Российский научно-исследовательский противочумный институт «Микроб», Саратов Цель. Исследование биологических свойств природных штаммов геновариантов V. cholerae биовара El Tor, влияющих на их адаптационные способности в условиях недостатка питательных веществ, при сравнительном анализе с типичными штаммами. Материалы и методы. Конкурентную пробу проводили путем посева смеси клеток двух сравниваемых штаммов в автоклавированную речную воду. Скорость роста штаммов определяли по значению оптической плотности. Экспрессию генов изучали методом ОТ-ПЦР с рассчитанными праймерами и зондами. Результаты. Установлено, что при совместном культивировании типичных штаммов и геновариантов V. cholerae биовара El Tor в условиях недостатка питательных веществ (автоклавированная речная вода) при комнатной температуре уровень выживаемости штаммов геновариантов выше, чем типичных штаммов, что указывает на их выраженные адаптационные преимущества в данных условиях. Показано, что селективные преимущества штаммов геновариантов обеспечиваются более высокой скоростью роста клеток и повышенной экспрессией гена rpoS. Заключение. Получены новые данные о способности бактерий штаммов геновариантов V. cholerae биовара El Tor в отличие от типичных изолятов к быстрому росту и повышенной экспрессии глобального регулятора стрессового ответа гена rpoS, что, возможно, способствует их лучшей адаптации не только в условиях недостатка питательных веществ, но при действии других стрессовых факторов.
Objective of the study was to identify the differences in the production of proteins in typical strain and genetically altered variant of V. cholerae O1, biovar El Tor.Materials and methods. Natural strains M1062 (Astrakhan, 1970) and M1509 (Moscow, 2010) were used as model strains in this work. Strains were cultivated on Luria Bertani agar at 37 °C. Electrophoresis was performed in accordance with W.K. Laemmli technique (1970), mass-spectrometric profiling – the method described by A. Shevchenko et al.Results and discussion. Mass-spectrometric scanning of cell lysates of the examined strains showed significant similarity of their proteomes (615 common proteins). The identified differences pertained to high expression of proteins in the strain M1062, participating in biosynthesis of DNA/RNA, included into “purines, pyrimidines, nucleosides” group, as well as regulatory proteins. In M1509 strain, biosynthesis of the proteins responsible for pathogenesis, adaptation under the influence of unfavorable environmental factors, included into “co-factors, vitamins, pigments” group, involved in lipid, carbohydrate, and protein metabolism, cellular processes, as well as proteins-transporters was increased. It has been suggested that the wide dissemination of El Tor genovariants is probably due to enhanced pathogenic and adaptive properties and also to the considerable transformation of cell metabolism.
Objective of the study was to conduct experiments on survival capacity of toxigenic genovariant strains as compared to typical strains of V. cholerae, biovar El Tor in autoclaved river water and in the organism of laboratory animals. Consequently, it was determined that both, genovariant and typical El Tor vibrios, sustained for a significant period of time (more than 5 months) in the river water, though a number of the bacterial cells in the population of the typical ones gradually decreased, while in the genovariant strains-the growth was observed in between the 7 th and 21 st day. CFU of the genovariants was 1.5-2.5 and 1.8-3.0 times higher than the amount of typical strain bacteria on the day 7 and day 21, respectively. Selective advantage of genovariants was also confirmed by competitive test in vitro. Furthermore, genovariants dominated over typical strains of V. cholerae, biovar El Tor in the mixed population in vivo (bioassay animals): CFU of the genovariants 1.25-84.0 times exceeded that of the typical strains when seeding out the contents of intestine and gastrointestinal wall homogenate. Put forward was an assumption that one of the factors for genovariant selective advantage might be enhanced adaptation ability affected by changes of cell metabolic activity. Identified capacity of genovariant toxigenic strains-not only to sustain in open water bodies of our country, but also to propagate in there, creates unfavorable epidemiological situation on cholera.
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