Screening method and application of anti-aspergillus niger lactic acid bacteria strain

Abstract

The invention discloses a screening method and application of an anti-Aspergillus niger lactic acid bacteria strain FOSU-ZX803, and the anti-Aspergillus niger lactic acid bacteria strain FOSU-ZX803 obtained by screening shows extremely strong inhibition ability and excellent probiotic characteristics and safety to Aspergillus niger, and meets the requirements of food safety strains. When the cell-free supernate of the strain FOSU-ZX803 is applied to bread preservation, the growth of aspergillus niger can be remarkably inhibited, and the shelf life of bread can be effectively prolonged. Besides, after the strain is primarily screened from the sour pickles and is subjected to induction culture, the screening efficiency and success rate are greatly improved, the aspergillus niger resistance activity of the strain is more stable, the performance is more remarkable, and the strain has higher application value.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of microorganisms, in particular to an anti-aspergillus niger lactic acid bacteria strain and a screening method and application thereof. BACKGROUND

[0002] Lactic acid bacteria (LAB) are a class of gram-positive, facultative anaerobic bacteria that exist widely in nature, especially in animals and plants. As common probiotics, lactic acid bacteria can synthesize organic acids, hydrogen peroxide and bacteriocins and other antibacterial compounds in the metabolic process. These metabolic products can effectively inhibit the growth and reproduction of fungi and spore germination, thereby playing an important role in food preservation. Among them, Aspergillus niger is a common filamentous fungus with strong adaptability, which can survive in high-acid, high-salt or low-temperature environments, easily contaminate food such as grains and flour, cause food moldy, and cause economic losses and food safety problems. Aspergillus niger

[0003] At present, the prevention and control of Aspergillus niger mainly relies on chemical, physical and biological methods. Chemical prevention and control often uses potassium sorbate and other preservatives, but there are risks of residues, potential health effects and easy induction of drug resistance, and may change the flavor and texture of food; physical prevention and control such as low-temperature refrigeration and radiation treatment can inhibit fungal growth, but may cause poor food taste and destruction of nutritional ingredients, and is difficult to prevent secondary pollution; biological control uses antagonistic strains (such as Bacillus) or natural antibacterial substances, but most of the existing strains are screened from soil or fermented food (such as soy sauce and brewing lees), which have insufficient compatibility with food systems such as bread, and have a narrow antibacterial spectrum and limited application conditions (such as requiring an acidic environment).

[0004] Therefore, there is an urgent need in the art for an anti-aspergillus niger lactic acid bacteria strain and a screening method thereof that is safe, efficient and compatible with food systems to solve the problems of the prior art and provide a green and sustainable solution for food preservation. SUMMARY

[0005] Therefore, in order to solve the above technical problems, the present application provides an anti-aspergillus niger lactic acid bacteria strain and a screening method and application thereof, and the specific technical solutions are as follows: An anti-aspergillus niger lactic acid bacteria strain FOSU-ZX803, the classification name of the anti-aspergillus niger lactic acid bacteria strain FOSU-ZX803 is Plantarum, the preservation number is GDMCC NO. 66234, the preservation date is April 28, 2025, and it is preserved in the Guangdong Microbial Culture Collection Center. Lactiplantibacillus plantarum

[0006] ​​In addition, the application further provides a screening method of the anti-aspergillus niger lactic acid bacterial strain FOSU-ZX803, which comprises the following steps: S1. isolating strains from fermented food, preliminarily screening strains with transparent calcium-dissolving rings through a calcium-dissolving ring experiment, streak culturing to obtain strains A, B and C; S2. performing gram staining and hydrogen peroxide enzyme experiment on the strains A, B and C screened in step S1 to screen strains A, B and C which are gram-positive and hydrogen peroxide enzyme-negative; S3. performing anti-aspergillus niger primary screening through a double-layer streak method, measuring the area of the inhibition zone to screen strain A with significant inhibition effect; S4. performing anti-aspergillus niger secondary screening through a 96-well plate method, calculating the inhibition rate to screen strain A with high inhibition rate; S5. performing physiological and biochemical identification and 16S rRNA gene sequence analysis on the strain A screened in steps S3 and S4 to determine the classification of the strain, thereby obtaining the anti-aspergillus niger lactic acid bacterial strain FOSU-ZX803.

[0007] Further, in step S1, the fermented food is pickled vegetable water; The temperature of the streak culture is 35-38 DEG C, and the time is 24-28 h.

[0008] Further, in step S3, the double-layer streak method is as follows: parallel lines are drawn on the lower plate with strains A, B and C, and then the lines are cultured at 35-38 DEG C for 20-28 h; then the PDA semi-solid medium containing aspergillus niger spores is shaken and poured on the lower plate with strains A, B and C, and the concentration of the aspergillus niger spores is 1x10 5 CFU / mL-5x10 5 CFU / mL; then the culture is further cultured in a constant-temperature incubator at 26-30 DEG C for 20-28 h, and the area of the inhibition zone formed around the streaked area is measured.

[0009] Further, in step S3, the significant inhibition effect is that the area of the inhibition zone is greater than or equal to 11 cm 2 .

[0010] Further, in step S4, the cell-free supernatant of strain D is mixed with the aspergillus niger spore suspension in the 96-well plate, and then the mixture is cultured at 26-30 DEG C for 68-76 h; then the absorbance value of the culture solution at 600 nm is measured, and the inhibition rate is calculated according to the formula to screen strain A with high inhibition rate. Further, the strain with high inhibition rate is greater than or equal to 98%.

[0011] The application further provides an application of the anti-aspergillus niger lactic acid bacterial strain FOSU-ZX803, and the application is that the anti-aspergillus niger lactic acid bacterial strain FOSU-ZX803 is applied in the field of inhibiting aspergillus niger.

[0012] Further, the field is food products, food preparation, and food preservation.

[0013] Compared with the prior art, the present application has the following beneficial effects: 1. The plant Lactobacillus FOSU-ZX803 screened by the present application not only has extremely strong inhibitory ability and excellent probiotic properties and safety to Aspergillus niger, but also meets the requirements of food safety strains. The cell-free supernatant of the strain FOSU-ZX803 of the present application applied to bread preservation can significantly inhibit the growth of Aspergillus niger and effectively prolong the shelf life of bread.

[0014] 2. The strain obtained by initial screening from sour pickles is induced and cultured, which greatly improves the efficiency and success rate of screening, has more stable anti-Aspergillus niger activity, more significant performance, and higher application value. BRIEF DESCRIPTION OF DRAWINGS

[0015] Figure 1 It is a schematic diagram of the colony morphology of the strain in Example 1; Figure 2 It is a schematic diagram of the microscope of the strain of gram-positive bacteria; Figure 3 It is a schematic diagram of the phylogenetic tree; Figure 4 It is a schematic diagram of the growth curve and acid production curve; Figure 5 It is a schematic diagram of hemolytic activity; Figure 6 It is a schematic diagram of antibiotic activity results; Figure 7 It is the growth condition of the strain in Example 6 under different pH; Figure 8 It is the survival rate of the strain in Example 6 in simulated artificial gastric juice (A) and artificial intestinal juice (B); Figure 9 It is a schematic diagram of the antibacterial experiment of the strain in Example 6; Figure 10 It is a schematic diagram of the application test results. DETAILED DESCRIPTION

[0016] In order to make the purpose, technical scheme and advantages of the present application more clear and explicit, the present application will be further described in detail below in combination with its embodiments. It should be understood that the specific embodiments described herein are only used to explain the present application, and do not limit the protection scope of the present application.

[0017] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. The terminology used in the description herein is for describing particular embodiments only and is not intended to be limiting of the application. As used herein, the term "and / or" includes any and all combinations of one or more of the associated listed items.

[0018] In an embodiment of the present application, an anti-aspergillus niger lactic acid bacterial strain FOSU-ZX803 is provided, which is classified as Lactiplantibacillus plantarum (Lactobacillus plantarum) and has a classification name of Lactiplantibacillus plantarum (Lactobacillus plantarum), a preservation number of GDMCC NO. 66234, and a preservation date of April 28, 2025, and is preserved in the Guangdong Microbial Culture Collection Center. Lactiplantibacillus plantarum

[0019] In an embodiment, the present application also provides a screening method of an anti-aspergillus niger lactic acid bacterial strain FOSU-ZX803, which comprises the following steps: S1. Isolating strains from fermented food, preliminarily screening strains with transparent calcium-dissolving rings through a calcium-dissolving ring experiment, streak culturing to obtain strains A, B, and C; S2. Carrying out Gram staining and hydrogen peroxide enzyme experiment on the strains A, B, and C screened in step S1 to screen strains A, B, and C that are Gram-positive and hydrogen peroxide enzyme-negative; S3. Carrying out anti-aspergillus niger primary screening through a double-layer streak method, measuring the area of the inhibition zone to screen strain A with significant antibacterial effect; S4. Carrying out anti-aspergillus niger secondary screening through a 96-well plate method, calculating the antibacterial rate to screen strain A with high antibacterial rate; and S5. Carrying out physiological and biochemical identification and 16S rRNA gene sequence analysis on the strain A screened in steps S3 and S4 to determine the classification of the strain and obtain the anti-aspergillus niger lactic acid bacterial strain FOSU-ZX803.

[0020] In an embodiment, in step S1, the fermented food is acid pickled vegetable water. The temperature of the streak culture is 35-38°C, and the time is 24-28h.

[0021] In an embodiment, in step S3, the double-layer streak method is as follows: parallel lines are drawn on the lower plate with strains A, B, and C, and then cultured at 35-38°C for 20-28h; then the PDA semi-solid medium containing aspergillus niger spores is shaken and poured onto the lower plate with strains A, B, and C, and the concentration of the aspergillus niger spores is 1×10 5 CFU / mL-5×10 5 CFU / mL; and then cultured in a constant-temperature incubator at 26-30°C for 20-28h to measure the area of the inhibition zone formed around the streaked area of the strain.

[0022] ​In one of the embodiments, in step S3, the significant bacteriostatic effect is that the bacteriostatic circle area is greater than or equal to 11 cm 2 .

[0023] In one of the embodiments, in step S4, the cell-free supernatant of the strain D is mixed with the Aspergillus niger spore suspension in a 96-well plate, and after 68-76 h of culture at 26-30℃, the absorbance value of the culture solution at 600 nm is measured, and the bacteriostatic rate is calculated according to the formula to screen the strain A with high bacteriostatic rate. In one of the embodiments, the strain with a high bacteriostatic rate of greater than or equal to 98% is screened.

[0024] In one of the embodiments, the application of the anti-Aspergillus niger lactic acid bacteria strain FOSU-ZX803 is provided, and the application is the anti-Aspergillus niger lactic acid bacteria strain FOSU-ZX803 in the field of inhibiting Aspergillus niger.

[0025] In one of the embodiments, the field is food products, food preparation, and food preservation.

[0026] In one of the embodiments, the food preservation is preferably bread, and the cell-free supernatant of the plant lactic acid bacteria FOSU-ZX803 is sprayed on the surface of the bread.

[0027] The above scheme induces the anti-Aspergillus niger lactic acid bacteria strain FOSU-ZX803 to have a significant anti-Aspergillus niger effect, which can be applied to anti-Aspergillus niger products, especially the preservation of bread, and gives the food a longer shelf life.

[0028] The embodiments of the application will be described in detail below with specific examples.

[0029] Example 1: Strains were isolated from sour pickles, and strains with transparent calcium-dissolving circles were preliminarily screened by calcium-dissolving circle experiments, and streak culture was performed to obtain strains A, B, and C. Strains A, B, and C stored in glycerol were inoculated in MRS liquid medium at an inoculation amount of 5%, and incubated at 37℃ for 24 h. After continuous activation twice, they were inoculated in MRS agar plates for dilution and counting, and the concentration of the bacterial solution was adjusted to 1×10 8 CFU / mL.

[0030] As Figure 1 shown, Figure 1 is a schematic diagram of the colony morphology of the strains screened in Example 1, and A, B, and C are three groups.

[0031] Example 2: The strains A, B and C screened in Example 1 were subjected to gram staining and hydrogen peroxide enzyme test, and the gram-positive and hydrogen peroxide enzyme-negative strains A, B and C were screened.

[0032] As shown in Figure 2 , Figure 2 The schematic diagram of microscope observation of the strains A, B and C screened in Example 2 is shown in A, B and C.

[0033] Example 3 MRS agar medium was poured into a culture dish to make a lower plate, and two 2 cm long parallel lines were drawn on the lower plate using activated lactic acid bacteria, with a parallel line spacing of 1 cm. The culture dish was placed in a 37℃ constant temperature and humidity incubator for 24 h. Fungal spore concentration of 1x10 5 spores / mL was inoculated into the PDA semi-solid medium with a 3% inoculation amount, and then poured into the lower plate. After complete solidification, the culture dish was inverted and cultured in a 28℃ constant temperature incubator for 24 h. The software Image was used to measure the inhibition area, which was marked as strain A, strain B and strain C. The inhibition zone is shown in Table 1.

[0034] Table 1: Inhibition zone of strains A, B and C

[0035] Note: Different letters represent significant differences in the inhibition effect of the strains on Aspergillus niger (p<0.05) The double-layer streaking method was used for the initial screening of Aspergillus niger resistance, and the inhibition zone area was measured. The strain A with significant inhibition effect was screened. Example 4 In a 96-well plate, 190 μL of lactic acid bacteria cell-free supernatant and 10 μL of Aspergillus niger spore suspension (1x10 5 CFU / mL~5x10 5 CFU / mL) were added to the 96-well plate. The control group was 190 μL of fresh MRS broth medium with 10 μL of Aspergillus niger spore suspension. The culture was incubated at 28℃ for 72 h, with three replicates. After the incubation was completed, the absorbance value of the culture at OD 600nm was measured using a microplate reader, and the inhibition rate was calculated according to formula (1). The strain A with high inhibition rate was screened. The inhibition rate is shown in Table 3.

[0036] )×100 (1) Table 3: Inhibition rate of strains A, B and C

[0037] Note: Different letters represent significant differences in the inhibition effect of the strains on Aspergillus niger (p<0.05).

[0038] Example 5 The strain was subjected to physiological and biochemical identification and 16S rRNA gene sequence analysis to determine the classification of the strain, and the antifungal Aspergillus niger lactic acid bacteria strain FOSU-ZX803 was obtained.

[0039] After identification, the strain was determined to be Lactiplantibacillus plantarum Lactiplantibacillus plantarum , which was named Lactiplantibacillus plantarum FOSU-ZX803 (L.P. FOSU-ZX803), and was deposited with the Guangdong Microbial Culture Collection Center on April 28, 2025, with the strain deposit number GDMCC NO. 66234.

[0040] The physiological and biochemical identification results are shown in Table 4 below, Figure 3 and the phylogenetic tree of Lactiplantibacillus plantarum is shown in Figure 2.

[0041] Table 4: Physiological and biochemical identification results

[0042] Note: "-" represents negative; "+" represents positive Example 6 The strain in Example 5 was inoculated into MRS liquid medium at an inoculation amount of 5%, and incubated at 37°C. The OD 600nm and pH values of the bacterial solution at 0, 2, 4, 6, 8, 10, 12, 15, 18, 21, and 24 h were measured, and the growth and pH value change curves of the bacterial solution were drawn to reflect its growth and acid production capacity. The results are shown in Figure 4 .

[0043] Example 7 The strain in Example 5 was subjected to safety tests, including hemolytic activity, antibiotic sensitivity, and amino acid decarboxylase experiments. The hemolytic activity is shown in Figure 5 ; the antibiotic sensitivity set the strain L.P.9010 as the control group, and L.P.FOSU-ZX803 as the strain in Example 5, and the antibiotic activity results are shown in Table 4 and Figure 6 .

[0044] The hemolytic activity: the activated bacterial cells were streaked onto blood agar plates and incubated at 37°C for 24 h. The presence or absence of hemolytic rings around the colonies was observed. Staphylococcus aureus was used as a positive control. Hemolysis was classified as alpha hemolysis and beta hemolysis: alpha hemolysis was due to incomplete rupture of red blood cells forming a grass green hemolytic ring around the colonies, and beta hemolysis was due to complete lysis of red blood cells forming a clear and complete hemolytic ring around the colonies.

[0045] Antibiotic sensitivity test: the drug resistance of the strain was determined by paper disc diffusion method. The strain was cultured to 10 8 CFU / mL and 100 μL of the bacterial suspension was taken and uniformly coated on the surface of MRS agar plate. After the surface of the medium was dried, the drug sensitivity test paper patch was placed and gently pressed, and incubated at 37°C for 24 h. The diameter of the inhibition zone was measured by digital vernier caliper Amino acid decarboxylase test: the strain was cultured to 10 8 CFU / mL, 0.08 mL was taken and added into arginine decarboxylase, ornithine decarboxylase, lysine decarboxylase and amino acid decarboxylase control tubes, respectively, covered with 0.2 ml of sterile liquid paraffin, and incubated at 37°C for 24 h. The color change was observed. If the control tube and the experimental tube were both yellow, it was negative, and if the control tube was yellow and the experimental tube was yellow-green, it was positive.

[0046] Table 4: Antibiotic sensitivity results

[0047] In Table 4: sensitive (S); moderately sensitive (I); resistant (R).

[0048] Table 5: Amino acid decarboxylase test results

[0049] As can be seen from Table 4 and Table 5, the strain of the present application has excellent results, which shows that L.P. FOSU-ZX803 and the control strain L.P. 9010 are both γ-hemolytic and do not have hemolytic activity; they are sensitive or moderately sensitive to β-lactam antibiotics and have resistance to aminoglycoside antibiotics; and they do not have ornithine, arginine and lysine amino acid decarboxylases.

[0050] Example 8: The strains in Example 5 were subjected to probiotic activity determination, including the growth conditions of lactic acid bacteria at different pH, survival rate in simulated artificial gastric juice and artificial intestinal juice, and bacteriostatic experiment. The results are shown in Figure 7~9 and Table 6, wherein, Figure 7 is the growth condition of the strain of Example 5 at different pH; Figure 8 is the survival rate of the strain of Example 5 in simulated artificial gastric juice (A) and artificial intestinal juice (B), Figure 9 is a schematic diagram of the bacteriostatic experiment of the strain of Example 5.

[0051] Among them, the growth conditions at different pH: the bacterial suspension was inoculated in MRS liquid medium at pH 2, pH 4 and pH 6 at a transfer amount of 5%, and incubated at 37°C. The OD 600nmThe growth curve of the bacteria was drawn by plotting the viable cell count, reflecting the growth status of the bacteria at different pH values.

[0052] Survival rate in simulated artificial gastric juice and artificial intestinal juice: the strain was inoculated in 5 mL MRS liquid medium and cultured at 37°C for 24 h, then centrifuged at 8000 rpm for 15 min, and the supernatant was discarded. The bacterial slurry was washed twice with sterile PBS and resuspended. Simulated artificial gastric juice: 0.3% pepsin was added to PBS buffer, adjusted to pH 2.5, and then sterilized by filtration. Simulated artificial intestinal juice: 0.1% (w / v) trypsin and 0.3% (w / v) bovine bile salt were added to PBS buffer, adjusted to pH 8.0, and then sterilized by filtration. 1 mL of bacterial suspension was added to 9 mL of simulated artificial gastric juice, mixed well, and then incubated at 37°C. At 0 h, 1 h, 2 h, and 3 h, the culture was diluted with sterile saline, and then plated on MRS agar medium. After incubation at 37°C for 24 h, the number of colonies was counted, and the survival rate was calculated. 1 mL of the culture treated in simulated artificial gastric juice for 3 h was inoculated in 9 mL of simulated artificial intestinal juice, and then incubated at 37°C. The survival rate was calculated at 0 h, 1 h, 2 h, and 3 h.

[0053] ) × 100% In the formula, N1 is the viable cell count after 1 h, 2 h, and 3 h of treatment with simulated artificial gastric juice and intestinal juice, and N2 is the viable cell count after 0 h of treatment with simulated artificial gastric juice and intestinal juice.

[0054] Inhibition of pathogenic bacteria: the inhibition of the strain was determined by agar diffusion method. Escherichia coli, Salmonella, Listeria monocytogenes, and Staphylococcus aureus were used as indicator bacteria. The OD value of the pathogenic bacteria was adjusted to 0.5, and the pathogenic bacteria were added to sterile and cooled LB agar medium to 50°C at a concentration of 0.2%. After the surface of the culture dish was dry, a 9 mm puncher was used to punch a hole, and 100 μL of bacterial suspension, culture medium, and cell-free supernatant of lactic acid bacteria were added to the hole. Sterile water was used as a blank group, and each group had 3 replicates. The liquid was allowed to diffuse into the agar, and then incubated at 37°C for 24 h. The diameter of the inhibition zone was measured.

[0055] It should be noted that strain L.P.9010 was used as a control group, and L.P.FOSU-ZX803 was the strain in Example 5.

[0056] Table 6: Inhibition experiment

[0057] Note: Different capital letters represent significant differences (p<0.05) in the same strain-related liquid under different pathogenic bacteria; lowercase letters represent significant differences (p<0.05) in the same pathogenic bacteria in different strain-related liquids.

[0058] From Figure 7~8 And the data analysis of Table 6 shows that the strain of the application has excellent antibacterial activity.

[0059] Example 9: Application test: Take an equal amount of bread samples, set up a control group and a treatment group. Control group: use a watering can to evenly spray 100 muL of Aspergillus niger spore suspension (Aspergillus niger spore suspension concentration is 2x10 6 CFU / mL) and 400 muL of sterile distilled water into each sample; treatment group: take bread samples with the same specifications as the control group, and also add 100 muL of Aspergillus niger spore suspension and 400 muL of cell-free supernatant of lactic acid bacteria (strains of the application). Place the above treated control group and treatment group samples in a 28 degree incubator and continuously observe for 5 days. The results are shown in Figure 10

[0060] Figure 10 The application test results are shown in the schematic diagram, and it can be clearly seen from Figure 10 that the bread treated by the strain of the application has a longer shelf life and can effectively resist Aspergillus niger.

[0061] The technical features of the above-mentioned examples can be combined in any way. In order to make the description concise, not all possible combinations of technical features in the above-mentioned examples are described, but as long as the combination of technical features does not exist contradictory, it should be considered as the scope of the present application.

[0062] The above-mentioned examples only express several embodiments of the application, and the description is more specific and detailed, but it should not be understood as a limitation on the scope of the patent. It should be pointed out that for ordinary skilled in the art, without departing from the concept of the present application, some modifications and improvements can be made, which are within the scope of the present application. Therefore, the protection scope of the patent of the present application should be subject to the appended claims.​

Claims

1. A strain of lactic acid bacteria resistant to Aspergillus niger, FOSU-ZX803, characterized in that, The Aspergillus niger-resistant lactic acid bacteria strain FOSU-ZX803 is classified as *Lactobacillus plantarum* (…). Lactiplantibacillusplantarum The accession number is GDMCCNO. 66234, the accession date is April 28, 2025, and it is deposited at the Guangdong Provincial Center for Microbial Culture Collection.

2. A method for screening Aspergillus niger-resistant lactic acid bacteria strain FOSU-ZX803, characterized in that, The screening method is used to screen for the Aspergillus niger-resistant lactic acid bacteria strain FOSU-ZX803 as described in claim 1, and the screening method includes the following steps: S1. Strains were isolated from fermented foods. Strains with transparent calcium dissolution zones were initially screened using a calcium dissolution zone experiment. Strand culture was then performed to obtain strains A, B, and C. S2. Gram staining and catalase experiments were conducted on strains A, B, and C screened in step S1 to screen for Gram-positive and catalase-negative strains A, B, and C. S3. A double-stripe method was used for initial screening against Aspergillus niger. The area of ​​the inhibition zone was measured, and strain A with significant antibacterial effect was selected. S4. A 96-well plate method was used for secondary screening against Aspergillus niger. The inhibition rate was calculated, and strain A with a high inhibition rate was selected. S5. Physiological and biochemical identification and 16S rRNA gene sequence analysis were performed on strain A selected in steps S3 and S4 to determine the bacterial species classification, resulting in the Aspergillus niger-resistant lactic acid bacteria strain FOSU-ZX803.

3. The screening method according to claim 2, characterized in that, In step S1, the fermented food is pickled vegetable brine; The streak incubation was carried out at a temperature of 35-38°C for 24-28 hours.

4. The screening method according to claim 2, characterized in that, In step S3, the double-layer streak method is as follows: Strawberries A, B, and C are streaked parallel to each other on the lower plate and incubated at 35-38℃ for 20-28 hours. Then, the PDA semi-solid medium containing Aspergillus niger spores is shaken well and poured onto the lower plate containing strains A, B, and C. The concentration of Aspergillus niger spores is 1×10⁻⁶. 5 CFU / mL ~5×10 5 CFU / mL; then continue to incubate in a constant temperature incubator at 26~30℃ for 20~28h, and measure the area of ​​the inhibition zone formed around the streaked area of ​​the strain.

5. The screening method according to claim 2, characterized in that, In step S3, the antibacterial effect is significant when the area of ​​the inhibition zone is ≥11cm². 2 .

6. The screening method according to claim 2, characterized in that, In step S4, the cell-free supernatant of strain D was mixed with the Aspergillus niger spore suspension in a 96-well plate and incubated at 26-30°C for 68-76 hours. The OD value of the culture medium was then measured. 600nm The absorbance value at the specified location was used to calculate the antibacterial rate, and strain A with a high antibacterial rate was selected.

7. The screening method according to claim 6, characterized in that, Screen for strains with an antibacterial rate of ≥98%.

8. The application of a strain of lactic acid bacteria resistant to Aspergillus niger, FOSU-ZX803, characterized in that, The application is the use of the Aspergillus niger-resistant lactic acid bacteria strain FOSU-ZX803 as described in claim 1 in the field of inhibiting Aspergillus niger.

9. The application according to claim 8, characterized in that, The field in question is food products, food preparation, and food preservation.

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