Microalgae fertilizer capable of improving salt and alkali resistance of cucumbers

Abstract

The invention provides a microalgae fertilizer capable of improving salt and alkali resistance of cucumbers, and the microalgae fertilizer is a composite microalgae bio-fertilizer and comprises scenedesmus obliquus, nitzschia grain and extracellular polymeric substances generated in the culture process of the nitzschia grain. The components of the microalgae compound fertilizer disclosed by the invention are all freshwater algae strains and natural active microalgae cells, so that sodium ions and metal cations cannot be accumulated, the microalgae compound fertilizer does not contain blue-green algae, and cytotoxin which is harmful to the environment cannot be generated; the used algae strains are easier to breed, and the biomass accumulation speed of the algae strains is far higher than that of other algae strains, so that the prepared microalgae biological fertilizer has the advantages of saving time and cost. And the polyunsaturated fatty acid generated by the microalgae fertilizer can enhance the plant resistance through an osmotic regulation system and an antioxidant enzyme ROS system.

Description

Technical Field

[0001] The present invention belongs to the technical field of plant growth regulators, and particularly relates to a microalgae fertilizer with improved saline-alkali resistance of cucumbers. Background Art

[0002] As an important factor in agricultural production, fertilizers play a crucial role in increasing crop yields and ensuring food security. Biological fertilizers are a new type of fertilizer that is efficient, pollution-free, and harmless. Long-term use helps to establish a virtuous cycle of the soil, reduce the use of chemical fertilizers, restore soil fertility, promote plant growth, and achieve the coordinated development of crop yield increase and environmental protection.

[0003] The organisms used to prepare biological fertilizers mainly include bacteria, fungi, microalgae, etc. Microalgae have become a research hotspot in biological fertilizers in recent years due to their high carbon fixation ability and good effect on promoting crop growth. Among them, microalgae fertilizer technology is even regarded as one of the directions encouraged for development in the future in China.

[0004] Through photosynthesis, microalgae can not only convert and accumulate inorganic nutrient elements in the environment into organic nutrients, but also secrete substances such as organic acids and extracellular polysaccharides into the environment, changing the environmental pH, promoting the redistribution of environmental nutrient elements. Some nitrogen-fixing cyanobacteria can also fix nitrogen (N 2 ) in the atmosphere using nitrogenase, convert it into organic nitrogen available to plants, and increase the soil organic matter content to improve soil fertility and increase crop yields. Currently, freshwater microalgae widely used in fertilizer production are mainly cyanobacteria and green algae.

[0005] Seaweed fertilizers are widely used because their source biomass is huge and can be obtained by fishing and harvesting. The types of seaweed fertilizers include brown algae, red algae, etc., which can provide a large amount of nutrient elements for crops. However, since the seaweed growth environment contains more salt ions, and the current production process of seaweed fertilizers usually directly composts seaweed or applies it after cell disruption by physical methods. Some studies have confirmed that after applying seaweed fertilizers, the content of cations such as sodium in the soil increases sharply. Further research found that both chemical fertilizers and seaweed fertilizers applied by traditional and intensive farming methods will increase the salt content in the soil. Therefore, in the process of planting salt-sensitive plants and in areas with more salt accumulation, when choosing algae fertilizers, it is necessary to carefully select seaweed-derived fertilizers according to local conditions.

[0006] Although cyanobacteria have been proven to have good biological nitrogen fixation effects in many crops, can improve crop yields and nutrient utilization rates, and have good application effects in crop yield increase and chemical fertilizer substitution, cyanobacteria are prokaryotes and have metabolic products different from eukaryotes. Research has found that most cyanobacteria can produce cyanotoxins, and a non-protein amino acid with neurotoxicity, P-methyl amino-L-alanine (BMAA), is widely distributed. Among the planktonic cyanobacteria discovered globally so far, more than 90% of the species have the ability to synthesize BMAA. It is generally believed that under suitable growth conditions such as temperature and nutrients, almost all cyanobacterial strains can synthesize the neurotoxin BMAA by themselves. Therefore, the direct application of cyanobacterial products, especially live cyanobacterial cells, in the environment poses potential risks to animals, the environment, and humans. Summary of the Invention

[0007] The object of the present invention is to provide a microalgae fertilizer with improved saline-alkali resistance for cucumbers, which can improve the saline-alkali resistance of cucumbers and enhance the quality of cucumbers cultivated under saline-alkali conditions.

[0008] The microalgae fertilizer provided by the present invention contains Nitzschia palea (Np) and Scenedesmus obliquus (Sp).

[0009] Furthermore, the microalgae fertilizer also contains the extracellular polymer of Nitzschia palea.

[0010] The extracellular polymer of Nitzschia palea is obtained by culturing Nitzschia palea, removing the algal cells, and collecting the filtrate.

[0011] The culturing is carried out using WC medium. After the biomass of the algal cells grows to 3.5 g·L -1 , the algal cells are removed by centrifugation.

[0012] Furthermore, in the microalgae fertilizer, the content of Nitzschia palea is 73.24 mg·L -1 , and the content of Scenedesmus obliquus is 401.27 mg·L -1 ;

[0013] The concentration of the extracellular polymer of Nitzschia palea is 61.06%.

[0014] Another aspect of the present invention also provides a use of the microalgae fertilizer, which is an application in improving the germination rate and growth under improved saline-alkali resistance of cucumbers.

[0015] The microalgae fertilizer provided by the present invention does not contain cyanobacteria and will not produce cytotoxins to harm the environment; moreover, the components of the microalgae compound fertilizer of the present invention are all freshwater algal strains and will not accumulate sodium ions and metal cations. In terms of the use effect, the microalgae fertilizer of the present invention has an enrichment effect on the soil microorganisms of crop roots. And the algal strains used in the present invention are easier to reproduce, and far exceed other algal strains in terms of the biomass accumulation rate, so the preparation of microalgae bio-fertilizer has the advantage of saving time cost. Moreover, the polyunsaturated fatty acids produced by the microalgae fertilizer of the present invention can enhance plant resistance through the osmotic adjustment system and the antioxidant enzyme ROS system. Brief Description of the Drawings

[0016] Figure 1 : Diagram showing the effect of different concentrations of Scenedesmus obliquus on the chlorophyll content of cucumber seedlings under saline-alkali stress, where Figure a is the chlorophyll a content diagram, Figure b is the chlorophyll b content diagram; Figure c is the carotenoid content diagram;

[0017] Figure 2 : Diagram showing the effect of different concentrations of Scenedesmus obliquus on the content of osmotic adjustment substances in cucumber seedlings under saline-alkali stress; where Figure a is the malondialdehyde content diagram; Figure b is the electrolyte permeability diagram; Figure c is the soluble sugar content diagram; Figure d is the soluble protein content diagram;

[0018] Figure 3 : Diagram showing the effect of different concentrations of Scenedesmus obliquus on the antioxidant enzyme activities of cucumber seedlings under saline-alkali stress; where Figure a is the peroxidase (POD) activity diagram; Figure b is the superoxide dismutase (SOD) activity diagram; Figure c is the catalase (CAT) activity diagram;

[0019] Figures 1 - 3 CK in [[ ]] represents the control group treated with clear water; SA represents the saline-alkali stress treatment group (SA);

[0020] Sp250 + SA represents the saline-alkali stress treatment group added with 250 mg·L -1 (干重) Scenedesmus obliquus treatment solution; Sp500 + SA represents the saline-alkali stress treatment group added with 500 mg·L -1 (干重) Scenedesmus obliquus treatment solution; Sp750 + SA represents the saline-alkali stress treatment group added with 750 mg·L -1 (干重) Scenedesmus obliquus treatment solution; Sp1000 + SA represents the saline-alkali stress treatment group added with 1000 mg·L -1 (干重) Scenedesmus obliquus treatment solution.

[0021] Figure 4 : Diagram showing the effect of different concentrations of Nitzschia paleacea on the chlorophyll content of cucumber seedlings under saline-alkali stress; where Figure a is the chlorophyll a content diagram; Figure b is the chlorophyll b content diagram; Figure c is the carotenoid content diagram;

[0022] Figure 5 : Effects of Nitzschia palea at different concentrations on the antioxidant enzyme activities of cucumber seedlings under salt-alkali stress; among which, Figure a is the peroxidase (POD) activity diagram; Figure b is the superoxide dismutase (SOD) activity diagram; Figure c is the catalase (CAT) activity diagram;

[0023] Figure 6 : Effects of Nitzschia palea at different concentrations on the contents of osmotic adjustment substances in cucumber seedlings under salt-alkali stress; among which, Figure a is the malondialdehyde content diagram; Figure b is the electrolyte leakage rate diagram; Figure c is the soluble sugar content diagram; Figure d is the soluble protein content diagram;

[0024] Figures 4 - 6 In which, CK represents the control group treated with clear water; SA represents the salt-alkali stress treatment group; Np25+SA represents the salt-alkali stress treatment group added with 25 mg·L -1 (干重) Nitzschia palea treatment solution; Np50+SA represents the salt-alkali stress treatment group added with 50 mg·L -1 (干重) Nitzschia palea treatment solution; Np75+SA represents the salt-alkali stress treatment group added with 75 mg·L -1 (干重) Nitzschia palea treatment solution; Np100+SA represents the salt-alkali stress treatment group added with 100 mg·L -1 (干重) Nitzschia palea treatment solution;

[0025] Figure 7 : Effects of extracellular polymers of Nitzschia palea at different concentrations on the chlorophyll contents of cucumber seedlings under salt-alkali stress; among which, Figure a is the chlorophyll a content diagram; Figure b is the chlorophyll b content diagram; Figure c is the carotenoid content diagram;

[0026] Figure 8 : Effects of extracellular polymers of Nitzschia palea at different concentrations on the antioxidant enzyme activities of cucumber seedlings under salt-alkali stress; among which, Figure a is the peroxidase (POD) activity diagram; Figure b is the superoxide dismutase (SOD) activity diagram; Figure c is the catalase (CAT) activity diagram;

[0027] Figure 9 : Effects of extracellular polymers of Nitzschia palea at different concentrations on the contents of osmotic adjustment substances in cucumber seedlings under salt-alkali stress; among which, Figure a is the malondialdehyde content diagram; Figure b is the electrolyte leakage rate diagram; Figure c is the soluble sugar content diagram; Figure d is the soluble protein content diagram;

[0028] Figures 7 - 9Among them, CK represents the fresh water control treatment group; SA represents the saline-alkali stress treatment group; Np-EPS1+SA represents the saline-alkali stress treatment group added with the extracellular polymer treatment solution of Nitzschia palea diluted by 25%; Np-EPS2+SA represents the saline-alkali stress treatment group added with the extracellular polymer treatment solution of Nitzschia palea diluted by 50%; Np-EPS3+SA represents the saline-alkali stress treatment group added with the extracellular polymer treatment solution of Nitzschia palea diluted by 75%; Np-EPS4+SA represents the saline-alkali stress treatment group added with the undiluted extracellular polymer treatment solution of Nitzschia palea;

[0029] Figure 10 : Diagram of the effect of microalgae fertilizer on the photosynthetic pigment content of cucumber seedlings; among them, Figure a is the chlorophyll a content diagram; Figure b is the chlorophyll b content diagram; Figure c is the carotenoid content diagram; in the figure, CK represents the fresh water control; SA represents the saline-alkali stress treatment; COM represents the microalgae bio-fertilizer treatment; COM+SA represents the treatment of applying microalgae bio-fertilizer under saline-alkali stress;

[0030] Figure 11 : Diagram of the effect of microalgae fertilizer on the growth of cucumber seedlings; among them, Figure a is the plant height of cucumber; Figure b is the stem diameter of cucumber;

[0031] Figure 12 : Diagram of the effect of microalgae fertilizer on the content of osmotic adjustment substances in cucumber leaves grown in saline-alkali soil; among them, Figure a is the malondialdehyde content diagram; Figure b is the electrolyte permeability diagram; Figure c is the soluble sugar content diagram; Figure d is the soluble protein content diagram;

[0032] Figure 13 : Diagram of the effect of applying microalgae bio-fertilizer on the antioxidant enzyme system in cucumber leaves grown in saline-alkali soil; among them, Figure a is the peroxidase (POD) activity diagram; Figure b is the superoxide dismutase (SOD) activity diagram; Figure c is the catalase (CAT) activity diagram; Figure d is the ascorbate peroxidase (APX) activity diagram;

[0033] Figure 14 : Diagram of the effect of microalgae bio-fertilizer on the fruit yield and size of cucumbers grown in saline-alkali soil; Figure a is the fresh weight of a single fruit diagram; Figure b is the fruit length diagram; Figure c is the fruit diameter diagram;

[0034] Figure 15 : Diagram of the effect of applying microalgae bio-fertilizer on the nutrient component content of cucumber fruits grown in saline-alkali soil; Figure a is the vitamin c content diagram; Figure b is the tartaric acid content diagram; Figure c is the soluble sugar content diagram; Figure d is the soluble sugar content diagram;

[0035] Figures 11 - 15 Among them, SA represents natural saline-alkali soil; SSA represents sterilized natural saline-alkali soil; COM+SA represents adding microalgae bio-fertilizer to natural non-sterilized saline-alkali soil; COM+SSA represents adding microalgae bio-fertilizer to sterilized saline-alkali soil for treatment. Detailed implementation mode

[0036] The experiments related to the present invention were completed in the Key Laboratory of Hydrobiology of Heilongjiang Province, Harbin Normal University. The plants used included cucumber (Cucumis sativus L.), Nitzschia palea, and Scenedesmus obliquus.

[0037] Among them, the Nitzschia palea used in the examples can be the Nitzschia strains preserved in the commercial microalgae library, such as the Nitzschia with the preservation number of CCTCC, NO: M 2021192.

[0038] Similarly, the Scenedesmus obliquus can also be the one preserved in the commercial microalgae library or sold by the company. The Scenedesmus obliquus used in this example was purchased from Shanghai Guangyu Biotechnology Co., Ltd., with the product number of GY-D13.

[0039] After screening, it was determined that the microalgae fertilizer prepared by combining Nitzschia palea (hereinafter abbreviated as Np), Scenedesmus obliquus (hereinafter abbreviated as Sp), and Nitzschia palea extracellular polymer (hereinafter abbreviated as Np-EPS) has the best effect of improving the saline-alkali resistance of cucumbers.

[0040] Among them, the Scenedesmus obliquus SP was cultured in a photobioreactor using BG11 medium. The BG11 medium can be a commercially available medium or prepared according to the components shown in Table 1.

[0041] The Nitzschia palea NP was cultured in a tubular photobioreactor using WC medium. The WC medium can be a commercially available medium or prepared according to the components shown in Table 2.

[0042] The Nitzschia palea extracellular polymer Np-EPS was cultured in a tubular photobioreactor using WC medium. The WC medium can be a commercially available medium or prepared according to the components shown in Table 1. After culturing until the algal cell biomass grew to 3.5 g·L -1 (dry weight), the algal cells were removed by centrifugation, and the filtrate was collected to obtain the Nitzschia palea extracellular polymer.

[0043] Table 1: Composition ratio table of BG11 medium

[0044]

[0045]

[0046] Table 2: WC Medium Component Ratio Table

[0047]

[0048] After taking out the two microalgae suspensions (Sp and Np) from the reactor respectively, they were filled into 50 mL screw-cap centrifuge tubes and centrifuged at 5000 r·min -1 for 15 min. Discard the supernatant, disperse the precipitate with ultrapure water and then enrich and centrifuge again. Repeat 3 times. After thoroughly removing the residual medium, transfer the algal cell precipitate to a sterilized Erlenmeyer flask. After adding ultrapure water to resuspend the microalgae precipitate in the Erlenmeyer flask, use a 0.45 μm filter membrane that has been dried to a constant weight and pre-weighed (record the dry weight of the filter membrane m 1 ) to filter 5 mL of the algal solution. Carefully transfer the filter membrane to a vacuum dryer and continue drying at 85 °C until a constant weight is reached. Weigh with an analytical balance and record the mass of the filter membrane carrying the dry algal powder at this time (m 2 ). The dry weight of the microalgae (m) is calculated by Equation (1), and the microalgae biomass concentration (c, g·L -1 ) is calculated by Equation (2). Adjust the concentrations of Scenedesmus obliquus and Nitzschia palea to the specified treatment concentrations for standby.

[0049] m = m 2 - m 1 (1)

[0050] c = m·0.005 -1 (2)

[0051] In the formula: m—the mass of the dry matter of the microalgae (g);

[0052] m 1 —the mass of the 0.45 μm filter membrane after drying (g);

[0053] m 2 —the mass of the filter membrane carrying the dry algal powder (g);

[0054] c—the microalgae biomass concentration (g·L -1 ).

[0055] The test concentrations and abbreviations of the components of each microalgae fertilizer are shown in Table 3.

[0056] Table 3: Test Concentrations and Abbreviations of 3 Components of Microalgae Fertilizer

[0057]

[0058] The screening process and effects of the microalgae fertilizer of the present invention will be described in detail below in conjunction with the examples and the drawings.

[0059] Example 1: Effect of Scenedesmus obliquus on Improving the Salt-alkali Resistance of Cucumbers

[0060] Cucumber seedlings at the two-leaf and one-heart stage were hydroponically cultured. After 7 days of culture, a saline-alkali stress treatment (SA) with a concentration of 90 mmol of Na prepared according to the ratio of NaCl:NaHCO 3 = 1:2 was applied. Non-stressed fresh water was used as the control group (CK). 0, 250 mg·L + (dry weight, the same below), 500 mg·L -1 , 750 mg·L -1 , 1000 mg·L -1 of *Scenedesmus obliquus* were added to the above experimental materials respectively. After 5 days of treatment, the chlorophyll content and the content of cellular osmoregulatory substances in cucumber seedling leaves were measured. -1

[0061] Under saline-alkali stress treatment, the changes in the content of photosynthetic pigments in cucumber leaves after applying *Scenedesmus obliquus* are as Figure 1 shown. Saline-alkali stress (SA group) significantly reduced the photosynthesis of cucumber leaves. The contents of chlorophyll a, chlorophyll b and carotenoids were 0.4 times, 0.47 times and 0.44 times that of CK respectively. After applying *Scenedesmus obliquus*, the degree of damage to leaf photosynthesis was alleviated. Especially in the treatment with *Scenedesmus obliquus* at a concentration of 500 mg·L -1 , the photosynthesis of the leaves increased significantly. The contents of chlorophyll a, chlorophyll b and carotenoids were 0.92 times, 1.03 times and 0.82 times that of the CK group respectively, and there was no significant difference in chlorophyll a and chlorophyll b compared with CK. In addition, when the concentration of *Scenedesmus obliquus* was 250 mg·L -1 , 750 mg·L -1 and 1000 mg·L -1 , although the contents of chlorophyll a, chlorophyll b and carotenoids were significantly lower than those of the CK group, they were significantly higher than those of the saline-alkali stress group (SA group), indicating that the photosynthesis parameters were improved to varying degrees after applying the algal solution. As Figure 1 shown in Fig. a, when the concentration of *Scenedesmus obliquus* was 250 mg·L -1 , the content of chlorophyll a was 0.69 times that of CK and 1.73 times that of the SA group; when the concentration reached 1000 mg·L -1 , the content of chlorophyll a was 0.58 times that of CK and 1.45 times that of the SA group. The content trend of chlorophyll b was the same. When the concentration was 250 mg·L -1 , they were 0.73 times and 1.55 times that of the CK group and the SA group respectively. When the concentration of *Scenedesmus obliquus* was 1000 mg·L -1 , it was 0.47 times that of the CK group and 0.99 times that of the SA group ( Figure 1 b). As Figure 1 shown in Fig. c, the carotenoid content was 0.68 times that of the CK group (algal solution concentration 250 mg·L​-1 ) and 0.58 times (algal solution concentration 750 mg·L -1 ), which were 1.54 times and 1.31 times that of the SA group. The above results indicate that under saline-alkali stress, applying *Scenedesmus obliquus* can effectively alleviate the damage to leaf photosynthesis caused by saline-alkali stress, and the algal solution concentration of 500 mg·L -1 has the best effect.

[0062] As Figure 2 shown, the conductivity of the SA group was significantly higher than that of the CK group, being 1.39 times that of the CK group. After applying the microalgal solution, the electrolyte leakage rate decreased significantly. When the concentration was 500 mg·L -1 , it was 82% of the control group and 0.59 times that of the saline-alkali stress group with a significant difference. The conductivity varied among different concentration treatment groups of the algal solution and was significantly lower than that of the SA group ( Figure 2 b). The soluble sugar content in the SA group was significantly lower than that in the CK group, being 0.45 times that of the CK group. After applying the algal solution, the content increased. When the algal solution concentration was 250 mg·L -1 , it was significantly higher than that of the SA group, being 1.71 times that of the SA group and 0.77 times that of the CK group (P < 0.05). When the algal solution concentration reached 500 mg·L -1 , the soluble sugar content reached the highest value, being 2.27 times that of the SA group. After that, as the algal solution concentration increased, the soluble sugar content gradually decreased. When the concentration reached 1000 mg·L -1 , it was 1.58 times that of the SA group (P < 0.05) and was significantly lower than that of the CK group, being 0.71 times that of the CK group ( Figure 2 c). The soluble protein content was as Figure 2 shown in d. Its trend was similar to that of the soluble sugar content. The content in the SA group was 0.54 times that of the CK group (P < 0.05). After applying the algal solution, it increased significantly and reached the highest value at 500 mg·L -1 , being 1.94 times that of the SA group with no significant difference compared with the CK group. After that, as the concentration increased, the soluble protein content gradually decreased. There was no significant difference between 750 mg·L -1 and 250 mg·L -1 . However, when the concentration was 1000 mg·L -1 , it was significantly lower than the other three algal solution concentration treatment groups and was still significantly higher than that of the SA group. Both 750 mg·L -1 and 1000 mg·L -1 were significantly lower than that of the CK group, being 0.85 times and 0.69 times that of the CK group respectively. The above results show that after applying *Scenedesmus obliquus*, it can effectively alleviate the oxidative damage of cucumber seedlings caused by saline-alkali stress and is beneficial to the osmotic adjustment and repair of seedlings under saline-alkali stress. Among them, the algal solution concentration of 500 mg·L -1 has a better effect.

[0063] AsFigure 3 As shown, the peroxidase (POD) activity was lower in the saline-alkali stress treatment group (SA), and there was no significant difference compared with the treatment group with 250 mg·L -1 of algal solution; when the algal solution concentration increased to 500 mg·L -1 , the POD activity increased significantly, which was 1.71 times that of the treatment group with 250 mg·L -1 of algal solution, but there was no significant difference compared with the CK group. With the further increase of the algal solution concentration, the POD activity decreased and was significantly lower than that of the treatment groups with 250 mg·L -1 of algal solution and the CK group ( Figure 3 a).

[0064] The superoxide dismutase (SOD) activity was the highest in the algal solution group with 500 mg·L -1 , and there was no significant difference compared with the algal solution group with 250 mg·L -1 , but it was significantly higher than that of the SA group, the algal solution groups with 750 mg·L -1 and 1000 mg·L -1 , being 1.54 times, 1.24 times and 1.23 times of them respectively. There was no significant difference in SOD activity between the algal solution groups with 750 mg·L -1 and 1000 mg·L -1 ( Figure 3 b).

[0065] The catalase (CAT) activity was also the highest in the algal solution group with 500 mg·L -1 , and there was no significant difference compared with the CK, but it was significantly higher than that of the other four treatment groups, being 2.17, 1.38, 1.29 and 1.80 times of the SA group, the groups with 250 mg·L -1 , 750 mg·L -1 and 1000 mg·L -1 respectively. There was no significant difference in SOD activity between the algal solution groups with 250 mg·L -1 and 750 mg·L -1 ; there was no significant difference in SOD activity between the SA group and the algal solution group with 1000 mg·L -1 ( Figure 3 c).

[0066] The above results show that under saline-alkali stress conditions, applying an appropriate amount of Scenedesmus obliquus algal solution can significantly affect the antioxidant enzyme system of cucumber seedlings. Among them, the algal solution concentration of 500 mg·L -1 is particularly effective in enhancing the activities of POD, SOD and CAT, indicating that the Scenedesmus obliquus algal solution at this concentration may have a significant promoting effect on the adaptability and viability of cucumber seedlings under saline-alkali stress conditions.

[0067] Example 2: Effect of Nitzschia palea on improving the saline-alkali resistance of cucumbers

[0068] Cucumber seedlings at the two-leaf and one-heart stage were hydroponically cultured. After 7 days of culture, a saline-alkali stress treatment (SA) with a concentration of 90 mmol of Na prepared according to the ratio of NaCl:NaHCO 3 =1:2 was applied. Non-stressed fresh water was used as the control group (CK). The above experimental materials were respectively added with treatment solutions of Nitzschia palea at 0, 25 mg·L + , 50 mg·L -1 , 75 mg·L -1 , 100 mg·L -1 . After 5 days of treatment, the chlorophyll content and osmotic adjustment substance content of cucumber seedling leaves were measured to screen the suitable application concentration of Nitzschia palea. -1

[0069] To study the effect of Nitzschia palea on the photosynthesis of cucumber leaves under saline-alkali stress, the contents of chlorophyll a, b and carotenoids in the leaves were detected, as Figure 4 shown. The content of chlorophyll a decreased significantly in the single saline-alkali treatment SA group, which was 0.53 times that of CK. After applying the algal solution in the saline-alkali treatment, the content of chlorophyll a gradually increased. When the concentration was 25 mg·L -1 , it was significantly higher than the SA group, which was 1.4 times that of the SA group. When the concentration reached 50 mg·L -1 , it reached the highest value, and there was no significant difference compared with CK, which was 2.03 times that of the SA group. After that, with the increase of the algal solution concentration, the content of chlorophyll a decreased significantly. When the concentration was 100 mg·L -1 , there was no significant difference compared with 25 mg·L -1 ([[]] Figure 4 a). The content of chlorophyll b in each treatment group decreased significantly compared with the CK group, and the content in the SA group was the lowest, which was 0.34 times that of CK. After applying the algal solution, the content of chlorophyll b under saline-alkali stress was increased. When the algal solution concentration was 25 mg·L -1 (1.52 times), 50 mg·L -1 (1.93 times), 75 mg·L -1 (1.71 times) and 100 mg·L -1 (1.48 times) were all significantly higher than the SA group, and 50 mg·L -1 was significantly higher than other algal solution treatment groups ([[]] Figure 4 b). The change trend of carotenoid content was similar to that of chlorophyll b content, that is, when the concentration of Nitzschia palea was 50 mg, it was significantly higher than each treatment group, but significantly lower than the CK group, which was 2.25 times that of the SA group. The content in the SA group was significantly lower than that in the CK group, indicating that the single saline-alkali treatment inhibited the synthesis of carotenoids, and the application of the algal solution alleviated this inhibitory effect. When the concentration was 25 mg·L -1 ​When treating the algal solution with the concentration, the carotenoid content was 1.67 times that of the SA group, and there was no significant difference compared with the treatment groups with concentrations of 75 mg·L -1 and 100 mg·L -1 (c). It can be seen from the above results that the exogenous application of *Nitzschia paleacea* can reduce the damage of saline-alkali stress to the photosynthesis of cucumber leaves. Among them, the effect is better when the concentration of the algal solution is 50 mg·L Figure 4 -1 -1 , and the alleviating effect on chlorophyll a is the most significant.

[0070] The antioxidant enzyme system is a system for plants to regulate reactive oxygen species by themselves when under stress, Figure 5 as shown in the effect of applying different concentrations of *Nitzschia paleacea* on the antioxidant enzyme system of cucumber seedlings under saline-alkali stress. The POD activity was the lowest in the SA group, significantly lower than that of other groups. The POD activities of the algal solution concentrations of 25 mg·L -1 , 50 mg·L -1 and 75 mg·L -1 showed no significant difference from that of the CK group. Among them, the POD activity of the algal solution concentration of 50 mg·L -1 was significantly higher than that of the algal solution concentration of 100 mg·L -1 group, being 1.11 times that of the latter ( Figure 5 a). The SOD activity of CK was the highest, followed by that of the algal solution concentration of 50 mg·L -1 group. The SOD activities of the algal solution concentrations of 25 mg·L -1 , 75 mg·L -1 , 100 mg·L -1 showed no significant difference from that of the SA group. The SOD activities in the algal solution concentration of 50 mg·L -1 group were 1.12, 1.14 and 1.10 times that of the SA, 25 mg·L -1 and 100 mg·L -1 groups respectively ( Figure 5 b).

[0071] The CAT activity was the highest in the CK group, significantly higher than that of other groups, and the lowest in the SA group. The CAT activities of the algal solution concentrations of 50 mg·L -1 and 75 mg·L -1 groups showed no significant difference and were higher than those of other algal solution treatment groups. Among them, the CAT activity of the algal solution concentration of 50 mg·L -1 was 1.22 times that of the SA group, 1.13 times that of the 25 mg·L -1 group, and 1.10 times that of the 100 mg·L -1 group ( Figure 5c). The above results indicate that applying *Nitzschia paleacea* under saline-alkali stress can significantly increase the antioxidant enzyme activity in cucumber leaves, resist the oxidative damage caused by stress, and the concentration of *Nitzschia paleacea* algal solution at 50 mg·L -1 has the best effect.

[0072] Figure 6 As shown, after applying different concentrations of *Nitzschia paleacea* under saline-alkali stress, the malondialdehyde (MDA) content in the SA group was significantly higher than that in other groups, 2.35 times that of CK. After applying *Nitzschia paleacea*, the MDA content decreased significantly. When the algal solution concentration was 25 mg·L -1 , 50 mg·L -1 , 75 mg·L -1 , 100 mg·L -1 , they were 0.68, 0.60, 0.65, and 0.73 times that of the SA group respectively. The lowest value of MDA content appeared in the treatment group with an algal solution concentration of 50 mg·L -1 . There was no significant difference in MDA content between the algal solution concentrations of 50 mg·L -1 and 75 mg·L -1 , and no significant difference between 25 mg·L -1 and 100 mg·L -1 , and these 4 algal solution treatment groups were all significantly higher than the CK group ( Figure 6 a). The electrical conductivity reached the highest value in the SA group, 1.65 times that of CK. After applying *Nitzschia paleacea*, the electrical conductivity decreased significantly. As the algal solution concentration increased, the electrical conductivity gradually increased, and there was no significant difference between the treatments with different algal solution concentrations. The electrical conductivities at concentrations of 75 mg·L -1 and 100 mg·L -1 were 1.36 times and 1.44 times that of the CK group respectively, but 0.83 times and 0.83 times that of the SA group; the electrical conductivities at 25 mg·L -1 and 50 mg·L -1 were 0.79 times and 0.76 times that of the SA group respectively ( Figure 6 b). Except for the treatment group with an algal solution concentration of 50 mg·L -1 , the soluble sugar content in the CK group was significantly higher than that in other groups. The content in the SA group was 0.71 times that of the CK group. With the application of the algal solution, the soluble sugar content showed a trend of first increasing and then decreasing. At a concentration of 25 mg·L -1 , it was significantly higher than the SA group, 1.12 times that of the SA group; it reached the maximum value at an algal solution concentration of 50 mg·L -1 , 1.38 times that of the SA group. There was no significant difference between 75 mg·L -1 and 25 mg·L -1 , and there was no significant difference from the SA group when the algal solution concentration reached 100 mg·L -1 ( Figure 6c). The soluble protein content was the highest in the treatment group with the algal solution concentration of 50 mg·L -1 and was 1.43 times that of the CK. Subsequently, it gradually decreased with the increase of the algal solution concentration. The differences between the algal solution concentrations of 100 mg·L -1 , 25 mg·L -1 and the CK group were not significant. They were all lower than those of other algal solution treatment groups but higher than those of the SA group, being 1.58 times, 1.42 times and 1.49 times the soluble protein content of the SA group, respectively. The above results indicate that when the application concentration of *Nitzschia palea* is 50 mg·L -1 , it can effectively increase the content of osmotic adjustment substances in cucumber seedlings and reduce the oxidative damage caused by saline-alkali stress to the plants ( Figure 6 d).

[0073] The results show that under saline-alkali stress conditions, applying the algal solution of *Nitzschia palea* at 50 mg·L -1 (dry weight) can significantly restore the formation of photosynthetic pigments in cucumber seedling leaves, increase the activity of the antioxidant enzyme protection system of cucumber seedlings, and reduce the degree of cell membrane damage in cucumber leaves. The above results indicate that the algal solution of *Nitzschia palea* at 50 mg·L -1 (dry weight) has a significant promoting effect on the adaptability and viability of cucumber seedlings under saline-alkali stress conditions.

[0074] Example 3: Effect of extracellular polymers of *Nitzschia palea* on improving the saline-alkali resistance of cucumbers

[0075] In this example, the effects of extracellular polymers of *Nitzschia palea* on the formation of photosynthetic pigments, antioxidant enzyme system and cell membrane damage in cucumber seedling leaves were further analyzed. The results are as Figure 7 — Figure 9 shown.

[0076] Cucumber seedlings at the two-leaf and one-heart stage were hydroponically cultured. After 7 days of culture, saline-alkali stress treatment (SA) with a concentration of 90 mmol prepared according to the ratio of NaCl:NaHCO 3 = 1:2, that is, the treatment solution ratio was 30 mmol of NaCl + 60 mmol of NaHCO + was applied. The non-stressed fresh water was used as the control group (CK). Different concentrations of the extracellular polymer treatment solution of *Nitzschia palea* were added to the above experimental materials. The extracellular polymer of *Nitzschia palea* was obtained by culturing *Nitzschia palea* to reach 3.5 g·L 3 -1 ​The supernatant without algal cells remaining after centrifugally enriching Nitzschia palea algal cells at (dry weight). The concentration of extracellular polymeric substances (Np-EPS) of Nitzschia palea was diluted to 25% concentration based on the collected stock solution and denoted as Np-EPS1, diluted to 50% concentration and denoted as Np-EPS2, diluted to 75% concentration and denoted as Np-EPS3, and the undiluted 100% stock solution was denoted as Np-EPS4. After 5 days of treatment, the chlorophyll content and osmotic adjustment substance content of cucumber seedling leaves were measured to screen the appropriate application concentration of extracellular polymeric substances of Nitzschia palea.

[0077] The results showed that the application of extracellular polymeric substances of Nitzschia palea had a mitigating effect on the damage caused by the synthesis of photosynthetic pigments in cucumber leaves under saline-alkali stress, and the results were as Figure 7 shown. Under saline-alkali stress (SA group), the chlorophyll a content decreased significantly, being 0.41 times that of CK. After applying the extracellular polymeric substances of the algal solution, the chlorophyll a content gradually increased. When the concentration was 25%, it was significantly higher than that of the SA group. There was no significant difference between the 50% concentration and the 25% concentration. When the concentration reached 75%, the content was significantly higher than the previous two groups, reaching the maximum value, which was 2.54 times that of SA. However, when the polymer concentration increased further, the chlorophyll a content showed a downward trend ( Figure 7 a). The chlorophyll b content was also the lowest under single saline-alkali stress, being 0.37 times that of the CK group. After applying the extracellular polymeric substances, it increased significantly. Among them, the chlorophyll b contents at the applied concentrations of 25%, 50%, and 75% were all significantly higher than that of the SA group, being 1.33, 1.66, and 2.47 times that of it respectively ( Figure 7 b). The content of carotenoids was similar to the change in chlorophyll a content. The SA treatment group was the lowest, being 0.33 times that of CK. After applying the extracellular polymeric substances, it increased significantly. There was no significant difference between the algal solution concentrations of 25% and 50%. When the concentration was 75%, the carotenoid content reached the maximum value, which was 3.05 times that of the SA group and not significantly different from that of CK. From the above results, it can be seen that the extracellular polymeric substances of Nitzschia palea can significantly reduce the damage to leaf photosynthesis caused by saline-alkali stress, enable the leaves to resume normal photosynthesis, and the appropriate concentration of the extracellular polymeric substances is 75% ( Figure 7 c).

[0078] The effects of extracellular polymeric substances of Nitzschia palea at different concentrations on the antioxidant enzyme system of cucumber seedlings were as Figure 8 shown. The POD activity of the saline-alkali stress group (SA) was the lowest. Subsequently, with the increase in the concentration of extracellular polymeric substances, the POD activity gradually increased. Among them, when the polymer concentration was 75%, the activity was the highest, being 1.42 times that of the POD activity of the SA group and about 1.15 times that of the polymer concentrations of 25% - 50%. There was no significant difference in the POD activity between 75% and 100% of the polymer concentration ( Figure 8a). The SOD activity showed a similar trend of change. The activity was also the highest when the polymer concentration was 75%, which was 1.43 times that of the SA group and significantly higher than that of other polymer concentration treatment groups. It was 1.28 times that of the 100% polymer concentration. Figure 8 b). The CAT activity was the lowest in the SA group and the highest in the CK group. With the increase in the polymer concentration, the CAT activity showed a trend of first increasing and then decreasing. When the polymer concentration was 75%, the CAT activity was 1.64 times that of the SA group and 1.49 times that of the 100% concentration group. Figure 8 c). From the above results, it can be seen that 75% of the extracellular polymer of Nitzschia palea can improve the antioxidant enzyme activity of plants and enhance their ability to resist saline-alkali stress.

[0079] Such as Figure 9 The figure shows the results of the effect of the extracellular polymer of Nitzschia palea on the osmotic adjustment of cucumber seedlings. The MDA content increased significantly in the SA group, which was 1.75 times that of the CK. After applying the extracellular polymer, the MDA content gradually decreased. At a concentration of 25%, it was significantly lower than that of the SA group, which was 0.85 times that of the SA. When the concentration reached 75%, the MDA content was the lowest and was significantly lower than that of the 25% group and the SA group, which were 0.83 times and 0.71 times respectively, and there was no significant difference compared with the 50% concentration group. Figure 9 a). The conductivity showed a similar trend of change. Among them, the saline-alkali SA treatment group was significantly higher than the CK, which was 1.54 times that of the CK. After applying the extracellular polymer of the algal solution, the conductivity decreased significantly. When the polymer concentration was 75%, the conductivity was the lowest, which was 0.78 times that of the SA, but was significantly higher than the CK group, which was 1.20 times that of the CK. Figure 9 b). Figure 9 c-d are the contents of soluble sugar and soluble protein. It can be seen that the accumulation of these two substances in the single saline-alkali SA treatment group decreased significantly compared with the CK, which were 0.47 times and 0.59 times that of the CK respectively. After applying the extracellular polymer of Nitzschia palea, the contents of soluble sugar and soluble protein gradually increased and reached the maximum value at a concentration of 75%, which were 1.89 times and 1.57 times that of the SA group respectively, and there was no significant difference in the soluble protein content compared with the CK. Figure 9 c). When the polymer concentration reached 100%, the contents of the two osmotic adjustment substances decreased significantly, but were also significantly higher than those of the SA group. The above results show that the extracellular polymer of Nitzschia palea can increase the content of soluble substances in cucumber seedlings, resist the damage caused by saline-alkali stress, reduce the degree of membrane lipid peroxidation of seedlings, and the effect is better at a concentration of 75%.

[0080] Under saline-alkali stress conditions, the application of the extracellular polymer of Nitzschia palea can significantly restore the formation of photosynthetic pigments in cucumber seedling leaves, improve the activity of the antioxidant enzyme protection system of cucumber seedlings, reduce the degree of cell membrane damage in cucumber leaves. The results measured after the application of this example show that the extracellular polymer of Nitzschia palea at a 75% dilution has a significant promoting effect on the adaptability and viability of cucumber seedlings under saline-alkali stress conditions.

[0081] Through the above example results, the following appropriate concentrations that can alleviate the saline-alkali stress damage of cucumber seedlings were found: Scenedesmus obliquus 500 mg·L -1 , Nitzschia palea 50 mg·L -1 , and the concentration of the extracellular polymer of Nitzschia palea is 75% of the stock solution.

[0082] Example 4: Detection of the effects of different concentrations of microalgae fertilizers

[0083] In order to better explore the effects of the concentrations of the components of microalgae fertilizers on the germination rate of cucumber seeds, based on the results of single-factor experiments, to better explore the effects of the concentration of algal fertilizer on the germination rate of cucumber seeds, Design-Expert V12.0.3 software was used for response surface design. Taking the concentration of Sp algal fertilizer (A), the concentration of Np algal fertilizer (B), and the concentration of Np-EPS algal fertilizer (C) as independent variables, and the germination rate of cucumber seeds as the response value (Y), according to the Box-Behnken principle, based on the concentration of Scenedesmus obliquus Sp 500 mg·L -1 , the concentration of Nitzschia palea 50 mg·L -1 , and the concentration of the extracellular polymer of Nitzschia palea being 75% as the intermediate value, the model Box-Behnken (BBD) was selected in the software Design-Expert V 12.0.3 to set up a 3-factor 3-level response surface experiment, and the experiment was carried out according to the experimental list, and the results were entered into the software.

[0084] The experimental results were analyzed by multiple regression fitting to obtain the quadratic multinomial regression equation of the cucumber seed germination rate with respect to the three factors as shown in Equation (3):

[0085] Y = 96.39 + 1.20A + 0.6125B + 1.62C + 0.7100AB -

[0086] 2.65AC + 1.54BC - 8.78A2 - 8.48B2 - 2.94C2. (3)

[0087] The variance analysis of the regression model was carried out, as shown in Table 4. This response surface model is significant, the regression is extremely significant (P < 0.0001), and the correlation coefficient R of the equation 2The value was 0.9998, and the adjusted determination coefficient Radj = 0.9995, indicating that the response surface model fitted well with the actual results and the experimental method was reliable; the lack-of-fit term was not significant (P = 0.2489), further indicating that the response surface model fitted well with the actual results and could be used for the comparison and analysis of the optimization of diatom culture conditions; the coefficient of variation (0.1898%) was low, proving that the response surface model had high reliability. From the F values of each item, it can be seen that the factors affecting the germination rate of cucumber seeds were in the order of: C > A > B, that is, Np-EPS > Sp > Np.

[0088] Table 4: Analysis of variance table of the optimal regression model for cucumber seed germination rate

[0089]

[0090]

[0091] By solving the regression equation (3) using Design-Expert V 12.0.3 software, the conditions for the maximum germination rate of cucumber seeds were obtained as follows: the concentration of Sp was 401.27 mg·L -1 , the concentration of Np was 73.24 mg·L -1 , and the concentration of Np-EPS was 61.06%. This was the optimal condition for cucumber seed germination. Under this condition, the germination rate of cucumber seeds was 84.9824%. According to the feasibility of preparing the fertilizer, for the convenience of later measurement, the value-taking conditions were adjusted as follows: the concentration of Sp was 400 mg·L -1 , the concentration of Np was 70 mg·L -1 , and the concentration of Np-EPS was 60%. Six repeated experiments as above were carried out according to the software, and the result showed that the germination rate of cucumber seeds was 96.45%. The difference from the predicted value was not significant, indicating that the microalgae fertilizer at this concentration could effectively improve the germination rate of cucumber seeds and was the optimal concentration that could alleviate the saline-alkali stress of cucumbers.

[0092] Example 5: Effect detection of the determined optimal ratio microalgae fertilizer

[0093] Figure 10 Abbreviation description: Clear water was used as the control (CK), saline-alkali stress treatment (SA), microalgae fertilizer treatment (COM), and microalgae fertilizer treatment under saline-alkali stress (COM + SA).

[0094] After cucumber seeds germinated, they were sown in a 9 cm diameter nutrient sowing, and the cultivation substrate was peat soil: vermiculite = 1:2. Saline-alkali stress treatment (SA) was carried out, and the treatment solution ratio was NaCl 30 mM + NaHCO 3 60 mM. Using clear water as the control (CK), after applying the microalgae bio-fertilizer (COM) with the above formula for 14 days, the chlorophyll content of cucumber leaves was measured. As Figure 10As shown in Figure a, the content of chlorophyll a was the highest in the COM group, showing no significant difference from that in the CK group, but significantly higher than those in the SA group and the COM+SA group, being 2.16 times and 1.37 times of them respectively. The content of chlorophyll a in the algal fertilizer applied group under saline-alkali treatment (COM+SA) was significantly higher than that in the SA group, being 1.57 times of it. The content of chlorophyll b reached the maximum in the control group, showing no significant difference from that in the COM group. The content of chlorophyll b in the CK group was 1.18 times of that in the COM+SA group and 1.64 times of that in the SA group. The content of chlorophyll b in the COM+SA group was 1.38 times of that in the SA group ( Figure 10 b). The content of carotenoid was also the highest in the CK group, being 1.77 times, 1.17 times and 1.37 times of those in the SA group, the COM group and the COM+SA group respectively. The content of carotenoid in the COM+SA group showed little difference from that in the COM group and also little difference from that in the SA group. The content of carotenoid in the algal fertilizer treated group (COM) was significantly higher than that in the SA group, being 1.51 times of it ( Figure 10 c). The above test results show that the microalgal fertilizer can significantly increase the contents of chlorophyll a and chlorophyll b in cucumber seedlings at the two-leaf and one-heart stage, and reduce the adverse effects of saline-alkali stress on leaf photosynthetic pigments.

[0095] Example 5: Comparison of the effects of microalgal fertilizers and other fertilizers

[0096] Figure 11 Abbreviation description: untreated saline-alkali soil treatment (SA), sterilized saline-alkali soil treatment (SSA), microalgal fertilizer applied to untreated saline-alkali soil treatment (COM+SA), microalgal fertilizer applied to sterilized saline-alkali soil treatment (COM+SSA)

[0097] The planting soil at a depth of 20 cm from the surface layer of saline-alkali soil in the Songnen Plain saline-alkali land was taken for the test. The sampling site was located in Anda City, Heilongjiang Province. The pH value of the soil sample was measured to be 8.5. After the above natural saline-alkali soil was passed through a 4-mm sieve and subjected to sterilization (SSA) and non-sterilization (SA) treatments, it was used for planting cucumber seedlings at the two-leaf and one-heart stage. 200 mL of microalgal bio-fertilizer (COM) was watered every 10 days. After 50 days of planting, plant samples were taken for the determination of plant height, plant stem diameter and other indicators.

[0098] The results of cucumber plant height traits showed that the cucumber plants grown after applying microalgal fertilizer to saline-alkali soil (COM+SA) had the highest plant height, being 1.17 times of that in the saline-alkali treatment group SA and 1.54 times of that in the sterilized saline-alkali soil SSA. The plant height of cucumber plants grown after applying algal fertilizer to sterilized soil (COM+SSA) was lower than that in the COM+SA, but the difference was not significant. Figure 11) The cucumber plants grown in the COM+SSA group had the thickest stems. There was no significant difference compared with the COM+SA group, but it was significantly higher than the SA group and the SSA group, being 1.06 times and 1.39 times that of the SA group and the SSA group respectively. The stem diameter of the cucumbers grown in the SA group was significantly higher than that in the SSA group, being 1.31 times that of the SSA group. The above results indicate that applying microalgae fertilizer to saline-alkali soil significantly increases the plant height and stem diameter of cucumber plants, and the cucumbers grow sturdily in saline-alkali soil after applying microalgae fertilizer.

[0099] The osmotic adjustment system reflects the degree of stress on the plant and its self-regulation ability. In this example, the alleviating effect of the microalgae fertilizer described in the present invention on cucumber saline-alkali stress was illustrated by detecting the contents of malondialdehyde, relative electrical conductivity, free proline, and soluble protein in cucumber leaves.

[0100] As Figure 12 shown, the content of malondialdehyde (MDA) was the highest in the SA group, significantly higher than that in other groups, in the order of SA>SSA>COM+SA>COM+SSA. The MDA content in the COM+SA group was 0.54 times that in the SA group, and the MDA content in the COM+SSA group was 0.50 times that in the SSA group ( Figure 12 a). There was no significant difference in the relative electrical conductivity between the SA group and the SSA group, and both were higher than the other two groups. Among them, the relative electrical conductivity of the SA group was 1.39 times that of the COM+SA group, the relative electrical conductivity of the SSA group was 1.41 times that of the COM+SA group, and 1.60 times that of the COM+SSA group. The relative electrical conductivity of the COM+SA group was significantly higher than that of the COM+SSA group, being 1.14 times that of the COM+SSA group ( Figure 12 b). The change in the free proline content of each treatment group was SSA>SA>COM+SSA>COM+SA. Among them, the proline content in the SSA group was significantly higher than that in other groups, being 1.11 times that of the SA group, 1.76 times that of the COM+SSA group, and 2.09 times that of the COM+SA group. The proline content in the COM+SSA group was significantly higher than that in the COM+SA group, being 1.19 times that of the COM+SA group ( Figure 12 c). The soluble protein content in the COM+SA group was significantly higher than that in the COM+SSA group, being 1.17 times that of the COM+SSA group. There was no significant difference in the soluble protein content between the SA group and the SSA group, and both were at a relatively low level, being 0.66 times and 0.59 times that of the COM+SA group respectively ( Figure 12 d). The above results indicate that after applying microalgae fertilizer to cucumber plants grown in saline-alkali soil, the osmotic adjustment ability of the leaves can be effectively improved by regulating the contents of free proline and soluble protein, reducing the MDA content and relative electrical conductivity, and alleviating the stress damage suffered by the plants.

[0101] At the same time, the changes in the antioxidant enzyme system of cucumber leaves after applying microalgae fertilizer to cucumbers grown in saline-alkali soil were detected.

[0102] After applying the microalgae bio-fertilizer, the enzyme activities of the antioxidant enzyme system in cucumber leaves are as follows Figure 13 shown. The results show that there is no significant difference in the POD activity between the SA group and the SSA group, and both are significantly higher than the other two groups. The POD activity of the COM+SA group is 0.65 times that of SA, while the POD activity of the COM+SSA group is 0.69 times that of the SSA group. There is no significant difference in the POD activity between the COM+SA and COM+SSA groups ( Figure 13 a). The SOD activity in the leaves shows a similar trend of change. Among them, the SOD activity of the SA group is 1.41 times that of the COM+SA group, and the SSA group is 1.35 times that of the COM+SSA group. There is no significant difference in the SOD activity between the COM+SA and COM+SSA groups ( Figure 13 b). The CAT activity is significantly different among different treatment groups, and the trend of change is SSA > SA > COM+SSA > COM+SA. Among them, the SSA group is 1.17 times that of the SA group, 1.58 times that of the COM+SSA group, and 2.22 times that of the COM+SA group. The CAT activity of the COM+SSA group is significantly higher than that of the COM+SA group, being 1.41 times that of the latter ( Figure 13 c). The APX enzyme activity is relatively high in the SA group and the SSA group, and there is no significant difference between the two, and both are significantly higher than the other two groups. There is little difference in the APX enzyme activity between the COM+SA group and the COM+SSA group. Among them, the APX activity of the SA group is 1.20 times that of the COM+SA group and 1.24 times that of the COM+SSA group; while the APX activity of the SSA group is 1.18 times that of the COM+SA group and 1.23 times that of the COM+SSA group ( Figure 13 d). From the above results, it can be seen that whether it is sterilized or unsterilized saline-alkali soil, cucumbers are affected by saline-alkali stress and activate the antioxidant enzyme system to resist the harm of excessive reactive oxygen species. After applying the microalgae bio-fertilizer, the damage caused by stress to the plants can be alleviated, and the activity of the antioxidant enzyme system decreases somewhat.

[0103] Example 6: Influence effect of microalgae fertilizer on cucumber growth

[0104] Figure 14 Abbreviation description: unsterilized saline-alkali soil treatment (SA), sterilized saline-alkali soil treatment (SSA), unsterilized saline-alkali soil treatment with microalgae fertilizer application (COM+SA), sterilized saline-alkali soil treatment with microalgae fertilizer application (COM+SSA)

[0105] After applying the microalgae fertilizer, the average single fruit weight, fruit diameter and length of cucumber fruits are as follows Figure 14As shown in the figure. The results of the fresh weight of single cucumber fruits showed that the SSA group and the SA group significantly inhibited fruit growth, and their fresh weight per fruit was significantly lower than that of other groups. The fresh weight per fruit of the SSA group was the lowest, which was 30.7% lower than that of the SA group. After applying microalgae bio-fertilizer, the fresh weight per fruit of the treatment group increased significantly (P < 0.05). The fresh weight per fruit of the COM+SA group increased by 41.5% compared with the SA group, by 85.0% compared with the SSA group, and by 12.6% compared with the COM+SSA group (P < 0.05), indicating that applying microalgae bio-fertilizer could significantly increase the cucumber fruit yield ( Figure 14 a). The fruit length was the highest in the COM+SA group, which was significantly higher than that of other treatment groups. The fruit length of the COM+SA group increased by 23.4% compared with the SA group, by 34.8% compared with the SSA group, and by 10.2% compared with the COM+SSA group (P < 0.05)( Figure 14 b). The change trend of the cucumber fruit diameter among the treatment groups was COM+SA group > COM+SSA group > SA group > SSA group. Among them, the fruit diameter of the SSA group decreased by 11.2% compared with the SA group, by 21.1% compared with the COM+SA group, and by 18.0% compared with the COM+SSA group (P < 0.05)( Figure 14 c). The above results showed that the fresh weight per fruit, fruit length, and fruit diameter were the lowest in the sterilized saline-alkali soil treatment group, and applying microalgae bio-fertilizer significantly increased the fresh weight per fruit, fruit length, and thickness.

[0106] The quality of cucumber fruits is as important as the yield. At the same time, the changes in the contents of various nutritional components of cucumber fruits were detected.

[0107] The change in the content of vitamin C (Vc) was COM+SA group > COM+SA group > SSA group > SA group. Among them, the Vc content of the COM+SA group increased by 42.7% compared with the SA group and by 19.3% compared with the SSA group (P < 0.05). The Vc content of the COM+SSA group was the highest, which was significantly higher than that of other groups, increasing by 93.7% compared with the SA group and by 62.0% compared with the SSA group (P < 0.05)( Figure 15 a). The tartaric acid content was the highest in the COM+SSA group, and there was no significant difference compared with the COM+SA group. The tartaric acid contents of both were significantly higher than those of the SA group and the SSA group. The tartaric acid content of the COM+SSA group increased by 12.4% compared with the SA group and by 19.5% compared with the SSA group. There was no significant difference in the tartaric acid content between the SA group and the SSA group (P > 0.05)( Figure 15b). There was no significant difference in the soluble protein content in the fruits between the COM+SSA group and the COM+SA group, and both were significantly higher than those in the SA group and the SSA group. Among them, the soluble protein content in the COM+SA group increased by 63.3% compared with the SA group and by 57.9% compared with the SSA group, while the soluble protein content in the COM+SSA group increased by 62.4% compared with the SA group and by 57.2% compared with the SSA group (P

[0108] < 0.05)( Figure 15 c). The soluble sugar content was the highest in the COM+SSA group, followed by the COM+SA group > SSA group > SA group. Among them, the soluble sugar content in the SSA group increased by 27.0% compared with the SA group, while the COM+SA group increased by 50.3% compared with the SA group, the COM+SSA group increased by 79.0% compared with the SA group and by 41.0% compared with the SSA group (P < 0.05)( Figure 15 d).

[0109] In summary, the microalgae bio-fertilizer provided by the present invention is composed of fresh microalgae cells and microalgae extracellular polymers. This microalgae bio-fertilizer can promote seed germination and seedling growth of cucumbers under both normal growth and saline-alkali stress conditions. Further, by applying it to the research on the important growth stages of cucumbers under saline-alkali stress, and comprehensively analyzing the effects of microalgae fertilizer on the growth of cucumber seedlings, fruit yield and quality under sterilized and non-sterilized saline-alkali soil planting conditions, it is found that this microalgae bio-fertilizer can promote the growth of cucumbers planted in sterilized and non-sterilized saline-alkali soils, reduce the saline-alkali damage of plants, and improve the cucumber yield and fruit quality. The microalgae bio-fertilizer of the present invention can provide material guarantee for cucumber planting in saline-alkali soils in China.

Claims

1. A microalgae fertilizer, characterized in that: The microalgae fertilizer contains Nitzschia gracilis and Scenedesmus obliquus.

2. The microalgae fertilizer according to claim 1, characterized in that The microalgae fertilizer contains 73.24 mg·L -1 , the content of Scenedesmus obliquus was 401.27 mg·L -1 .

3. The microalgae fertilizer according to claim 1, characterized in that The microalgae fertilizer also contains extracellular polymers of Nitzschia granatum.

4. The microalgae fertilizer according to claim 3, characterized in that The extracellular polymer of Nitzschia gracilis is obtained by culturing Nitzschia gracilis, removing algal cells and collecting filtrate.

5. The microalgae fertilizer according to claim 4, characterized in that: The culture is carried out using WC medium until the algae cell biomass grows to 3.5 g·L -1 Afterwards, the algal cells were removed by centrifugation.

6. The microalgae fertilizer according to claim 3, characterized in that The microalgae fertilizer has a concentration of extracellular polymers of Nitzschia granatum of 61.06%.

7. Use of the microalgae fertilizer according to claim 1 or claim 3 in improving the salt-alkali resistance of cucumber.

8. A method for promoting cucumber germination, characterized in that: The method is to use the microalgae fertilizer according to claim 1 or claim 3 in the cucumber germination process.

9. A method for promoting the growth of cucumbers in saline-alkali soil, characterized in that: The method is to apply the microalgae fertilizer according to claim 1 or claim 3 during planting.

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