Bacillus thiaminolyticus and application of bacillus thiaminolyticus in antagonizing streptococcus agalactiae

Abstract

The application discloses a bacillus pumilus strain and application of the bacillus pumilus strain in antagonizing streptococcus agalactiae. Aneurinibacillus aneurinilyticus ) JZK-136, which is preserved in the Guangdong Microbial Culture Collection Center (GDMCC) and is located at No. 59, Building 5, 100, Middle Jianli Road, Guangzhou, with a postal code of 510070, on June 23, 2026, and has a preservation number of GDMCC No: 68496. The bacillus pumilus strain JZK-136 has an antagonistic effect on streptococcus agalactiae GDMCC1.768 through an Oxford cup antibacterial circle experiment, and the diameter of the antibacterial circle is 8.3 mm. Therefore, the bacillus pumilus strain JZK-136 can be applied to antagonize streptococcus agalactiae.

Description

Technical Field

[0001] This invention belongs to the field of microbiology, specifically relating to the application of a strain of thiamine-degrading Bacillus and its antagonistic Streptococcus agalactiae. Background Technology

[0002] agalactococcus ( 无乳链球菌 Group B Streptococcus (GBS) is a Gram-positive opportunistic streptococcus that commonly resides in the human digestive tract, reproductive tract, and nasal cavity. It is an important pathogen causing invasive infections in newborns and pregnant women, and is also a significant pathogen in livestock and farmed fish, often causing large-scale fish diseases.

[0003] Thiamine Bacillus ( 解硫胺素芽孢杆菌 ) is a genus of Bacillus thiamine ( 芽孢杆菌属 The type species is a group of strictly aerobic, Gram-positive, sporolytic bacteria capable of decomposing thiamine. Summary of the Invention

[0004] The purpose of this invention is to provide a strain of Bacillus thiamine (Bacillus). 芽孢杆菌属 解硫胺素 ) and its application in antagonizing Streptococcus agalactiae.

[0005] The first objective of this invention is to provide a strain of Bacillus thiamine dehydrogenase (B. thiamine dehydrogenase). 芽孢杆菌属 解硫胺素 JZK-136 is deposited at the Guangdong Provincial Microbial Culture Collection Center (GDMCC), located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, 510070, China. The deposit date is June 23, 2026, and the accession number is GDMCC No. 68496.

[0006] A second objective of this invention is to provide a microbial agent containing the aforementioned Bacillus thiamine (Bacillus). 解硫胺素芽孢杆菌 JZK-136 is used as the active ingredient.

[0007] This invention demonstrates through experiments that Bacillus thiamine decongestant (Bacillus) 解硫胺素芽孢杆菌 JZK-136 can antagonize Streptococcus agalactiae, wherein the Streptococcus agalactiae is Streptococcus agalactiae (… 链球菌属 无乳的 ‎)GDMCC1.768.

[0008] A second objective of this invention is to provide Bacillus thiamine dehydrogenase (Bacillus) 芽孢杆菌属 解硫胺素 Application of JZK-136 or the above-mentioned bacterial agents in the preparation of products antagonizing Streptococcus agalactiae.

[0009] Preferably, the agalactococcus is agalactococcus (Streptococcus agalactiae). 无乳链球菌‎)GDMCC1.768.

[0010] Preferably, the product is a drug, disinfectant, water quality improver, feed or feed additive.

[0011] The present invention also provides a product for inhibiting Streptococcus agalactiae, containing the aforementioned Bacillus thioamine JZK-136 or the aforementioned bacterial agent.

[0012] The present invention also provides a method for in vitro inhibition of Streptococcus agalactiae, by applying the aforementioned Bacillus thioamine JZK-136 or the aforementioned bacterial agent.

[0013] This invention also provides a drug antagonizing Streptococcus agalactiae, containing Bacillus thiamine (Bacillus). 解硫胺素芽孢杆菌 JZK-136 or the aforementioned bacterial agent.

[0014] This invention, through the Oxford cup inhibition zone experiment, found that Bacillus thiamine-releasing agent JZK-136 has an antagonistic effect against Streptococcus agalactiae GDMCC1.768, with an inhibition zone diameter of 8.3 mm. Therefore, Bacillus thiamine-releasing agent JZK-136 can be used to antagonize Streptococcus agalactiae.

[0015] Thiamine Bacillus ( 解硫胺素芽孢杆菌 JZK-136 is deposited at the Guangdong Provincial Microbial Culture Collection Center (GDMCC), located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, 510070, China. The deposit date is June 23, 2026, and the accession number is GDMCC No: 68496. Attached Figure Description

[0016] Figure 1 These are photographs of the Oxford cup inhibition zone experiment of Bacillus thiamine JZK-136 against Streptococcus agalactiae GDMCC1.768. A, B, C: Plate photographs of Streptococcus agalactiae GDMCC1.768. a, Escherichia coli ATCC8739; b, Bacillus thiamine JZK-136; c, cefepime; d, Bacillus thiamine JZK-136. Detailed Implementation

[0017] The following embodiments are further illustrations of the present invention, but not limitations thereof.

[0018] Example 1:

[0019] Thiamine Bacillus ( 解硫胺素芽孢杆菌JZK-136 was isolated from industrial spoilage products. The strain was identified using 16S rDNA molecular identification technology. The universal primers for PCR amplification were: F27 5'-AGAGTTTGATCCTGGCTCA-3', R1492 5'-GGTTACCTTGTTACGACTT-3'.

[0020] Thiamine Bacillus ( 解硫胺素芽孢杆菌 JZK-136 is deposited at the Guangdong Provincial Microbial Culture Collection Center (GDMCC), located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, 510070, China. The deposit date is June 23, 2026, and the accession number is GDMCC No: 68496.

[0021] Example 2:

[0022] agalactococcus ( 无乳链球菌 GDMCC1.768 was inoculated into BHI medium (10.0 g / L peptone, 12.5 g / L dehydrated calf brain extract, 5.0 g / L dehydrated calf heart extract, 5.0 g / L sodium chloride, 2.0 g / L glucose, 2.5 g / L disodium hydrogen phosphate, pH 7.4±0.2) and cultured at 37 ℃ with shaking. Preparation of bacterial suspension: Samples of the exponential growth phase of Streptococcus agalactiae GDMCC1.768 strain were taken, centrifuged, washed once with PBS buffer, resuspended in PBS, and diluted to 2×10⁻⁶. 9 CFU / mL was used to obtain a suspension of Streptococcus agalactiae GDMCC1.768.

[0023] Thiamine Bacillus ( 解硫胺素芽孢杆菌 JZK-136 and Escherichia coli ( 大肠杆菌 ATCC8739 was inoculated into TSB medium (tryptone 17.0 g / L, plant peptone 3.0 g / L, sodium chloride 5.0 g / L, dipotassium hydrogen phosphate 2.5 g / L, glucose 2.5 g / L, pH 7.3±0.2) and cultured at 37 ℃ with shaking for 24 h. Cell-free supernatants were prepared: samples of the 24 h culture broths of Bacillus thiamine-degrading strains JZK-397 and Escherichia coli ATCC8739 were taken, centrifuged, and the supernatants were collected. The supernatants were then filtered through a 0.22 µm filter membrane to remove bacteria, yielding cell-free supernatants for Bacillus thiamine-degrading strains JZK-136 and Escherichia coli ATCC8739, respectively.

[0024] Cefepime (CAS: 123171-59-5, C302666-1g, purity ≥ 98%) was purchased from Aladdin Biochemical Technology Co., Ltd. (Shanghai, China). Preparation of cefepime solution: Cefepime hydrochloride hydrate was weighed and dissolved in sterile saline solution, and then serially diluted to prepare a 0.5 µg / mL cefepime solution.

[0025] 1. Oxford cup inhibition zone experiment of Bacillus thiamine JZK-136

[0026] Take 100 µL of Streptococcus agalactiae GDMCC1.768 bacterial suspension and spread it onto BHI agar plates, with three replicates per group. Punch four wells in each Oxford cup. Add 100 µL of cell-free supernatant of Escherichia coli ATCC8739 to well a as a negative control, add 100 µL of cefepime (0.5 µg / mL) to well c as a positive control, and add 100 µL of cell-free supernatant of Bacillus thiamine JZK-136 to wells b and d respectively. Incubate overnight at 37 ℃ for 24 h, then observe and measure the diameter of the inhibition zone.

[0027] 2. Experimental Results:

[0028] For detailed results of the Oxford cup inhibition zone assay of Bacillus thiamine JZK-136 against Streptococcus agalactiae GDMCC1.768, please refer to [link to assay]. Figure 1 See Table 1. The experimental results showed that cell-free culture medium of *Bacillus thiamine* JZK-136 had a good antagonistic effect against *Streptococcus agalactiae* GDMCC1.768, with inhibition zone diameters of 8.0±0.2 mm and 8.5±0.4 mm in the two wells, and an average inhibition zone diameter of 8.3 mm. The cell-free supernatant of the negative control *Escherichia coli* ATCC8739 showed an inhibition zone diameter of 6.0±0 mm against *Streptococcus agalactiae* GDMCC1.768, with no inhibitory effect. The positive control cefepime showed an inhibition zone diameter of 13.7±0.6 mm against *Streptococcus agalactiae*. The experimental results indicate that *Bacillus thiamine* JZK-136 has a good antagonistic effect against *Streptococcus agalactiae* GDMCC1.768.

[0029] Table 1. Statistics on the diameter of the inhibition zone of Bacillus thiophanate-methyl against Streptococcus agalactiae in Oxford cups.

Claims

1. Thiamine-degrading Bacillus ( Aneurinibacillus aneurinilyticus JZK-136, accession number: GDMCC No: 68496.

2. A microbial agent, characterized in that, Contains the thiamine-degrading Bacillus as described in claim 1 ( Aneurinibacillus aneurinilyticus JZK-136 is used as the active ingredient.

3. The Thiamine-Degrading Bacillus as described in claim 1 ( Aneurinibacillus aneurinilyticus The application of the microbial agent described in JZK-136 or claim 2 in the preparation of products antagonizing Streptococcus agalactiae.

4. The application according to claim 3, characterized in that, The agalactococcus mentioned is agalactococcus (Streptococcus agalactiae) Streptococcus agalactiae ‎)GDMCC1.

768.

5. The application according to claim 3, characterized in that, The products mentioned are medicines, disinfectants, water quality improvers, feed or feed additives.

6. A product for inhibiting Streptococcus agalactiae, characterized in that, It contains Bacillus thiocyanate JZK-136 as described in claim 1 or the bacterial agent as described in claim 2.

7. A method for inhibiting Streptococcus agalactiae in vitro, characterized in that, Apply the Bacillus thioamnione JZK-136 as described in claim 1 or the bacterial agent as described in claim 2.

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