Adaptation to growth at low temperature of psychrotrophic bacteria isolated from soil of the northern region of the Perm Krai

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Abstract

Halotolerant bacteria of the genera Bacillus, Microbacterium, Glutamicibacter, Brevibacterium, Rhodococcus, Pseudomonas, Thalassospira, as well as halophilic bacteria of the genus Halomonas, previously isolated from saline ecotopes of the northern region of Perm Krai, were studied for their ability to grow at low temperatures. It was found that strains of bacteria of the genera Rhodococcus, Glutamicibacter, Microbacterium, Pseudomonas, Brevibacterium, Thalassospira and Halomonas grew at a temperature of 5°C. Bacteria belonging to the genera Brevibacterium and Rhodococcus grew at 0°C. Cryoprotective substances were studied using proton magnetic resonance spectroscopy. Since in natural habitats bacterial cells can attach to the surfaces of solid particles and form microcolonies, cryoprotectors were studied by growing bacterial strains on the surface of an agar medium. Using the psychrotrophic culture of Rhodococcus sp. SMB38 as an example, the dependence of the osmolyte composition on cultivation conditions was studied. When grown on an agar medium without adding sodium chloride at 25°C, the cells of the strain Rhodococcus sp. SMB38 synthesized ectoine, hydroxyectoine, trehalose and alanine. The proportion of trehalose was higher than other compounds and amounted to 48%. With a decrease in temperature, the intracellular amount of trehalose, ectoine and alanine increased. In this case, the proportion of trehalose was higher than other compounds and was equal to 55%. The same patterns were noted when growing the strain on a dense medium containing 2% NaCl. However, under cold conditions, a change in the ratio of compounds was revealed: the largest share, equal to 48%, belonged to ectoine. In this regard, the study of cryoprotective substances was carried out by culturing cold-resistant strains on a dense medium containing 2% NaCl. It was shown that the cells of bacteria of the genera Halomonas and Thalassospira accumulated ectoine. In the cells of bacteria of the genera Glutamicibacter and Pseudomonas, trehalose performed the function of a cryoprotector. Psychrotrophic culture of the genus Brevibacterium accumulated trehalose, ectoine and alanine in cells. In this work, an increase in the amount of alanine in bacterial cells with a decrease in temperature is described for the first time, which may indicate the participation of this amino acid in adaptation to cold.

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L. N. Anan’ina

Perm Federal Research Center of the UB RAS

Author for correspondence.
Email: ludaananyina@mail.ru

Institute of Ecology and Genetics of Microorganisms UB RAS

Russian Federation, 614081, Perm

A. A. Gorbunov

Perm Federal Research Center of the UB RAS

Email: ludaananyina@mail.ru

Institute of Technical Chemistry UB RAS

Russian Federation, 6614013, Perm

V. S. Aleev

Perm Federal Research Center of the UB RAS

Email: ludaananyina@mail.ru

Institute of Ecology and Genetics of Microorganisms UB RAS

Russian Federation, 614081, Perm

O. V. Yastrebova

Perm Federal Research Center of the UB RAS

Email: ludaananyina@mail.ru

Institute of Ecology and Genetics of Microorganisms UB RAS

Russian Federation, 614081, Perm

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2. Fig. 1. Amounts of osmolytes accumulated by Rhodococcus sp. SMB38 cells under different cultivation conditions. Incubation was carried out on a solid medium. Glucose served as a source of carbon and energy. Cultivation conditions (medium salinity, temperature): 1 – without sodium chloride, 25°C; 2 – without sodium chloride, 5°C; 3 – 2% NaCl, 25°C; 4 – 2% NaCl, 5°C.

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3. Fig. 2. 1H NMR spectra of ethanol extracts from cells of cold-resistant bacterial strains isolated from the VMKMS area of ​​Perm Krai. The bacterial strains were cultured at 5°C on a dense medium containing 2% sodium chloride. Legend: alanine (A); glutamate (G); glutamine (Q); trehalose (T); ectoine (E); unidentified compound (X); monodeuterium water (HDO); tert-butyl alcohol (t-BuOH); SMB31, SMB32, SMB34, SMB38, SN101, U1 – designations of bacterial strains.

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4. Fig. 3. Sections of the 1H NMR spectra of ethanol extracts from cells of the Pseudomonas sp. SN101 strain grown at 5°C on an agar medium containing 2% sodium chloride (a) and without it (b). Glucose served as a source of carbon and energy in both variants. Legend: glutamate (G); trehalose (T); unidentified compounds (X, Y).

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