Polysaccharide-producing talsalonema and application thereof in treatment of high-nitrogen and high-phosphorus wastewater

Abstract

The invention discloses a strain of polysaccharide-producing Trichothrix sp. And application of the polysaccharide-producing Trichothrix sp. ZSL-NS5 in high nitrogen and phosphorus wastewater treatment. The strain named as the Trichothrix sp. ZSL-NS5 is preserved in the China Center for Type Culture Collection on November 7, 2024, and the preservation number is CCTCC NO.M 20242470. The invention further discloses a preparation method of the polysaccharide-producing Trichothrix sp. ZSL-NS5. The application of the polysaccharide-producing Trichothrix sp. ZSL-NS5 in the high nitrogen and phosphorus wastewater treatment of the polysaccharide-producing Trichothrix sp. The algal strain is separated from wastewater of an aerobic pond of a dairy farm, has relatively strong wastewater tolerance and pollutant purification capacity, and has a total phosphorus removal rate of more than 95%, an ammonia nitrogen removal rate of nearly 100%, a TN removal rate of more than 85% and a COD removal rate of more than 70%. Besides, the talsalonema has the capability of producing microalgae EPS and polysaccharide, the polysaccharide content of the talsalonema reaches 63%, the yield of the talsalonema is obviously higher than that of common microalgae, and the talsalonema has the application value of preparing chemical raw materials and drugs.

Description

Technical Field

[0001] The invention belongs to the field of aquaculture wastewater treatment technology and biotechnology, and specifically relates to a polysaccharide-producing Talsa filamentosa and application thereof in dairy farm wastewater treatment. Background Art

[0002] Microalgae are tiny microorganisms whose cell morphology can only be seen under a microscope. Microalgae can use sunlight to convert carbon dioxide and water in water into oxygen and organic matter. This process consumes inorganic carbon sources in the water and helps reduce the eutrophication of water bodies. Microalgae can efficiently absorb nitrogen (such as ammonia nitrogen, nitrate) and phosphorus (such as phosphate) in wastewater and convert them into nutrients required for their own growth, thereby reducing the nutrient load in the water body. In addition, microalgae remove organic pollutants such as surfactants and oils from wastewater through adsorption on the cell surface and internal metabolic processes. Some microalgae have the ability to accumulate heavy metals. Through biosorption and bioaccumulation, heavy metal ions such as cadmium, chromium, mercury and lead can be effectively removed from water. Microalgae have a fast growth rate, which enables them to respond quickly in sewage treatment and effectively remove pollutants. Microalgae biomass is rich in protein, polysaccharides and other nutrients, and can be used as raw materials for high-value products such as biofuels, medicines, fertilizers, etc., to achieve resource conversion in wastewater. In addition, microalgae have efficient carbon fixation ability, which helps to mitigate greenhouse gas emissions. In summary, microalgae technology is widely used in various wastewater treatment fields due to its advantages such as "pollution reduction and carbon reduction, resource transformation, and turning waste into treasure".

[0003] Dairy farm manure is characterized by large wastewater output, complex composition, and difficulty in treatment. The water quality of dairy farm wastewater varies greatly, and the ammonia nitrogen content is high, resulting in an imbalance in the carbon-nitrogen ratio, which makes wastewater treatment complex and poses a challenge to wastewater treatment. At present, the treatment technology for dairy farm wastewater is mainly based on biological combination processes, but there are still high concentrations of organic matter and high concentrations of ammonia nitrogen in the effluent. In addition, the existing process usually produces a large amount of residual sludge, which still needs further treatment. However, microalgae biological treatment technology does not require the addition of additional carbon sources, thereby reducing the investment cost of the wastewater treatment process; and does not produce residual sludge and its secondary treatment problems; microalgae biomass can be used as chemical raw materials to generate economic value. Therefore, microalgae biotechnology has gradually been favored by scholars in the field of livestock and poultry wastewater.

[0004] At present, there are more and more studies on microalgae treatment of dairy wastewater, but there are fewer studies on screening native microalgae from wastewater habitats and applying the microalgae to wastewater treatment and biomass recovery. Traditional commercially available microalgae are difficult to adapt to the high concentration of organic matter, high ammonia nitrogen and toxic and harmful components in dairy wastewater. Therefore, it is very important to find new microalgae varieties that can adapt to the characteristics of dairy wastewater. On the other hand, the subsequent utilization value of microalgae biomass is very important. The polysaccharides in microalgae have many uses. They can be used as gel materials in soft materials, sensors and other fields; in the medical field, they are used to prepare drug carriers such as injections and tablets; they can also prepare polymer scaffold materials and composite materials for repairing and replacing human tissues. However, how to cultivate microalgae with high polysaccharide yield is still a problem that needs to be solved. Summary of the invention

[0005] In view of the problems that dairy farm wastewater has complex composition, low pollutant removal efficiency, and difficulty in returning to the field, the present invention provides a self-isolated Desertifilum sp. ZSL-NS5 for treating dairy farm wastewater and producing polysaccharides, providing technical support for aquaculture wastewater treatment.

[0006] The present invention solves the technical problem by adopting the following technical solutions:

[0007] A polysaccharide-producing strain of Tarsha filamentous algae, named Tarsha filamentous algae ZSL-NS5, classified and named as Desertifilum sp, deposited on November 7, 2024, deposited in China Center for Type Culture Collection, with a deposit number of CCTCC M 20242470, and deposited at Wuhan University, Wuhan, China.

[0008] Moreover, the formula of the culture medium of Tarsha filamentous algae ZSL-NS5 is as follows: 1.5 g sodium nitrate, 0.04 g dipotassium hydrogen phosphate, 0.075 g magnesium sulfate heptahydrate, 0.036 g calcium chloride dihydrate, 0.006 g ammonium ferric citrate, 0.006 g citric acid, 0.001 g disodium ethylenediaminetetraacetic acid, 0.02 g sodium carbonate, 1 mL of trace element mother solution, add water to make up to 1000 mL, adjust the pH to about 7.1, and sterilize at 121°C under high pressure for 20 min.

[0009] Moreover, the formula of the trace element mother solution is: 2.86g / L boric acid, 1.86g / L manganese chloride tetrahydrate, 0.22g / L zinc sulfate heptahydrate, 0.08g / L copper sulfate pentahydrate, 0.39g / L sodium molybdate dihydrate; 0.05g / L cobalt nitrate hexahydrate, add water to make up to 1000mL.

[0010] Application of polysaccharide-producing Tharsa filamentosa ZSL-NS5 in dairy wastewater.

[0011] A method for treating dairy farm wastewater using polysaccharide-producing Tarsa filamentosa. The wastewater needs to be treated through four steps of coarse filtration, centrifugation, refiltration, and dilution. The inoculation ratio of ZSL-NS5 algae liquid to wastewater is 1:5 by volume. The algae liquid is centrifuged at 7000-8000r / min for 10 minutes. After centrifugation, the upper culture medium is discarded, and Tarsa filamentosa algae mud is added to the dairy farm wastewater and shaken or stirred evenly.

[0012] Moreover, the treatment conditions are: light intensity 4000-5000Lux, light-dark ratio 12:12 hours, temperature 25°C, the treatment cycle can be 8-12 days, and air circulation or shaking is used to prevent microalgae from settling or sticking to the wall.

[0013] Moreover, in the treatment of dairy farm wastewater by Tarsa filamentous algae ZSL-NS5, the wastewater can be sterilized or not, the dilution multiples of anaerobic tank effluent and oxidation pond wastewater are 1-5 times, and the dilution multiple of stabilization pond wastewater is 1-2 times.

[0014] Application of polysaccharide-producing Tarsa filamentosa ZSL-NS5 as biofertilizer.

[0015] The advantages and positive effects of the present invention are:

[0016] 1. The present invention screens out a strain of Talsa filamentosa with strong pollution resistance and purification ability from the wastewater of aerobic ponds in dairy farms. Its growth performance in wastewater is better than that in culture medium, its growth rate in wastewater is faster, and it can adapt to high-concentration dairy farm wastewater.

[0017] 2. The Talsa filamentous algae in the present invention has a strong ability to purify wastewater. Due to its rich extracellular polymers and its own filamentous characteristics, the filamentous algae has obvious effects on removing chromaticity, COD, ammonia nitrogen, TN and TP in wastewater, which is better than traditional pure cultured microalgae varieties.

[0018] 3. The Tharsa microalgae provided by the present invention has a carbohydrate content of more than 60%, which is higher than that of conventional microalgae, and can provide high-quality carbohydrate raw materials. The microalgae can also produce other products, such as proteins, oils, etc., and can achieve the purpose of biomass recovery while treating wastewater.

[0019] 4. The present invention provides a wastewater treatment method that does not require additional material and energy consumption and can significantly reduce the economic cost of dairy farm wastewater treatment. Compared with other methods of treating dairy farm wastewater, the treatment method of the present invention is more efficient, has more stable performance, is economically beneficial, and meets the requirements of clean production. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 Microscopic photograph of microalgae cell morphology.

[0021] Figure 2 Maximum Parsimony tree constructed based on 16S rDNA fragment sequences (microalgae of the present invention marked with a red frame).

[0022] Figure 3 Growth indicators of Talsa filamentosa (a: dry weight; b: chlorophyll; c: component content; d: component ratio).

[0023] Figure 4 Purification effect of Talsa filamentosa on dairy wastewater with different concentrations. DETAILED DESCRIPTION

[0024] The present invention is further described in detail below through specific examples. The following examples are only illustrative and not restrictive, and the protection scope of the present invention cannot be limited thereto.

[0025] The experimental methods used in the following examples are all conventional methods unless otherwise specified, and the reagents, methods and equipment used are all conventional reagents, methods and equipment in the technical field unless otherwise specified.

[0026] The dairy farm wastewater of the present invention is taken from the anaerobic fermentation effluent of a dairy farm in Tianjin. The Tharsha silk algae of the present invention is separated from the aerobic pond wastewater of the dairy farm. A polysaccharide-producing Tharsha silk algae isolated from the dairy farm wastewater has better tolerance and treatment effect on wastewater, and has been preserved (preservation number is CCTCC NO.M20242470), and has very good application prospects.

[0027] The Talsa filamentosa of the present invention is separated from the above-mentioned dairy farm aerobic pond wastewater.

[0028] Example 1

[0029] Isolation and identification of Talsa filamentosa

[0030] 1. Isolation of Talsa algae

[0031] Take the aerobic pond wastewater from the dairy farm (COD is 4562.3 mg / L, total nitrogen is 560.8 mg / L, ammonia nitrogen is 507.5 mg / L, total phosphorus is 64.1 mg / L, salinity is 1.9‰, pH is 7.7), spread it on the plate after gradient dilution, and after single algae grow on the plate, pick up the single algae and put it on the growth medium.

[0032] The culture was carried out for 15 days at a temperature of 25°C and a light ratio of 6000 Lux, 3000 Lux and darkness of 8:8:8 hours to obtain a Talsa silk algae liquid. After obtaining a single algae liquid, it was identified as Talsa silk algae ZSL-NS5. Then, the culture was expanded until there was enough algae liquid (volume ratio during expansion: algae liquid: culture medium = 1:5).

[0033] The formula of growth medium (BG11): sodium nitrate 1.5g, potassium dihydrogen phosphate 0.04g, magnesium sulfate heptahydrate 0.075g, calcium chloride dihydrate 0.036g, ammonium ferric citrate 0.006g, citric acid 0.006g, disodium ethylenediaminetetraacetic acid 0.001g, sodium carbonate 0.02g, trace element mother solution 1mL, add water to make up to 1000mL, adjust the pH to about 7.1, and sterilize at 121℃ for 20min.

[0034] The formula of trace element mother solution: boric acid 2.86g / L, manganese chloride tetrahydrate 1.86g / L, zinc sulfate heptahydrate 0.22g / L, copper sulfate pentahydrate 0.08g / L, sodium molybdate dihydrate 0.39g / L; cobalt nitrate hexahydrate 0.05g / L, add water to make up to 1000mL.

[0035] 2. Morphological identification of microalgae

[0036] A small amount of sample was observed under an Olympus BX53 microscope and the image was acquired using an Olympus DP80 ( Figure 1 ). The filaments are solitary or intertwined, often curved, unbranched, thinly sheathed, blue-green. The transverse wall is not constricted or slightly constricted, the filaments are pointed at the end, without cap-like structures, heterocysts and thick-walled spores. The cells are columnar, 3.0-8.0μm long and 2.2-4.0μm wide.

[0037] The Tarsha filamentous algae ZSL-NS5 obtained by the above screening is classified and named: Desertifilum sp., the preservation date is: November 7, 2024, the preservation unit is: China Center for Type Culture Collection, and the preservation number is CCTCCNO.M20242470.

[0038] Example 2

[0039] Growth performance of Talsa algae in dairy wastewater

[0040] (1) The isolation and expansion of the Talsa filamentous algae were the same as in Example 1.

[0041] (2) Wastewater needs to be filtered through gauze to remove large impurities, centrifuged at 4000rpm for 10min, and then filtered through ordinary filter paper, and diluted according to different gradients. Take the algae liquid of Tarsha filamentous algae ZSL-NS5 in the logarithmic growth period. The volume ratio of algae liquid to wastewater is 1:5. Add 50mL of algae liquid to 250mL of wastewater, corresponding to a dry weight of 0.2g / L. The algae liquid is centrifuged at 7000-8000r / min for 10 minutes. After centrifugation, the upper culture medium is removed, and Tarsha filamentous algae balls are added to the wastewater, shaking or stirring evenly to avoid Tarsha filamentous algae aggregation or precipitation. The wastewater dilution is 25%, 50% and 75% V / V. Set the light intensity to 4000-5000Lux, the light-dark ratio to 12:12 hours, and the temperature to 25℃. Treat for a total of 12 days. Use air circulation or shaking to prevent microalgae from settling or sticking to the wall. Sampling was performed to measure dry weight, chlorophyll, biomass yield and composition, etc.

[0042] On the 12th day, the dry weight and chlorophyll production of Tarsha filamentous algae were 2.307 g / L and 5.46 mg / L, and the protein, polysaccharide and oil production were 99.1 μg / mL, 176.2 μg / mL and 25.6 μg / mL, respectively.

[0043] Example 3

[0044] Effect of Talsa algae on the treatment of dairy wastewater

[0045] 1. The separation and expansion of the Talsa algae were the same as in Example 1.

[0046] 2. Treatment of dairy farm wastewater: The treatment steps were similar to those in Example 2. After 12 days of treatment, samples were taken to determine the chemical oxygen demand, total nitrogen, total phosphorus and ammonia nitrogen content.

[0047] Its total phosphorus removal rate can reach 99.9%; the ammonia nitrogen removal rate is up to 95.6%; the COD removal rate is as high as 75.8% and the TN removal rate is as high as 85.6% ( Figure 4 ).

[0048] Table 1 Variation of various parameters of dairy farm wastewater

[0049]

[0050]

[0051] Example 4

[0052] Experimental study on the effect of returning the algae liquid of Tarsha filamentous algae ZSL-NS5 to the field

[0053] The algae liquid after harvest can be used as biofertilizer and returned to the field. In order to verify the promoting effect of Tarsha silk algae on crops, four groups of fertilization experiments were set up: CK was a control group without fertilizer, T1 was a group with chemical fertilizer (25 kg / mu), and T2 was a group with Tarsha silk algae fertilizer (40 L (algae concentration 1.35×10 7 T3 is a group of chemical fertilizer + Talsa silk algae fertilizer (25kg chemical fertilizer / mu + 40L organic fertilizer / mu). The crop is rapeseed (Brassica chinensis L.).

[0054] The experimental results show that compared with CK, algae fertilizer can significantly promote the increase of nutrient alkaline nitrogen, but reduce the total nitrogen content of the soil; however, the total nitrogen content of rapeseed increases, which shows that algae fertilizer can promote the absorption and utilization of nutrients by crops. The soil nutrients and rapeseed basic nutrients of the group applying organic fertilizer + algae fertilizer have increased. For example, compared with CK and the group applying chemical fertilizer T1, the soil alkaline nitrogen of the T3 group increased by 20.5% and 6.9%, respectively.

[0055] Table 1 Effects of algae fertilizer on soil nutrients and rapeseed basic nutrients

[0056]

[0057] From the above examples, it can be seen that the provided strain of Talsa algae with excellent growth can show excellent pollution tolerance and pollutant purification ability in dairy wastewater of different dilutions. In addition, the Talsa algae can produce higher levels of sugar substances and extracellular polymers while purifying pollutants. The present invention provides an excellent algae strain for the production of microalgae sugars and the microalgae in the process of aquaculture wastewater treatment.

[0058] The above description is only a preferred embodiment of the present invention and does not limit the present invention in any form. Any technician familiar with the profession, without departing from the scope of the technical solution of the present invention, according to the technical essence of the present invention, any simple modification, equivalent replacement and improvement made to the above embodiment still falls within the protection scope of the technical solution of the present invention.

[0059] Although the embodiments of the present invention are disclosed for illustrative purposes, those skilled in the art will appreciate that various substitutions, changes and modifications are possible without departing from the spirit and scope of the present invention and the appended claims. Therefore, the scope of the present invention is not limited to the contents disclosed in the embodiments.

Claims

1. A polysaccharide-producing strain of Talsa algae, characterized by: The name is Tarsha filamentous algae ZSL-NS5, the classification name is Desertifilum sp, the preservation date is November 7, 2024, the preservation unit is China Center for Type Culture Collection, the preservation number is CCTCC NO.M20242470, and the preservation address is Wuhan University, Wuhan, China.

2. The polysaccharide-producing Talsa algae according to claim 1, characterized in that: The formula of the culture medium of Tarsha filamentous algae ZSL-NS5: 1.5g sodium nitrate, 0.04g dipotassium hydrogen phosphate, 0.075g magnesium sulfate heptahydrate, 0.036g calcium chloride dihydrate, 0.006g ammonium ferric citrate, 0.006g citric acid, 0.001g disodium ethylenediaminetetraacetic acid, 0.02g sodium carbonate, 1mL trace element mother solution, add water to make up to 1000mL, and adjust the pH to 7-8.

3. The polysaccharide-producing Talsa algae according to claim 2, characterized in that: The formula of trace element mother solution: boric acid 2.86g / L, manganese chloride tetrahydrate 1.86g / L, zinc sulfate heptahydrate 0.22g / L, copper sulfate pentahydrate 0.08g / L, sodium molybdate dihydrate 0.39g / L; cobalt nitrate hexahydrate 0.05g / L, add water to make up to 1000mL.

4. Use of the polysaccharide-producing Tarsa filamentosa ZSL-NS5 according to claim 1 in dairy farm wastewater.

5. A method for treating dairy farm wastewater using the polysaccharide-producing Talsa algae according to claim 1, characterized in that: Wastewater needs to be treated through four steps: coarse filtration, centrifugation, refiltration, and dilution. The inoculation ratio of ZSL-NS5 algae liquid and wastewater is 1:5 by volume. The algae liquid is centrifuged at 7000-8000r / min for 10 minutes. After centrifugation, the upper culture medium is discarded and the Talsha filamentous algae mud is added to the dairy farm wastewater and shaken or stirred evenly.

6. The method for treating dairy farm wastewater according to claim 5, characterized in that: The treatment conditions are: light intensity 4000-5000Lux, light-dark ratio 12:12 hours, temperature 25°C, treatment cycle can be 8-12 days, air circulation or shaking is used to prevent microalgae from settling or sticking to the wall.

7. The method for treating dairy farm wastewater according to claim 5, characterized in that: In the treatment of dairy farm wastewater by Tarsa filamentous algae ZSL-NS5, the wastewater can be sterilized or not, the dilution multiple of anaerobic tank effluent and oxidation pond wastewater is 1-5 times, and the dilution multiple of stabilization pond wastewater is 1-2 times.

8. Use of the polysaccharide-producing Tharsa filamentous algae according to claim 1 as a biofertilizer.

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