координирует работу множества других генов, необходимых для нормальной жизнедеятельности С. diphtheriae. Ген, кодирующий DtxR, можно использовать для анализа популяционной структуры вида, так как он присутствует в геноме как токсигенных, так и нетоксигенных штаммов. В работе было изучено 45 штаммов С. diphtheriae, выделенных на территории Российской Федерации в 2010-2015 гг. Анализ этих штаммов показал, что ген dtxR обнаруживается у всех представителей вида и является высококонсервативным участком генома С. diphtheriae. Большинство выявленных полиморфизмов были синонимичны (16 из 18 однонуклеотидных замен). Несмотря на низкий уровень филогенетического сигнала, аллельный вариант dtxR был ассоциирован с биологическими признаками штамма (биовар, токсигенность). Полученные данные свидетельствуют о высокой активности отрицательного отбора, направленного на поддержание в популяции существующей последовательности белка, и позволяют рекомендовать наработку фрагментов гена dtxR методом полимеразной цепной реакции в качестве дополнительного метода идентификации возбудителя, что особенно актуально в условиях растущего числа заболеваний, ассоциированных с нетоксигенными штаммами С. diphtheriae.

AIM:Study the prevalence and mechanisms of resistance in circulating C. diphtheriae strains.MATERIALS AND METHODS:664 C. diphtheriae strains isolated in 1987 - 2013 in various regions of Russia and sent to the reference center of Gabrichevsky Moscow Research Institute of Epidemiology and Microbiology were the object of the study. Antibiotic sensitivity of the strains was studied by disk-diffusion and E-test methods using 10 antimicrobial preparations. Nucleotide sequence analysis was carried out by using BLAST program and EMBL/GenBank database.RESULTS:Most of the studied strains turned out to be sensitive to all the antibacterial preparations used. 1.2% of C. diphtheriae strains turned out to be resistant to penicillin and 6.0% had intermediate level of resistance. 0.4 - 0.6% of the strains had intermediate level of resistance to macrolides, and 4.0 - 4.4% were resistant. 2.0% of the strains had multiple resistance. Erm(X)-specific PCR carried out in this study showed that all the C. diphtheriae strains resistant to macrolide antibiotics carry erm(X) gene.CONCLUSION:The results of the study indicate a fairly high level of prevalence for C. diphtheriae strains resistant to antibiotics.

AIM:Characterization of contemporary C. diphtheriae strains isolated in Russia by using multilocus DNA sequencing (MLST).MATERIALS AND METHODS:28 toxigenic C. diphtheriae strains isolated in Russia in 2002-2012 and sent to diphtheria and pertussis reference center of Gabrichevsky Research Institute of Epidemiology and Microbiology were studied. C. diphtheriae strain genotyping was performed by using MLST based on atpA, dnaE, dnaK, fusA, leuA, odhA and rpoB gene fragments. Identification of alleles and ST was carried out according to EMBL/GenBank and PubMLST, eBurst approach was used for cluster analysis.RESULTS:By using MLST contemporary toxigenic C. diphtheriae strains isolated in Russia in 2002 - 2012 were characterized. 8 genotypes (ST41, ST5, ST8, ST28, ST25, ST44, ST-new1 and ST-new2) were identified, 3 among them were dominating--ST8, ST28 and ST-new1. Most of the toxigenic strains belong to biovar gravis and ST8. Among biovar mitis strains a higher heterogeneity by ST membership was noted, but with prevalence of ST28 strains.CONCLUSION:Use of MLST allowed to characterize contemporary circulating population of toxigenic C. diphtheriae strains isolated in Russia and showed perspective of application of this method for characterization of diphtheria causative agent population and detection of epidemically significant strains, as well as juxtaposing of them with genetic structure of foreign strains.