Population density and size of bacteria in the course of cultivation of their small formsEurasian Soil Sc.


L. M. Polyanskaya, R. B. Gorodnichev, E. A. Vorob’eva, D. G. Zvyagintsev
Earth-Surface Processes / Soil Science


Max born medal and prize

The Institute of Physics

Rape in marriage

Lee H. Bowker, o̊Dean of the Graduate School and Research


ISSN 10642293, Eurasian Soil Science, 2015, Vol. 48, No. 4, pp. 395–399. © Pleiades Publishing, Ltd., 2015.

Original Russian Text © L.M. Polyanskaya, R.B. Gorodnichev, E.A. Vorob’eva, D.G. Zvyagintsev, 2015, published in Pochvovedenie, 2015, No. 4, pp. 447–451. 395


It is supposed that the small size of the bacteria in a soil is determined by the natural conditions, which limit the bacterial development by a deficit of nutrients or by stress situations. Small bacteria cells with a min imal size of 0.2 µm, which is considered to be the smallest [14], were found with the help of an epifluo rescent microscope in many samples from lakes, riv ers, soils, snow, and rainwater as a response of the cells to unfavorable environment and stress factors, which were studied in laboratory experiments [10]. It was found that the cultivation of the obtained nanoforms on rich nutrient media resulted in the return of the ini tial forms [1, 6, 11, 20].

An earlier study with rearing cells (0.23 µm in diameter) in the filtrates of a soil suspension in a ther mostat at a temperature of 28°C demonstrated that the bacterial cells reproduced intensely during incuba tion, and this was recorded with the classic method by the increase of the population density on glasses [9]. It should be noted that the cell rearing was carried out in a soil suspension without introducing additional nutri ent substrates.

The aim of this work was to study the number and size of the bacterial cells before and after cultivation of their small forms (0.23 and 0.38 µm) under favorable moisture and temperature conditions.


Two samples of soils developing under xerophytic communities were studied. A sample from the surface layer (0–5 cm) of a mountain meadowsteppe shal low subalpine soil (Mollic Leptosol) was taken 20 km northwest of Dushanbe (Tajikistan) at the altitude of 1000 m a.s.l. in November 2010. It was placed into a sterile bag, stored for 8 h at –4°C, and then taken to the laboratory (within 8 h), where it was stored at ⎯18°C.

A sample from a loesslike residualsolonchakous subtropical light sierozem (Haplic Calcisol (Endosalic,

Yermic)) was taken in the Negev Desert (Israel) at the altitude of 600–800 m a.s.l. This is an area with an arid subtropical climate with mean monthly temperatures ranging from +12°C in January to +27°C in July and annual precipitation from 50 to 300 mm; no rains take place from June to October. The soil was formed under conditions of a nonpercolative water regime and was characterized by considerable accumulations of car bonates, sulfates, and chlorides in the subsoil.

The samples were subjected to ultrasonic pretreat ment using a lowfrequency UZDN1 (22 kHz, 0.44 A, 2 min) disperser [4].

The method of cascade filtration was applied to determine the numbers of bacteria in separate size fractions: 1 mL of soil suspension (1 : 100) was filtered through nuclear filters (Dubna Scientific Center Pro duction) with pore diameters of 1.85, 0.43, 0.38, and


Population Density and Size of Bacteria in the Course of Cultivation of Their Small Forms

L. M. Polyanskaya, R. B. Gorodnichev, E. A. Vorob’eva, and D. G. Zvyagintsev

Faculty of Soil Science, Moscow State University, Vorob’evy gory, Moscow, 119991 Russia email: lpolyanskaya@mail.ru

Received April 14, 2014

Abstract—The population density, size, and biomass of the soil bacteria from a mountain meadowsteppe soil of Tajikistan and a light sierozem of the Negev Desert have been analyzed using the method of “cascade” filtration. It was shown that, when cultivating small fractions of soil bacteria, the total number of bacteria increased by 1.5 times and the bacterial size became greater. The number of coarse cells with a size of 1.85 and 0.43 μm essentially increased in both soils. If the contribution of these fractions was about 10–20% in the initial soils, it increased up to 50–60% in the incubated filtrates. The cells with a size of 0.38 and 0.23 μm accounted for about 70% of the total bacteria in the initial soils, while, in the incubated filtrates, the share of 0.23 μm cells composed about 30% in the filtrate and that of 0.38 μm cells reached 45–50% in the filtrate.

The average diameter of the bacteria increased from 0.4 to 0.8–0.9 μm; the biomass of bacteria in these fil trates increased by 7–8 times in comparison with the initial soils at the expense of an increasing number of large cells after cultivation.

Keywords: Mollic Leptosols, Haplic Calcisols (Endosalic, Yermic), bacteria, cell size, cultivation, biomass

DOI: 10.1134/S1064229315040079 396


POLYANSKAYA et al. 0.23 µm and a membrane filter (Synpore) with a pore diameter of 0.17 µm. The filtration was performed with a vacuum pump and a Bunsen flask. The lumines cence of the filters was extinguished by staining with a saturated alcohol solution of Sudan black (Germany).

The nuclear filters were placed into this solution for several days and the membrane filters for several hours; then, they were washed out in sterile water, dried, and used for filtering [13].

Four layers of filtering paper were placed on the surface of a metallic screen in a Bunsen flask, a filter (nuclear or membrane) was placed atop and forced against the surface of the device with a metallic ring, and the suspension was added. The suspension was fil tered stepby step from the filter with greater pore size to that with a smaller pore size. The number of bacte ria was counted on every filter (three replicate filters were used), and the population density was calculated taking conventionally that the cell size was equal to or slightly greater than the pore diameter of the filter on which the bacteria were retained. The calculations were performed on the basis of the assumption that the cells had a spherical shape, and this was confirmed by the data of electron and scanning microscopy [8].