[MALDI-TOF mass-spectrometric analysis of cell proteins during identification of leptospira genus members].

AIM: Development of methodological approaches for identification of leptospira by using MALDI-TOF direct protein profiling technology. MATERIALS AND METHODS: Analysis of cell proteins of 34 leptospira strains was carried out in Microflex LT by using "MALDI Biotyper 3.0 for identification and classification of microorganisms" program. RESULTS: 19 reference spectra of reference leptospira strains from 7 species were generated and imported into MALDI Biotyper 3.0 database. Identification of 6 strains with undetermined taxonomic position was carried out. CONCLUSION: The approved method allows determination of leptospira species with accuracy that depends on their adaptation to nutrient media, preparation approach and sample storage conditions for mass-spectrometry.

[Epidemiological situation and prophylaxis of zoonotic and natural-focal infectious diseases in Siberia and the Far East].

Analysis of zoonotic and natural-focal infectious disease morbidity in 2009 - 2011 in Siberia and the Far East is presented, and a complex of measures aimed at their prophylaxis is proposed. Analysis is carried out based on the data received by Reference Center of Monitoring of Natural-Focal Infection Causative Agents and Regional Center of Monitoring of I-II Pathogenicity Group Causative Agents at the Irkutsk Research Institute of Plague Control from departments and Centers of Hygiene and Epidemiology of Siberian, Far Eastern, 3 subjects of Urals Federal District and 5 Stations of Plague Control of Federal Service for Control in the Sphere of Protection of Consumers' Rights and Well-Being of Humans. In the morbidity structure in this region "tick-borne" infections were established to predominate--69.4%, among bacterial--yersiniosis dominates. Deterioration of epizootic situation on rabies is observed in the Republics of Tuva and Buryatia.

[Problems of epidemiology and diagnostics of leptospiroses in Siberia and Far East].

Multi-year literature data as well as materials of the Reference centre of monitoring of natural foci infections of Irkutsk Research Institute of Plague Control of Siberia and the Far East regarding epidemiology, epizootology and laboratory diagnostics of leptospiroses in Siberia and the Far East are analyzed and summarized in the review. Situation on leptospiroses in the region has changed significantly. In 50 - 70s of the 20th century diseases were registered ubiquitously in the form of outbreaks, group and single cases. Currently a low level of sporadic morbidity is noted in separate subjects of the Russian Federation. The contemporary state of the problem remains insufficiently clear, this demands the expansion of studies, creation of modern databases, as well as introduction into the practice of highly sensitive methods of express diagnostics in a complex with bacteriologic and serologic methods.

[Modeling of interaction between Yersinia pestis and Tetrahymena pyriformis in experimental ecosystems].

Modeling of interaction Yersinia pestis-Tetrahymena pyriformis in artificial soil ecosystem (ASE) containing soil of burrows of main carrier from Gorno-Altayski natural plague reservoir, as well as in physiological solution (PS) and in Hottinger broth (HB). Optimal proportion of bacterial and protozoa cells was possible to obtain and depended from virulence of Y. pestis and environmental conditions. In ASE at 18-22 degrees C association was the most stable under the microbial burden of 100 microbial cells (m.c.) per infusorian. Resistance of plague agent to phagocytosis by T. pyriformis was determined by strain's virulence. Avirulent strain Y. pestis [cyrillic letter: see text]-2377 was rapidly eliminated by protozoan in HB, PS and in ASE under the burden of 10 m.c per infusorian. Y. pestis [cyrillic letter: see text]-3443 with selective virulence compared with [cyrillic letter: see text]-2377 preserved in association longer in any tested medium. Highly virulent Y. pestis [cyrillic letter: see text]-3448 was the most resistant to phagocytosis by T. pyriformis.

[Clonal structure of Yersinia pestis populations in experimental soil ecosystems].

Two basic tendencies--formation of latent (uncultivable) form (LF) and hemin storage variability--has been revealed during study of clonal structure dynamics of Y. pestis populations in artificial soil ecosystems in long-term incubation conditions. Y. pestis populations disappeared within 3 - 6 months at 18 - 22 degrees C, whereas at 4 - 8 degrees C a subsequent replacement of vegetative cells on LF, which are capable to prolonged survival (up to 22 months) in soil with ability to reversion in the presence of abundance of nutrients, has been observed. Bacteria of virulent strain retained all determinants of pathogenicity when reverted to LF, whereas bacteria of avirulent strain (defective on plasmid of Ca-dependence), on the contrary, undergo further degradation that resulted in loss of a pgm locus and gradual disappearance of population. LF revertants of highly virulent strain restored properties of initial population and were highly virulent.

[The population variability of Yersinia pestis in soil samples from the natural focus of plague].

Three Y. pestis strains were found to exist in the experimental soil ecosystem at a temperature of 4 degrees - 8 degrees C for a longer period (10 months, the term of observation) than at room temperature (3.5 months). Y. pestis population structure was characterized by relative stability in strains of the subspecies altaica and heterogeneity in the strain of the main subspecies, manifested by the loss of the pgm locus by vegetative cells and the preservation of pgm+ variants in the latent (uncultivable) form (LF). In the populations of all strains uniformity in calcium dependence, the tendency towards a decrease in the synthesis of factor 1, nutritional requirements in amino acids was observed. An important factor of the preservation of Y. pestis in the soil was LF formation. At room temperature this process quickly resulted in the death of the population. At 4 degrees - 8 degrees C A. pestis altaica avirulent strain could be inoculated onto solid nutrient media for a two-fold longer period (for 4 month) than the strain with selective virulence and for 5.5 months longer than Y. pestis pestis highly virulent strain.

[Ecological regularities of the existence of pathogenic Yersinia in soil ecosystems].

In this review the data on the ecology of pathogenic Yersinia in soil ecosystems, based on prolonged observations, were analyzed and summarized. In contrast to saprophytic species, ubiquitously spread in nature, pathogenic representatives of the genus Yersinia occurred only in the soil of natural foci and of these, Y. pestis were found only in the soil of burrows of the main carriers. The complex of abiotic and biotic factors (temperature, humidity, chemical composition, interactions in biocenosis) which determined the possibility of the existence of Yersinia in the soil environment and the preservation of their pathogenic properties was considered. Special attention was paid to their geno-phenotypic variability as the main factor of the adaptation of the causative agents of plague, pseudotuberculosis and intestinal yersiniosis in the environment.

[Evaluation of the polymerase chain reaction effectiveness in the investigation of out-breaks of pseudotuberculosis]. / Otsenka éffektivnosti polimeraznoi tsepnoi reaktsii pri rassledovanii vspyshek psevdotuberkuleza.

Data on the investigation of pseudotuberculosis epidemic outbreaks with the use of the polymerase chain reaction (PCR), are presented. Specific fragments of Yersinia pseudotuberculosis DNA were detected in 81.25% of patients, in 46.83% of cases confirmed by the isolation of Y. pseudotuberculosis cultures. The study of washings from vegetables and equipment in vegetable stores and kitchens yielded positive results in PCR in 8.52% and the survey of rodents--in 3.85% of cases. In the course of the bacteriological study of these specimens 6 Y. pseudotuberculosis cultures were isolated: 3--from vegetable products, 1--from a Norway rat and 2--from house mice. The coincidence of the data obtained by bacteriological study and PCR showed that the latter method gave objective results, while being capable of ensuring rather rapid analysis. PCR should be regarded as a signal test for the bacteriological search of the definite infective agent in the material under study.

[Mode of sample preparation for pseudotuberculosis laboratory diagnosis by polymer chain reaction method]. / Sposob podgotovki prob dlia laboratornoi diagnostiki psevdotuberkuleza metodom polimeraznoi tsepnoi reaktsii.

A mode of feces sample preparation was developed for polymerase chain reaction (PCR) assay. It was based on alkaline treatment of the material. This treatment killed the most part of indigenous microflora, whereas Yersinia survived, because it was relatively resistant to alkaline. The mode was tested using human feces artificially contaminated with Yersinia pseudotuberculosis. Positive responses in samples containing 10(3)-10(8) microbial cells per ml were obtained by PCR assay with Yersi and Yers2, Invl and Inv2, YP3 and YP4 primers. Diagnostic efficiency of PCR for patients, small mammals, and washings from environmental objects was 4.75, 1.66, and 2.12 times higher than diagnostic efficiency of bacteriological analysis of these samples, respectively. Positive results in PCR were obtained at the day of the material collection and treatment, whereas Y. pseudotuberculosis was isolated only after 8-20 days. Positive samples in PCR and in bacteriological analysis were found to coincide. A brief scheme of the Y. pseudotuberculosis laboratory diagnosis is suggested. According to this scheme, target-oriented bacteriological assay is performed only in those samples, in which preliminary PCR assay after 1-3 days of incubation gave positive results of Y. pseudotuberculosis DNA detection.