RU2370458C2 - Method of water cyanobacteriae bloom control - Google Patents

Abstract

FIELD: biotechnology.

SUBSTANCE: invention concerns biotechnology and can be applied for activation of open water pool and stream self-clarification. Method involves water pool algolisation by suspension of Chlorella vulgaris strain #C-111 of Plant Physiology Institute in winter, spring and summer periods. In winter and spring the suspension undergoes adaptation before algolisation. Algolisation is performed at least for 3 years.

EFFECT: prevented water bloom caused by cyanobacteriae.

Description

The invention relates to biotechnology and can be used to enhance the self-cleaning of open water bodies and streams.

As you know, algae play an important, positive role in self-cleaning processes. In the process of photosynthesis, they produce oxygen, which helps to increase the level of oxygen saturation of water and the mineralization of organic substances. For metabolic processes associated with the formation of organic substances, algae absorb various nutrients and soluble organic substances from water, which also contributes to self-purification processes. However, the positive function of algae in the processes of self-purification largely depends on their concentration and the implementation of active life. When one of the algae species is propagated to high concentrations, their positive role as the primary producing element is replaced by a negative one and they turn from a self-cleaning factor into a factor of self-pollution or biological pollution, and also simplify the ecosystem and make it less resistant to extreme factors (Kuzmenko M. I. The role of organic nutrition in the productivity of algae. - Gidrobiol. Zh., 1970, 6, No. 1, pp. 109-124).

With an increase in the concentration of algae biomass to 100 mg / l dry matter and more dramatically decreases the oxygen productivity of cells, the decomposition processes that occur with the absorption of oxygen are enhanced. Organic matter and its decomposition products accumulate in the medium. As a result of the excessive development of algae and their subsequent death, secondary biological pollution of water bodies and a significant deterioration in water quality occur (water BOD ₅ increases, bichromate oxidizability and the number of suspended particles increase, color increases and transparency decreases, harmful substances accumulate).

Phenols appear in water in concentrations exceeding the permissible sanitary norm up to 200 times. During fermentation from the organic matter of blue-green algae, acetone, alcohols, and organic acids are formed in clusters. The oxygen content in the water during the decay of the masses of blue-green algae drops to zero, especially in places of surge, increases BOD ₅ , resulting in local fish kills, especially juveniles.

The danger of excessive "flowering" is further aggravated by the fact that, during the course of life, blue-green algae alkalize the environment and create favorable conditions for the development of pathogenic microflora and pathogens of intestinal diseases, including cholera vibrio. In addition, during the “bloom” period, algae more actively absorb the short-wave part of visible light, heat up and are sources of infrared radiation, which can significantly affect the thermal regime of the reservoir.

A known method of dealing with the "flowering" of reservoirs with blue-green algae, providing for the stocking of reservoirs with herbivorous fish and algolization in winter with a suspension of the strain Chlorella vulgaris BIN (RU 2263141, C12N 1/12, publ. 02/10/2005).

The method was carried out in the Penza reservoir (dam section), with an area of shallow water - 20-25%, average depth - 5.1 m, maximum depth -15 m. “Blooming” of water in summer is caused by the blue-green alga Merismopedia tenuissima, which prefers coastal zone and does not produce toxins.

The oxygen regime of the Penza reservoir during the research period was at a relatively stable level in all parts of the reservoir.

The known method involves the stocking of a reservoir of herbivorous fish - white and motley silver carp to increase the effect of algolization.

However, it is known that white and motley silver carp use various types of algae for food, but they primarily prefer representatives of protococcal and diatoms. However, blue-green algae can make up a significant part of its food lump, despite the fact that their assimilation practically does not occur (Borutsky, EB Materials on the nutrition of the Amur silver carp. Works of the Amur. Ichthyol. Expedition, 1949, v. 1. M ., 1950, pp. 287-303. Borutsky EB Food white and colorful silver carp in natural ponds and ponds of the USSR. - In the book: trophology of aquatic animals. M., 1973, pp. 199-322).

Moreover, through the intestines of the white silver carp, the cells of the main causative agent of the “bloom” of Microcystis aeruginosa water (this particular type of algae dominates the others during the “bloom” of water in the Volga reservoirs) passes through a large mass without damage and are alive, capable of germination.

In this case, the Microcystis colonies are crushed already in the first part of the intestine into separate parts, the cells of the latter are compacted and take the form of a ball. Changing the nature of cell packings in the colony and the formation of spherical shapes can be considered as an adaptation to maintaining viability in extreme conditions. This form reduces contact with the acidic digestive juice of the intestinal tract of fish. In the middle part of the intestine, the packing density of cells increases even more, and their aggregation occurs. In the posterior intestine, the number of spherical aggregates decreases as a result of their decay into individual living cells that are secreted outward. Algae cells that have passed through the intestines are capable of reproduction and after 3-6 days of their stay in water, young intensively growing colonies form from them (Sirenko L.A., Gavrilenko M.Ya. “Flowering” of water and eutrophication. Kiev: Naukova Dumka, 1978 p. 60-62, 96-97).

Thus, the number of spores of blue-green algae wintering in the upper layers of silt can significantly increase.

In addition, it should be noted that water temperature is the most important factor in the seasonal cycles of phytoplankton. The temperature optimum of different types of algae does not coincide, which determines the change in the species composition by season. The general scheme of the annual cycle of phytoplankton in ponds of temperate latitudes has the following form. In winter, under ice (especially when ice is covered with snow), phytoplankton is almost absent due to lack of solar radiation. The vegetation cycle of phytoplankton begins in March-April, when solar radiation is sufficient for photosynthesis of algae, even under ice. In this period, small flagellate and cold-water species of diatoms begin to develop, then moderately warm diatoms, green and blue-green. In summer, there is a maximum productivity of blue-green and green algae. Diatoms in the summer, as a rule, occupy a subordinate position and are represented by warm-water species. In autumn, with a decrease in water temperature to (+10) - (+ 12) ° С and lower, the productivity of cold-water species of diatoms again increases (Plant Life, vol. 3 algae. Lichens, edited by M.M. Gollerbakh. M.: Education, 1977, pg. 7-100).

The known method involves the use as an algolysant suspension of a strain of Chlorella vulgaris BIN. This strain of green algae has a wide temperature range and exhibits pronounced antagonistic properties to algoflora. This means that the coincidence of the temperature optimums of diatoms and Chlorella vulgaris BEST can cause inhibition of the former and even loss of certain diatoms from the biocenosis. In addition, the ratio of the biomass of planktonic and benthic forms of diatoms may be disturbed.

Diatoms occupy an absolutely exceptional place in the general circulation of substances in nature. Being a powerful and inexhaustible source of organic matter, they serve as a constant feed base and the initial link in the food chains of many organisms. When they die, they give a mass of detritus and soluble organic substances that go to the nutrition of bacteria and protozoa. By themselves, they are excellent food for many invertebrate animals and fish. A direct relationship has been established between the quantity and distribution of phyto- and zooplankton, phyto- and zoobenthos, and fish productivity.

Microcystis aeruginosa, Anabaena, and Aphanizomenon flos-aguae occupy a dominant position in the "bloom" of water in the reservoirs of the Volga cascade. These types of algae cause "bloom" in almost all climatic zones, produce toxins.

The technical result is the prevention of "flowering" of water by blue-green algae.

The technical result is achieved by the fact that in the known method, which provides for the algolization of reservoirs in winter - February with a suspension of green algae Chlorella vulgaris, according to the invention, the algolization of reservoirs is additionally carried out in the spring and summer periods of the year, suspension of the strain Chlorella vulgaris IGF No. C-111 is used as an algolizing agent. with a transmittance of 1.4-6 in the amount of 15-25 l per 1 million m ^{3 of} water, and in bays, beams and places of wind surges at the rate of 15-25 l / ha, adaptation is carried out before algolization in winter and spring algolysant.

In the spring, the algolization of water bodies is carried out twice in the first phase of spring: March-April, during ice-freeing - after breaking the ice and when the upper layer of water is warmed up to (+10) - (+ 12) ° С, in the second phase: when the average depth is reached water temperature in the reservoir (+5) - (+ 10) ° С, in the summer months - monthly, and in bays, gullies and places of wind surges in the summer period at least 2 times a month.

Adaptation of the algolizant is carried out by cooling it from the temperature of cultivation to the temperature of the water in the pond at the time of algolization at the rate of 3-5 ° C per day and holding at this temperature for 1.5-2 days.

Algolization is carried out for at least 3 years according to the same scheme.

In the proposed method to obtain an algolysant, a strain of unicellular green alga Chlorella vulgaris IGF No. C-111 (Su 1751981, C12N 1/12, publ. 02.10.1997) is used. The strain was isolated from water samples of the Nurek reservoir by the method of accumulative cultures. Young cells are weakly ellipsoid, ranging in size from 0.5 to 2.0 microns. Adult spherical with a diameter of 6-9 microns, do not settle to the bottom. The optimum cultivation temperature is (+26) - (+ 36) ° С.

The development cycle of the strain is as follows: during the daytime, an active process of photosynthesis takes place, as a result of which the cells intensively gain biomass. Cell sizes from 6 to 21 hours increase from 3 to 9 microns. Starting from 22 to 4 hours, they are actively divided. By 5 hours, young cells are ready for photosynthesis. The development cycle is persistent, it can be violated by artificially changing the light regime.

The dominant position in the “flowering” of water in the Volga reservoirs is occupied by the blue-green alga Microcystis aeruginosa, which belongs to the class Chroococceae, order Chroococcales, order Stereometreae, family Microcystidaceae Elenk, genus Microcystis. The order Chroococcales is characterized by unicellular and colonial or only colonial algae, with a thick, mucous, structureless gelatinous mass. Colonies are free, spherical, ellipsoidal or shapeless. Spherical cells with gas vacuums, 3-7 microns in diameter, densely located in the colony. Reproduction - most often occurs by cell division, less often - the formation of nannocytes.

The optimum temperature for growth and development is 30-34 ° C.

Thus, the temperature optimum for strain Chlorella vulgaris IGF No. C-111 and Microcystis aeruginosa is at the same level.

In the cells of Chlorella vulgaris of both strains (used in the prototype and in the proposed solution), there are cell structures - lysosomes containing enzymes from the number of hydrolases that can cleave the most important classes of chemical compounds, including proteins, carbohydrates, nucleic acids.

The basis of the shell of diatoms and blue-green algae is a protein-carbohydrate complex, which dissolves under the action of hydrolytic enzymes Chlorella vulgaris.

However, the active isolation of enzymes in different strains of Chlorella vulgaris occurs at different temperatures: 20 ° C in BIN and 26-30 ° C in IGF No. C-111. Therefore, in the prototype, there may be lysis of diatom cells, and in the proposed method, blue-green, in particular Microcystis aeruginosa.

The adaptation of the algolizant to the environmental conditions of the reservoir is due to the following.

The cultivation temperature of the strain Chlorella vulgaris IGF No. C-111 (+26) - (+ 36) ° C. The temperature of the water in the pond at the time of algolization in the winter, under the ice is in the range (+0.5) - (+ 1.5) ° С. The introduction of a suspension containing cells in a vegetative state, i.e. capable of active reproduction, the water causes severe stress, as a result of which biochemical changes occur in a living organism, aimed at overcoming the factor causing stress. First of all, the activity of oxidative enzymes sharply increases, the energy metabolism of the cell changes.

Using an electron microscope, it was found that young cells of the strain Chlorella vulgaris IGF No. C-111, whose size is 0.5-2.0 μm, are compressed with a sharp change in temperature. The cell membrane, not adapted to low temperatures, becomes rigid, the pores that connect the cell with the outside world are closed. The higher the intensity and duration of exposure, the greater the severity of these changes until the death of the cell. Adult cells, the size of which is 6-9 microns, placed after 30 days in conditions of a slow rise in temperature to + 26 ° C, partially restore the ability to reproduce.

Studying the cells of the strain of Chlorella vulgaris IGF No. C-111 found that the cells in the vegetative state (just obtained during cultivation) have a primary membrane. In the process of slow cooling of the suspension, the cells naturally transition to a state of rest with the formation of a secondary membrane, the density of which is much higher than the density of the primary membrane. This gives the cell strength. Moreover, for about 1.5-2 days after reaching the required temperature, the compaction of the shell continues.

A slow rise in the temperature of water containing a suspension of strain Chlorella vulgaris IGF No. C-111, at a rate of 3-5 ° C per day to a temperature of + 26 ° C, made it possible to establish that the secondary shell stretches, becomes elastic, as if dissolved. The cell gradually goes into a vegetative state with high activity for reproduction.

The method is as follows.

From a reservoir intended for algolization of Chlorella vulgaris, 250-300 l of water are taken and the strain of Chlorella vulgaris IGF No. C-111 was cultivated on it.

For the cultivation of the strain, a nutrient medium of the following composition is prepared:

ammonium nitrate 1 kg superphosphate, 10% solution 1 liter ferric chloride, 1% solution 150 ml cobalt nitrate 1 g copper sulfate 1 g bacterial suspension 40 l

Cultivation was carried out until a transmittance of 1.4-6% was achieved, which was determined on a Spekol spectrocolorimeter.

Studies of the effectiveness of the method were carried out on the Bereslavsky reservoir.

The main parameters of the morphometry of the reservoir:

Mirror area, thousand ha 1,52 The volume of water, million m ³ 52,5 Maximum depth, m 8.4 Average depth, m 3,5 The greatest width, km 2.9 Length, km 9.0

The climate of the reservoir area is sharply continental: summers are hot, not sufficiently humid, winters are cold, with little snow. Strong east and south-east winds and dry winds prevail.

The bulk of the water enters the reservoir due to its artificial supply from the Tsimlyansk reservoir by pumping stations. First, the Karpovskoye reservoir is filled with Tsimlyanskaya water, then water is pumped to Bereslavskoye, then to Varvarovskoye. During the operation of locks, water partially from the Varvarovsky reservoir falls into the river. The Volga, the other part (about 80-85% of its discharge) through the Bereslavskoe and Varvarovskoe reservoirs again returns to the Tsimlyansk reservoir. From spring to autumn, during the navigation period, the “winter” water of the reservoirs is replaced by Tsimlyanskaya water.

During the “bloom” of water in the Bereslavsky reservoir, the content of blue-green algae in the phytoplanktocenosis was 62-70%. Aphanizomenon, Anabaena and Microcystis accounted for up to 80%.

Algolization of the Bereslavsky reservoir was carried out in 2005 at certain points according to the proposed method.

Twice a month, water samples were taken at various points in the reservoir. The analysis of the total abundance and biomass of planktonic algae was carried out on a Synpor membrane filter No. 5 with a pore diameter of 0.6 μm by the deposition method. At the same time, phytoplankton material was collected to determine the species composition.

The presence of Chlorella vulgaris was determined monthly in water samples. To identify the strain Chlorella vulgaris IGF No. C-111, one water sample was grown on an selective nutrient medium (N - 64 mg / L; P - 8 mg / L; Fe - 0.1 mg / L; Co - 0.001 mg / L and Cu - 0.001 mg / l), the other without adding a nutrient medium. The experiment was carried out at the optimum temperature (30 ° C) and lighting (DRI-250 lamp) for 10-12 days. After the specified period, the test samples were microscopic to determine the species composition of phytoplankton.

As a result of studies, a decrease in the intensity of reproduction of blue-green algae was found. In August 2005, the share of the latter amounted to 30% in the composition of phytoplanktocenosis. Moreover, such representatives of blue-green algae as Gomphosphaeria lacustris Chod and Merismopedia tenuissima Lemm dominated. The share of Aphanizomenon, Anabaena and Microcystis in the composition of blue-green algae was 12.7%.

At the same time, in June and July, biomass was dominated by diatoms, up to 45%, and in August, green algae, up to 35%.

The participation of strain Chlorella vulgaris IGF No. C-111 in the numerical composition of green algae was maximized in August - 3.3% of the biomass.

Repeated algolization was carried out in 2006. In August 2006, the share of blue-green algae was 14% in the composition of phytoplankton. Representatives of Gomphosphaeria lacustris Chod dominated.

The proportion of toxic algae Aphanizomenon, Anabaena and Microcystis in the composition of blue-green algae was 6.1%.

At the same time, in June and July, diatoms of 45-46% dominated in biomass, and in August, green algae dominated to 37%.

The participation of strain Chlorella vulgaris IGF No. C-111 in the numerical composition of green algae was maximized in August - 3.9% of the biomass.

Claims (1)

A method of controlling the “bloom” of water by blue-green algae, which provides for the algolization of water bodies in winter - February with a suspension of green algae Chlorella vulgaris, characterized in that the algolization of water bodies is additionally carried out in the spring period - twice in the first phase of spring: March-April during freezing - after opening ice and when the upper layer (0-0.3 m) of water is heated to 10-12 ° C, in the second phase: when the average water temperature in the pond reaches a depth of 5-10 ° C, and in the summer - monthly, and in bays, beams and places of wind surges in summer the first period at least 2 times a month, as an algolysant use a suspension of strain Chlorella vulgaris IGF No. C-111 with a transmittance of 1.4-6 in the amount of 15-25 l per 1 million m ^{3 of} water, and in bays, beams and in places of wind surges at the rate of 15-25 l / ha, before algolization in the winter and spring periods, the algolizant is adapted by cooling it from the cultivation temperature to the temperature of the water in the reservoir at the time of algolization at the rate of 3-5 ° C per day and holding at that temperature for 1.5-2 days, and algolization is carried out for at least ie 3 years in the same way.