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Standardization of Effective Dose of Fowl Cholera Vaccine in Pigeon in Bangladesh

M. T. Islam
Department of Microbiology and Hygiene, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh-2202, Bangladesh. Present: Asssiant Commissioner, Ministry of Public Administration, People’s Republic of Bangladesh, Dhaka, Banglades

M. H. Ali
Department of Microbiology and Hygiene, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh-2202, Bangladesh. Present: Department of Livestock Services, Ministry of Fisheries and Livestock, People’s Republic of Bangladesh, Dhaka, Bangladesh.

A. Chandra
Department of Microbiology and Hygiene, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh-2202, Bangladesh. Present: Department of Livestock Services, Ministry of Fisheries and Livestock, People’s Republic of Bangladesh, Dhaka, Bangladesh.

S. Saha
Department of Microbiology and Hygiene, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh-2202, Bangladesh

M. A. Islam
Department of Microbiology and Hygiene, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh-2202, Bangladesh

Abstract/ Executive Summary:

An experiment was conducted to determine the effective dose of formalin killed (FK) fowl cholera (FC) vaccines prepared with virulent avian Pasteurella multocida (PM 38) serotype 1 (X-73) collected from the laboratory of the Department of Microbiology and Hygiene, BAU, Mymensingh. To determine the effective dose of vaccine, 7 weeks old 30 pigeons were immunized and each group consists of 5 birds. The groups are represented by A, B, C, D, E and F. The birds belonging to groups (A-E) were vaccinated with different doses of vaccine, after two weeks of first, second immunization and challenge experiment, blood was collected from all vaccinated birds, and serum was analyzed to determine antibody titer against P. multocida by passive hemagglutination test (PHA). The PHA titer after two weeks of first vaccination were 16±3.92, 17.6±3.92, 25.6±3.92, 32±8.76, 35.2±7.84 of group A,B,C,D and E, respectively at the dose of 0.2ml (0.26×10⁸ CFU)/birds, 0.4ml (0.5×10⁸ CFU)/birds, 0.8 ml (1.04×10⁸ CFU)/birds, 1ml (1.3×10⁸ CFU)/birds, respectively. The PHA titer of prevaccination and control birds was <4. The PHA titer after 2 weeks of second vaccination or boostering were 32±8.76, 35.2±7.84, 44.8±7.84, 57.6±18.66, 70.4±15.68, of group A,B,C,D and E, respectively. After 2 weeks of challenge infection, the mean PHA titer were 44.8±7.84, 51.2±7.84, 70.4±15.68, 102.4±15.68 and 140.8±31.34 of group A,B,C,D and E, respectively. In this experiment, the antibody titer of the vaccinated pigeons with 0.4, 0.6, 0.8 and 1ml per bird via intramuscular route were higher than that of the pigeons vaccinated with 0.4ml/bird, 0.6ml/bird, 0.8ml/bird and 1ml/bird were satisfactory in terms of protective potential against P. multocida. For prevention and control of avian pasteurellosis 0.4ml to o.6ml (0.52×10⁸ CFU to 0.78×10⁸ CFU)/birds of vaccine may be used instead of 1ml (1.3×10⁸ CFU)/birds for better immunization of pigeon against fowl cholera infection.

Keywords: Fowl cholera, Pigeon, Vaccine, Dose

Location: Department of Microbiology and Hygiene, Bangladesh agricultural University, Mymensingh, Bangladesh

Start Date:

End Date:

Program Area: Animal Health and Management

Commodity: Pigeon, Vaccine

Objectives:

To compare the different doses of vaccine and prevent economic losses from vaccination and to standardize an appropriate dose of vaccine effective against fowl cholera in pigeon.

Methodology:

Bacterial strain: A stock culture of P. multocida (PM-38), serotype 1 (X-73) organism was obtained from the laboratory of Department of Microbiology and Hygiene, Bangladesh agricultural University, Mymensingh. The bacteria were originally isolated from a diseased duck from an outbreak of duck cholera (Choudhury et al., 1985). Experimental birds: A total of 30 non-vaccinated healthy pigeons of seven weeks of age of local breed of either sex were obtained from some household of Trisal upazila of Mymensingh district and used for this experiment. The birds had no previous history of infection or vaccination against fowl cholera. Adequate balanced ration and water were given to the pigeons. Vaccine preparation: P. multocida bacteria were cultured in nutrient broth enriched with yeast extract and incubated for overnight at 37°C, shaken at 4-5hrs interval manually. It was stocked in 80% glycerol and CFU was counted. Formalin was added @8ml/1000ml and kept in room temperature for 24 hrs. Alum was added as adjuvant @35gm/1000ml and kept at room temperature. The sterility was tested on blood agar media and safety test was done in chicken and mice. Finally formalin killed fowl cholera vaccine was prepared. Experimental immunization: Thirty birds of local breed were divided into six groups consisting of five birds in each group. The birds of group A, B, C, D and E were vaccinated subcutaneously (SC) with different set doses of experimentally prepared formalin killed fowl cholera vaccine. The initial doses of vaccine were given to the birds of groups A, B, C, D and E at the age of seven weeks. These birds were re-vaccinated with same doses of vaccine after two weeks of primary vaccination at the age of nine weeks. The birds of group F served as control. Collection of serum from the immunized birds: 3 ml of blood was collected either from the jugular vein or wing vein of all vaccinated pigeons of each group without anticoagulant and was poured gently in sterile glass tubes. The sera was collected in pre-vaccination (7 weeks of age), after first vaccination (at 9 weeks), after booster vaccination (11 weeks) and after challenge (13 weeks). The sera were collected according to the procedure of Tripathy et al. (1970). Passive hemagglutination assay (PHA): The test was performed to determine antibody titer of the vaccinated (primary, booster and challenge) pigeons according to the method described by Tripathy et al. (1970) with slight modification. Collection and preservation of serum as complement from male guineapig: Adult healthy male guineapig was obtained from the Department of Microbiology and Hygiene, BAU, Mymensingh. An amount of 10 ml of blood was collected directly from the heart without using anticoagulant. Aseptically collected blood was immediately poured into a 100 ml volume of conical flask previously kept on ice. As complement is labile even at room temperature, refrigeration temperature was strictly maintained in all the steps of preparation and preservation. The prepared serum of guinea pig was dispensed in small volume in sterile vials and preserved at -20°C until used (Tripathy et al., 1970). Collection and preservation of sheep red blood cells (SRBC) suspension: The collection and preservation of SRBE suspension and rabbit serum was done according to the procedure of Tripathy et al. (1970). Determination of the humoral immune response (HIR) of pigeons was performed according to Cheesbrough (1985). Statistical analysis: The antibody titers of vaccinated and control pigeons were analyzed by Student t test. The survivability percentage of the pigeons was analyzed by Mantel log rank test. P value of <0.05 was considered significant.

Source of Information: Bangl. J. Vet. Med. (2017). 15 (2): 97-105 ISSN: 1729-7893 (Print), 2308-0922 (Online)

Article Link (Online Journal):

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

Clinical findings after challenge: All the experimentally vaccinated and control birds that were challenged with virulent strain of. P. multocida PM-38, serotype 1 (X-73) by intramuscular route at the dose of 1.4 ×107 CFU/ml manifested the following clinical changes: Control birds: Non-vaccinated control birds showed no clinical signs at 3 hours of post inoculation (PI). The clinical signs first appeared at 6 hours PI-included dullness and depression. At 12 hours PI, there were dullness, depression, slight rise of body temperature (42.5°c) and increased respiratory rate (40-45/min). The observed clinical signs at 24 hours PI were severe weakness, drowsiness, anorexia, rise of body temperature (43.6°c), increased respiratory rate (45-55/min), lameness, whitish (chalky) diarrhoea with mucus. The clinical signs at 48 hours PI were almost similar to that or 24 hours. The other signs included anorexia, lameness, greenish diarrhoea with mucus, subnormal temperature (41°C), decreased respiratory rate (15-25/min). Death of one bird occurred first at 48 hours PI, one died at 72 hours and the rest died at 96 hours PI. The vaccinated pigeons did not exhibit any clinical signs and there was no death within 96 hours of PI. After 7 days of PI, in some birds of group A, although there was formation of cyst like structure at elbow joint of the legs, but after 30 days of PI the cysts got diminished.

The results of protection test after challenge infection revealed that all the pigeons of group B, C, D and E vaccinated with 0.4 ml, 0.6 ml, 0.8 ml and 1 ml/bird respectively were well protected. Whereas, few pigeons of group A (0.2 ml/bird) showed chronic type of localize infection after challenge. This protection test indicated that all doses of vaccine protected 100% vaccinated birds.

Knowledge Management: Journal