Analysis of electrophysical profiles of plankton and biofilm cells on the model of Azospirillum baldaniorum bacteria

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Abstract

Biofilm formation is a widespread phenomenon in the world of microbes. They can affect human and animal health, cause damage to various industries, and at the same time can be useful in areas such as wastewater treatment or increasing the bioavailability of nutrients for plants. This actualizes the development of biofilm research methods. In this paper, an optical sensor method for indicating bacterial biofilm formation taking into account biological variability is described for the first time using the example of plant growth-stimulating rhizobacteria of the genus Azospirillum. A correlation was found between changes in the electrophysical parameters recorded by the sensor system and morphological features of bacteria from planktonic and/or biofilm cultures: the presence of motor organelles (flagella), polymorphism and ultrastructure of cellular forms. It was found that the profile of microbial cells recorded by the optical system in planktonic and biofilm forms differs significantly. When comparing cells of different strains (parent strain and its derivatives) or planktonic and biofilm bacteria, the variables recorded by the electro-optical sensor system are consistent with the changes in the micro- and ultrastructure of bacteria recorded by us using other methods. The results of the analysis of the electrophysical profiles of A. baldaniorum Sp245 can be used as a reference for identifying the specificity of the interaction of biofilm cells of this strain with various components of the root surface of the putative plant partner using an optical sensor system.

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About the authors

A. V. Sheludko

Federal Research Center “Saratov Scientific Center of the Russian Academy of Sciences”

Author for correspondence.
Email: shel71@yandex.ru

Institute of Biochemistry and Physiology of Plants and Microorganisms

Russian Federation, Saratov, 410049

S. S. Evstigneeva

Federal Research Center “Saratov Scientific Center of the Russian Academy of Sciences”

Email: shel71@yandex.ru

Institute of Biochemistry and Physiology of Plants and Microorganisms

Russian Federation, Saratov, 410049

E. M. Telesheva

Federal Research Center “Saratov Scientific Center of the Russian Academy of Sciences”

Email: shel71@yandex.ru

Institute of Biochemistry and Physiology of Plants and Microorganisms

Russian Federation, Saratov, 410049

Yu. A. Filip’echeva

Federal Research Center “Saratov Scientific Center of the Russian Academy of Sciences”

Email: shel71@yandex.ru

Institute of Biochemistry and Physiology of Plants and Microorganisms

Russian Federation, Saratov, 410049

L. P. Petrova

Federal Research Center “Saratov Scientific Center of the Russian Academy of Sciences”

Email: shel71@yandex.ru

Institute of Biochemistry and Physiology of Plants and Microorganisms

Russian Federation, Saratov, 410049

D. I. Mokeev

Federal Research Center “Saratov Scientific Center of the Russian Academy of Sciences”

Email: shel71@yandex.ru

Institute of Biochemistry and Physiology of Plants and Microorganisms

Russian Federation, Saratov, 410049

I. V. Volokhina

Federal Research Center “Saratov Scientific Center of the Russian Academy of Sciences”

Email: shel71@yandex.ru

Institute of Biochemistry and Physiology of Plants and Microorganisms

Russian Federation, Saratov, 410049

I. V. Borisov

Federal Research Center “Saratov Scientific Center of the Russian Academy of Sciences”

Email: shel71@yandex.ru

Institute of Biochemistry and Physiology of Plants and Microorganisms

Russian Federation, Saratov, 410049

V. D. Bunin

EloSystem GbR

Email: shel71@yandex.ru
Germany, Berlin 13407

O. I. Guliy

Federal Research Center “Saratov Scientific Center of the Russian Academy of Sciences”

Email: guliy_olga@mail.ru

Institute of Biochemistry and Physiology of Plants and Microorganisms

Russian Federation, Saratov, 410049

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2. Fig. 1. A ‒ Transmission electron microscopy of A. baldaniorum Sp245 and Sp245.1063 cells from 20 h liquid cultures (1) or biofilms (2) formed on glass under liquid medium during 6 days of cultivation. B ‒ Transmission electron microscopy of ultrathin sections of A. baldaniorum Sp245 cells from biofilm fragments formed under liquid medium on the glass surface during 6 days of cultivation. Panel (a) shows “vegetative” and “long” cells, panel (b) shows “cyst-like forms”. Scale bar ‒ 1 μm.

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3. Fig. 2. A ‒ Effect of medium composition on the rate of movement of planktonic cells of A. baldaniorum Sp245 in liquid medium. B ‒ Electrophysical profiles of bacteria from planktonic 20-h liquid cultures and 6-day biofilms. C ‒ Distribution of cells by length. The average length of A. baldaniorum Sp245 and Sp245.1063 cells, respectively, in 20-h planktonic liquid cultures was (2.6 ± 0.3) and (2.1 ± 0.2) μm, (2.8 ± 0.3) and (2.8 ± 0.2) μm in the case of 6-day biofilms. D ‒ Dynamics of biomass accumulation in biofilms formed by A. baldaniorum Sp245 and Sp245.1063 on glass under liquid LB. OP 590 – optical density of crystal violet desorbed after biofilm staining. D – Results of changes in the electro-optical signal of cells at a frequency of 740 kHz. E – Effect of pronase and sodium periodate on the biomass of 6-day-old biofilms formed on glass under liquid LB medium. % – percentage ratio of the optical density of the dye desorbed from stained films after their incubation in pronase or sodium periodate solution to the same indicator without treatment. One-way analysis of variance (ANOVA) of the data in panels (D) and (E) was performed comparing the indicators for each strain (lowercase letters) or comparing the indicators of Sp245 and Sp245.1063 (uppercase letters); different letters denote statistically significant differences; a, a, A and A are the mean values ​​with the smallest magnitude.

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4. Fig. 3. Atomic force microscopy results. AFM images of 6-day biofilms of the A. baldaniorum Sp245 strain formed on the liquid LB/glass (A, B, D, E) and liquid LB/air (B, E) interfaces.

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