Strain XY-144 as well as fermentation supernatant and application thereof

Abstract

The invention relates to the technical field of microorganisms, and discloses a strain XY-144 as well as fermentation supernate and application thereof. Wherein the Latin classification name of the strain XY-144 is Fangia hongkongensis, the preservation unit is China General Microbiological Culture Collection Center, the preservation address is No.3, No.1 Yard, Beichen West Road, Chaoyang District, Beijing, the preservation number is CGMCC No: 31090, and the preservation date is June 26, 2024. Fermentation supernate prepared from the strain XY-144 can obviously inhibit formation of a methicillin-resistant staphylococcus aureus biological membrane and remove the methicillin-resistant staphylococcus aureus biological membrane, the thermal stability of the fermentation supernate is good, and antibacterial activity and toxicity are not found. Therefore, the strain XY-144 and its fermentation supernatant are safe and reliable in application in food industry, and have great advantages in food application.

Description

Technical Field

[0001] The invention relates to the technical field of microorganisms, and in particular to a strain XY-144 and a fermentation supernatant thereof and application thereof. Background Art

[0002] Food safety has long been one of the main factors affecting public health, and the incidence of foodborne diseases caused by microbial pathogens has been on the rise worldwide. Methicillin-resistant Staphylococcus aureus (MRSA) is a common foodborne pathogen, and the foodborne diseases caused by it have become a global public health issue.

[0003] Currently, the prevention and control strategies for foodborne diseases mainly use microbial disinfectants or exposure to low doses of ultraviolet A (UVA). However, direct killing methods may cause problems such as the development of antibiotic resistance. In addition, these foodborne pathogens often have the ability to form biofilms, which can protect the pathogens from the cleaning process and continue to survive in the environment.

[0004] Some strategies used in the food industry to control biofilms, such as cleaning and disinfection, plasma treatment, and ultrasonic treatment, are effective but still difficult to completely control biofilms. Therefore, more effective and sustainable strategies are urgently needed to combat foodborne pathogens. Summary of the invention

[0005] In view of the above-mentioned deficiencies in the prior art, the present invention provides a strain XY-144 capable of removing biofilm or inhibiting biofilm formation and its fermentation supernatant, aiming to solve the problem of the lack of more effective and sustainable strategies in the prior art to combat foodborne pathogens.

[0006] The technical solution of the present invention is as follows:

[0007] In the first aspect, a strain XY-144 is provided. The Latin classification name of the strain XY-144 is Fangia hongkongensis. The preservation unit is the General Microbiology Center of the China General Microbiological Culture Collection Administration. The preservation address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing. The preservation number is CGMCC No: 31090 and the preservation date is June 26, 2024.

[0008] In a second aspect, a fermentation supernatant of strain XY-144 is provided, wherein the fermentation supernatant is prepared by fermenting the strain XY-144 as described in the first aspect.

[0009] A preferred technical solution is that the method for preparing the fermentation supernatant comprises the steps of:

[0010] A single colony of strain XY-144 was placed in a SGTYP liquid culture medium and cultured by shaking to obtain a fermentation broth; the fermentation broth was centrifuged to obtain the fermentation supernatant.

[0011] According to a further preferred technical solution, the shaking culture conditions are: temperature 20-37°C, rotation speed 100-300 rpm, and time 2-4 days.

[0012] According to a further preferred technical solution, the centrifugal treatment conditions are: temperature 0-10°C, rotation speed 8000-12000 rpm, and time 10-30 minutes.

[0013] In a third aspect, there is provided a use of the strain XY-144 as described in the first aspect or the fermentation supernatant as described in the second aspect in the preparation of a product for removing biofilm.

[0014] In a preferred technical solution, the biofilm removal product is specifically a product for removing methicillin-resistant Staphylococcus aureus biofilm.

[0015] In a fourth aspect, there is provided a use of the strain XY-144 as described in the first aspect or the fermentation supernatant as described in the second aspect in the preparation of a product for inhibiting biofilm formation.

[0016] In a preferred technical solution, the product for inhibiting biofilm formation is specifically a product for inhibiting biofilm formation of methicillin-resistant Staphylococcus aureus.

[0017] Beneficial effects: The present invention provides a new strain XY-144, which is classified and named Fangiahongkongensis. The fermentation supernatant prepared by the strain can significantly inhibit the formation of methicillin-resistant Staphylococcus aureus biofilm and remove the methicillin-resistant Staphylococcus aureus biofilm, and has good thermal stability, and can better maintain activity and efficacy in practical applications. In addition, the experimental results show that the fermentation supernatant of XY-144 has no antibacterial activity and is not easy to cause pathogens to develop drug resistance; at the same time, it is not toxic and has no effect on the germination rate and sprout length of alfalfa seeds; therefore, the application of strain XY-144 and its fermentation supernatant in the food industry is safe and reliable, which has great advantages for food applications. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 It is the evolutionary tree of strain XY-144 and the reference strain in Example 1.

[0019] Figure 2 This is a graph showing the antibacterial activity test results of the fermentation supernatant of XY-144 against MRSA in Example 2.

[0020] Figure 3This is a diagram showing the verification results of the fermentation supernatant of XY-144 in Example 3 for the removal and inhibition activity of MRSA biofilm.

[0021] Figure 4 This is a graph showing the results of the activity verification of each component of the fermentation supernatant of XY-144 in Example 4 in removing MRSA biofilm.

[0022] Figure 5 This is a graph showing the thermal stability test results of the fermentation supernatant of XY-144 in Example 5.

[0023] Figure 6 This is a graph showing the toxicity test results of the salting-out component of XY-144 in Example 6. DETAILED DESCRIPTION

[0024] The present invention provides strain XY-144 and fermentation supernatant thereof and application thereof. To make the purpose, technical scheme and effect of the present invention clearer and more specific, the present invention is further described in detail below.

[0025] Biofilm is a polymer of microorganisms. Biofilm can enhance the resistance or tolerance of bacteria to adverse environmental conditions. The extracellular polymer (EPS) matrix can limit the diffusion of antimicrobial drugs. The formation of biofilm greatly enhances the survival ability of bacteria. Compared with inhibiting the formation of biofilm, it is particularly difficult to remove mature biofilm. Studies have found that the formation of biofilm is positively correlated with the increase in EPS production. The destruction of EPS can achieve the effect of anti-biofilm. Therefore, the development of anti-biofilm with quorum sensing quenching as the breakthrough point has certain prospects. At present, the methods for removing biofilm are mainly some plant-derived active products, new antimicrobial peptides, surfactants, quorum sensing inhibitors or quorum sensing quenching enzymes. There are few reports on active substances for removing biofilm from marine microorganisms, and even fewer active substances for fighting foodborne pathogens.

[0026] Based on this, the embodiment of the present invention provides strain XY-144, the Latin classification name of the strain XY-144 is Fangia hongkongensis, the preservation unit is the General Microbiology Center of the China Microorganism Culture Collection Administration, the preservation address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, the preservation number is CGMCC No: 31090, and the preservation date is June 26, 2024.

[0027] Specifically, the inventors collected a sample of a sea cow from the shallow waters of Shenzhen Dapeng Nature Reserve and isolated strains to obtain about 200 strains. After screening the isolated strains for activity, they found that strain XY-144 can produce substances that can remove biofilms and inhibit biofilm formation. Comparison analysis of 16S rRNA sequences and whole genome sequences showed that the closest evolutionary relationship of strain XY-144 is Fangia hongkongensis FSC776 T The similarity of 16S rRNA sequence was 99.37% and the core gene homology was 78%, so it was speculated that XY-144 was a new strain.

[0028] The embodiment of the present invention provides a fermentation supernatant of strain XY-144, wherein the fermentation supernatant is prepared by fermenting the strain XY-144 as described above.

[0029] In one embodiment, the method for preparing the fermentation supernatant comprises the steps of:

[0030] A single colony of strain XY-144 was placed in a SGTYP liquid culture medium and cultured by shaking to obtain a fermentation broth; the fermentation broth was centrifuged to obtain the fermentation supernatant.

[0031] In a more specific embodiment, the shaking culture conditions are: temperature 20-37° C., rotation speed 100-300 rpm, and time 2-4 days.

[0032] In a more specific embodiment, the centrifugal treatment conditions are: temperature 0-10°C, rotation speed 8000-12000 rpm, time 10-30 minutes.

[0033] The embodiments of the present invention provide the use of the strain XY-144 or the fermentation supernatant described above in preparing a product for removing biofilm.

[0034] In one embodiment, the biofilm removal product is specifically a product for removing methicillin-resistant Staphylococcus aureus biofilm.

[0035] The embodiments of the present invention provide the use of the strain XY-144 or the fermentation supernatant described above in preparing a product that inhibits biofilm formation.

[0036] In one embodiment, the product for inhibiting biofilm formation is specifically a product for inhibiting biofilm formation of methicillin-resistant Staphylococcus aureus.

[0037] The present invention will be further described below by means of specific examples.

[0038] In the embodiment of the present invention, the culture medium involved is prepared with deionized water, and the formula is as follows:

[0039] LB liquid culture medium: tryptone 10 g / L, yeast powder 1 g / L, sodium chloride 5 g / L.

[0040] LB solid medium: tryptone 10 g / L, yeast powder 1 g / L, sodium chloride 5 g / L, agar 15 g / L.

[0041] MA solid medium: peptone 5g / L, yeast extract 1g / L, ferric citrate 0.1g / L, sodium chloride 19.45g / L, magnesium chloride 5.98g / L, sodium sulfate 3.24g / L, calcium chloride 1.8g / L, potassium chloride 0.55g / L, sodium carbonate 0.16g / L, potassium bromide 0.08g / L, strontium chloride 0.034g / L, boric acid 0.022g / L, sodium silicate 0.004g / L, sodium fluoride 0.0024g / L, sodium nitrate 0.0016g / L, disodium hydrogen phosphate 0.008g / L, agar 15g / L.

[0042] SGTYP liquid culture medium: glucose 5 g / L, soluble starch 5 g / L, tryptone 1 g / L, yeast extract 1 g / L, peptone 1 g / L.

[0043] SGTYP solid culture medium: glucose 5g / L, soluble starch 5g / L, tryptone 1g / L, yeast extract 1g / L, peptone 1g / L, agar 15g / L.

[0044] In the embodiment of the present invention, the preparation process of the fermentation supernatant of XY-144 and the overnight seed solution of MRSA is as follows:

[0045] Fermentation supernatant of XY-144: Scrape a small amount of frozen bacterial cells of strain XY-144 onto MA solid medium for streaking and culture at 28°C for 3 days. Take a single colony and place it in 3 mL of SGTYP liquid medium, shake culture at 28°C and 200 rpm for 3 days, and centrifuge at 10,000 rpm for 20 minutes at 4°C to obtain the fermentation supernatant of XY-144.

[0046] Overnight seed solution of MRSA: Scrape a small amount of frozen cells of strain MRSA onto LB solid medium for streaking and culture at 37°C for 1 day. Take a single colony and place it in 3 mL of LB liquid medium, shake and culture overnight at 37°C and 200 rpm to obtain the overnight seed solution of MRSA.

[0047] Example 1 Isolation and identification of strain XY-144

[0048] Strain XY-144 was collected from Shenzhen Dapeng Nature Reserve in March 2017 and is a symbiotic bacterium of the marine invertebrate Dugong.

[0049] The DNA of XY-144 was extracted using TIANamp Bacteria DNA Kit (TIANGEN), and the 16S rDNA region of XY-144 was amplified by PCR using conventional forward primer 27F and reverse primer 1492R. The samples were sent for sequencing to obtain the 16S rRNA sequence data of XY-144. After comparison analysis, the 16S rRNA sequence of XY-144 was similar to that of Fangia hongkongensis FSC776. T The similarity of 16S rRNA sequences of XY-144 was 99.37%, so XY-144 was identified as Fangia hongkongensis. The 16S rRNA sequence of XY-144 has been submitted to the GenBank database with the accession number OR262794.1.

[0050] The DNA of XY-144 was extracted using TIANamp Bacteria DNA Kit (TIANGEN) and sent for sequencing to obtain the whole genome data of XY-144. The genome evolution tree was established based on the sequencing results. Figure 1 As shown. Figure 1 It can be seen that XY-144 belongs to the genus Fangia, and the closest evolutionary strain is Fangia hongkongensis FSC776 T The core gene homology was 78%, so it was speculated that XY-144 was a new strain.

[0051] XY-144 was deposited in the China General Microbiological Culture Collection Center (CGMCC) on June 26, 2024. The deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, and the deposit number is CGMCC No: 31090.

[0052] Example 2 Antibacterial activity of strain XY-144

[0053] The overnight seed solution of MRSA was diluted 1000 times with LB liquid medium, and then 40 μL was spread on the LB solid medium, and then a circular paper was placed on it. 5 μL of each of the fermentation supernatant of XY-144, SGTYP liquid medium (negative control), and SGTYP liquid medium containing 5 μg / mL vancomycin (positive control) were spotted on the circular paper, and then cultured at 37°C for 1 day. The experimental results are as follows: Figure 2 shown. Figure 2 In the figure, 1 refers to the addition of the fermentation supernatant of XY-144; 2 refers to the addition of SGTYP liquid culture medium; 3 refers to the addition of SGTYP liquid culture medium containing 5 μg / mL vancomycin. Figure 2 It can be seen that the fermentation supernatant of XY-144 has no antibacterial activity against MRSA.

[0054] Example 3 Verification of biofilm removal and inhibitory activity of strain XY-144

[0055] The overnight seed solution of MRSA was diluted 100 times by volume with LB liquid medium containing 0.5% (w / v) glucose, and 1 mL / well was added to a 24-well plate. After culturing at 37°C for 1 day, the bacterial solution was slowly aspirated with a spray gun to obtain a 24-well plate containing MRSA biofilm. The fermentation supernatant of XY-144 was diluted with LB liquid medium containing 0.5% (w / v) glucose to 75%, 50%, 25%, 12.5%, and 6.25% of the fermentation supernatant of XY-144, and different concentrations of dilutions were obtained. The dilution was slowly added to a 24-well plate containing MRSA biofilm at 1 mL / well. The SGTYP liquid medium was also diluted in the same proportion and added as a negative control. After culturing at 37°C for 1 day, the bacterial solution was slowly aspirated with a spray gun, washed once with saline, dried at 37°C for 30 minutes, and then fixed at 65°C for 30 minutes before being taken out and cooled to room temperature. Each well was stained with 1 mL of 0.1% (w / v) crystal violet solution. After 5 minutes, the stain was washed off and dried. The removal effect of the biofilm was observed. Figure 3 As shown in (a). Figure 3 In (a), 144 75% refers to the addition of a dilution containing 75% (v / v) XY-144 in a fermentation supernatant; 144 50% refers to the addition of a dilution containing 50% (v / v) XY-144 in a fermentation supernatant; 144 12.5% ​​refers to the addition of a dilution containing 12.5% ​​(v / v) XY-144 in a fermentation supernatant; 144 6.25% refers to the addition of a dilution containing 6.25% (v / v) XY-144 in a fermentation supernatant; SG 75% refers to the addition of a dilution containing 75% (v / v) SGTYP liquid culture medium; SG 50% refers to the addition of a dilution containing 50% (v / v) SGTYP liquid culture medium; SG 12.5% ​​refers to the addition of a dilution containing 12.5% ​​(v / v) SGTYP liquid culture medium; SG 6.25% refers to the addition of a diluent containing 6.25% (v / v) SGTYP liquid culture medium. Figure 3 As shown in (a), the fermentation supernatant of XY-144 has a clearing effect on MRSA biofilm, and the clearing effect increases with the increase in the proportion of fermentation supernatant.

[0056] The overnight seed solution of MRSA was diluted 100 times by volume with LB liquid medium containing 0.5% (w / v) glucose, and 1 mL / well was added to a 24-well plate to obtain a 24-well plate containing MRSA. The fermentation supernatant of XY-144 was diluted with LB liquid medium containing 0.5% (w / v) glucose to 75%, 50%, 25%, 12.5%, and 6.25% of the fermentation supernatant of XY-144, and dilutions of different concentrations were obtained. The dilution was slowly added to the 24-well plate containing MRSA at 1 mL / well, and the SGTYP liquid medium was also diluted in the same proportion and added as a negative control. After culturing at 37°C for 1 day, the bacterial solution was slowly aspirated with a spray gun, washed once with saline, dried at 37°C for 30 minutes, and then fixed at 65°C for 30 minutes and then taken out and cooled to room temperature. Each well was stained with 1 mL of 0.1% (w / v) crystal violet solution. After 5 minutes, the stain was washed off and dried. The biofilm inhibition effect was observed. Figure 3 As shown in (b). Figure 3 In (b), 144 75% refers to the addition of a dilution containing 75% (v / v) XY-144 in the fermentation supernatant; 144 50% refers to the addition of a dilution containing 50% (v / v) XY-144 in the fermentation supernatant; 144 12.5% ​​refers to the addition of a dilution containing 12.5% ​​(v / v) XY-144 in the fermentation supernatant; 144 6.25% refers to the addition of a dilution containing 6.25% (v / v) XY-144 in the fermentation supernatant; SG 75% refers to the addition of a dilution containing 75% (v / v) SGTYP liquid culture medium; SG 50% refers to the addition of a dilution containing 50% (v / v) SGTYP liquid culture medium; SG12.5% ​​refers to the addition of a dilution containing 12.5% ​​(v / v) SGTYP liquid culture medium; SG 6.25% refers to the addition of a diluent containing 6.25% (v / v) SGTYP liquid culture medium. Figure 3 As shown in (b), the fermentation supernatant of XY-144 has an inhibitory effect on the formation of MRSA biofilm, and the inhibitory effect increases with the increase in the proportion of the fermentation supernatant.

[0057] Example 4 Isolation and purification of active ingredients from strain XY-144

[0058] Tris-HCl buffer (1M, pH 8) was added to the fermentation supernatant of XY-144 until the volume of Tris-HCl buffer was 10%, and then ammonium sulfate powder was slowly added until its mass concentration was 50%, and then placed at 4°C overnight to precipitate protein to obtain a precipitated protein solution. The precipitated protein solution was centrifuged at 10000 rpm for 20 minutes at 4°C to remove the supernatant, and then heated with ddH 2O to resuspend the precipitate to obtain a precipitate sample. The precipitate sample was subjected to salting out. Before salting out, the dialysis bag (Biyuntian, 44 mm, 14 kDa) was sequentially treated with a dialysis bag treatment solution (2% (w / v) NaHCO 3 , 1M EDTA, pH 8), ddHO 2 O, 1M EDTA and ddH 2 O for 10 minutes and then cool. After leak testing, add the above precipitate sample into the dialysis bag and dialysis with ddH 2 O was used as the dialysate and dialyzed on a horizontal shaker at 80 rpm for 20 hours, during which the dialysate was replaced 3 times. After the last replacement, the dialysate was dialyzed for another 2 hours to obtain the XY-144 salting-out fraction.

[0059] The XY-144 salting-out fraction was ultrafiltered and concentrated using an ultrafiltration tube (Millipore, 3 kDa). The filtrate and concentrate obtained were recorded as XY-144 filtrate and XY-144 ultrafiltration fraction, respectively. The XY-144 ultrafiltration fraction was then further purified by anion exchange column. The DEAE homogenate resin was loaded into the column (column volume was 3 mL), and 3 times the column volume of 2M NaCl solution, ddH 2 The column was washed with 20 mM Tris-HCl buffer (pH = 8) with 3 column volumes, and then equilibrated with 20 mM Tris-HCl buffer (pH = 8) with 3 column volumes, and then 6 mL of XY-144 ultrafiltration fraction was loaded. The XY-144 loading flow-through was obtained by washing with 20 mM Tris-HCl (pH 8) solution containing 0.1 M NaCl with 3 column volumes, and then eluted with 20 mM Tris-HCl (pH 8) solution containing 0.1 M, 0.25 M, 0.5 M, and 1 M NaCl with 3 column volumes, and XY-144 elution fractions were obtained, which were recorded as E and E respectively. 0.1 、E 0.25 、E 0.5 and E 1 ; Finally, the column was regenerated with 4 column volumes of 20 mM Tris-HCl (pH 8) solution containing 2 M NaCl.

[0060] The collected components and SGTYP liquid culture medium (negative control) were diluted with LB liquid culture medium containing 0.5% (w / v) glucose to verify the activity of removing MRSA biofilm. The specific operation is shown in Example 3. The experimental results are shown in Figure 4 shown. Figure 4In (a), salting out 20% refers to the addition of a diluent containing 20% ​​(v / v) XY-144 salting out component; salting out 10% refers to the addition of a diluent containing 10% (v / v) XY-144 salting out component; ultrafiltration 20% refers to the addition of a diluent containing 20% ​​(v / v) XY-144 ultrafiltration component; ultrafiltration 10% refers to the addition of a diluent containing 10% (v / v) XY-144 ultrafiltration component; negative control 20% refers to the addition of a diluent containing 20% ​​(v / v) SGTYP liquid culture medium; negative control 10% refers to the addition of a diluent containing 10% (v / v) SGTYP liquid culture medium; filtrate 10% refers to the addition of a diluent containing 10% (v / v) XY-144 filtrate; and the blank control did not add any component. Figure 4 In (b), FT 10% refers to the addition of a diluent containing 10% (v / v) XY-144 load flowthrough; E 0.1 5% means adding elution fraction E containing 5% (v / v) XY-144 0.1 dilution; E 0.25 5% means adding elution fraction E containing 5% (v / v) XY-144 0.25 dilution; E 0.5 5% means adding elution fraction E containing 5% (v / v) XY-144 0.5 dilution; E 1 5% means adding elution fraction E containing 5% (v / v) XY-144 1 The blank control had no components added. Figure 4 It can be seen that the XY-144 salting-out fraction after salting-out purification and the XY-144 ultrafiltration fraction after salting-out ultrafiltration concentration still have a clearing effect on MRSA biofilm. However, the XY-144 filtrate after ultrafiltration did not show a clearing effect on MRSA biofilm. The results of anion exchange purification of the ultrafiltration fractions showed that among the XY-144 elution fractions after the DEAE column, only the XY-144 elution fraction E eluted with 20mM Tris-HCl (pH8) solution containing 0.25M NaCl 0.25 It has activity in clearing MRSA biofilm.

[0061] Example 5 Thermal stability test of active ingredients of strain XY-144

[0062] The fermentation supernatant of XY-144 was incubated at 28°C, 40°C, 60°C, 80°C and 100°C for 30 minutes, cooled to room temperature, and then the activity of removing MRSA biofilm was verified. The overnight seed solution of MRSA was diluted 100 times by volume with LB liquid culture medium containing 0.5% (w / v) glucose, and 1 mL / well was added to a 24-well plate. After culturing at 37°C for 1 day, the bacterial solution was slowly sucked away with a spray gun to obtain a 24-well plate containing MRSA biofilm. The fermentation supernatant of XY-144 treated at different temperatures was diluted with LB liquid medium containing 0.5% (w / v) glucose to a volume ratio of 50% of the fermentation supernatant of XY-144 treated, and then 1 mL / well was slowly added to the 24-well plate containing MRSA biofilm. The SGTYP liquid medium was also diluted in the same proportion and added as a negative control. After culturing at 37°C for 1 day, the bacterial solution was slowly aspirated with a spray gun, washed once with saline, dried at 37°C for 30 minutes, and then fixed at 65°C for 30 minutes before being taken out and cooled to room temperature. 1 mL of 0.1% (w / v) crystal violet solution was added to each well for staining, and the stain was washed off after 5 minutes and dried. The removal effect of the biofilm was as follows: Figure 5 Then, 5 mL of 30% (v / v) acetic acid solution was used to dissolve the pigment, and 100 μL was taken after mixing to measure OD using an enzyme marker. 550 , the test results are as follows Figure 5 As shown in (b). Figure 5 It can be seen that the fermentation supernatant of XY-144 has good thermal stability and can eliminate MRSA after incubation at 60°C for 30 minutes... The activity of the biofilm is still very good.

[0063] Example 6 Toxicity test of active ingredients of strain XY-144

[0064] Tris-HCl buffer (1M, pH 8) was added to the fermentation supernatant of XY-144 until the volume of Tris-HCl buffer was 10%, and then ammonium sulfate powder was slowly added until its mass concentration was 50%, and then placed at 4°C overnight to precipitate protein to obtain a precipitated protein solution. The precipitated protein solution was centrifuged at 10000 rpm for 20 minutes at 4°C to remove the supernatant, and then heated with ddH 2 O to resuspend the precipitate to obtain a precipitate sample. The precipitate sample was subjected to salting out. Before salting out, the dialysis bag (Biyuntian, 44 mm, 14 kDa) was sequentially treated with a dialysis bag treatment solution (2% (w / v) NaHCO 3 , 1M EDTA, pH 8), ddHO 2 O, 1M EDTA and ddH 2 O for 10 minutes and then cool. After leak testing, add the above precipitate sample into the dialysis bag and dialysis with ddH 2O was used as the dialysate and dialyzed on a horizontal shaker at 80 rpm for 20 hours. During this period, the dialysate was replaced 3 times. After the last change of the liquid, the dialysate was dialyzed for another 2 hours to obtain the XY-144 salting-out component. After soaking alfalfa seeds in water, rinse them continuously with water on the filter screen, then soak them in sodium hypochlorite (84 disinfectant diluted 10 times) for 10 minutes, wash them with sterile water 4 to 5 times, then air-dry them and put them into a 50 mL EP tube, add a desiccant and store them for later use. Weigh 50 seeds in a culture dish, add 20 mL of sterile water containing 10% (v / v) XY-144 salting-out component, and culture them at 25°C. The negative control group was 20 mL of sterile water. Observation and recording were performed at the 24th and 48th hours. The results are as follows Figure 6 As shown. Figure 6 It can be seen that the salting-out component of XY-144 has no effect on the germination of alfalfa, whether it is the germination rate or the length of the sprout, and is not toxic.

[0065] It should be understood that the application of the present invention is not limited to the above examples. For ordinary technicians in this field, improvements or changes can be made based on the above description. All these improvements and changes should fall within the scope of protection of the claims attached to the present invention.

Claims

1. Strain XY-144, characterized in that The Latin classification name of the strain XY-144 is Fangia hongkongensis. The preservation unit is the General Microbiology Center of China Microbiological Culture Collection Administration. The preservation address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing. The preservation number is CGMCC No: 31090 and the preservation date is June 26, 2024.

2. The fermentation supernatant of strain XY-144, characterized in that The fermentation supernatant is prepared by fermenting the strain XY-144 as claimed in claim 1.

3. The fermentation supernatant according to claim 2, characterized in that The method for preparing the fermentation supernatant comprises the steps of: A single colony of strain XY-144 was placed in a SGTYP liquid culture medium and cultured by shaking to obtain a fermentation broth; the fermentation broth was centrifuged to obtain the fermentation supernatant.

4. The fermentation supernatant according to claim 3, characterized in that The shaking culture conditions are: temperature 20-37° C., rotation speed 100-300 rpm, and time 2-4 days.

5. The fermentation supernatant according to claim 3, characterized in that The centrifugal treatment conditions are: temperature 0-10°C, rotation speed 8000-12000 rpm, and time 10-30 minutes.

6. Use of the strain XY-144 according to claim 1 or the fermentation supernatant according to any one of claims 2 to 5 in the preparation of a product for removing biofilm.

7. The use according to claim 6, characterized in that: The biofilm removal product is specifically a product for removing methicillin-resistant Staphylococcus aureus biofilm.

8. Use of the strain XY-144 according to claim 1 or the fermentation supernatant according to any one of claims 2 to 5 in the preparation of a product that inhibits biofilm formation.

9. The use according to claim 8, characterized in that: The product for inhibiting biofilm formation is specifically a product for inhibiting methicillin-resistant Staphylococcus aureus biofilm formation.

Images