TWI394579B - Compositions for treating or preventing infection of riemerella anatipestifer - Google Patents

Description

Composition for treating or preventing infection with zerm bacteria

The present invention relates to a composition against infection by Riemerella anatipestifer .

Reptilii is one of the major infectious agents of poultry and can infect ducks, turkeys, chickens, crickets, crickets, swan, geese, etc. ( Avian Dis 2005; 49: 104-7). In the case of ducks, the infection with R. cerevisiae is common in young ducks, often accompanied by septicemia and polyserositis, and the mortality rate is high, which has a great impact on the duck industry ( Avian Dis 1985; 29: 128). -35; and Avian pathol 1997; 26:791-802). However, there is currently limited knowledge of defense systems against duck-resistant pathogens, and even if vaccines have been developed using genotypic variation, they are limited in their ability to produce cross-protection in different serotypes of R. falciparum. The use of vaccines to prevent infection with Rep. Therefore, most of the current breeders still use antibiotics to control R. falciparum infections. Commonly used antibiotics include ceftriofur, enrofloxacin, novobiocin, lincomycin, sulphonamides, and the like. However, these antibiotics can cause side effects and can lead to the development of drug-resistant strains, and even these drug-resistant strains may enter the human body via the food chain, adversely affecting humans ( J Vet Diagn invest 2003; 15:26-9; and Antimicrob Agents Chemother 2005; 49: 690-8).

Therefore, there is still a need in this field to develop an agent that is effective against R. falciparum infection, particularly a non-antibiotic agent, for use in the poultry farming industry as a medicament for controlling infection with R. eutropha.

The present invention finds that a specific fragment of the antibacterial protein has an effect against infection by a bacterium. The invention breaks through the current limited effect of the vaccine for preventing and treating Atrabacterium infection in poultry breeding, and the antibiotic has side effects and drug resistance, and provides an antibacterial protein which can effectively resist the infection of the bacterium, and is greatly beneficial for the prevention and treatment of the infection of the bacterium

Accordingly, in one aspect, the present invention provides a composition for treating or preventing a Reuters infection comprising an effective amount of a peptide and a carrier selected from the group consisting of:

(1) a peptide having the amino acid sequence of SEQ ID NO: 1;

(2) a peptide having the amino acid sequence of SEQ ID NO: 2;

(3) a peptide having the amino acid sequence of SEQ ID NO: 3;

(4) a peptide having the amino acid sequence of SEQ ID NO: 4;

(5) A combination of one or more of the above (1) to (4).

In another aspect, the invention provides the use of the above-described peptide for the preparation of a medicament for the treatment or prevention of a Re retrovirus infection.

In still another aspect, the present invention provides a method for killing a bacterium of the bacterium in vitro, comprising administering an effective amount of the above-described peptide and a carrier to an article or environment that may contain a bacterium .

Various specific embodiments of the invention are described in detail below. Other features of the present invention will be apparent from the following detailed description of various embodiments.

It is believed that those skilled in the art of the invention can <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; Accordingly, the following description is to be considered as illustrative and not restrictive

All technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to the invention.

The article "a" as used herein refers to one or more (ie, at least one) grammatical terms of the article.

In one aspect, the invention provides a composition for treating or preventing a Re retrovirus infection comprising an effective amount of a peptide and a carrier selected from the group consisting of:

(1) a peptide having the amino acid sequence of SEQ ID NO: 1;

(2) a peptide having the amino acid sequence of SEQ ID NO: 2;

(3) a peptide having the amino acid sequence of SEQ ID NO: 3;

(4) a peptide having the amino acid sequence of SEQ ID NO: 4;

(5) A combination of one or more of the above (1) to (4).

In another aspect, the invention provides the use of the above-described peptide for the preparation of a composition for the treatment or prevention of a Re retrovirus infection.

In still another aspect, the present invention provides a method for killing a bacterium of the bacterium of the bacterium, comprising administering an effective amount of the above-mentioned peptide and a carrier to an article or environment which may contain a bacterium .

As used herein, the terms "protein", "polypeptide" or "peptide" are used interchangeably and refer to a biological polymer obtained by linking amino acids via peptide bonds. A particular sequence of proteins or polypeptides can be prepared using chemical synthesis or genetic engineering techniques.

The peptides according to the invention each have the amino acid sequence of SEQ ID NO: 1, 2, 3, or 4 and can be prepared by conventional chemical synthesis or genetic engineering techniques and with reference to the teachings and teachings herein. Related technologies, including polymerase chain reaction (PCR) amplification techniques, nucleic acid purification, plastids, enzyme processing of nucleic acid fragments, sequencing techniques, and protein expression and synthesis, can be found in Molecular Cloning: A Laboratory Manual, Second Edition, Cold Spring Harbor Laboratory Press, edited by Sambrook J et al., 1989, and Current Protocols in Molecular Biology, edited by Frederick MA et al., 2001, John Wiley & Sons, Inc.

The peptide according to the invention may also be isolated from a living organism (for example, fish or shrimp). For example, grouper antibacterial protein-1 (epinecidin-1) having the amino acid sequence of SEQ ID NO: 1 can be isolated from Epinephelus coioides (see DNA Cell Biol 2007; 26: 403-13) ). Furthermore, an anti-lipopolysaccharide factor (ALF) having the amino acid sequence of SEQ ID NO: 2 can be isolated from Penaeus monodon ; the anti-lipopolysaccharide molecule is a Gram-negative bacterium Endotoxin, including a linear structure or a cyclic structure, wherein the cyclic structure is formed by a disulfide bond between the C-terminal cystine acid and the N-terminal cystic acid in the amino acid sequence of the anti-lipopolysaccharide factor (see Int) Immunopharmacol 2007;7: 687-700). In one embodiment, the linear structure of the molecule is placed in 10% dimethyl sulfoxide (DMSO, Dimethyl sulfoxide), and the cyclic structure of the anti-lipopolysaccharide factor is obtained after reacting at room temperature for 24 hours. Further, hepatidin can be isolated from Oreochromis mossambicus , including TH1-5 and TH2-3, having the amino acid sequences of SEQ ID NOS: 3 and 4, respectively (see Mol immunol 2007). ;44: 1922-34).

A peptide according to the present invention includes a variant which refers to a peptide which is substantially homologous to the original peptide, but which results in an amino acid different from the peptide of the present invention due to one or more deletions, insertions or substitutions. The sequence, but retains substantially the same biological activity as the original peptide. The substitution may include a sequence of retention substitutions, meaning that the amino acid group is known to be substituted with a group having similar physiochemical properties. Suitable retention of the amino acid in the peptide or protein is well known to those of ordinary skill in the art and can be accomplished without altering the biological activity of the resulting molecule. Preferably, the variant has the same steric structure as the original protein peptide.

As used herein, the term "active" refers to the efficacy against R. vivax. In one embodiment, the activity can be demonstrated in accordance with the Minimization Inhibition Concentrations (MIC) of the R. cerevisiae according to the American National Laboratory for Clinical Microbiological Laboratory Standards. As shown in the following Table 1, the MIC of the peptide having the amino acid sequence of SEQ ID NO: 1 of the present invention is about 6.25-50 μg/ml for the R. cerevisiae; the present invention has the amino group of SEQ ID NO: 2. The cyclic structure of the acid sequence has a MIC of about 12.25-25 μg/ml, and the linear structure of the peptide has a MIC of about 6.25-25 μg/ml; the peptide having the amino acid sequence of SEQ ID NO: 3 of the present invention The MIC is about 25-400 μg/ml; and the MIC of the peptide having the amino acid sequence of SEQ ID NO: 4 of the present invention is about 100-450 μg/ml.

As used herein, "treatment or prevention" includes both medical treatment or prophylactic treatment for a disease. Those in need of such treatment include individuals who have developed the disease and individuals who need to prevent the disease from occurring. Therefore, in a specific embodiment, treating or preventing a disease according to the present invention means administering the peptide of the present invention to an individual who has been infected with R. vaginalis or an individual having the possibility and tendency to infect R. vaginalis, or is administered to the aforementioned The environment in which an individual may be exposed to the individual's tendency to heal, treat, alleviate, alleviate, alter, correct, improve, promote, or influence the symptoms of the infection.

As used herein, "effective amount" refers to a dose of the peptide of the present invention that achieves the above-described therapeutic or prophylactic effect in an individual to which it is treated. The dosage of the composition of the present invention may vary depending on various factors such as the route of administration, the weight and variety of the individual receiving the agent, and the purpose of administration. The skilled person can determine the dosage of the case based on experience and methods established by the art.

For carrying out the above treatment, the compositions of the present invention may be further formulated with an acceptable carrier in the desired dosage form. As used herein, "acceptable carrier" means that the carrier is compatible with the active ingredient contained in the compositions of the present invention, preferably to stabilize the active ingredient and not deleterious to the individual to be administered or the environment to be administered. The carrier can act as a diluent, carrier, excipient or base for the active ingredient. The compositions of the present invention can be formulated into the desired dosage forms using a variety of known conventional methods and suitable carriers.

In a specific embodiment, the compositions of the present invention are formulated into a pharmaceutical composition with a pharmaceutically acceptable carrier. Examples of pharmaceutically acceptable carriers include, but are not limited to, excipients such as lactose, dextrose, sucrose, sorbose, mannose, starch, gum arabic, calcium phosphate, alginate, tragacanth, gelatin , calcium citrate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, sterilized water, syrup and methylcellulose; lubricants, for example, talc, magnesium stearate and mineral oil; wetting agents; Suspending agents; preservatives, for example, methyl and propyl hydroxybenzoates; sweeteners; and flavoring agents.

The pharmaceutical compositions may be in the form of lozenges, pills, powders, lozenges, sachets, troches, elixirs, suspensions, emulsions, solvents, syrups, soft and hard gelatin capsules, suppositories, injection solutions and packaging powders. In a particular implementation, the compositions of the invention are in the form of an injectable solution.

The pharmaceutical composition can be delivered by any physiologically acceptable route, for example, orally, parenterally (e.g., muscle, intravenous, subcutaneous, intraperitoneal), transdermal, rectal, inhaled, and the like. In a particular implementation, the compositions of the invention are administered by injection.

In addition to the above formulation, the compositions of the present invention may also be formulated as feed supplements, added to feeds of animals (eg, poultry), or formulated as environmental antibacterial agents, for application to an environment in which the animal may be exposed (eg, poultry Growing farms or waters) to control infection by R. eutropha.

The peptide of the invention has a remarkable effect against R. rees, and can reach a concentration of bacteriostatic effect at a low concentration, which directly destroys the cell membrane of the cell, causes the intracellular substance to flow out of the cell, causes cell death, and has been confirmed to improve survival in the living body of the animal. Rate, with excellent treatment or prevention of infection with Rep.

In a specific example, the peptide of the present invention has a minimum inhibitory concentration against Streptococcus in the range of 6.25-450 μg/ml.

In yet another specific example, testing against common cultured ducks demonstrates that, whether the peptides and bacteria of the present invention are co-administered, the peptide of the present invention is applied in advance to receive a bacterial infection, or a bacterial infection is first applied. With the peptide of the present invention, the survival rate of the young ducks was significantly higher than that of the control group which only applied the bacteria.

Another aspect of the present invention provides a method for killing a bacterium of the genus of the bacterium, which comprises administering an effective amount of the above-described peptide and a carrier to an article or environment which may contain a bacterium.

In a specific example, the treatment of the bacterium with the peptide of the present invention, using a transmission electron microscope, clearly observes that after the treatment of the bacteria, the cell membrane exhibits an irregular shape and is broken, causing the intracellular material to flow out of the cell, causing the cell to be caused. death.

In one embodiment, the compositions of the present invention are formulated in a liquid, granular, or powder form. In particular, the compositions of the invention can be applied to farms or farmed waters of animals.

The invention will be further illustrated by the following examples, but the actual invention is not limited by the examples shown.

Example 1: Synthesis of Antibacterial Protein of the Invention

The antibacterial protein used in the present invention was synthesized by Gil Biochemical Corporation (Shanghai, China) using a solid phase procedure of Fmoc/tBu chemistry. Table 1 lists the names of the respective antibacterial proteins and the amino acid sequence.

The synthesized polypeptide is extracted, lyophilized and purified by reverse-phase high-performance liquid chromatography (RP-HPLC), and then the purified peptide is analyzed by mass spectrometry and high performance liquid chromatography. Molecular weight and purity. The purified polypeptide was reconstituted with phosphate buffer (pH 7.4) for subsequent experiments.

Example 2: Determination of minimum inhibitory concentration

This example tests the minimum inhibitory concentration of antibacterial proteins against several strains of R. vaginalis, including the A2, B2, T6, RA16, and T10 strains provided by Professor Yu Zhangyou from the Department of Veterinary Medicine, National Chiayi University, and National Taiwan University. The CFC27, CFC437, CFC363, RA3 and RA9 strains provided by Prof. Zhang Zhaofu from the Veterinary Research Institute and the MRS strain isolated from the Cherry Valley Duck of the private duck farm in Jiaoxi, Taiwan. The culture solution containing each strain is incubated with the antibacterial protein to be tested, and the minimum inhibitory concentration is determined by referring to the previously published microculture dilution method ( Int Immunopharmacol 2007; 7: 687-700). Table 2 shows the test results.

As shown in Table 2, the above antibacterial protein has antibacterial effects against various strains of Listeria species, and the minimum inhibitory concentration range is 6.25-450 μg/ml.

Example 3: Penetrating electron microscopy analysis

The experimental procedure for the transmission electron microscope is the same as that of the previously published literature ( Int Immunopharmacol 2007; 7: 687-700). Briefly, the bacterial cultures of the R. smegmatis strains T6, MRS and CFC27 were passed through a centrifuge (Beckman Coulter, Avanti J-20XP, JA-25.50, Palo Alto, CA, USA) at 3000 rpm at 4 °C. Centrifuge for 5 minutes. The precipitate was reconstituted with PBS at a concentration of 1.0 at OD550 nm. 5 ml of the bacterial reconstituted material was mixed with various antibacterial proteins (100 μg/ml) for 16 hours. The control group contained only bacterial reconstituted material (not treated with antibacterial protein) suspended in PBS or culture medium. After sectioning, observation was performed by a transmission electron microscope (Hitachi, H-7000, Tokyo, Japan) under conditions of 75 kV.

The results showed that the three R. reesei strains MRS (Fig. 1A), CFC27 (Fig. 1B) and T6 (Fig. 1C) treated with the antibacterial protein EPI, CA, LA, or TH1-5 of the present invention were clearly observed from the TEM. The treated control group showed significant differences in cell type; in the untreated control group, the cell membrane was intact without damage, while the bacteria treated with the antibacterial protein showed irregular shape and damage to the cell membrane, resulting in the outflow of intracellular substances. Cells that cause cell death.

Example 4: Animal Infection Experiment

This example used a pumpkin duck ( C. moschata duck) and a cherry valley duck for animal infection experiments. The source of the animals is a four-day-old duckling hatched from a general farm, fed with generally commercially available feed and sterile water, and the young ducks are kept in cages in an isolated room to maintain the temperature of the isolated room in accordance with standard operating procedures. The bacteria were injected directly into the abdominal cavity of the duck using a syringe (29 G x 1/2, Terumo Medical, Elkton, MD, USA) and the mortality was recorded daily.

In the first set of experiments, there were 30 tested ducklings in each group, and the control group received R. vaginalis (1 × 10 ⁸ colony forming units (cfu), each duck) or PBS; the experimental group received each Injection of a mixture of antibacterial proteins (100 μg per duckling) and R. brevis (1 x 10 ⁸ cfu per duckling).

The second set of experiments was followed by 336 hours of the first experiment, followed by additional injections of R. vaginalis (1 × 10 ⁶ cfu, each young duck).

In the third set of experiments, each group had 20 tested young ducks, first injected with R. rees (1 × 10 ⁸ cfu, each duckling), and then injected with PBS or each antibacterial protein after 2 or 24 hours. (100 μg, each duckling).

In the fourth set of experiments, each group had 20 tested young ducks, first injected with PBS or each antibacterial protein (100 μg per duckling), and then injected with R. falciparum after 2 or 24 hours (1× 10 ⁸ cfu, each duckling).

The above animal test, in the Cherry Valley duck, the strain of R. serrata is MRS, the tested antibacterial proteins are EPI, CA, LA, and TH1-5, and the interval between the antibacterial protein and the bacterium is 2 hours. In the pumpkin duck, the strain of the bacterium used is T6, the antibacterial proteins tested are EPI, CA, LA, TH1-5, and TH2-3, and the interval between the antibacterial protein and the bacterium is 24 hour.

Statistical analysis was performed on the data for each injection experiment using the t test to compare the differences between the two groups. Comparisons between groups were tested using SPSS software variant analysis (ANOVA). The different letters on the graph show significant differences between the two groups; the same letters show no significant differences between the two groups.

The results showed that EPI, CA, LA, and TH1-5 significantly reduced the mortality of MRS-infected Cherry Valley ducks (Figure 2); while EPI, CA, LA, TH1-5, and TH2-3 significantly reduced the mortality Mortality of T6-infected pumpkin ducks (Figure 3).

As shown in Fig. 2(A), the mortality of Cherry Valley ducks was extremely high within 14 days after infection with MRS strain (1×10 ⁸ cfu/duck) (23 out of 30 animals in 10 days died); The 14-day survival rate of animals with antibacterial protein LA or TH1-5 (100 μg/ml) and MRS strains reached 93.33% (2 out of 30 animals in three days), and the antibacterial protein EPI was applied together. The 14-day survival rates of animals with either CA and MRS strains were also 76.66% (7 out of 30 animals in nine days) and 73.33% (8 out of 30 animals in nine days died). In the above experiments in which antibacterial proteins and bacteria were commonly applied, the survival rate of the young ducks was significantly higher than that of the control group (p<0.05).

As shown in Fig. 2(B), after the animals received the above-mentioned common antibacterial protein and bacteria, the MRS strain (1 × 10 ⁸ cfu/duck) was again applied after 14 days, and the results showed that only the bacteria were injected. The survival rate of ducks was only 16.66%. The survival rate of ducklings that had previously received antibiotics EPI, CA, LA, or TH1-5 and bacteria increased to 47.82%, 63.63%, 71.42% and 75%, respectively. .

As shown in Figures 2(C) and 2(D), the survival rate of young ducks is significantly higher than that of the bacteria, whether it is a bacterial infection or an antibacterial protein, or an antibacterial protein is applied in advance. And PBS control group.

Similarly, as shown in Fig. 3(A), the survival rate of pumpkin ducks was only 30% within 14 days after infection with T6 strain (1×10 ⁸ cfu/duck), and the survival rate of young ducks administered only with PBS was 14-day survival rate of animals with 100% antibacterial protein EPI, TH1-5, LA, TH2-3, or CA (100 μg/ml) and T6 strain (1×10 ⁸ cfu/duck) They are 100%, 75%, 75%, 60% and 60% respectively. In the above experiments in which antibacterial proteins and bacteria were jointly applied, the survival rate of the young ducks was significantly higher than that of the control group only.

Further, as shown in Fig. 3(B), after the pumpkin duck received the above-mentioned common antibacterial protein and bacteria, the T6 strain (1 × 10 ⁸ cfu/duck) was again applied after the 14th day, and the results showed that only the bacteria were received. The survival rate of the injected ducklings was only 40%, while the young ducks who had previously received the antibacterial protein TH2-3, EPI, TH1-5, LA, or CA and bacteria had a higher survival rate of 91.7%. 90%, 86.7%, 86.7%, and 83.3%.

Furthermore, as shown in Fig. 3(C), animals that received only T6 strains caused high mortality within 14 days (20 of 20 animals died within 4 days); in contrast, if After 24 hours of T6 strain application, the antibacterial proteins EPI, TH2-3, TH1-5, LA or CA were applied, and the survival rate of the animals was higher, 100%, 95%, 90%, 85%, and 80%, respectively. .

Finally, as shown in Fig. 3(D), the animals that received only T6 strains had a mortality rate of 80% in 14 days; in contrast, if the antibacterial proteins EPI, CA, LA, TH1-5 were applied in advance, And TH2-3 (100 μg), T6 strain (1 × 10 ⁸ cfu / duck) was applied after 24 hours, the animal survival rate was higher, 70%, 50%, 60%, 55%, and 65, respectively. %.

Example 4: Bactericidal activity test of antibacterial protein in animal liver

Young ducks receive different ways of applying antibacterial proteins and/or strains. In the first set of experiments, the animals were first beaten with T6 strain (1×10 ⁸ cfu/duck), and after 24 hours, the antibacterial proteins EPI, CA, LA, TH1-5 or TH2-3 (100 μg/duck) were applied. . In the second set of experiments, the animals were first given antibacterial protein EPI, CA, LA, TH1-5 or TH2-3 (100 μg/duck), and T6 strain (1×10 ⁸ cfu/duck) was applied 24 hours later. . In the third set of experiments, animals were co-administered with T6 strain (1 x 10 ⁸ cfu/duck) and antibacterial proteins EPI, CA, LA, TH1-5 or TH2-3 (100 μg/duck). After the animals received 1, 3, 5, and 7 days of application, liver tissues were taken for analysis. The liver tissue was first homogenized, placed in PBS, and spread on a sheep blood agar plate, and then the plate was placed at 37 ° C for 24 hours, and finally the number of bacterial colonies was counted separately. Significant differences were calculated using the SAS analysis program (p < 0.05).

The results showed that whether the antibacterial proteins EPI, CA, LA, TH1-5 or TH2-3 were applied simultaneously with the bacteria (Fig. 4A), after the infection (Fig. 4B), or before the infection (Fig. 4C), the injection After 7 days, the number of bacteria in the liver of the animals was significantly reduced (p<0.05).

Summarizing the above test, the present invention confirmed that the antibacterial proteins EPI, CA, LA, TH1-5 or TH2-3 have bactericidal activity against R. vaginalis, and can significantly improve the survival rate of infected animals. In view of the various side effects of antibiotics and the increasing number of resistant strains, the present invention demonstrates that the above-mentioned antibacterial proteins have the potential to develop into agents or adjuvants for the prevention of infection with R. vaginalis, particularly in the waterfowl farming industry.

Those skilled in the art will be able to make changes and modifications in accordance with the embodiments without departing from the spirit of the invention. It is to be understood that the invention is not to be limited by the scope of the embodiments disclosed herein,

The foregoing description and the embodiments may achieve better illustrative effects by the additional drawings. To enhance the description of the invention, the drawings of the appropriate embodiments are set forth herein. It is to be noted that the present invention is not limited to the descriptions set forth herein.

Figure 1 shows a transmission electron microscope image of R. eutropha after treatment with an antibacterial protein. Figure 1 (A) is for the MAS strain, (a) to (e) are untreated MAS strain, MAS strain treated with antibacterial protein EPI, MAS strain treated with antibacterial protein CA, treated with antibacterial protein LA The MAS strain, and the MAS strain treated with the antibacterial protein TH1-5, and (f) to (j) are the results of repeated experiments under the same conditions of (a) to (e). Figure 1 (B) is for CFC27 strain, (a) to (e) are untreated CFC27 strain, CFC27 strain treated with antibacterial protein EPI, CFC27 strain treated with antibacterial protein CA, treated with antibacterial protein LA The CFC27 strain, and the CFC27 strain treated with the antibacterial protein TH1-5, and (f) to (j) are repeated test results under the same conditions of (a) to (e). Figure 1 (C) is for the T6 strain, (a) to (e) are untreated T6 strain, T6 strain treated with antibacterial protein EPI, T6 strain treated with antibacterial protein CA, treated with antibacterial protein LA The T6 strain, and the T6 strain treated with the antibacterial protein TH1-5, and (f) to (j) are repeated test results under the same conditions of (a) to (e).

Figure 2 shows that the antibacterial protein of the present invention can increase the survival rate of Cherry Valley duck infected by the RES. In Fig. 2(A), RA indicates that only the MAS strain was applied; PBS indicates that only PBS was applied; EPI, CA, LA, and TH1-5 indicated that each of the antibacterial proteins and R. faecalis were co-administered. In Fig. 2(B), each line shows the result of additionally injecting R. eutropha under the same conditions as in Fig. 2(A). In Fig. 2(C), RA-2hr-PBS was first administered with R. vaginalis, and then PBS was administered 2 hours later. RA-2hr-TH1-5 was first administered with R. vaginalis and then administered 2 hours later. Antibacterial protein TH1-5; RA-2hr-LA is first applied to the bacterium, and then the antibacterial protein LA is applied after 2 hours; RA-2hr-CA is first applied to the bacterium, and then the antibacterial protein is applied after 2 hours. CA; and RA-2hr-EPI were first administered with R. rapae, and the antibacterial protein EPI was applied after 2 hours. In Fig. 2(D), PBS-2hr-RA is first administered with PBS, and then 2 days after the application of Rep.; TH1-5-2hr-RA is first applied antibacterial protein TH1-5, and then 2 hours After the application of R. serrata; LA-2hr-RA is first applied antibacterial protein LA, and then 2 days after the application of Rep.; CA-2hr-RA is first applied antibacterial protein CA, and then 2 hours after the application of Rey Bacillus; and EPI-2hr-RA was first applied with the antibacterial protein EPI, and then treated with R. falciparum 2 hours later.

Figure 3 shows that the antibacterial protein of the present invention can increase the survival rate of pumpkin duck infected by the R. serovar T6 strain. In Fig. 3(A), RA indicates that only R. serrata was administered; PBS indicates that only PBS was applied; EPI, CA, LA, TH1-5, and TH2-3 indicated that each antibacterial protein and R. rees were co-administered. In Fig. 3(B), each line shows the result of additionally injecting R. eutropha under the same conditions as in Fig. 3(A). In Fig. 3(C), RA-24hr-PBS is first administered with R. vaginalis, and then PBS is administered after 24 hours; RA-24hr-TH2-3 is first applied to R. rapae, and then applied after 24 hours. Antibacterial protein TH2-3; RA-24hr-TH1-5 is first applied to the bacterium, and then the antibacterial protein TH1-5 is applied after 24 hours; RA-24hr-LA is first applied to the bacterium, and after 24 hours Apply antibacterial protein LA; RA-24hr-CA is first applied to R. rapae, and then apply antibacterial protein CA after 24 hours; and RA-24hr-EPI is first applied to R. serrata, and then applied antibacterial after 24 hours. Protein EPI. In Fig. 3(D), PBS-24hr-RA was first applied with PBS, and then strained with T. reesei after 24 hours; TH2-3-24hr-RA was first applied with antibacterial protein TH2-3, then 24 After the hour, Streptococcus faecalis was administered; TH1-5-24hr-RA was first applied to the antibacterial protein TH1-5, and then 24 hours later, the bacterium was killed; LA-24hr-RA was first applied antibacterial protein LA, then 24 After 24 hours, the antibiotics CA was applied. CA-24hr-RA was first applied with antibacterial protein CA, and then 24 hours later; and EPI-2hr-RA was first applied with antibacterial protein EPI, and then applied 24 hours later. Hitella bacillus.

Figure 4 shows the results of bactericidal activity tests of antibacterial proteins against the R. reuteri T6 strain in the liver of animals. In Fig. 4(A), RA indicates that only R. serrata is administered; and EPI+RA, CA+RA, LA+RA, TH1-5+RA, and TH2-3+RA indicate that each antibacterial protein and thunder are jointly applied. Bacillus. In Fig. 4(B), RA indicates that only R. serrata is administered; RA-24hr-TH2-3 is first applied to R. rapae, and after 24 hours, antibacterial protein TH2-3 is applied; RA-24hr-TH1-5 It is first applied to the bacterium, and then the antibacterial protein TH1-5 is applied after 24 hours; the RA-24hr-LA is first applied to the bacterium, and then the antibacterial protein LA is applied after 24 hours; the RA-24hr-CA is the first The antibacterial protein CA was applied after 24 hours; and the RA-24hr-EPI was first applied to the bacterium, and the antibacterial protein EPI was applied after 24 hours. In Fig. 4(C), RA indicates that only R. serrata was administered; TH2-3-24hr-RA was first applied to the antibacterial protein TH2-3, and then 24 hours after the administration of R. rees; TH1-5-24hr-RA The antibacterial protein TH1-5 was first applied, and then the bacterium was killed by 24 hours; LA-24hr-RA was first applied with the antibacterial protein LA, and then the bacterium was killed after 24 hours; CA-24hr-RA was the first The antibacterial protein CA was applied, and the R. eutropha was administered after 24 hours; and the EPI-2hr-RA was first applied with the antibacterial protein EPI, and then the R. eutropha was administered 24 hours later.

Claims (8)

Use of a peptide selected from the group consisting of a composition for treating or preventing a infection with a Re retrovirus: (1) a peptide having the amino acid sequence of SEQ ID NO: 1; a peptide of the amino acid sequence of SEQ ID NO: 2; (3) a peptide having the amino acid sequence of SEQ ID NO: 3; (4) having the amino acid sequence of SEQ ID NO: a peptide; and (5) a combination of one or more of the above (1) to (4). The use according to the first aspect of the patent application, wherein the peptide having the amino acid sequence of SEQ ID NQ: 2 has a linear or cyclic structure. The use according to the first aspect of the patent application, wherein the composition is for treating or preventing a Reyebacterium infection of a bird. According to the use of the third aspect of the patent application, wherein the bird is a duck. A method for killing a bacterium of the bacterium of the genus in vitro, comprising administering to a substance or environment which may contain a bacterium of the genus: a composition comprising an effective amount of a peptide selected from the group consisting of: 1) a peptide having the amino acid sequence of SEQ ID NO: 1; (2) a peptide having the amino acid sequence of SEQ ID NO: 2; (3) having an amino group of SEQ ID NO: a peptide of an acid sequence; (4) a peptide having the amino acid sequence of SEQ ID NO: 4; and (5) a combination of one or more of the above (1) to (4). The method of claim 5, wherein the peptide having the amino acid sequence of SEQ ID NO: 2 has a linear or cyclic structure. The method of claim 5, wherein the composition is formulated into a liquid, a granule, or a powder. The method of claim 5, wherein the environment is a farm or aquaculture water.