KR101209964B1 - Vaccine composition for swine polyserositis and manufacturing method thereof - Google Patents

Abstract

Porcine multiple meningitis preventive vaccine of the present invention is very excellent in safety, preservation and vaccine effect. In addition, the vaccine for preventing swine multiple meningitis of the present invention can be directly administered to pigs exposed to the risk of swine multiple meningitis to prevent pigs from multiple meningitis, and can be administered to pregnant sows by being capable of maternal transfer antibodies. It is possible to prevent multiple meningitis from piglets.

Description

Vaccine composition for swine polyserositis and manufacturing method

The present invention relates to a vaccine composition for preventing swine polymeningitis and a method for producing the same, and more particularly, to a vaccine composition for preventing swine polymenoritis and a method for producing the same, which can effectively prevent multiple meningitis caused by Streptococcus.

Multiple meningitis, one of the most common diseases in domestic pig farms, is caused by various areas such as joints, pleura, pericardium, and peritoneum. It is accompanied by serous effusion and causes systemic inflammation of the intestinal membrane.

The causative agent of multiple meningitis is present in most pig farms, and due to the development of infection, delayed growth of broilers, mortality, delayed delivery date, and decreased feed efficiency, and administration of drugs for treatment. It is causing a great economic loss to the farm.

Meanwhile, the representative causative agent of multiple meningitis is Haemophilus parasuis ) and Streptococcus suis ). Among the cases of Glacier's disease caused by Haemophilus parasuis, it occurs nationwide regardless of region, and its prevalence is mainly fibrosis peritonitis, pleurisy, pericarditis and meningitis. Symptoms of multiple meningitis have been developed, and a number of therapeutic or preventive vaccines have been developed and sold for the purpose of preventing or treating the disease.

On the other hand, Streptococcus ( Streptococcus) in many pig farms in the domestic pig industry in which a large number of broilers are concentrated and raised. suis ) is assumed to be distributed. Streptococcus serotypes are a group of strains that each form an independent specific antibody. Although various streptococcal strains that could not be separated by other serotypes have been found, studies on swine multiple meningitis caused by streptococci have been insufficient.

Therefore, it is urgent to research streptococci that cause multiple meningitis and to solve the damage caused by multiple meningitis in domestic pig farms every year.

In addition, streptococci causing multiple meningitis in Korea and streptococci causing multiple meningitis in foreign countries have different serotypes, and thus, effective application of imported vaccines prevents effective protection against streptococci, which causes multiple meningitis in Korea. There is a need for new identification and new development of vaccines to prevent multiple meningitis caused by streptococci.

The present invention has been made in order to solve the above problems, the first problem to be solved of the present invention is to identify streptococci causing multiple meningitis in domestic pig farms, using the causative bacteria vaccine that can prevent multiple meningitis To provide.

It is to protect the pigs from multiple meningitis by using the above-prepared porcine multiple meningitis vaccine of the present invention.

In order to achieve the first object of the present invention, the present invention is a streptococcus ( Streptococcus) causing multiple meningitis of pigs suis serotype 2 ) Attenuated or inactivated serotype 2 provides a vaccine composition for the prevention of swine multiple meningitis containing it as an active ingredient.

According to a preferred embodiment of the present invention, the Streptococcus suis serotype 2 may comprise a high antigenic Mrp gene and an Epf gene which is a high pathogenic gene.

According to a preferred embodiment of the present invention, the Streptococcus suis serotype 2 may comprise the 16S rRNA sequence of SEQ ID NO.

According to another preferred embodiment of the present invention, the Streptococcus suis ) serotype 2 can be used as deposited with the Korean National Center for Microbiological Conservation under Accession No. KCCM 11111P.

According to another preferred embodiment of the present invention, the inactivation may be inactivated by formalin treatment.

According to another preferred embodiment of the present invention, the vaccine composition may further comprise an aluminum hydroxide gel as an adjuvant.

According to another preferred embodiment of the present invention, there is provided a method for preventing multiple meningitis disease by administering the vaccine composition of the present invention to swine, more preferably the vaccine composition is oral, nasal, anal, intramuscular , Intradermal, intravenous or subcutaneous administration.

According to another preferred embodiment of the present invention, streptococci of Accession No. KCCM 11111P, which causes multiple meningitis in pigs, and a vaccine composition comprising the same as an active ingredient are provided.

According to another preferred embodiment of the present invention, (1) taking a sample from a pig infected with multiple meningitis and culturing it to produce a colony; And (2) PCR using the forward primer represented by SEQ ID NO: 1 and the reverse primer represented by SEQ ID NO: 2 to the cultured colonies, and have a homologous sequence of 95% or more with 16S rRNA sequence of SEQ ID NO: It provides a method for producing a vaccine composition for preventing swine polysclerosis comprising the step of: identifying the Streptococcus suis serotype 2 strain.

According to another preferred embodiment of the present invention, the method may further include selecting colonies having the morphological characteristics of Streptococci from the colonies cultured between the steps (1) and (2).

According to another preferred embodiment of the present invention, (3) selecting a strain comprising the Mrp gene in the identified strain; (4) selecting a strain containing the Epf gene from the strain containing the Mrp gene may further include.

According to another preferred embodiment of the present invention, the step (3) may be carried out PCR using the forward primer represented by SEQ ID NO: 3 and the reverse primer represented by SEQ ID NO: 4.

According to another preferred embodiment of the present invention, the step (4) may be carried out PCR using the forward primer represented by SEQ ID NO: 5 and the reverse primer represented by SEQ ID NO: 6.

Porcine multiple meningitis preventive vaccine of the present invention is very excellent in safety, preservation and vaccine effect. In addition, the vaccine for preventing swine multiple meningitis of the present invention can be directly administered to swine exposed to the risk of swine multiple meningitis to prevent swine from multiple meningitis caused by Streptococcus serotype 2 strains, and may be a maternal transfer antibody. By administering to pregnant sows, multiple meningitis caused by Streptococcus serotype 2 strains occurring in the piglets can be prevented.

1 is a gel photograph taken by electrophoresis after PCR reaction with Streptococcus having Mrp gene which is a high antigenic gene (M; marker, 1; mrp positive strain, 2; mrp negative strain).
2 is a gel photograph taken by electrophoresis after PCR reaction with Streptococci having Epf gene, a gene having pathogenicity and antigenicity. (M; marker, 1; mrp positive strain, 2; mrp) Negative strain).

As described above, not only studies on streptococcal polycysticitis caused by streptococcus have been conducted properly, but also streptococci causing multiple meningitis in Korea and streptococci causing multiple meningitis in foreign countries have been identified. Different from each other, the application of imported vaccines did not provide effective protection.

Therefore, the present invention provides a vaccine composition for the prevention of swine multiple meningitis, which attenuates or inactivates Streptococcus suis serotype 2 serotype 2, which causes multiple meningitis in pigs, and contains the same as an active ingredient. Sought solution.

Here, the multiple meningitis is a disease in which inflammation occurs in various parts of the pleura, pericardium, peritoneum, accompanied by serous effusion, and fibrinous pericarditis in which fibrin is deposited in the pericardium of the heart. ), And in the peritoneum, fibrinous peritonitis in which fibrin is deposited in the intestinal membrane of the peritoneum causes systemic inflammation. However, in the present invention, the multiple meningitis is Streptococcus suis ), and is limited to multiple meningitis caused by pigs.

Streptococcus suis ) serotype 2 pathogen is attenuated or inactivated streptococci corresponding to serotype 2 according to the classification of serotypes of ordinary streptococci, and used as an active ingredient of a vaccine composition for the prevention of multiple meningitis in pigs. such, the vaccine compositions prepared via this is administered directly to a pig exposed to a pig multiple tent salt risk streptococci (Streptococcus suis ) can prevent pigs from multiple meningitis caused by infection with serotype 2, and maternal transfer antibody is possible, and administration to pregnant sows can prevent multiple meningitis from piglets.

Specifically, the vaccine composition for the prevention of swine polymeningitis of the present invention preferably comprises the steps of: (1) collecting a sample from a pig infected with multiple meningitis and culturing it to produce colonies; And (2) PCR using the forward primer represented by SEQ ID NO: 1 and the reverse primer represented by SEQ ID NO: 2 to the cultured colonies, and have a homologous sequence of 95% or more with 16S rRNA sequence of SEQ ID NO: Identification of Streptococcus suis serotype 2 strain can be prepared.

First, as a step (1), a sample is taken from a pig infected with multiple meningitis and cultured to produce colonies. Thereafter, the method may further include selecting colonies having morphological characteristics of streptococci from the colonies cultured between the steps (1) and (2). In this case, the morphological characteristics of streptococci in colonies generally mean the morphological characteristics of streptococci, which are widely known. Preferably, when the colonies have passed 48 hours, the colonies are as small as about 1 mm, the edges are circular, the color is milky, and the colonies. You can choose to create a green or transparent hemolytic area around you.

Then (2) PCR can be performed using the forward primer represented by SEQ ID NO: 1 and the reverse primer represented by SEQ ID NO: 2 to the cultured colonies to identify Streptococcus suis.

(3) may further comprise the step of selecting a strain comprising Mrp gene (Genbank No. AM493987.1) in the identified strain. Specifically, streptococci containing high antigenic Mrp genes may express more antigens required for the immune response of the vaccine when cultured. On the other hand, step (3) can be carried out PCR using the forward primer represented by SEQ ID NO: 3 and the reverse primer represented by SEQ ID NO: 4.

Thereafter, step (4) may further include selecting a strain including the Epf gene (Genbank No. X64450) from the strain containing the Mrp gene. The Epf gene is a gene related to both pathogenicity and antigenicity possessed only by Streptococcus suis type 2, and thus it is possible to select strains that are more favorable for antibody production. In step (4), PCR may be performed using the forward primer represented by SEQ ID NO: 5 and the reverse primer represented by SEQ ID NO: 6.

In order to find suitable strains for vaccine production among the isolated streptococci, the present inventors finally isolated strains containing both the highly antigenic Mrp gene and the highly pathogenic gene Epf gene, and died in Korea by multiple meningitis. Newly Isolated Streptococcus from Streptococcus subtype of the suis strain was identified and identified as a strain belonging to serotype 2 through an antiserum aggregation reaction.

Thus, the identified streptococci have more than 95%, preferably the same homology with the 16S rRNA sequence of SEQ ID NO: 7, and deposited with the Korean microbial conservation center (KCCM) as accession number KCCM 11111P.

Porcine multiple meningitis preventive vaccine of the present invention is characterized in that it is prepared by the conventional attenuated or inactivated (inactvation) of the streptococcal serotype 2.

Attenuation or inactivation of the causative agent of the disease can be carried out in a conventional manner. That is, the antigen can be exposed to chemical agents such as formaldehyde, paraformaldehyde, beta-propiolactone, ethyleneimine or derivatives thereof, or inactivated by heat, irradiation or the like. Attenuation can be achieved by ultraviolet (UV) irradiation of disease-causing organisms, by chemical treatment or by high-order serial passage in vitro. Attenuation can also be achieved by making clear genetic changes, for example by deleting certain sequences of the sequences of strains known to provide toxicity or by inserting sequences into the genome of the strain.

In the present invention, the inactivation of the streptococcal serotype 2 can be preferably used with formalin. Inactivation with formalin (formaldehyde) has already been demonstrated to be reliable, certainty, and stable in achieving inactivation compared to inactivation with other chemicals, providing a stable product. This inactivation method is suitable for mass production, saving manpower and production costs, and has been recognized worldwide for its Japanese encephalitis virus vaccine, which has been used for decades.

The vaccine may further comprise a pharmaceutically acceptable carrier in addition to the inactivated streptococcal serotype type 2 antigen. A pharmaceutically acceptable carrier herein means a carrier medium that does not interfere with the effectiveness of the biological activity of the active ingredient, is chemically inert and not toxic to the subject to be administered. Various carriers are known in the art and may include distilled, brine or mineral water.

The compositions of the present invention may also include one or more pharmaceutically or veterinary acceptable preservatives. For example, it may include an EDTA salt, methiolate, and the like.

Adjuvants can be added to vaccines comprising inactivated streptococcal serotype 2 according to techniques known to those skilled in the art. Examples of adjuvants are mineral gels such as aluminum hydroxide, surfactants such as resorcithin, glycosides such as saponin or saponin derivatives, lantitidine, Avridine, carbopol, polyphosphazene, dimethyldioctadecylammonium bromide (DDA), quill Quill A, ampigen, mineral oil, etc. are mentioned.

Vaccines comprising the inactivated streptococcal serotype type 2 can be prepared according to standard techniques well known to those skilled in the pharmaceutical and veterinary arts. The content of such vaccines can be determined by those skilled in the medical and veterinary arts, taking into account such factors as the age, sex, weight, species and condition of the subject to be administered and the route of administration. This composition may be administered alone or in combination with other immunological antigens or vaccine compositions or therapeutic compositions or for subsequent administration.

Examples of formulations in which the vaccine of the invention may be used include liquid preparations such as oral, nasal, anal, vaginal, intragastric suspensions, syrups and the like and sterile suspensions or emulsions for subcutaneous, endothelial, intramuscular, or intravenous administration. It may include a formulation. It may also be a solid preparation of soluble components or particulates which is resuspended in a pharmaceutically acceptable diluent prior to use. Soluble components or particulates can be prepared according to techniques known to those skilled in the pharmaceutical and veterinary arts, such as coacervation, biacervation, coagulation, spray drying, bubble drying, precipitation, lyophilization and the like.

The present invention also provides a method for preventing multiple meningitis by administering the vaccine to pigs exposed to the risk of developing multiple meningitis caused by Streptococcus serotype 2 strain. On the other hand, since the vaccine composition is difficult to apply cross-prevention, it is difficult to use to prevent multiple meningitis of pigs caused by streptococcal serotypes 3 and 4 infection.

The vaccine can be administered to pigs through a variety of veterinary acceptable routes. For example, it may be administered by oral administration in addition to feed or drinking water, instillation into the eye mucosa, intramuscular injection, intravenous injection, intradermal injection and the like. Depending on the route of administration, those skilled in the veterinary art can determine the dosage. For example, 0.1 to 1 ml can be inoculated per 1 kg of body weight, and after the first inoculation, a certain time (eg, 1 to 2 months) has elapsed, and the second inoculation can be performed.

At this time, when streptococcal polysclerosis is induced in piglet farms during the 7 to 18-day-old young piglets, there is no time for direct inoculation of the vaccine into the young sows, so the pregnant sows are inoculated with the vaccine. Thus, antibodies formed in sows can be transferred to mammalian pigs through colostrum, thereby preventing multiple meningitis caused by Streptococcus serotype 2.

When multiple meningitis is induced in weaning pigs or growing pigs of 4 weeks or older, the vaccine can be directly inoculated to mammalian pigs, weaning pigs, or growing pigs to prevent multiple meningitis caused by Streptococcus serotype 2.

Hereinafter, the present invention will be described in detail with reference to the preferred embodiments and experimental examples so that those skilled in the art can easily practice the present invention. However, the following Examples and Experimental Examples are provided only to explain the present invention more easily, and the content of the present invention is not limited by the Examples.

[ Example  One]

Step 1: Colony  Primary separation using morphological features

A case of fibrosis epicarditis, pleurisy, and peritonitis was examined among pigs infected with multiple meningitis at a domestic pig farm and referred to the Seoul National University Pathology Department.

The necropsy mass was carefully opened with a dissected autopsy mass to prevent organ contamination during autopsy, and samples were taken from the intestinal membrane site where fibroplastic lesions were observed with a sterile swab. The collected samples were applied to agar medium containing 5% cotton blood and then evenly applied using platinum teeth.

The medium was placed in an incubator at 37 ° C., expanded for 24 hours in aerobic culture, and observed under a microscope. The morphological characteristics of streptococci were as small as 1 mm, rounded at the edges, milky, and around the colonies after 48 hours. About 50 independent colonies with green or transparent hemolytic sites were selected and second cultured with platinum in TSA (Trypticase soy agarose) medium.

Step 2: 16S rRNA  Secondary Separation Using Sequences

In order to isolate only Streptococci among 50 strains cultured in step 2 above, PCR reactions were performed using primer sequences designed from known Streptococcus 16S rRNA sequences.

In order to extract the strain DNA required for the PCR reaction, the secondary cultured strain was dissolved in 1 ml of sterile PBS buffer, suspended and heated in boiling water for 10 minutes. After centrifugation at 3000 rpm for 5 minutes, the supernatant from which DNA was extracted was separated, and DNA for each strain was extracted.

In addition, in order to prepare a primer specific for Streptococcus 16s rRNA, a 5'-AGAGTTTGATCCTGGCTCAG-3 'sequence (SEQ ID NO: 1) was used as a forward primer sequence using a known Streptococcus 16s rRNA sequence. As the reverse primer sequence, a 5'-CGGGATCCCAGGCCCGGGAACGTATTCAC-3 'sequence (SEQ ID NO: 2) was prepared and used.

5 μl of DNA extracted from the strain, forward primer 10 nM, reverse primer 10 nM, TLA polymerase (Bioneer, Daejeon, Korea) 1U, KCl 50mM, MgCl ₂ After mixing 1 mM and Tris-HCl (pH8.3) 10 mM, PCR reaction (GeneAmp PCR System 9700, Applied Biosystems, Foster city, CA, USA) was performed with 50 µl of the reaction solution prepared by adding distilled water. .

The PCR reaction was repeated for 5 minutes at 95 ° C. for 5 minutes, followed by 30 seconds at 94 ° C., 1 minute at 60 ° C., 1 minute at 72 ° C., and finally at 72 ° C. for 5 minutes. Under conditions.

Through the above PCR reaction, 28 strains with 16S rRNA PCR products were identified, and they were genetically identified as streptococci.

Step 3: Isolation and Identification of Streptococci

Streptococci suitable for vaccine production were isolated and identified from 28 isolates of streptococci.

Suitable strains for the preparation of such vaccines were isolated and identified strains having Mrp gene, which is a high antigenic gene, and Epf gene, a gene involved in both pathogenicity and antigenicity.

First, in order to select a strain having an Mrp gene, 5 μl of DNA extracted from each strain and an omnidirectional primer (5′-ATTGCTCCACAAGAGGATGG-3 ′) (SEQ ID NO: 3) in the same manner as in the DNA extraction method of Example 2 above. ) 10nM, reverse primer (5'-TGAGCTTTACCTGAAGCGGT-3 ') (SEQ ID NO: 4) 10nM, TLA polymerase (Bioneer, Daejeon, Korea) 1U, KCl 50mM, MgCl2 1mM, and Tris-HCl (pH8.3) 10mM After mixing, a PCR reaction (GeneAmp PCR System 9700, Applied Biosystems, Foster city, CA, USA) was performed with 50 µl of the reaction solution prepared by adding distilled water to select strains having the Mrp gene.

At this time, the PCR reaction was heated for 2 minutes at 95 ℃, then repeated 40 times the reaction of 30 seconds at 94 ℃, 30 seconds at 58 ℃, 30 seconds at 72 ℃, and finally at 72 ℃ for 2 minutes It was carried out under the reaction conditions.

As shown in FIG. 1, after the PCR reaction, 10 μl was electrophoresed on 2% agarose gel to which 0.02% ET-Br was added for 40 minutes to select a strain having the 188 bp Mrp gene. It was.

In order to select a strain having the Epf gene among the first selected strains, 5 μl of DNA extracted from each strain and an omnidirectional primer (5′-GCTACGACGGCCTCAGAAATC-) were selected in the same manner as the DNA extraction method of Example 2 above. 3 ') (SEQ ID NO: 5) 10 nM, reverse primer (5'-TGGATCAACCACTGGTGTTAC-3') (SEQ ID NO: 6) 10 nM, TLA polymerase (Bioneer, Daejeon, Korea) 1U, KCl 50 mM, MgCl2 1 mM, and Tris-HCl (pH8.3) After mixing 10mM, a PCR reaction (GeneAmp PCR System 9700, Applied Biosystems, Foster city, CA, USA) was carried out with 50 µl of the reaction solution prepared by adding distilled water, and the strain containing the Epf gene. Was selected.

In this case, the PCR reaction is heated for 10 minutes at 95 ℃, then repeated 40 times 1 minute at 94 ℃, 55 seconds at 60 ℃, 2 minutes at 72 ℃ 40 times, and finally for 10 minutes at 72 ℃ It was carried out under the reaction conditions.

As shown in FIG. 2, after the PCR reaction, 10 μl was electrophoresed on 2% agarose gel to which 0.02% ET-Br was added for 40 minutes to finally isolate a strain containing 626 bp Epf gene. I identified it.

The strain finally isolated by the above method was newly isolated streptococcus ( Streptococcus) from pigs killed by multiple meningitis in Korea. suis ) is a subtype of the strain, and the present inventors used the University of Montreal, Canada (Groupe de Recherche sur les Maladies Infectieuses du Porc, Faculte de medecine veterinaire, Universite de Montreal) to verify the serotypes of the final isolated streptococci. As a result of requesting the antiserum aggregation reaction, it was confirmed that it is serotype 2 and deposited with KCCM 11111P at the Korea Microorganism Conservation Center.

The deposited KCCM 11111P was confirmed to have a 16s rRNA sequence represented by SEQ ID NO: 7.

Step 4 : Streptococcus serotype 2  Preparation of the Used Vaccine

The 1 x 10 ⁵ CFU (colony forming unit) Streptococcus serotype 2 strain (Accession No. KCCM 11111P) identified in step 3 contains 1% horse serum and 1% yeast extract. After inoculation into 100 ml of brain heart infusion (BHI) broth, the cultures were stirred with an incubator at 37 ° C. for 12 hours.

The Streptococcus serotype 2 strain was cultured until propagated at 1 × 10 ⁸ CFU, and then 0.8 ml of formalin was added to 100 ml of the grown culture, followed by stirring for 24 hours or more to inactivate Streptococcus type 2.

To 100 ml of the inactivated culture solution, 8 ml of aluminum gel was added and stirred for at least 24 hours to prepare a vaccine. After the vaccine was prepared, the presence or absence of sterilization was confirmed that there was no growth of mold and bacteria.

Experimental Example  1. General Criteria Test of Vaccine

1-1. Characteristic test

As a result of testing according to the general standard for animal biological preparations, the vaccine of the above preparation was a milky white suspension, which had no foreign matter, no odor, and a uniform content of the divided contents.

1-2. Sterility test

Aseptic test results for the vaccine of the above preparation was obtained by culturing for 7 days at 22 ° C. and 37 ° C. after culturing in Nutrient agar, Nutrient broth, Thioglycollate medium, and any bacteria and fungi. The development of was not recognized.

Experimental Example  2. Prevention of Multiple Meningitis Using Maternal Transition Antibodies

If streptococcal polysclerosis is caused in a piglet period between 7 and 18 days of delivery in a hog farm, it is too young to give vaccines to pregnant sows. We tested whether the immunity formed in sows was transferred through colostrum and prevented streptococcal multiple meningitis in lactating piglets.

Specifically, streptococci isolated from Example 3 above were inoculated with the vaccine by intramuscular injection of 2cc of the vaccine of the above preparation 5 weeks before delivery and 3 weeks before delivery to the pregnant sows. Streptococcus suis ) serotype 2 (Accession No. KCCM 11111P) was challenged with a syringe at a dose of 1 × 10 ⁷ CFU, followed by observation for 5 weeks, and the results are shown in Table 1 below.

group Experimental sow head Sow vaccination Piglets
Attack
Age Piglets
Attack
Head Multiple meningitis onset Multiple meningitis
Incidence (%) Vaccination group 5 5 weeks before delivery 3 weeks before delivery
(Two inoculations) 3 weeks old
30 4 13.3 Positive control group 5 Unvaccinated 3 weeks old 30 26 86.6 Negative control group 5 Unvaccinated Unvaccinated 30 0 0

In Table 1, the positive control group refers to a group that is not challenged with vaccination only, and the negative control group means a group not to be vaccinated and challenged, and the vaccination group is a group that has performed both vaccination and challenge vaccination. Means.

As a result, the piglets (vaccinated groups) delivered from the vaccinated sows had received sufficient antibody from the colostrum of the sows, and no obvious clinical symptoms were observed even after the vaccination at 3 weeks of age. Did not even die. However, in 4 heads, the symptoms were slightly reduced at 6 weeks of age 3 weeks after challenge inoculation, and only mild meningitis was observed at autopsy at 8 weeks of age when the challenge was terminated.

In contrast, the piglets (positive controls) delivered from the unvaccinated sows had no clinical antibodies through colostrum, so clear clinical symptoms were observed after challenge vaccination. In the positive control group, anorexia was observed 1 week after challenge vaccination, and at 6 weeks of age 3 weeks after challenge vaccination, severely reduced piglets were observed. At fifteen weeks of age at the end of challenge, severe fibrosis was found in 26 of 30 patients.

In the negative control group which was not vaccinated and not challenged, no symptoms and lesions were observed at all until 8 weeks of age at the end of the test.

The inactivated Streptococcus of the present invention through the results shown in the Streptococcus (Streptococcus suis) pig multiple tent salt preventive vaccine containing serotypes Type 2 as the active ingredient is to minimize the damage caused by Streptococcus effective in the prevention of multiple tents It was found that it was possible to prevent multiple meningitis in the piglet by using the parental transfer antibody by directly administering it to pregnant sows.

Experimental Example  3. In a piglet  Preventing Multiple Meningitis by Vaccination

If multiple mastitis is induced in weaning pigs older than 4 weeks on the farm, it is effective to inoculate the piglets with vaccination. Vaccination was performed on the piglets to examine whether the piglets had a protective effect against streptococcal multiple meningitis.

Specifically, the vaccine of the production example was inoculated by intramuscular injection at 1 week of age in the piglet at 1 cc, and injected by intramuscular injection at 2 weeks of 3 weeks of age.

Then, after the second vaccination the identified Streptococcus separation in Example 3 to 5 weeks after two weeks Streptococcus (Streptococcus suis ) serotype 2 was challenged with a syringe at a dose of 1 × 10 ⁷ CFU, followed by observation for 5 weeks, and the results are shown in Table 2 below.

group Experiment head Vaccination Age Attack
Age Multiple meningitis onset Multiple meningitis
Incidence Vaccination group 30 1st week 2nd 3rd week
(2 doses) 5 weeks old 2 6.7 Positive control group 30 Unvaccinated 5 weeks old 29 96.6 Negative control group 30 Unvaccinated Unvaccinated 0 0

In Table 2, the positive control group means a group that is not vaccinated, but only the vaccination, and the negative control group is a group that does not have both vaccination and vaccination. Means.

As a result, no significant clinical symptoms related to multiple meningitis were observed in the group of two doses of the vaccine and two weeks after the second vaccination (vaccinated group) until the end of the experiment. No one died until this was terminated. However, two heads showed a slight contraction at 8 weeks of age 3 weeks after challenge, and mild automatitis was observed at 10 weeks of age when the challenge was completed.

On the contrary, in the group inoculated with streptococci at 5 weeks of age without vaccination (positive control group), distinct clinical symptoms related to multiple meningitis were observed after challenge vaccination.

First, in the positive control group, anorexia was observed from 1 week after challenge vaccination, and very reduced piglets were observed at 8 weeks of age 3 weeks after challenge vaccination. At 10 weeks of age at the end of challenge, very severe fibrosis was found in 29 of 30 patients.

In the negative control group which was not vaccinated and not challenged, no symptoms and lesions were observed at all until 8 weeks of age at the end of the test.

Inactivated streptococcus ( Streptococcus ) of the present invention through the above results suis ) Swine MS vaccine containing serotype 2 as an active ingredient effectively increases the immunity to streptococci, which are administered to pigs, to cause MS, thereby effectively preventing MS. It can be seen that it can be useful for domestic pig farms vulnerable to the risk of developing multiple meningitis.

Korea Microorganism Conservation Center (overseas) KCCM11111P 20101011

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Claims (15)

Attenuated or inactivated, vaccine composition for the prevention of swine multiple meningitis containing Streptococcus suis serotype 2, which causes multiple meningitis in pigs having the 16S rRNA sequence of SEQ ID NO: 7, as an active ingredient. delete delete The method of claim 1,
Streptococcus suis ) serotype 2 is a vaccine composition for the prevention of swine polymeningitis, characterized in that deposited with the Korea microbial conservation center with the accession number KCCM 11111P. The method of claim 1,
The inactivation is inactivated by formalin treatment Porcine multiple meningitis prevention vaccine composition, characterized in that. The method of claim 1,
A vaccine composition for preventing porcine multiple meningitis, further comprising aluminum hydroxide gel as an excipient. A method for preventing multiple meningitis disease by administering a vaccine composition according to any one of claims 1, 4, 5 and 6 to pigs. 8. The method of claim 7, wherein the vaccine composition is administered orally, nasal, anal, intramuscular, intradermal, intravenous or subcutaneous. Streptococcus of Accession No. KCCM11111P, causing multiple meningitis in pigs suis ). A vaccine composition for preventing swine polysclerosis containing Streptococcus suis of Accession No. KCCM11111P, which causes multiple meningitis in attenuated or inactivated pigs, as an active ingredient.
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