CS213847B1 - Inactivated adsorbate vaccine against infectious bovine keratoconjuctivitis - Google Patents

Description

The invention provides for active protection of bovine vaccines against infectious keratoconjunctivitis caused by Moraxella bovis.

Attempts to control infectious bovine keratoconjunctivitis by immunoprophylactic measures were based on the observation that calves formerly infected with infectious bovine keratoconjunctivitis have resisted infection with homologous strains of Moraxella bovia (HUGHES, DE-PUGH, GW-McDONALECTIVE INFRACTIONAL CULTURALCONSUNCTIONAL CULTURAL INFECTIVE CULTURE). and Moraxella bovis infection: Resistance to re-exposure with homologous and heterologous Moraxella bovis (Am. J. vet. Res., 29, 1968, pp. 829-833). Later, several types of vaccines were tested. Relatively favorable results were obtained when Moraxella bovis (HUGHES, DE - FUGH, GW) was used to vaccinate a live culture: Experimentally Induced Bovine Infeetious Keratoconjunctivitis: Effectiveness of Intramuscular Vaccination with Moraxella Bovis Cultura Am. 32, 1971, pp. 879-886. It was further verified that partial protection efficacy could also be achieved with a formaldehyde inactivated vaccine. The optimal interval between vaccinations of 14 days was also determined (HUGHES, DE - PUGH, GW: Experimentally Induced Infectious Bovine Keratoconjunctivitis: Relationship of Vaccination Schedule to Export with Homologous Moraxella Bovie Culture. Am.J.vet.Res., 36, 1975, 9, 263 265.)

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In Brazil, live vaccines prepared from individual strains of Moroxella bovis and combinations thereof have been tested against infectious bovine keratoconjunctivitis. Good protection was achieved when a homologous strain was used for infection. If ae strains were heterologous, the protection was very variable (BAPTISTA, PJHP: Querato - conjunctivitis infecciosa dos bovinos: (I) Resistance of bovinos vacinados with cultures of vivfiB and aggression of experimental homologs and heterologues of Moraxella bovis. (II). (III) Immunology.

Bol. Inst. Pesquisas vet. Desidério Pinamor, 3, 1975, pp. 5–35. In the US in Qregon, infectious bovine keratoconjunctivitis was similarly used to control infectious bovine keratoconjunctivitis similarly to a good outcome of an autogenous vaccine prepared from a culture of Moraxella bovis isolated from a fresh case (STOREY, RC - STODDARD, P. - BARGER, R .: An Autogenous Vaccine in Succesful Treatment of Infectious Keratitis in Cattle (Vet, Med. Small. Ali.Clin., 72, 1977, pp. 1050-1051.) Despite some good results of a vaccine containing both living and dead Moraxella bovis, provided only partial protection against the disease. Later, another vacoin containing a concentrate of Moraxella bovis fimbriae, ultrasonically broken and suspended in water or oil, was tested. The calves vaccinated with the fimbium vaccine with the olive adjuvant had protective immunity against infection with the homologous strain of Moraxella bovis (PUGH, GW-HUDGES, DS-BOOTH, GDi). Experimentally induced infectious bovine keratoconjunctivitis: Effectivenees of a pilus vaccine against homologous disease. J.vet. Res., 38,

1977, pp. 1519 - 1522.) Attempts to enhance protective antibody response by vaccine enriched with incomplete Freund's adjuvant (HUGHES, DE-PUGH, GW-BOOTH, GD: Induced infectious bovine keratoconjunctiivitis: Vaccination with whole cell bacetrins of Moraxella bovis adjuvant mixed with Freund Am.J.vet.Res., 38, 1977, a. 1905-1907) or Mycobacterium paratuberculosis vaccine have not been successful (PUGH, GW-McDonald, TJ LARSEN, AB: Experimentally Induced Infectious Bovine Keratoconjunctivitis: Potentation of and MCraxella bovis pilus vaccine and immunogenicity by a vaccination with Mycobacterium paratuberculosis bacterin. Am.J.vet.Res., 39,1978, p. 1656-1661. A different type of vaccine against infectious bovine keratoconjunctivitis has been reported from Uruquay. It contained autolysed cultures of Moraxella bovis, Staphylococcus aureus & Corynebacterium pyogenea, was inactivated with formaldehyde and aluminum hydroxide was added as adjuvant. The vaccine had a very good protective and therapeutic effect (QUINONES SOWERBY, CA: Nueva vacuna confra la queratoconjunctivitie de los bovinos. Vet. Uruqay, 5, 1978, pp. 27-34). A similar composition is a vaccine of French origin KERRAB, manufactured by IFFA Mérieux . Information about its protective efficacy in our cattle breeds varies. Good results have been reported in the treatment of infectious bovine keratoconjunctivitis. Can be treated either locally (FLEMING, GA: Bobine infectious keratoconjunctivitia. Vet. Rec., 97, 1975, p. 486.), or overall, and recommends that both treatments be combined (PUGH, GW - McDonald, TJ): Infectious bovine In the control of infectious keratoconjunctivitis, the efficacy of treatment and vaccination was also compared, and it was found that if the cattle herd was treated with ocular bovine animals. furazolide spray every week from May to August, reducing the incidence of the disease more than vaccination with autogenous Moraxella bovis with dead formaldehyde (HUGHES, DE

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- KOHLMEIER, R.H. - PUGHj G.W .: Comparison of Vaccination and Treatment in Natural Controlling Infectious Bovine Keratoconjunctivitis. Am. J.vet. Res., 40, 1979, p.

241 - 244 *) Control of infectious bovine keratoconjunctivitis by treatment is, however, laborious and impracticable in conditions of large-scale breeding and grazing. A more feasible route is to immunize with an effective vaccine prepared from homologous, fully virulent Moraxella bovis strains.

The increasing problem of infectious bovine keratoconjunctivitis has led to the development of a vaccine whose good protective efficacy has been verified in experiments (ŠIMON, V. - HOŘAVOVÁ, P. - KREJČÍ, 3 ·· Immunoprophylaxis of infectious bovine keratoconjunctivitis. 20s. ^?

An inactivated adsorbate vaccine against infectious bovine keratoconjunctivitis has been invented to replace a vaccine of French provenance KÉRABOV, which is not effective enough in our conditions * o * 10

The essence of the vaccine is that it contains a suspension of Moraxella bovis of 10 to 10 GFU / l ml in an amount of 30 to 50 vol%, a suspension of fimbriae of the same bacterial strain with a density of up to 80% at 540 nm in an amount of 30 to 50 vol. % and the remainder to 100% by volume are adjuvant and inactivating agent.

The high protective efficacy of the vaccine is achieved by the properties of a selected bacterial strain, fully virulent, which, in addition to the haemolytic and dermonecrotic toxin, forms fimbriae, which is a critical sign of the pathogenicity and virulence of Moraxella bovis strains. The efficacy of the vaccine was verified in two successive experiments. In the first experiment, 37 were included in the vaccinated group and 49 in the calves. After experimental ipfection 27 days after the last vaccination, only one case of slight corneal scar was found in the vaccinated calves group. In the control group of unvaccinated calves, 10 cases, 38.8% of keratitis of varying grade were found. After an experimental infection carried out 28 days after the last vaccination, keratitis was found in only one calf, ie 5.3% · In the control group of unvaccinated calves, 10 cases of keratitis, ie 23.8%, were diagnosed. significant to the significance level p - 0.05.

The vaccine is prepared from the bacterial strain Moraxella bovis, which has the following properties: Gram-negative short strong sticks, Hemolytic, immovable, non-reducing nitrates in nitrites, Non-inducing non-fermenting carbohydrates, Proteolytic ability, Positive oxidase test in three litmus milk zones - upper zone dark blue, middle zone formed by lavender color cassel, lower zone from coagulated casein, not growing on Simons citric medium. On agar medium with 5-10% sheep blood it grows in shiny, gray-white convex colonies, sharp edges, 1-3 mm in diameter, slightly pressed into the medium. Cultures isolated from pathological material that have not been passaged multiple times on blood agar form solid, adhering colonies, artoagglutinating in saline. The cause of autoagiutinability is the formation of fimbrios. Moraxella bovis strains from different sites are antigenically different.

Example:

The vaccine contains:

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Example:

The vaccine contains:

a suspension of Moraxella bovis of 10 $ CFU / ml ..... 40% by volume of fimbriae suspensions having a density of 80% at a wavelength of 540 π »..... 40% by volume sterile aluminum hydroxide gel. ... 19.6% formaldehyde ..... 0.4%

The bacterial suspension is generally obtained by washing the growth from the surface of the blood agar with saline, the 10 cm agar plate being washed with 2 ml of sterile saline.

Fimbriae are obtained by homogenizing the bacterial suspension for 20 minutes at 10,000 rpm while cooling. The homogenate was centrifuged at 6000 rpm for 60 minutes. at 3-5 ° C. Fimbriae in the supernatant ae precipitate at 4 ° C for 18-24 hours after the addition of magnesium chloride to a concentration of 0.1 mol / l. The precipitated fimbriae are centrifuged at 6000 rpm. for 60 minutes. The sediment is dissolved in six times less sterile distilled water than the original volume of the bacterial suspension. The fimbria suspension is adjusted to a density of 80% at 540 m. Higher permeability of the fimbria suspension is possible, but unnecessarily expensive to manufacture.

The bacterial suspension is checked for the presence of the fimbriae either electronically or by the hemagglutination assay of tSAKDHU, T.S. - WHITE, F.K. - SIMPSON, C.F .: Asaocletion of pili with rough colony type of Moraxella bovis. Am.J. Res. Res., 35, 1974, No. 3, pp. 437-439).

Vaccine sterility is checked after storage at 4-5 ° C for 72 hours by plating on blood agar.

The harmlessness of the vaccine is controlled by intramuscular injection at a dose of 5 ml of calf at about 6-10 weeks of age. There must be no inflammation at the injection site.

The vaccine has protective efficacy against Moraxella bovis infections that are antigenically homologous and the strain used in the vaccine. The antigenic identity of field strains and vaccine strains of Moraxella bovis is checked either by precipitation in an agar gel or by passive hemagglutination. The antigen for these reactions is prepared by disintegrating the suspension of the fimbriae by ultrasound. The liquid antigen is adjusted to a density of 50% transmittance at 540 nm. Antibodies are prepared by sequential intravenous injections on rabbits weighing about 2 kg according to the following schedule:

Week 1 3 days in a row 0.5 ml, 2-3 weeks following 3 days in a row 1 ml antigen. Rabbits are bled 14 days after the last vaccination. * Antigenic identity is indicated by distinct precipitation lines on agar gel precipitation, or nearly identical high titers in passive hemagglutination.

Calves are vaccinated at the lowest age of 8 weeks with a dose of 5 ml in cervical muscle twice every 14 - 21 days. Young cattle from 150 kg upwards are vaccinated with a dose of 8 ml also twice every 14 21 days into the neck muscles.

The vaccine may be stored at 5 ° C for six months. Immunity after vaccination lasts about months. The vaccine has autoagglutinating properties and therefore agglutinin should be used before use.

213 847 shake. The efficacy of the vaccine has been demonstrated in the previously described experiments. So far, 390 calves have been vaccinated without signs of health disorders.

Claims (1)

Inactivated adeorbate vaccine against infectious keratoconjunctivitis of acota, characterized by: containing a suspension of Moraxella bovia of 10 to 10 ° CFU / ml in an amount of 30 to 50 vol%, a suspension of fimbriae of the same bacterial strain having a density of up to 80% at a wavelength of 540 nm in an amount of 30 up to 50% by volume and the remainder up to 100% by volume are adjuvant and inactivating agent.