Journal of microbiology, epidemiology and immunobiologyJournal of microbiology, epidemiology and immunobiology0372-93112686-7613Central Research Institute for Epidemiology99810.36233/0372-9311-93UnknownEvaluation of azoximer bromide (polyoxidonium) influence on the adhesive properties of the Yersinia pestis EV NIIEG vaccine strain by atomic force microscopyShchukovskayaT. N.<p><strong>Tatyana N. Shchukovskaya</strong> — D. Sci. (Med.), Prof., main researcher, Department of immunology </p><p><em>Saratov</em></p>tatyanaschuk@mail.ruhttps://orcid.org/0000-0001-8995-0894GoncharovaA. Y.<p><strong>Anastasiya Y. Goncharova</strong> — Cand. Sci. (Med.), researcher, Department of immunology </p><p><em>Saratov</em></p>fake@neicon.ruhttps://orcid.org/0000-0002-9994-7936BugorkovaS. A.<p><strong>Svetlana A. Bugorkova</strong> — D. Sci. (Med.), deputy head, Department of immunology </p><p><em>Saratov</em></p>fake@neicon.ruhttps://orcid.org/0000-0001-7548-4845ErokhinP. S.<p><strong>Pavel S. Erokhin</strong> — Cand. Sci. (Phys.-Math.), researcher, Laboratory of diagnostic technologies </p><p><em>Saratov</em></p>fake@neicon.ruhttps://orcid.org/0000-0001-9525-8327KudryavtsevaO. M.<p><strong>Olga M. Kudryavtseva</strong> — Cand. Sci. (Biol.), senior researcher, Department of immunology </p><p><em>Saratov</em></p>fake@neicon.ruhttps://orcid.org/0000-0002-9894-3394Russian Research Anti-Plague Institute «Microbe»030720219832983071003202110032021Copyright © 2021, Shchukovskaya T.N., Goncharova A.Y., Bugorkova S.A., Erokhin P.S., Kudryavtseva O.M.2021<p><strong>Aim.</strong> To characterize the influence of azoximer bromide (polyoxidonium, PO) in cultivation conditions on the morpho- and nanomechanical cell surface properties of Y. pestis EV NIIEG vaccine strain and its derivatives Y. pestis КМ218 (pYT, pYV, pYP), Y. pestis КМ216 (pYT, pYV, pYP+), Y. pseudotuberculosis, Y. enterocolitica by atomic force microsopy (AFM), as well as on the adhesion of cells Y. pestis EV NIIEG to human collagen type IV.</p>
<p><strong>Materials and methods. </strong>The measurements were carried out using the Solver P47-PRO probe microscope (NT-MDT, Russia), standard methods of semi-contact AFM and AFM imaging analysis program. The adhesion of Y. pestis EV NIIEG to type IV collagen was determined by the number of cells binding to glass slides covered with human collagen type IV.</p>
<p><strong>Results.</strong> The introduction of PO in the cultivation environment caused changes in the morphometric parameters of the cells of Y. pestis EV NIIEG vaccine strain and its isogenic derivatives (increase in volume, flatten ingested (S/H), index I (W/H). These changes were accompanied by the transformation of nanomechanical properties of the cell surface (reducing the root mean square, adhesion force), which countenance was associated with the plasmid profile. The lesser decrease of adhesion force in the absence of changes of the index I was observed in cells Y. pseudotuberculosis and Y. enterocolitica with plasmid pYV. In the strain Y. enterocolitica KM383 (pYV) PO did not induce significant changes in the indicators studied. The introduction of the PO into the cultivation environment decreased the ability of Y. pestis EV cells to bind to human collagen type IV. Modification by PO the adhesive properties of the vaccine strain Y. pestis EV NIIEG was accompanied by an increase in its immunogenicity.</p>Yersinia pestisazoximer bromidebacterial cellatomic force microscopymorphometric analysissurface structureYersinia pestisазоксимера бромидбактериальная клеткаатомно-силовая микроскопияморфометрический анализповерхностная структура[1. Ribet D., Cossart P. How bacterial pathogens colonize their hosts and invade deeper tissues. Microbes Infect. 2015; 17(3): 173–83. https://doi.org/10.1016/j.micinf.2015.01.004][2. Tsang T.M., Annis D.S., Kronshage M., Fenno J.T., Usselman L.D., Mosher D.F., et al. Ail protein binds ninth type III fibronectin repeat (9FNIII) within central 120-kDa region of fibronectin to facilitate cell binding by Yersinia pestis. J. Biol. Chem. 2012; 287(20): 16759–67. https://doi.org/10.1074/jbc.m112.358978][3. Vaca D.J., Thibau A., Schütz M., Kraiczy P., Happonen L., Malmström J., et al. Interaction with the host: the role of fibronectin and extracellular matrix proteins in the adhesion of Gram-negative bacteria. Med. Microbiol. Immunol. 2020; 209(3): 277–99. https://doi.org/10.1007/s00430-019-00644-3][4. Куклева Л.М. Адгезины возбудителя чумы. Проблемы особо опасных инфекций. 2018; (2): 14–22. https://doi.org/10.21055/0370-1069-2018-2-14-22][5. Eddy J.L., Gielda L.M., Caulfield A.J., Rangel S.M., Lathem W.W. Production of outer membrane vesicles by the plague pathogen Yersinia pestis. PLoS One. 2014; 9(9): e107002. https://doi.org/10.1371/journal.pone.0107002][6. Qing G., Gong N., Chen X., Chen J., Zhang H., Wang Y., et al. Natural and engineered bacterial outer membrane vesicles. Biophys. Rep. 2019; 5(4): 184–98. https://doi.org/10.1007/s41048-019-00095-6][7. Дубровина В.И., Мухтургин Г.Б., Балахонов С.В., Витязева С.А., Старовойтова Т.П., Иванова Т.А. и др. Изучение иммунофизиологических свойств штаммов чумного микроба с различным плазмидным составом. Бюллетень Восточно-Сибирского научного центра Сибирского отделения Российской академии медицинских наук. 2013; (6): 136–9.][8. Петров Р.В., Хаитов Р.М., Некрасов А.В., Аттаулаханов Р.И., Пучкова Н.Г., Иванова А.С. и др. Поликсидоний: Механизм действия и клиническое применение. Медицинская иммунология. 2000; 2(3): 271–8.][9. Омельченко Н.Д., Иванова И.А., Беспалова И.А., Филиппенко А.В. Иммуномодуляторы и специфическая профилактика инфекционных болезней. Проблемы особо опасных инфекций. 2017; (3): 21–6. https://doi.org/10.21055/0370-1069-2017-3-21-2][10. Shakya A.K., Nandakumar K.S. Applications of polymeric adjuvants in studying autoimmune responses and vaccination against infectious diseases. J. R. Soc. Interface. 2013; 10(79): 20120536. https://doi.org/10.1098/rsif.2012.0536][11. Крылов Д.А., Чайникова И.Н., Перунова Н.Б., Челпаченко О.Е., Паньков А.С., Смолягин А.И. и др. Влияние иммуномодулятора полиоксидония на биологические свойства микроорганизмов. Журнал микробиологии, эпидемиологии и иммунобиологии. 2003; (4): 74–8.][12. Харсеева Г.Г., Москаленко Е.П., Алутина Э.Л., Бреадо А.М. Влияние полиоксидония на адгезивные свойства Corynebacterium diphtheria. Журнал микробиологии, эпидемиологии и иммунобиологии. 2009; (2): 11–5.][13. Сафенкова И.В., Жердев А.В., Дзантиев Б.Б. Применение атомно-силовой микроскопии для характеристики единичных межмолекулярных взаимодействий. Успехи биологической химии. 2012; 52: 281–314.][14. Beaussarta A., El-Kirat-Chatel S. Microbial adhesion and ultrastructure from the single-molecule to the single-cell levels by Atomic Force Microscopy. Cell Surf. 2019; 5: 10003. https://doi.org/10.1016/j.tcsw.2019.100031][15. Yamashita S., Lukacik P., Barnard T.J., Noinaj N., Felek S., Tsang T.M., et al. Structural insights into Ail-mediated adhesion in Yersinia pestis. Structure. 2011; 19(11): 1672–82. https://doi.org/10.1016/j.str.2011.08.010][16. Chauvaux S., Dillies M.A., Marceau M., Rosso M.L., Rousseau S., Moszer I., et al. In silico comparison of Yersinia pestis and Yersinia pseudotuberculosis transcriptomes reveals a higher expression level of crucial virulence determinants in the plague bacillus. Int. J. Med. Microbiol. 2011; 301(2): 105–16. https://doi.org/10.1016/j.ijmm.2010.08.013][17. Уткин Д.В., Булгакова Е.Г., Ерохин П.С., Кузнецов О.С., Куклев В.Е., Осина Н.А. Исследование морфологических особенностей клеток бактерий Yersinia pestis, выращенных при различных температурных условиях, методом атомносиловой микроскопии. Известия Саратовского университета. Новая серия. Серия: Химия. Биология. Экология. 2019; 19(1): 87–93. https://doi.org/10.18500/1816-9775-2019-19-1-87-93][18. Arseni L., Lombardi A., Orioli D. From structure to phenotype: impact of collagen alterations on human health. Int. J. Mol. Sci. 2018; 19(5): 1407. https://doi.org/10.3390/ijms19051407][19. Karsdal M.A., ed. Biochemistry of Collagens, Laminins and Elastin. Oxford: Academic Press; 2019. https://doi.org/10.1016/C2018-0-00074-2][20. Бывалов А.А., Конышев И.В. Адгезины Yersinia pseudotuberculosis. Инфекция и иммунитет. 2019; 9(3–4): 437–48. https://doi.org/10.15789/2220-7619-2019-3-4-437-448][21. Ma C.D., Acevedo-Vélez C., Wang C., Gellman S.H., Abbott N.L. Interaction of the hydrophobic tip of an atomic force microscope with oligopeptides immobilized using short and long tethers. Langmuir. 2016; 32(12): 2985–95. https://doi.org/10.1021/acs.langmuir.5b04618][22. Iqbal K.M., Bertino M.F., Shah M.R., Ehrhardt C.J., Yadavalli V.K. Nanoscale phenotypic textures of Yersinia pestis across environmentally-relevant matrices. Microorganisms. 2020; 8(2): 160. https://doi.org/10.3390/microorganisms8020160][23. Плескова С.Н., Дубровин Е.В., Голубева И.С., Горшко- ва Е.Н., Пудовкина Е.Е. Нанотехнологическая АСМ-морфометрия бактериальных клеток. Вестник Нижегородско- го университета им. Н.И. Лобачевского. 2013; (2-2): 34–8.][24. Yuan Y., Hays M.P., Hardwidge P.R., Kim J. Surface characteristics influencing bacterial adhesion to polymeric substrates. RSC Advances. 2017; 7: 14254–61. https://doi.org/10.1039/c7ra01571b][25. Евсеева В.В., Платонов М.Е., Копылов П.Х., Дентовская С.В., Анисимов А.П. Активатор плазминогена чумного микроба. Инфекция и иммунитет. 2015; 5(1): 27–36. https://doi.org/10.15789/2220-7619-2015-1-27-36][26. Куклева Л.М., Бойко А.В. Активатор плазминогена – много- функциональный белок возбудителя чумы. Проблемы особо опасных инфекций. 2016; (3): 13–20. https://doi.org/10.21055/0370-1069-2016-3-13-20][27. Kienle Z., Emody L., Svanborg C., O’Tool P.W. Adhesive properties conferred by the plasminogen activator of Yersinia pestis. J. Gen. Microbiol. 1992; 138(Pt. 8): 1679–87.][28. Zhang P., Skurnik M., Zhang S., Schwartz O., Kalyanasundaram R., Bulgheresi S., et al. Human dendritic cell-specific intercellular adhesion molecule-grabbing nonintegrin (CD209) is a receptor for Yersinia pestis that promotes phagocytosis by dendritic cells. Infect. Immun. 2008; 76(5): 2070–9. https://doi.org/10.1128/IAI.01246-07][29. Yang K., He Y., Park C.G., Kang Y.S., Zhang P., Han Y., et al. Yersinia pestis interacts with SIGNR1 (CD209b) for promoting host dissemination and infection. Front. Immunol. 2019; 10: 96. https://doi.org/10.3389/fimmu.2019.00096][30. Byvalov A.A., Kononenko V.L., Konyshev I.V. Single-cell force spectroscopy of interaction of lipopolysaccharides from Yersinia pseudotuberculosis and Yersinia pestis with J774 macrophage membrane using optical tweezers. Biochem. Moscow Suppl. Ser. A. 2018; 12: 93–106. https://doi.org/10.1134/S1990747818020058][31. Книрель Ю.А., Анисимов А.П. Липополисахарид чумного микроба Yersinia pestis: cтруктура, генетика, биологические свойства. Acta Naturae. 2012; 4(3): 49–61. https://doi.org/10.32607/20758251-2012-4-3-46-58][32. Wang C., Stanciu C.E., Ehrhardt C.J., Yadavalli V.K. The effect of growth temperature on the nanoscale biochemical surface properties of Yersinia pestis. Anal. Bioanal. Chem. 2016; 408(40): 5585–91. https://doi.org/10.1007/s00216-016-9659-9][33. Montminy S., Khan N., McGrath S., Walkowicz M. J., Sharp F., Conlon J.E., et al. Virulence factors of Yersinia pestis are overcome by a strong lipopolysaccharide response. J. Nature Immun. 2006; 7(10): 1066–73. https://doi.org/10.1038/ni1386]