Application of mutagenized strain of bacillus subtilis in killing oncomelania and mutagenizing method

Abstract

The present application relates to the technical field of parasite prevention and treatment, in particular to the application of a mutant strain of bacillus subtilis in killing oncomelania and a mutagenesis method. The present application adopts a mutant strain of bacillus subtilis to kill oncomelania. The mutant strain is any one or combination of ARTP-129 or ARTP-154. ARTP-129 was preserved in China Center for Type Culture Collection on January 6, 2025, with the preservation number of CCTCC M 2025036. ARTP-154 was preserved in China Center for Type Culture Collection on January 6, 2025, with the preservation number of CCTCC M 2025037. Through the above technical solution, the mutant strain of bacillus subtilis is adopted to kill oncomelania. The present application is conducive to improving the efficiency of killing oncomelania and has the advantages of green environmental protection, no harmful residues and environmental improvement.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of parasitic prevention and treatment, in particular to application of a mutant strain of Bacillus subtilis in killing Oncomelania and a mutagenesis method. BACKGROUND

[0002] Since the 1950s, China has begun to study microorganisms for killing Oncomelania, and currently, dozens of microorganisms for killing Oncomelania have been found, such as Pseudomonas corrugata, Bacillus cereus B59, Streptomyces violaceus, and autotrophic Xanthomonas, and some of the microorganisms for killing Oncomelania have been studied thoroughly, but there are still great difficulties in field application and low efficiency in killing Oncomelania due to the harsh use conditions of most microorganisms for killing Oncomelania or the problems that in-vitro synthesis and mass production are difficult to achieve by using existing technologies.

[0003] Atmospheric and Room Temperature Plasma (ARTP) mutagenesis technology is a physical mutagenesis technology, and the mutagenesis principle thereof is to cause damage to genetic material of a strain by using active energy particles, so as to induce the strain cell to start the SOS mechanism, and finally obtain a genetically stable mutant strain. Compared with traditional physical mutagenesis and chemical mutagenesis, the ARTP mutagenesis technology has the advantages of higher efficiency, wider application range, more diverse genetic material damage mechanism, safe operation, and mild environment, and has been successfully applied to mutagenic breeding of bacteria, fungi, microalgae and other microorganisms. The successful application of the ARTP mutagenesis technology in microbial breeding can be used in the research on improving the killing effect of microorganisms for killing Oncomelania, and the technology is introduced to develop a new type of microorganism for killing Oncomelania. SUMMARY

[0004] In order to solve the problems in the related art, the present application provides application of a mutant strain of Bacillus subtilis in killing Oncomelania and a mutagenesis method, and the mutant strain of Bacillus subtilis is used to kill Oncomelania, which is conducive to improving the efficiency of killing Oncomelania and has the advantages of green environmental protection, no harmful residues and environmental improvement.

[0005] To solve the above problems, the following technical solutions are provided:

[0006] The application of the mutant strain of Bacillus subtilis in killing Oncomelania uses the mutant strain of Bacillus subtilis to kill Oncomelania, and the mutant strain is any one or a combination of ARTP-129 or ARTP-154.

[0007] By the above scheme, the mutagenized strain of Bacillus subtilis is used to kill the oncomelania, the mutagenized strain of Bacillus subtilis is obtained by mutagenic treatment, and the mutagenized strain is used to kill the oncomelania; Bacillus subtilis is a commonly used biocontrol bacteria, which has the advantages of green environmental protection, no harmful residues and environmental improvement, and the present application takes Bacillus subtilis 3-4 as the experimental object, and carries out ARTP mutagenesis on it, and obtains the mutagenized strains ARTP-129 and ARTP-154 with high oncomelania killing effect, thereby greatly improving the efficiency of oncomelania killing.

[0008] The ARTP-129 is preserved in the China Center for Type Culture Collection on January 6, 2025, and the preservation number is CCTCC M 2025036, and the classification name is Bacillus subtilis ARTP-129.

[0009] The ARTP-154 is preserved in the China Center for Type Culture Collection on January 6, 2025, and the preservation number is CCTCC M 2025037, and the classification name is Bacillus subtilis ARTP-154.

[0010] The ARTP-129 solution is diluted from the ARTP-129 fermentation liquor by using water.

[0011] In the above scheme, the ARTP-129 solution is used to soak and kill the oncomelania, and the mortality rate of the oncomelania reaches 85%.

[0012] The ARTP-154 solution is diluted from the ARTP-154 fermentation liquor by using water.

[0013] In the above scheme, the ARTP-154 solution is used to soak and kill the oncomelania, and the mortality rate of the oncomelania reaches 85%.

[0014] The mutagenesis method of the mutagenized strain of Bacillus subtilis comprises the following steps:

[0015] S1: The activated Bacillus subtilis bacterial liquid is inoculated into an LB liquid culture medium shaking flask at an inoculation amount of 1%, and is placed in a constant temperature shaker for shaking culture; samples are taken every 2h within 32h, and samples are taken every 4h after 32h;

[0016] S2: The bacterial liquid in S1 is centrifuged to obtain a bacterial liquid precipitate, the bacterial liquid precipitate is washed with sterile normal saline, and the concentration of the bacterial liquid precipitate is diluted with sterile normal saline, and the diluted bacterial liquid precipitate is used as a bacterial suspension;

[0017] S3: sterilize the metal carrier, draw the bacterial suspension and uniformly coat the surface of the sterile carrier, place the metal carrier in the normal pressure room temperature plasma mutagenesis instrument, and mutagenize the bacterial suspension to obtain a mutagenized bacterial suspension;

[0018] S4: dilute the mutagenized bacterial suspension, uniformly coat the dilute mutagenized bacterial suspension on a solid culture medium, and culture in a constant temperature incubator, and obtain a mutagenized lethal rate curve according to the growth of the strain;

[0019] S5: according to the mutagenized lethal rate curve, select the best mutagenesis treatment time to obtain a mutagenized strain.

[0020] The temperature of the constant temperature shaker in S1 is 30 DEG C, and the rotation speed is 200 rpm / min.

[0021] The centrifugation time in S2 is 5 min, the rotation speed is 5000 rpm / min, and the dilution bacterial liquid concentration is 10 6 ~10 8 CFU / mL.

[0022] The sterilization method in S3 is to burn the metal carrier on the flame for 10 s, and then cool it down; the incident power of mutagenesis treatment is 100 W, the gas flow is 10 SLM, the irradiation distance is 2 mm, and the treatment time is 20-25 s.

[0023] In S4, the mutagenized bacterial suspension is diluted by 10 -3 ~10 -6 times; the culture temperature in the constant temperature incubator is 30 DEG C, and the culture time is 24 h.

[0024] The above scheme has the following advantages:

[0025] The mutagenized strain of the Bacillus subtilis of the application kills the Oncomelania, adopts the biological snail killing method, compared with the physical snail killing method and the chemical snail killing method, has smaller investment, smaller engineering quantity, saves time and labor, faster snail killing effect, relatively wider application range, and the mutagenized strain of the Bacillus subtilis 3-4 has the advantages of green, non-toxic, high efficiency and stability, and the mutagenized strain of the Bacillus subtilis can kill the Oncomelania, thereby controlling the spread of schistosomiasis. BRIEF DESCRIPTION OF DRAWINGS

[0026] In order to make the content of the application more easily understood, the application will be further described in detail below according to specific embodiments of the application and in combination with the drawings, in which:

[0027] Figure 1 The growth curve of the Bacillus subtilis 3-4;

[0028] Figure 2The lethality curve of ARTP mutagenesis against Bacillus subtilis 3-4 is shown.

[0029] Figure 3 The image shows the snail-killing effect of the LB fermentation broth of the mutant strain after 24 hours. Detailed Implementation

[0030] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0031] In specific embodiment 1, Bacillus subtilis 3-4 was selected as the raw material. Bacillus subtilis 3-4 was deposited at the China Center for Type Microbiology on January 6, 2025, with accession number CCTCC M 2025035 and classified as Bacillus subtilis 3-4.

[0032] Step 1: Selecting materials and preparation

[0033] The experimental strain was Bacillus subtilis 3-4, and the experimental subject was Oncomelania hupensis. After the Oncomelania hupensis captured in the wild was transported to the laboratory, it was rinsed three times with dechlorinated water and then placed in a tray lined with a moist sponge pad and straw paper. It was kept at room temperature (26±1℃) for 7-10 days to adapt to the environment. Adult Oncomelania hupensis with 6-8 turns, no infection and good vitality were selected for the experiment. Before the experiment, the Oncomelania hupensis used for the experiment were washed three times with ultrapure water.

[0034] Experimental reagents: peptone, yeast extract, sodium chloride, and agar powder;

[0035] Experimental instruments: constant temperature incubator, constant temperature shaking incubator, multifunctional microplate reader, and ambient pressure room temperature plasma mutagenesis instrument.

[0036] Step 2: Preparation of ARTP-129 and ARTP-154

[0037] 2.1. Plot the growth curves of Bacillus subtilis 3-4:

[0038] The activated bacterial culture was inoculated into LB liquid medium shake flasks at a 1% (v / v) inoculation rate, with a volume of 100 mL / 250 mL LB. The flasks were incubated at 30 ℃ and 200 rpm / min using a shaker. Samples were taken every 2 h, and then every 4 h after 32 h. Uninoculated LB liquid medium was used as a control. The OD of the fermentation broth was measured at 600 nm using a microplate reader. 600 Value, record sampling time and OD600 The OD values were repeated three times, and the growth curve of Bacillus subtilis 3-4 was plotted with time (h) as the abscissa and the OD values as the ordinate. 600 The OD values were repeated three times, and the growth curve of Bacillus subtilis 3-4 was plotted with time (h) as the abscissa and the OD values as the ordinate.

[0039] As shown in Figure 1 , the growth curve of Bacillus subtilis 3-4 was determined, and the growth of Bacillus subtilis 3-4 can be divided into different periods; the growth curve of Bacillus subtilis 3-4 shows that the first 6 h is in the stationary phase, and after 6 h it enters the logarithmic phase, and the cell concentration reaches the maximum at the 44th hour; the bacterial activity in the logarithmic phase is relatively high, which is more conducive to the screening of positive mutagenesis strains, so the bacterial suspension for mutagenesis treatment is prepared by culturing the bacterial liquid to 30 h.

[0040] 2.2, Preparation of bacterial suspension:

[0041] According to the growth curve, the bacterial liquid in the middle and late logarithmic growth phase was taken, centrifuged at 5000 rpm / min for 5 min, the bacterial liquid precipitate was washed twice with sterile physiological saline, and then the bacterial liquid precipitate was diluted with sterile physiological saline, and the dilution concentration was 10 6 ~10 8 CFU / mL as the bacterial suspension, and the concentration of the bacterial suspension was determined by plate counting method.

[0042] 2.3, ARTP mutagenesis:

[0043] The metal chip was sterilized by burning on the flame for 10 s, and after cooling, 10 μL of bacterial suspension was uniformly coated on the surface of the sterile chip, and the metal chip was placed in the normal pressure room temperature plasma mutagenesis instrument, the incident power was 100 W, the gas flow was 10 SLM, the irradiation distance was 2 mm, and the bacterial suspension was treated, and the treatment time was 0 s, 10 s, 20 s, 30 s, 40 s, 50 s, 60 s, respectively. The mutagenized chip was placed in 990 μL of sterile physiological saline, and the bacteria were washed off on a vortex shaker to obtain a mutagenized bacterial suspension.

[0044] 2.4, Calculation of mutagenesis lethality

[0045] The mutagenized bacterial suspension was diluted by 10 -3 , 10 -4 , 10 -5 , 10 -6Take 100 μL of each appropriately diluted mutant bacterial suspension and spread it evenly on a solid culture medium, with three replicates for each gradient. Invert the medium and incubate it in a 30 ℃ constant temperature incubator for 24 h. Then, count the colonies using the colony counting method to obtain the number of colonies at different mutagenesis times. Use the unmutated bacterial solution as a control. Calculate the mutagenesis lethality rate according to formula (1) to determine the optimal mutagenesis time. Plot the mutagenesis lethality rate curve and select the appropriate mutagenesis time.

[0046] (1),

[0047] 2.5 Evaluation of the molluscicidal effect of the mutagenic strain

[0048] 2.5.1 Preparation of the sample solution to be tested

[0049] The activated mutant strain was inoculated into LB liquid medium and cultured in a constant temperature shaker at 30 ℃ and 200 rpm / min for 24-30 h to obtain LB fermentation broth. The OD of the LB fermentation broth was then adjusted. 600 The value was 0.4 ± 0.1. The LB fermentation broth was prepared into 50.00 mL / L test solutions.

[0050] 2.5.2 Immersion method

[0051] The molluscicidal effect of the mutant strain was evaluated using an immersion method. The test sample solution prepared in section 2.5.1 was loaded into different wells of a six-well plate (13 mL / well), with 20 test snails placed in each well. An ultrapure water control was also included. After immersion for 72 h, the snails were removed, rinsed three times with ultrapure water, and then placed in a six-well plate lined with a moistened sponge pad and straw paper. After a 72 h recovery period at 26°C, snail mortality was assessed. This process was repeated three times.

[0052] 2.5.3 Methods for identifying dead Oncomelania snails

[0053] After the experiment in 2.5.2, the snails were placed in ultrapure water. The presence or absence of soft tissue protrusion was observed. Snails with protruding soft tissue were considered alive. Snails without protruding soft tissue were further judged by tapping to determine their lifespan. They were divided into three categories: dead snails (no soft tissue protrusion, and no shrinkage reaction after being broken), dying snails (no soft tissue protrusion, but shrinkage reaction after being broken), and alive snails (soft tissue protrusion, and shrinkage reaction after being broken). The number of dead snails in each group was recorded, and the snail mortality rate was calculated. The formula for calculating the snail mortality rate (2) is as follows:

[0054] (2),

[0055] The calculation of the molluscicidal effect enhancement rate of the mutant strain is based on formula (3):

[0056] (3),

[0057] 2.6. Lethality curve of Bacillus subtilis 3-4 under ARTP mutagenesis treatment

[0058] The lethality rate is an important reference in mutagenesis experiments. After ARTP mutagenesis treatment for different durations, strains 3-4 of Bacillus subtilis exhibited varying degrees of mortality. Figure 2 As shown, the lethality of Bacillus subtilis 3-4 increased with prolonged treatment time. The average lethality was 20.12% at 10 s and as high as 95.38% at 20 s. A positive correlation existed between the bacterial mutagenesis rate and the mutagenic dose; a higher lethality (above 90%) indicated a higher positive mutagenesis rate. However, excessively long mutagenesis times might kill some desirable mutant strains, affecting the screening results. Therefore, 20 s was selected as the optimal mutagenesis treatment time, yielding 214 mutant strains.

[0059] Step 3: Experimental Data

[0060] like Figure 3 As shown in Table 1, the mortality rate of *Oncomelania hupensis* snails after 72 h of immersion in a 50 mL / L solution of the mutagenic strain and *Bacillus subtilis* 3-424 h LB fermentation broth was determined. The mortality rate was calculated by χ²... 2 The mortality rates of Oncomelania hupensis in all treatment groups were statistically significant compared to the ultrapure water control group. When the mortality rate of the mutant strain was ≤ 33.33% or ≥ 70.00%, the average mortality rate of Bacillus subtilis 3-4 was significantly different from that of the mutant strain (χ²). 2 = 4.126, P = 0.042; χ² 2 = 4.232, P = 0.040). Among the 72 mutant strains with better snail-killing effects than Bacillus subtilis 3-4, 15 mutant strains showed statistically significant differences in snail mortality rates compared to Bacillus subtilis 3-4. Among them, strains ARTP-129 and ARTP-154 had the best snail-killing effects, with an average snail mortality rate of 85.00%, representing a 64.51% improvement in snail-killing effect compared to Bacillus subtilis 3-4.

[0061] Table 1. Molluscicidal effect of LB fermentation broth of mutant strains after 24 hours

[0062] Sample name Snail mortality (%) Sample name Snail mortality (%) Ultrapure water control 1.32±0.64 Bacillus subtilis 3-4 51.67±6.01 ARTP-1 71.67±10.93 ARTP-2 38.33±1.67 ARTP-3 50.00±8.66 ARTP-4 26.67±16.91 ARTP-5 35.00±7.64 ARTP-6 28.33±4.41 ARTP-7 56.67±21.28 ARTP-8 26.67±4.41 ARTP-9 23.33±8.33 ARTP-10 48.33±9.28 ARTP-11 40.00±10.41 ARTP-12 28.33±7.26 ARTP-13 33.33±18.56 ARTP-14 45.00±10.00 ARTP-15 23.33±13.64 ARTP-16 40.00±5.00 ARTP-17 40.00±18.03 ARTP-18 51.67±8.82 ARTP-19 73.33±10.93 ARTP-20 58.33±10.93 ARTP-21 61.67±6.01 ARTP-22 33.33±4.41 ARTP-23 58.33±18.56 ARTP-24 55.00±13.23 ARTP-25 58.33±9.28 ARTP-26 63.33±4.41 ARTP-27 41.67±13.64 ARTP-28 40.00±16.07 ARTP-29 56.67±3.33 ARTP-30 40.00±5.77 ARTP-31 30.00±7.64 ARTP-32 45.00±5.77 ARTP-33 58.33±15.90 ARTP-34 25.00±8.66 ARTP-35 38.33±7.26 ARTP-36 25.00±10.00 ARTP-37 26.67±6.01 ARTP-38 25.00±10.00 ARTP-39 15.00±10.41 ARTP-40 38.33±11.67 ARTP-41 63.33±10.14 ARTP-42 45.00±20.21 ARTP-43 41.67±10.93 ARTP-44 45.00±20.21 ARTP-45 48.33±13.02 ARTP-46 58.33±6.01 ARTP-47 53.33±8.82 ARTP-48 58.33±7.26 ARTP-49 66.67±10.14 ARTP-50 53.33±11.67 ARTP-51 51.67±11.67 ARTP-52 51.67±7.26 ARTP-53 48.33±13.64 ARTP-54 60.00±18.03 ARTP-55 50.00±18.03 ARTP-56 43.33±4.41 ARTP-57 38.33±1.67 ARTP-58 50.00±12.58 ARTP-59 63.33±18.78 ARTP-60 51.67±10.93 ARTP-61 70.00±20.21 ARTP-62 63.33±20.48 ARTP-63 56.67±8.82 ARTP-64 61.67±16.91 ARTP-65 53.33±14.24 ARTP-66 65.00±5.77 ARTP-67 43.33±15.90 ARTP-68 58.33±6.67 ARTP-69 43.33±18.33 ARTP-70 48.33±4.41 ARTP-71 66.67±18.56 ARTP-72 65.00±7.64 ARTP-73 60.00±5.00 ARTP-74 60.00±7.64 ARTP-75 41.67±8.82 ARTP-76 65.00±12.58 ARTP-77 70.00±18.03 ARTP-78 50.00±13.23 ARTP-79 55.00±12.58 ARTP-80 48.33±9.28 ARTP-81 53.33±13.02 ARTP-82 25.00±8.66 ARTP-83 61.67±10.14 ARTP-84 38.33±11.67 ARTP-85 53.33±12.02 ARTP-86 30.00±17.56 ARTP-87 53.33±13.33 ARTP-88 36.67±16.41 ARTP-89 61.67±11.67 ARTP-90 56.67±4.41 ARTP-91 73.33±6.01 ARTP-92 63.33±8.82 ARTP-93 25.00±10.00 ARTP-94 18.33±8.82 ARTP-95 60.00±5.00 ARTP-96 55.00±11.55 ARTP-97 33.33±7.26 ARTP-98 63.33±7.26 ARTP-99 53.33±11.67 ARTP-100 31.67±21.67 ARTP-101 41.67±7.26 ARTP-102 46.67±14.24 ARTP-103 30.00±7.64 ARTP-104 26.67±16.91 ARTP-105 45.00±7.64 ARTP-106 70.00±10.41 ARTP-107 46.67±4.41 ARTP-108 56.67±12.02 ARTP-109 31.67±19.22 ARTP-110 46.67±7.26 ARTP-111 68.33±9.28 ARTP-112 31.67±16.67 ARTP-113 51.67±15.90 ARTP-114 36.67±14.81 ARTP-115 35.00±10.41 ARTP-116 60.00±10.41 ARTP-117 63.33±14.24 ARTP-118 63.33±17.40 ARTP-119 31.67±17.64 ARTP-120 66.67±15.90 ARTP-121 55.00±7.64 ARTP-122 46.67±10.14 ARTP-123 58.33±15.90 ARTP-124 75.00±11.55 ARTP-125 68.33±11.67 ARTP-126 76.67±13.33 ARTP-127 56.67±13.64 ARTP-128 73.33±9.28 ARTP-129 85.00±5.77 ARTP-130 68.33±9.28 ARTP-131 61.67±10.14 ARTP-132 36.67±4.41 ARTP-133 78.33±14.24 ARTP-134 60.00±7.64 ARTP-135 36.67±13.02 ARTP-136 73.33±12.02 ARTP-137 28.33±16.41 ARTP-138 71.67±20.88 ARTP-139 53.33±13.02 ARTP-140 36.67±6.67 ARTP-141 41.67±8.82 ARTP-142 63.33±6.67 ARTP-143 31.67±6.01 ARTP-144 31.67±10.14 ARTP-145 68.33±11.67 ARTP-146 46.67±14.81 ARTP-147 51.67±3.33 ARTP-148 35.00±2.89 ARTP-149 60.00±8.66 ARTP-150 26.67±16.91 ARTP-151 30.00±10.41 ARTP-152 33.33±13.02 ARTP-153 80.00±10.00 ARTP-154 85.00±8.66 ARTP-155 33.33±10.14 ARTP-156 25.00±5.00 ARTP-157 35.00±2.89 ARTP-158 26.67±7.26 ARTP-159 35.00±11.55 ARTP-160 48.33±8.82 ARTP-161 50.00±18.93 ARTP-162 38.33±18.78 ARTP-163 43.33±9.28 ARTP-164 33.33±3.33 ARTP-165 28.33±6.01 ARTP-166 31.67±12.02 ARTP-167 23.33±9.28 ARTP-168 36.67±19.65 ARTP-169 35.00±11.55 ARTP-170 38.33±20.88 ARTP-171 30.00±10.00 ARTP-172 36.67±4.41 ARTP-173 46.67±18.78 ARTP-174 51.67±9.28 ARTP-175 48.33±8.33 ARTP-176 51.67±22.05 ARTP-177 50.00±7.64 ARTP-178 40.00±15.28 ARTP-179 51.67±10.14 ARTP-180 45.00±2.89 ARTP-181 51.67±8.82 ARTP-182 28.33±7.26 ARTP-183 36.67±13.64 ARTP-184 28.33±6.01 ARTP-185 38.33±16.41 ARTP-186 33.33±17.40 ARTP-187 40.00±2.89 ARTP-188 50.00±5.00 ARTP-189 31.67±6.01 ARTP-190 31.67±1.67 ARTP-191 46.67±1.67 ARTP-192 48.33±14.53 ARTP-193 48.33±7.26 ARTP-194 28.33±7.26 ARTP-195 45.00±15.28 ARTP-196 23.33±13.64 ARTP-197 31.67±10.14 ARTP-198 45.00±11.55 ARTP-199 40.00±20.21 ARTP-200 60.00±15.28 ARTP-201 28.33±4.41 ARTP-202 25.00±10.41 ARTP-203 23.33±8.82 ARTP-204 25.00±5.77 ARTP-205 25.00±5.77 ARTP-206 28.33±8.33 ARTP-207 20.00±7.64 ARTP-208 10.00±2.89 ARTP-209 15.00±10.00 ARTP-210 41.67±14.81 ARTP-211 21.67±4.41 ARTP-212 16.67±7.26 ARTP-213 21.67±3.33 ARTP-214 28.33±6.01

[0063] In summary, in the experiment of killing Oncomelania snails by immersion, the mutagenic strains of Bacillus subtilis 3-4 were used to kill Oncomelania snails. Among them, strains ARTP-129 and ARTP-154 had the best snail-killing effect. When using ARTP-129 to kill Oncomelania snails, the mortality rate reached 85%, and when using ARTP-154 to kill Oncomelania snails, the mortality rate reached 85%.

[0064] Obviously, the above-mentioned embodiments are only examples for clearly illustrating the present application, and are not intended to limit the present application. Based on the above description, other different forms of changes or variations can be made by those skilled in the art, and all the changes or variations made without departing from the spirit of the present application should fall within the scope of the present application.

Claims

1. The application of mutagenic strains of Bacillus subtilis in the killing of Oncomelania hupensis, characterized in that, Mutated strains of Bacillus subtilis were used to kill Oncomelania snails; The mutagenic strain is any one or a combination of Bacillus subtilis ARTP-129 or Bacillus subtilis ARTP-154; The Bacillus subtilis ARTP-129 was deposited at the China Center for Type Culture Collection on January 6, 2025, with accession number CCTCC M 2025036. The Bacillus subtilis ARTP-154 was deposited at the China Center for Type Culture Collection on January 6, 2025, with accession number CCTCC M 2025037.

2. The application of the mutagenic strain of Bacillus subtilis as described in claim 1 in the killing of Oncomelania hupensis snails, characterized in that, Oncomelania snails were killed by immersion in a Bacillus subtilis ARTP-129 solution, which was prepared by diluting the Bacillus subtilis ARTP-129 fermentation broth with water.

3. The application of the mutagenic strain of Bacillus subtilis as described in claim 1 in the killing of Oncomelania hupensis snails, characterized in that, Oncomelania snails were killed by immersion in a Bacillus subtilis ARTP-154 solution, which was prepared by diluting the Bacillus subtilis ARTP-154 fermentation broth with water.

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