Scenedesmus euryales and application thereof in degradation of nitrogen and phosphorus in aquaculture wastewater

Abstract

The invention relates to the field of biological treatment of nitrogen and phosphorus in aquaculture wastewater, in particular to a Scenedesmus euryales strain and application thereof in degradation of nitrogen and phosphorus in aquaculture wastewater. The scenedesmus is preserved in the China General Microbiological Culture Collection Center (CGMCC), the preservation number is CGMCC No.46073, and the preservation date is August 12, 2024. The scenedesmus obtained through separation and screening can well adapt to the euryhaline wastewater after being subjected to salinity adaptability domestication, is stable in growth and has the application potential in the aspect of removing nitrogen and phosphorus in the mariculture wastewater.

Description

Technical Field

[0001] The invention relates to the field of biological treatment of nitrogen and phosphorus in aquaculture wastewater, and in particular to a strain of Scenedesmus euryhaline and application thereof in degradation of nitrogen and phosphorus in aquaculture wastewater. Background Art

[0002] Nitrogen and phosphorus in marine aquaculture wastewater have always been environmental problems, posing potential risks to aquatic ecosystems and human health. High-density intensive aquaculture brings about excess feed, drugs, animal carcasses and feces, resulting in large amounts of nitrogen and phosphorus. Studies have shown that nitrogen and phosphorus levels exceeding certain standards can cause toxic effects on aquatic organisms and cause eutrophication of water bodies, polluting the natural environment. Microalgae-based removal technology has the advantages of low carbon, low cost, eco-friendly and sustainable, and is considered to be a promising alternative to traditional wastewater treatment. In recent years, the application potential of microalgae in removing nitrogen and phosphorus from aquaculture wastewater has received widespread attention. Summary of the invention

[0003] The purpose of the present invention is to overcome the shortcomings and deficiencies of the prior art and provide a strain of Scenedesmus euryhaline and its application in the degradation of nitrogen and phosphorus in aquaculture wastewater.

[0004] The object of the present invention is achieved through the following technical scheme: a strain of Scenedesmus sp. and its application in the degradation of nitrogen and phosphorus in aquaculture wastewater, which is based on the fact that Scenedesmus sp. after salinity adaptation has the effect of efficiently removing nitrogen and phosphorus from wide-salt aquaculture wastewater. The Scenedesmus sp. is deposited in the General Microbiological Center of China Microbiological Culture Collection Administration, with a deposit number of CGMCC No.46073 and a deposit date of August 12, 2024.

[0005] Furthermore, the Scenedesmus sp. is a unicellular microalgae isolated from a wastewater sample from a certain aquaculture wastewater discharge outlet in Chaohe, Rizhao City, Shandong Province.

[0006] Furthermore, the salinity adaptive domestication method is characterized in that BG11 culture medium configured with all seawater in a ratio of seawater to freshwater is set at 1:25, 1:20, 1:10, 1:2, respectively, and the separated Scenedesmus is cultured in sequence until Scenedesmus adapted to BG11 culture medium configured with different seawater and freshwater is obtained.

[0007] Furthermore, the above-mentioned salinity-adaptive domesticated Scenedesmus algae solution is added to aquaculture wastewater with different gradient salinities for cultivation to obtain nitrogen and phosphorus-free water.

[0008] Furthermore, the adaptively domesticated Scenedesmus algae solution is a single-cell microalgae solution cultured in a salinity culture medium to a logarithmic growth phase.

[0009] Compared with the prior art, the present invention has the following advantages and effects:

[0010] The present invention discovers a strain of Scenedesmus sp. and its application in the degradation of nitrogen and phosphorus in aquaculture wastewater. The Scenedesmus sp. separated and screened by the present invention comes from aquaculture wastewater, can adapt well to the eurysalinity wastewater after salinity adaptability acclimation, has stable growth, and has application potential in removing nitrogen and phosphorus from marine aquaculture wastewater. BRIEF DESCRIPTION OF THE DRAWINGS

[0011] Figure 1 This is a cell morphology diagram of Scenedesmus sp. in BG11 medium with different salinities after acclimation at different salinities.

[0012] In the figure: A. Optical microscope image of Scenedesmus sp. in 0‰ solid culture medium, B. Cell morphology of algae liquid of Scenedesmus sp. in 0‰ salinity BG11 culture medium after acclimation at a salinity ratio of 1:25 between seawater and fresh water, C. Cell morphology of algae liquid of Scenedesmus sp. in 5‰ salinity BG11 culture medium after acclimation at a salinity ratio of 1:20 between seawater and fresh water, D. Cell morphology of algae liquid of Scenedesmus sp. in 10‰ salinity BG11 culture medium after acclimation at different salinities of 1:10 between seawater and fresh water, E. Cell morphology of algae liquid of Scenedesmus sp. in 15‰ salinity BG11 culture medium after acclimation at a salinity ratio of 1:2 between seawater and fresh water, F. Cell morphology of algae liquid of Scenedesmus sp. in 20‰ salinity BG11 culture medium after acclimation at all seawater salinities;

[0013] Figure 2 This is the growth curve of Scenedesmus sp. after acclimation in different salinities in BG11 medium;

[0014] Figure 3 The chlorophyll a content of Scenedesmus sp. in simulated seawater aquaculture wastewater culture medium with different salinities after acclimation with different salinities;

[0015] Figure 4 The chlorophyll b content of Scenedesmus sp. in simulated marine aquaculture wastewater culture medium with different salinities after acclimation with different salinities;

[0016] Figure 5 The carotene content of Scenedesmus sp. in simulated seawater aquaculture wastewater culture medium with different salinities after acclimation with different salinities;

[0017] Figure 6 It is the protein content of Scenedesmus sp. in simulated seawater aquaculture wastewater culture medium with different salinities after acclimation with different salinities;

[0018] Figure 7 It is the soluble polysaccharide content of Scenedesmus sp. in the simulated seawater aquaculture wastewater culture medium with different salinities after acclimation with different salinities;

[0019] Figure 8 It is the total lipid content of Scenedesmus sp. after acclimation at different salinities in simulated marine aquaculture wastewater culture medium at different salinities;

[0020] Fig. 9 The effect of Scenedesmus sp. in removing ammonia nitrogen from simulated marine aquaculture wastewater with different salinities after being acclimated to different salinities;

[0021] Fig.10 The effect of Scenedesmus sp. in removing nitrite nitrogen in simulated seawater aquaculture wastewater with different salinities after being acclimated to different salinities;

[0022] Fig.11 The effect of Scenedesmus sp. in removing nitrate nitrogen from simulated marine aquaculture wastewater with different salinities after being acclimated to different salinities;

[0023] Fig.12 The effect of Scenedesmus sp. in removing active phosphate in simulated seawater aquaculture wastewater with different salinities after being acclimated to different salinities. DETAILED DESCRIPTION

[0024] The present invention is further described in detail below in conjunction with embodiments and drawings, but the experimental methods of the present invention are not limited thereto.

[0025] Example 1: Isolation and screening of microalgae with strong nitrogen and phosphorus removal capabilities and salinity adaptation

[0026] The BG11 medium used in this example is the literature "Touloupakis, E., Zittelli, GC, Benavides, AMS, & Torzillo, G. (2023). Growth and photosynthetic performance of Nostoc linckia (formerly N. calcicola) cells grown in BG11 and BG110 media. Photochemical &

[0027] Photobiological Sciences, 22(4), 795-807." The disclosed culture medium.

[0028] Water samples with excessive nitrogen and phosphorus contents were taken from the sewage outlet of Chaohe Aquaculture Area in Rizhao City, Shandong Province for microalgae enrichment pre-culture, and the wastewater samples were added to BG11 culture medium for separation and screening of unicellular microalgae, among which Scenedesmus sp. S1 was isolated by itself, and the Scenedesmus was deposited in the General Microbiology Center of China National Microbiological Culture Collection Committee with a deposit number of CGMCC No.46073, a deposit date of August 12, 2024, and a deposit address of No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, and its nucleotide sequence is shown in SEQ ID NO.1.

[0029]

[0030] Prepare BG11 liquid culture medium with different ratios of seawater to freshwater, setting the ratios to 1:25, 1:20, 1:10, and 1:2 with all seawater;

[0031] The purified Scenedesmus spp. was cultured and the number of cells was 1×10 5 The cells / mL were inoculated into a 500mL conical flask containing 250mL of BG11 liquid medium with a ratio of seawater to fresh water of 1:25, and cultured in a light incubator under the conditions of 8000lux illumination, 12h:12h light-dark cycle, 25℃, and manual shaking three times at 8:00, 14:00, and 20:00 every day.

[0032] The purified Scenedesmus spp. was cultured and the number of cells was 1×10 5 The cells / mL were inoculated into a 500mL conical flask containing 250mL of BG11 liquid medium with a ratio of seawater to fresh water of 1:20, and cultured in a light incubator under the conditions of 8000lux illumination, 12h:12h light-dark cycle, 25℃, and manual shaking three times at 8:00, 14:00, and 20:00 every day.

[0033] The purified Scenedesmus spp. was cultured and the number of cells was 1×10 5 1000 mL / mL was inoculated into a 500 mL conical flask containing 250 mL of BG11 liquid medium with a ratio of seawater to fresh water of 1:10, and cultured in a light incubator under the conditions of 8000 lux illumination, 12h:12h light-dark cycle, 25°C, and manual shaking three times at 8:00, 14:00, and 20:00 every day.

[0034] The purified Scenedesmus spp. was cultured and the number of cells was 1×10 5 The cells / mL were inoculated into a 500mL conical flask containing 250mL of BG11 liquid medium with a ratio of seawater to fresh water of 1:2, and cultured in a light incubator under the conditions of 8000lux illumination, 12h:12h light-dark cycle, 25℃, and manual shaking three times at 8:00, 14:00, and 20:00 every day.

[0035] The purified Scenedesmus spp. was cultured and the number of cells was 1×10 5 The cells were inoculated into a 500 mL conical flask of BG11 liquid culture medium containing 250 mL of seawater and cultured in a light incubator under the conditions of 8000 lux illumination, 12 h:12 h light-dark cycle, 25 °C, and manual shaking three times at 8:00, 14:00, and 20:00 a.m. each day.

[0036] Finally, Scenedesmus sp. was obtained which could adapt to different seawater and freshwater configurations of BG11 culture medium.

[0037] The cell morphology of Scenedesmus sp. after adaptation to different salinities (ratio of seawater to freshwater: 1:25, 1:20, 1:10, 1:2, all seawater) was inoculated into BG11 medium with different salinities (0‰, 5‰, 10‰, 15‰, 20‰). Figure 1 As shown, the morphology is stable.

[0038] The growth curves of Scenedesmus sp. algae solutions that were adapted to different salinities (ratios of seawater to freshwater were 1:25, 1:20, 1:10, 1:2, and all seawater) were inoculated into BG11 medium with different salinities (0‰, 5‰, 10‰, 15‰, and 20‰). Figure 2 As shown, the growth curve of Scenedesmus sp. algae solution at each salinity presents an S-shape, indicating that the salinity adaptation effect has been achieved.

[0039] Example 2: Nitrogen and phosphorus removal and growth effect of microalgae in simulated aquaculture wastewater with different salinities

[0040] The adaptability of salinity-acclimated Scenedesmus sp. to simulated aquaculture wastewater with different salinities and its nitrogen and phosphorus degradation effect.

[0041] There were 5 experimental groups, divided into five salinity simulated aquaculture wastewater groups: 0‰, 5‰, 10‰, 15‰, and 20‰, with 3 replicates in each group. 200 mL of simulated aquaculture wastewater was added to a 250 mL conical flask, and the initial algae density was 1×10 5 The Scenedesmus sp. acclimated with the corresponding salinity in Example 1 was 10 cells / mL. All the conical flasks of the treatment groups were continuously cultured in a light incubator with an illumination of 8000 lux, a light-dark cycle of 12h:12h, and 25°C. They were manually shaken three times at 8:00, 14:00, and 20:00 every day, and their positions were exchanged to make the algae cells evenly suspended. When the culture reached the exponential growth phase on the fourth day, samples were taken to determine the photosynthetic pigments and nutrients.

[0042] The results are as follows Figure 3-8 As shown, the results showed that Scenedesmus sp. has good adaptability to euryhaline environments.

[0043] The culture was continued for 8 days under the above conditions, and samples were taken at the same time every day. 15 mL of algae liquid was centrifuged at 4000 rpm for 10 min, and the supernatant was filtered through a 0.45 μm filter membrane for the determination of nitrite nitrogen, ammonia nitrogen, nitrate nitrogen, and phosphate.

[0044] The results are as follows Figure 9-12As shown, the results showed that Scenedesmus sp. has a strong ability to remove nitrogen and phosphorus from simulated aquaculture wastewater.

Claims

1. Application of a strain of Scenedesmus euryhaline in the degradation of nitrogen and phosphorus in aquaculture wastewater, characterized by: The described Scenedesmus is deposited in the General Microbiology Center of China Microorganism Culture Collection Administration, with the deposit number CGMCC No.46073 and the deposit date August 12, 2024.

2. The use of a strain of Scenedesmus euryhaline in the degradation of nitrogen and phosphorus in aquaculture wastewater according to claim 1, characterized in that: After being domesticated for salinity adaptation, the Scenedesmus has the function of removing nitrogen and phosphorus from euryhaline aquaculture wastewater.

3. The use of a strain of Scenedesmus euryhaline in the degradation of nitrogen and phosphorus in aquaculture wastewater according to claim 1, characterized in that: The Scenedesmus is a unicellular microalgae.

4. The use of a strain of Scenedesmus euryhaline in the degradation of nitrogen and phosphorus in aquaculture wastewater according to claim 2, characterized in that: The salinity adaptive domestication method is to set a BG11 culture medium with a ratio of seawater to fresh water of 1:25, 1:20, 1:10, 1:2 and all seawater, and culture the separated Scenedesmus in sequence until a Scenedesmus adapted to BG11 culture medium with different seawater and freshwater configurations is obtained.

5. The use of a strain of Scenedesmus euryhaline in the degradation of nitrogen and phosphorus in aquaculture wastewater according to claim 4, characterized in that: The algae liquid of the salinity-adaptive domesticated Scenedesmus is added to aquaculture wastewater with different gradient salinities for cultivation to obtain water bodies free of nitrogen and phosphorus.

6. The use of a strain of Scenedesmus euryhaline in the degradation of nitrogen and phosphorus in aquaculture wastewater according to claim 5, characterized in that: The adaptive domesticated Scenedesmus algae liquid is a single-cell microalgae liquid cultured in a salinity culture medium to a logarithmic growth phase.

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