A bacteriophage composition for preventing and treating salmonella abortus, a biological preparation and application thereof

Abstract

The application belongs to the technical field of biotechnology, and particularly relates to a bacteriophage composition for preventing and treating Salmonella abortus equi, a biological preparation and application thereof. The bacteriophage composition comprises bacteriophage 4FS1, bacteriophage DS-BP2 and bacteriophage DS2; the preservation number of the bacteriophage 4FS1 is CCTCC NO: M 20242020; the preservation number of the bacteriophage DS-BP2 is CCTCC NO: M 202402021; and the preservation number of the bacteriophage DS2 is CGMCC No. 17093. The application isolates and screens three virulent bacteriophages 4FS1, DS-BP2 and DS2 for different host spectra of Salmonella, and combines the bacteriophages into a bacteriophage cocktail, so that the emergence of bacteriophage-resistant strains can be effectively prevented or delayed, and the screened bacteriophages have a synergistic effect. The three bacteriophages isolated by the application can be used alone or combined into a bacteriophage cocktail, and are used for preventing or treating infections caused by Salmonella abortus equi, such as abortion of female donkeys.

Description

Technical Field

[0001] The invention belongs to the field of biotechnology, and in particular relates to a bacteriophage composition for preventing and treating equine Salmonella abortus, a biological preparation and applications thereof. Background Art

[0002] There are numerous serotypes of Salmonella, most of which are pathogenic and have a wide host spectrum, including mammals, birds, and humans. However, different serotypes of Salmonella infect different hosts. Among them, Salmonella equi abortus mainly infects equine animals. Donkey paratyphoid is an infectious disease of equine animals, mainly caused by Salmonella equi abortus, characterized by abortion in pregnant donkeys. The main symptoms are abortion in female donkeys, sepsis in newborn foals, and polyarthritis. First-time pregnant female donkeys and newborn foals are the most susceptible. If not treated in time, it can lead to secondary infection and severe metritis, including purulent and foul-smelling bloody vaginal discharge. This metritis may persist for a long time, leading to repeated infertility or frequent abortions.

[0003] To date, all approved antibiotic classes, whether natural, semisynthetic, or synthetic compounds, have developed resistance in at least some of the pathogens they target. With few antibiotics available for clinical use, developing effective alternatives to antibiotics to overcome the antibiotic resistance crisis is urgent.

[0004] Bacteriophages are widely present in nature. They are a type of virus that primarily infects bacteria and are natural predators of bacteria. Compared to antibiotics, phages offer advantages such as high lysis efficiency, strong specificity, minimal side effects, protection from the host's normal microbiome, and low production costs. However, single phages can easily generate phage-resistant strains during their interaction with bacteria. Therefore, the search for a phage combination with diverse host profiles and a broad lysis spectrum has become an urgent challenge. Summary of the Invention

[0005] In view of the problems existing in the prior art, the object of the present invention is to provide a phage composition for preventing and treating equine Salmonella abortus that has a broad lysis spectrum, is not prone to producing phage-resistant strains, and has a significant synergistic lysis effect.

[0006] Another object of the present invention is to provide a biological preparation containing the above-mentioned phage composition.

[0007] The present invention also provides the use of the phage composition and biological preparation in preventing and treating equine abortion Salmonella.

[0008] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0009] The invention provides a phage composition for preventing and treating equine Salmonella abortus. The phage composition comprises two or more of phage 4FS1, phage DS-BP2 and phage DS2; the phage 4FS1 has a deposit number of CCTCC NO: M 20242020; the phage DS-BP2 has a deposit number of CCTCC NO: M 202402021; and the phage DS2 has a deposit number of CGMCC No. 17093.

[0010] Furthermore, the total content of phages in the phage composition is 10 8 -10 10 PFU / mL.

[0011] Furthermore, the mass ratio of the bacteriophage 4FS1, bacteriophage DS-BP2 and bacteriophage DS2 is 1:1:1.

[0012] The present invention also provides the use of the bacteriophage composition in preparing a product for preventing and treating equine abortion Salmonella infection.

[0013] Furthermore, in the phage composition, the multiplicity of infection of phage 4FS1 is 0.001-0.1; the multiplicity of infection of phage DS-BP2 is 0.01-0.1; and the multiplicity of infection of phage DS2 is 0.001-0.01.

[0014] Another object of the present invention is to provide a biological preparation for preventing and treating equine Salmonella abortion infection containing the above-mentioned bacteriophage composition.

[0015] The biological preparation provided by the present invention is in any veterinary acceptable dosage form; preferably, the preparation is a liquid dosage form.

[0016] The present invention also provides the use of the biological preparation in preparing medicine for preventing and treating equine abortion Salmonella infection.

[0017] Preferably, the incubation periods of the phage composition, phage 4FS1, phage DS-BP2, and phage DS2 are 20 min, 15 min, and 5 min, respectively, and all three last until 120 min to reach a plateau phase.

[0018] Preferably, in the phage composition, the pH value at which the activity of phage 4FS1 is stable is 4-11; the pH value at which the activity of phage DS-BP2 is stable is 4-10; and the pH value at which the activity of phage DS2 is stable is 4-12.

[0019] Preferably, in the phage composition, the temperature at which the activities of phages 4FS1, DS-BP2 and DS2 are stable is no higher than 70°C.

[0020] Beneficial effects of the present invention:

[0021] (1) The present invention isolates and screens three virulent phages, 4FS1, DS-BP2, and DS2, targeting different host profiles of Salmonella. Combining them into a phage cocktail can effectively prevent or delay the emergence of phage-resistant strains, and the phages screened by the present invention have a synergistic effect;

[0022] (2) The three phages isolated and obtained in the present invention are all broad-spectrum phages. In addition to lysing Salmonella abortus, they can also lyse serotypes of Salmonella such as Salmonella Enteritidis and Salmonella Typhimurium. They have high lysis activity and have good lysis effects at different multiplicities of infection (MOI). They have good temperature tolerance and acid-base tolerance and have a wide range of applicability in actual production applications.

[0023] (3) The three phages isolated and obtained in the present invention can be used alone or in combination to form a phage cocktail, which can be used to prevent or treat infections caused by Salmonella enterica equine abortion, such as abortion in donkeys.

[0024] Collection information 1

[0025] Biological material: Salmonella phage 4FS1

[0026] Storage time: November 4, 2024

[0027] Preservation unit: China Center for Type Culture Collection,

[0028] Deposit number: CCTCC NO: M 20242421,

[0029] Address of the collection unit: Wuhan University Collection Center, No. 299 Bayi Road, Wuchang District, Wuhan City, Hubei Province.

[0030] Collection Information 2

[0031] Biomaterial: Salmonella phage DS-BP2

[0032] Storage time: November 4, 2024

[0033] Preservation unit: China Center for Type Culture Collection,

[0034] Deposit number: CCTCC NO: M 20242420,

[0035] Address of the collection unit: Wuhan University Collection Center, No. 299 Bayi Road, Wuchang District, Wuhan City, Hubei Province.

[0036] Collection Information 3

[0037] Biomaterial: bacteriophage DS2

[0038] Collection time: January 18, 2019

[0039] Depository: General Microbiology Center of China Culture Collection Administration of Microorganisms,

[0040] Deposit number: CGMCC No.17093,

[0041] Address of the depository: Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing. BRIEF DESCRIPTION OF THE DRAWINGS

[0042] Figure 1 Transmission electron microscopy morphology of the three phages screened for the present invention;

[0043] Figure 2 The plaque morphology of three phages on a double-layer plate;

[0044] Figure 3 The optimal MOI diagram of the three phages screened in the present invention;

[0045] Figure 4 One-step growth curves of three phages;

[0046] Figure 5 pH stability of the three phages;

[0047] Figure 6 Thermostability of the three phages;

[0048] Figure 7 UV stability of three phages;

[0049] Figure 8 Phage in vitro lysis curve; ***: P < 0.001;

[0050] Figure 9 Detection results of bacterial loads in four tissues; ***: P < 0.001, **: P < 0.01, *: P < 0.05;

[0051] Figure 10 Phage titers of four tissues; **: P < 0.01, *: P < 0.05. DETAILED DESCRIPTION

[0052] The technical solutions provided by the present invention are described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0053] The invention provides a Salmonella phage composition. The phage composition comprises at least one of phage 4FS1, phage DS-BP2 and phage DS2. The phage 4FS1 has a deposit number of CCTCC No. M20242421, the phage DS-BP2 has a deposit number of CCTCC No. M20242420, and the phage DS2 has a deposit number of CGMCC No. 17093.

[0054] In the present invention, bacteriophage 4FS1, bacteriophage DS-BP2 and bacteriophage DS2 are obtained by separation of donkey farm feces and sewage. Bacteriophage 4FS1 and bacteriophage DS-BP2 were deposited in the China Center for Type Culture Collection on January 20, 2024, and bacteriophage DS2 was deposited in the General Microbiology Center of the China Association for the Collection of Microorganisms on January 18, 2019. The phage composition provided by the present invention can effectively prevent or delay the emergence of phage-tolerant strains, and the synergistic effect is obvious. The present invention has no special restrictions on the dosage ratio of the above-mentioned various phages, and any ratio can be adopted, preferably an equal proportion. In the present invention, the total content of phages in the phage composition is preferably 10 8 PFU / mL-10 10 PFU / mL, more preferably 10 9 In the present invention, the Salmonella phage composition is preferably a combination of any two of phage 4FS1, phage DS-BP2 and phage DS2, and more preferably a combination of phage 4FS1, phage DS-BP2 and phage DS2.

[0055] The present invention also provides a use of the bacteriophage composition in preparing a product for preventing and treating Salmonella infection.

[0056] The phage composition of the present invention has a wide lysis spectrum and can not only lyse equine abortion Salmonella, but also lyse various serotypes of Salmonella such as Enteritidis and Salmonella typhimurium.

[0057] In the phage composition of the present invention, the multiplicity of infection of phage 4FS1 is preferably 0.001-0.1, more preferably 0.01; the multiplicity of infection of phage DS-BP2 is preferably 0.1-0.01, more preferably 0.1; and the multiplicity of infection of phage DS2 is preferably 0.001-0.01, more preferably 0.01.

[0058] In the phage composition of the present invention, the incubation periods of phage 4FS1, phage DS-BP2, and phage DS2 are 20 min, 15 min, and 5 min, respectively, and all three last until 120 min to reach a plateau phase.

[0059] In the phage composition described herein, the activity of phage 4FS1 is stable at a pH of 4-11; the activity of phage DS-BP2 is stable at a pH of 4-10; and the activity of phage DS2 is stable at a pH of 4-12. In the phage composition described herein, the temperature at which the activity of phages 4FS1, DS-BP2, and DS2 is stable is no higher than 70°C. In the phage composition described herein, phages 4FS1, DS-BP2, and DS2 maintain a high titer even after 40 minutes of ultraviolet irradiation.

[0060] The present invention also provides a biological preparation for preventing and treating Salmonella infection, wherein the biological preparation comprises the above-mentioned equine abortion Salmonella phage composition.

[0061] In the biological preparation of the present invention, the titer of the above-mentioned Salmonella abortus phage composition is preferably 10 8 PFU / mL-10 10 PFU / mL, more preferably 10 9 PFU / mL.

[0062] In the present invention, the biological preparation can be any veterinary acceptable dosage form; preferably, the biological preparation is a liquid dosage form.

[0063] Example 1

[0064] (1) Isolation of bacteriophage:

[0065] Fecal and sewage samples were collected from donkey farms in different areas of Shandong Province. The host bacteria for phage isolation was Salmonella equi abortus S1, which was isolated and preserved in our laboratory from aborted donkey fetuses.

[0066] The collected sewage was initially filtered with filter paper to remove large solid impurities. 20 mL of the filtered sewage sample was taken out and placed in a 50 mL centrifuge tube. It was centrifuged at 10,000 g for 10 min. The supernatant was filtered with a 0.45 μm pore size sterile filter as the mother liquid for phage proliferation and stored at 4°C for later use.

[0067] Salmonella abortus S1 was inoculated into LB medium and cultured at 37°C with shaking until the logarithmic growth phase. 10 mL of the filtered supernatant was mixed with 5 mL of the host bacteria and 20 mL of LB medium. The culture was shaken at 37°C and 220 rpm for 12 hours to allow the phage in the fecal filtrate to fully proliferate. The cultured sample was centrifuged at 12,000 g for 3 minutes, and the supernatant was removed and filtered through a 0.22 μm sterile filter. This filtrate was the phage stock solution. The presence of phage in the supernatant was confirmed by spot analysis. 3 mL of the melted semi-solid medium was added to 100 μl of the host bacteria, mixed evenly, and poured into a standard LB solid culture dish. After solidification, 20 μl of the phage stock solution was added dropwise to the poured double-layer plate. After solidification, the plate was placed in a 37°C incubator and observed for the appearance of plaques. The filtrate with plaques was diluted appropriately in gradients, 100 μl of the diluted liquid and 100 μl of the host bacteria were added to 3 mL of semi-solid culture medium, poured onto NA solid culture medium, spread flat, and inverted cultured at 37°C for 8 h after solidification.

[0068] (2) Phage purification

[0069] Use a small pipette to pick up plaques and add them to 1 mL of saline. Shake on a shaker for 5 minutes. Perform appropriate gradient dilutions. Mix 100 μl of the diluted phage solution with 100 μl of host bacteria and add to thawed semi-solid culture medium. Pour the mixture onto LB solid medium and incubate at 37°C for 8 hours. Observe for the appearance of individual plaques. Repeat this process at least three times to purify the phage.

[0070] The LB liquid culture medium: weigh 25 g of commercial LB culture medium powder (product of Qingdao Haibo Biotechnology Co., Ltd.), dissolve it in 1 L of distilled water, sterilize it by high pressure at 121° C. for 20 min, and refrigerate it at 4° C. until use.

[0071] The NA solid culture medium: weigh 33 g of commercial NA culture medium powder (product of Qingdao Haibo Biotechnology Co., Ltd.), dissolve it in 1 L of distilled water, sterilize it by high pressure at 121°C for 20 min, cool it to about 45°C, pour it into a sterile plate, and refrigerate it at 4°C until use.

[0072] The semi-solid culture medium is as follows: 7.5 g agar powder is added to 1 L of LB liquid culture medium, sterilized by high pressure at 121° C. for 20 min, stored at 4° C. after aliquoting, and heated to melt completely before use.

[0073] (3) Phage proliferation

[0074] In order to improve the number of phages in unit volume, the method of liquid amplification was used. The single spot purified in step (2) was taken out and put into 1 mL of nutrient broth medium, 37°C water bath for 10 min, then centrifuged at 10000 r / min for 5 min, 100 μl of supernatant was taken and added to 100 μl of Salmonella abortus liquid medium, 37°C constant temperature incubator 220 r shaking for 4 h, then the mixture was clarified to obtain phage proliferation liquid. The supernatant was filtered with a sterile filter with a pore size of 0.22 μm, 10 times gradient dilution, and then the phage titer was determined by double layer plate method. According to the growth of different phages, the phage supernatant was diluted to different concentration gradients for pre-experiment, and the optimal growth ratio of phage was calculated according to the phage titer, and stored at 4°C for standby. After the phage was purified and amplified, the titer of the phage could reach 10 9 PFU / mL.

[0075] (4) Electron microscope observation of phage morphology

[0076] Phosphotungstic acid negative staining method was used, one drop of purified high titer phage liquid was absorbed on the copper mesh for 10 min, the residual suspension was absorbed, 2% phosphotungstic acid (PTA, 2% w / v) was used for staining for 3 min, the copper mesh was dried by infrared lamp, and transmission electron microscope observation was carried out. The results showed that the three phages belonged to the tail phages, and the results were shown in Figure 1 The whole genome sequencing results showed that 4FS1 phage belonged to long tail phage of Jerseyvirus genus, DS2 phage belonged to long tail phage of Epseptimavirus genus, and DS-BP2 phage belonged to short tail phage of Berlinvirus genus. None of the three phages contained lysogenic genes, drug resistance genes and virulence genes.

[0077] Example 2

[0078] 100 μL of overnight culture of Example 1 was mixed with semi-solid medium and poured into LB solid medium to make double layer plate, the phage was diluted 10 times to different concentrations, 20 μL of phage with different concentrations was dropped on the double layer plate, dried and then cultured in 37°C incubator for 8 h, and the gradient dilution of the plate was observed. The results are shown in Figure 2 The selected three strains of Salmonella phage grew well on the double layer plate, and were clear and transparent. With gradient dilution, the single spot of phage was finally observed, and phage 4FS1 and DS-BP2 appeared obvious halo around the phage plaque.

[0079] Example 3

[0080] The host spectrum of phages was determined by the droplet method. Different strains to be tested were cultured to the logarithmic growth phase. 100 μL of overnight culture solution was added to the semi-solid medium, mixed evenly, and poured into the NA solid medium. After solidification, different dilutions (10 1 ~10 7 10 μL of phage was spotted onto a double-layer plate. After drying, the plate was incubated upside down at 37°C for 8 hours. The phage lysis of the test strains was observed, and the phage plaque efficiency (EOP) was calculated. The results, as shown in Table 1, showed that 4FS1, DS-BP2, and DS2 phages exhibited a broad lysis spectrum, infecting 64.2% of the test strains with high infection efficiency.

[0081] Table 1 Phage lysis spectrum

[0082]

[0083]

[0084] EOP refers to the ratio of virus particles that can form visible plaques. Equine abortion Salmonella S1 was selected as the standard indicator bacteria to compare the titer of phage lysis of different host bacteria with the ratio of indicator bacteria. +++ means EOP ≥ 0.5, ++ means 0.1 ≤ EOP < 0.5, and + means 0.001 <EOP<0.1,-为EOP≤0.001。

[0085] Example 4

[0086] Host spectrum of phage The host bacteria S1 was cultured to the logarithmic growth phase (OD 600 =0.6), and the concentration of the bacterial solution was adjusted to 1×10 7 CFU / mL. Then, different concentrations of phage were added according to different MOI (phage / bacteria) = 10, 1, 0.1, 0.01, 0.001, and placed in a 37°C shaker for 4 hours. After centrifugation at 12000g for 1 minute, the culture solution was serially diluted in physiological saline, and the phage titer corresponding to different MOIs was determined by the double-layer plate method. The experiment was repeated three times, and the phage / bacteria ratio that produced the highest titer was the optimal MOI. The results are shown in the figure. Figure 3 As shown in the figure, when the optimal MOI of 4FS1, DS-BP2, and DS2 phages were 0.01, 0.1, and 0.01, respectively, the titer of each phage reached the highest, so this value was the optimal MOI of each phage.

[0087] Example 5

[0088] Host bacteria S1 was cultured to the logarithmic growth phase (OD 600 =0.6), and the concentration of the bacterial solution was adjusted to 1×108 About CFU / mL. The phage and host bacteria were mixed at an MOI of 10, and placed on a 37°C shaker for 10 minutes to allow the phage to fully adsorb to the host bacteria. After centrifuging the mixture at 10,000g for 10 minutes, the supernatant was discarded, and the precipitate was resuspended with the same volume of LB, centrifuged again, and the supernatant was discarded. The precipitate was washed twice. Finally, the resuspension was placed on a 37°C shaker for oscillation culture, and samples were taken at intervals to detect the phage titer until 240 minutes. The filtered phage liquid was serially diluted in physiological saline, and the phage titer corresponding to different times was determined by the double-layer plate method. The experiment was repeated three times. With the infection time as the horizontal axis and the phage titer as the vertical axis, a one-step growth curve of the phage-infected host bacteria was drawn. The results are shown in the figure. Figure 4 As shown, the incubation periods of phage 4FS1, phage DS-BP2, and phage DS2 were 20 min, 15 min, and 5 min, respectively, and all three lasted until 120 min to reach a plateau phase.

[0089] Example 6

[0090] Take 100 μL (1×10 9 PFU / mL) phage was placed in a 1.5 mL sterile EP tube, and 900 μL of SM buffer solution with different pH values ​​(2-13) was added. The titer was determined by double-layer plate method at 37°C water bath for 1, 2, and 3 hours. The experiment was repeated 3 times. Figure 5 As shown, 4FS1 phage is relatively stable at pH 4-11; DS-BP2 phage is relatively stable at pH 4-10; and DS2 phage is relatively stable at pH 4-12.

[0091] Take the phage suspension (10 9 PFU / mL) and placed them in test tubes. Four tubes were taken at each temperature, each with 500 μL. These test tubes were placed in a water bath at 37°C, 40°C, 50°C, 60°C, 70°C, and 80°C for 20 min, 40 min, and 60 min, respectively. At each time point, one tube was taken, 100 μL of the sample was aspirated for continuous gradient dilution, and the effect of different temperatures on phage titer was immediately determined using the double-layer agar plate method. The results are shown in Figure 2. Figure 6 As shown, the titers of phages 4FS1 and DS2 remained essentially unchanged (>10 9 PFU / mL); DS-BP2 phage titer > 10 in a 50°C water bath for 60 min 6 PFU / mL or above.

[0092] Take 4mL of phage proliferation solution (10 9PFU / mL) were placed under UV light at a distance of 40 cm and irradiated. Every 10 minutes, 200 μL of phage proliferation fluid was taken and the titer was measured using the double-layer plate method. Figure 7 As shown in the figure, phages 4FS1, DS-BP2 and DS2 can still maintain 10 4 With titers above PFU / mL, phages 4FS1, DS-BP2, and DS2 showed strong resistance to UV within 30 min.

[0093] Example 7

[0094] Host bacteria S1 was cultured overnight, and the activated bacteria solution was inoculated into 96-well cell plates and shaken at 37°C (220r) until the early exponential phase (OD 600 =0.3-0.4), at this time, single phage and different combinations of phage were added to each well with an MOI of 1. The culture was continued for 24 hours, and the changing trend of single phage and different combinations of phage in lysing host bacteria in vitro was detected. From the lysis curve ( Figure 8 ) It can be observed that the combination of the three phages can inhibit the growth of bacteria. Compared with the control group without phages, OD 600 The value decreased by 0.7, and the synergistic effect was significant (P < 0.001).

[0095] Example 8

[0096] Sixty mice aged about 6 weeks were randomly divided into 6 groups, with 10 mice in each group. The bacteria cultured to the logarithmic growth phase were diluted into 10 7 CFU / mL, 200 μL was inoculated via intraperitoneal injection. 24, 48, and 72 hours after bacterial infection, mice were euthanized, and the liver, spleen, lung, and uterus were harvested to measure the bacterial load and phage titer in the tissues.

[0097] Bacterial challenge group: mice were intraperitoneally injected with PBS 1 hour after bacterial infection, serving as the positive control group;

[0098] The phage treatment groups were divided into single phage treatment group and phage cocktail treatment group: Single phage treatment group: mice were intraperitoneally injected with 200 μL of three single phages (10 8 Phage cocktail treatment group: 1 hour after infection, mice were intraperitoneally injected with 200 μL of phage cocktail (10 8 PFU / mL).

[0099] The PBS control group of healthy mice was intraperitoneally injected with PBS throughout the whole process as a control.

[0100] The results are as follows Figure 9As shown in the figure, both the single phage treatment group and the phage cocktail treatment group could reduce the bacterial load in each tissue, among which the phage cocktail group reduced the tissue bacterial load more significantly. In each tissue 72 hours after the virus challenge, the bacterial load in the cocktail group was reduced by 2-6 gradients compared with the virus challenge group (P < 0.05); Figure 10 As shown, the phage titer in the tissue was also the highest in the cocktail phage group.

[0101] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as within the scope of protection of the present invention.

Claims

1. A bacteriophage composition for preventing and treating equine Salmonella abortion, characterized in that: The phage composition includes a phage ( Salmonella phage)4FS1、phage( Salmonella phage)DS-BP2 and phage ( Salmonella The invention relates to two or more phage) DS2; the deposit number of the phage 4FS1 is: CCTCC NO: M 20242021; the deposit number of the phage DS-BP2 is: CCTCC NO: M 202402020; the deposit number of the phage DS2 is CGMCC No.17093.

2. The phage composition according to claim 1, characterized in that The total content of phages in the phage composition is 10 8 -10 10 PFU / mL.

3. The phage composition according to claim 1 or 2, characterized in that The mass ratio of the bacteriophage 4FS1, bacteriophage DS-BP2 and bacteriophage DS2 is 1:1:

1.

4. Use of the bacteriophage composition according to any one of claims 1 to 3 in preparing a product for preventing and treating Salmonella abortion infection in horses.

5. The use according to claim 4, characterized in that In the phage composition, the multiplicity of infection of phage 4FS1 is 0.001-0.1; the multiplicity of infection of phage DS-BP2 is 0.01-0.1; and the multiplicity of infection of phage DS2 is 0.001-0.

01.

6. A biological preparation for preventing and treating equine Salmonella abortus infection, comprising the bacteriophage composition according to any one of claims 1 to 3.

7. The biological preparation according to claim 6, characterized in that The preparation is in any veterinary acceptable dosage form.

8. Use of the biological preparation according to claim 6 or 7 in the preparation of a medicament for preventing and treating Salmonella abortus infection in horses.

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