RU2355756C1 - Desulfovibrio oxamicus BACTERIA STRAIN, USED FOR PURIFYING SEWAGE WATER FROM SULPHATES AND IONS OF HEAVY METALS - Google Patents

Abstract

FIELD: biology.

SUBSTANCE: Desulfovibrio oxamicus VKM "В-2465Д" bacteria strain is extracted from river bottom sediments. Degree of purity from hexavalent chrome is 99.9%, and 80% from sulphates.

EFFECT: strain is characterised by high degree of purity of sewage water from ions of heavy metals with high concentration of sulphates.

3 cl

Description

The invention relates to industrial microbiology and relates to the production of a new strain of bacteria producing sulfide, which can be used in biotechnology for the treatment of industrial wastewater from engineering, instrument-making, and electrical enterprises from elevated concentrations of sulfate (over 3.0 g / l) and from iron and copper ions chrome.

The use of sulfate reducing bacteria (SBB) for the treatment of wastewater from heavy metal ions is widespread. SVBs grow due to the reduction of sulfates, this process leads to the formation of hydrogen sulfide. Biogenic hydrogen sulfide interacts with heavy metal ions, resulting in the formation of insoluble sulfides or the restoration of some soluble toxic metals with their transition into less toxic and less soluble forms. This principle is based on the use of SVB for biochemical wastewater treatment (Patent RU 2135422, C02F 3/34, 1999.08.27).

To increase the efficiency of biochemical purification methods using SVB, a high level of bacterial growth activity is required. In order to create the desired concentration of SFS in the process of wastewater treatment from sulfates and heavy metal ions, different approaches are used: add easily digestible organic food for SFS (Aut. St. No. 857013, class C02F 3/28, 1979) or use it specifically selected consortia of microorganisms (Vainshtein M., P. Kuschk, J. Mattusch, A. Vatsourina, A. Wiessner. 2003. "Model experiments on the microbial removal of chromium from contaminated groundwater." Water Research. V.37 (6), pp. 1401-1405).

However, the microorganisms used do not provide special activity for the growth of SFS in industrial wastewater, characterized by an increased concentration of sulfates and a different composition of polluting components.

One of the most closely related strains to the proposed facility is the bacterial strain Desulfovibrio vulgaris VKM B-1970 D (UV-3), used to treat wastewater from heavy metal ions. This strain is a highly active strain of sulfate-reducing bacteria, providing a stable biomass increase during wastewater treatment from heavy metal ions (RF Patent No. 2055817, CL C02F 3/34, C12N 1/20, 1996).

However, the VKM strain V-1970 D was not used for wastewater treatment at elevated sulfate concentrations.

The bacterial strains UFZ B 378, UFZ B 406 and UFZ B 407, reducing sulfates in sulfate and metal-containing waters, have been previously described (Patent RU 2174554, C12N 1/20, 1995.07.28).

The description does not indicate the taxonomic positions of these bacterial strains. Strains are used to improve acidic (pH 1-2) mine water and water filling closed mines, strains are anaerobic methylotrophic sulfate reducing agents. The method of bacterial decontamination of sulfate and metal-containing water using strains UFZ B 378, UFZ B 406 and UFZ B 407 differs in that only methanol is added as the sole source of carbon and energy.

However, methanol is a toxic compound, and its use should be limited.

The objective of the invention is to obtain a strain of bacteria for wastewater treatment from sulfates and heavy metal ions.

The technical result that can be achieved by using the present invention is that it provides for the treatment of wastewater from sulfates at their elevated concentrations and the possibility of conjugate removal of heavy metals. In addition, the range of bacteria strains used for wastewater treatment is expanding.

The invention is achieved by the use of a strain of bacteria Uf2v for wastewater treatment from sulfates and heavy metal ions.

The bacterial strain Uf2v was isolated from the bottom sediments of the river below the releases of industrial wastewater from a mining and processing enterprise and a paper mill.

The proposed strain of bacteria Uf2v, identified as Desulfovibrio oxamicus, is deposited in the All-Union Collection of Microorganisms (VKM) of the Institute of Biology and Physiology of Microorganisms named after G.K.Skryabin RAS under the number VKM V-2465 D.

The proposed bacterial strain Desulfovibrio oxamicus VKM B-2465 D is an active producer of hydrogen sulfide for the precipitation of heavy metal ions from solutions and is characterized by the following cultural-morphological and physiological-biochemical properties:

1. Cultural and morphological characteristics of the proposed strain is determined during its cultivation on a modified medium N 92M. The base environment N 92 is taken from the catalog of the All-Russian collection of microorganisms "All-Russian collection of microorganisms. Catalog". Pushchino - Moscow, VNITI, 1992, 362 p. The recipe for the modified medium N 92M is given below. Cultivation is carried out under anaerobic conditions at 30 ° C for two days.

2. Cultural signs: with the growth of the strain, the liquid medium opalesces, a black precipitate of iron sulfide accumulates at the bottom of the cultivation vessel, and the smell of hydrogen sulfide is present.

3. Morphological features and cell cytology: under microscopy, cells are represented by single vibrios with a size of 0.5 μm × 2-3 μm; cells are motile, have one polar flagellum. The cell wall is a gram-negative type (according to electron microscopy). They do not form a dispute.

Features of growth: reduction of sulfate to sulfide (hydrogen sulfide) with the accumulation of black ferrous sulfide in the standard medium, as well as (after exhaustion of iron) hydrogen sulfide.

4. Physico-biochemical characteristics of the proposed strain. Strict anaerobic. Able to sulfate breathing: uses sulfate, thiosulfate as an electron acceptor. As hydrogen donors and carbon sources use: sodium lactate, sodium acetate with yeast extract, pyruvate, malate. Ammonium and amino acids of the yeast extract are able to use nitrogen sources. Does not need growth stimulants. The pH range is from 6.5 to 8.0 with an optimum at 7.2-7.4; the medium used is 63DSM. With growth, a slight alkalization of the medium is observed, which correlates with the replacement of sulfate by sulfide. The temperature range for strain growth from 25 to 35 ° C, optimum at 30 ° C. Cells contain cytochrome C. Storage conditions: maintained by reseeding on culture medium. It can be stored in the refrigerator at 4-10 ° C for 3-6 months.

5. A comparative analysis of the sequenced fragment of the 16S rRNA gene showed that the proposed strain belongs to the species Desulfovibrio oxamicus with 98% similarity of the nucleotide sequences of the 16S rRNA genes.

On the basis of the data obtained in accordance with the determinant of Bergey bacteria (The determinant of Bergey bacteria. In 2 volumes T. 1: Transl. From English / Ed. J. Hole and others - M .: Mir, 1997) and according to the results a comparative analysis of the nucleotide sequence of the 16S rRNA gene with data from the NCBI database (GenBank - http://www.ncbi.nlm.nih.gov) the proposed strain was identified as Desulfovibrio oxamicus.

The use of bacteria strains of the species Desulfovibrio oxamicus for wastewater treatment was not previously known.

The proposed strain reduces the concentration of sulfates by 80% and heavy metals, in particular chromium (VI), by 99%.

The possibility of carrying out the invention is confirmed by the following examples, but is not limited to.

Example 1. Obtaining a strain of Desulfovibrio oxamicus VKM B-2465 D

The proposed strain is obtained by selecting bottom sediments and isolating from them an accumulative culture of bacteria by growing under anaerobic conditions at 30 ° C.

Lactic acid (lactate) is used as the sole carbon source for growth. Then, strain selection is continued at increasing concentrations of sulfates in the culture medium. The final selection of the pure culture of the resulting strain is produced by the method of serial dilutions, followed by seeding in an agarized version of the nutrient medium and reseeding of single colonies.

Work on the isolation and cultivation of the proposed strain is carried out on a modified medium N 92M.

Prescription medium N 92M:

Solution 1:

K ₂ HPO ₄ - 0.2 g

NH ₄ Cl - 0.3 g

NaCl - 1.0

CaCl ₂ × 2H ₂ O - 0.16

KCl - 0.5

MgSO ₄ × 7H ₂ O - 0.4 g

Na ₂ SO ₄ - 3.0 g

Yeast extract - 0.1 g

Distilled water - 973.0 ml

Solution 2:

Lactic acid (40%) - 1 ml

Solution 3:

NaHCO ₃ - 1.0 g

Distilled water - 10.0 ml

Solution 4:

Na ₂ S × 9H ₂ O - 0.3 g

Distilled Water - 6.0 ml

Solution 5:

FeSO ₄ × 7H ₂ O - 0.4 g

Distilled water - 10.0 ml

Preparation of modified medium N 92M for isolation and cultivation of the proposed strain.

Solution 1 was brought to a boil while blowing through this solution an oxygen-free gas mixture consisting of 97% N ₂ and 3% H ₂ . They are sterilized in the atmosphere of these gases. Solutions 3, 4 and 5 are sterilized in an atmosphere of N ₂ . After sterilization, all five solutions are combined. The pH value of the medium after mixing is set to 7.2-7.4.

Get a pure culture of the proposed strain of Desulfovibrio oxamicus VKM B-2465 D.

Example 2. The use of a strain of bacteria Desulfovibrio oxamicus VKM B-2465 D for water purification from high concentrations of sulfates

The process is carried out in a batch reactor. The reactor is seeded with a pre-grown culture of bacteria Desulfovibrio oxamicus VKM B-2465 D. The temperature of the cultivation is maintained in the range of 28-30 ° C. As a result of the life of the proposed strain, hydrogen sulfide is released and a precipitate of iron sulfide is formed, the sulfate content is reduced. The efficiency of removing sulfate from a liquid medium is increased by removing from the liquid medium the dissolved hydrogen sulfide produced by the proposed strain in the form of insoluble iron sulfide.

The efficiency of the reactor for water purification from increased concentrations of sulfates is up to 80% with an initial concentration of sulfates of 3 g / l in 14 days.

Example 3. The use of a strain of bacteria Desulfovibrio oxamicus VKM B-2465 D for water purification from hexavalent chromium ions

The process is carried out in a batch reactor. A reactor culture of the Desulfovibrio oxamicus VKM B-2465 D bacterial culture previously grown and immobilized on a porous support is seeded. The culture temperature in the reactor is maintained in the range of 28-30 ° C. A medium simulating wastewater with a chromium (VI) concentration of 17.5 mg / L and sulfate 0.5 g / L are introduced into the reactor. Lactic acid is used as a carbon source. As a result of the life of the SRB, hydrogen sulfide is released, which combines with metal ions, the accumulation of hydrogen sulfide in the medium is noted only after the complete disappearance of chromium (VI).

The efficiency of the reactor to remove 17.5 mg / l of hexavalent chromium is 99.9% in 5 days.

Thus, wastewater containing increased concentrations of sulfates (more than 3 g / l) and some heavy metal ions can be effectively treated with the proposed bacterial strain Desulfovibrio oxamicus VKM B-2465 D. The range of bacterial strains used for wastewater treatment is expanding. . No toxic components are used.

Claims (1)

The bacterial strain Desulfovibrio oxamicus BKM B-2465 D, used for wastewater treatment from sulfates and heavy metal ions.