Development and evaluation of outer membrane protein vaccine against duck pasteurellosis

Abstract

A research work was undertaken to prepare effective vaccines against P. multocida grown under different conditions and their immunopotency was assessed in one month old ducklings. The purity of the P. multocida A:l strain (DP1) was confirmed as per standard procedure. Pathogenicity of the isolate was assessed in six to eight weeks old mice. The isolate killed the mice with in 8 h intra peritoneally and within 24 h when injected subcutaneously. Whole cell protein of P. multocida (DP1) was extracted and they were subjected to discontinuous system of SDS- PAGE which revealed 20 to 26 visible protein bands of molecular weight ranging from 102 to 19 kDa. After growing P .multocida in iron sufficient and iron restricted medium its OMPs were extracted and they were analysed by SDS PAGE. In iron sufficient medium, 10 protein bands of MW which ranging from 91.84 to 19.02 kDa were revealed. In addition to the above a protein band with MW of 97.8 kDa were detected in iron restricted medium.The protein concentration was estimated by Modified Lowry method,and it was found to be 4mg/ml. The Median Lethal Dose (LDso) of P. multocida when determined in one, month old ducklings was found to be 10 -7.13 which contained 13 cells. Oil adjuvanted formalin inactivated vaccines were prepared from DPl grown under three different conditions viz., TSB, BHIB and BHIB supplemented with 100 u M 2,2' dipyridyl. Sterility, safety and potency test of the vaccines were done as per standard procedures. A total of 120 one month old ducklings were divided in to four groups with 30 birds in each group. The first three groups were vaccinated with ordinary bacterin, OMP vaccine prepared under iron sufficient condition and OMP vaccine prepared under iron restricted condition respectively and the fourth group served as control.The birds were vaccinated with 0.5millilitre of vaccine intramuscularly. The blood was collected from all the ducks on day 0,7, 14,21,28,45 and 60 day PV. Passive haemagglutination was done and the increase in antibody titre was observed from day 7 PV onwards for groups I,.Il and In. The highest antibody titre was obtained at 14th day PV and 21 st day PV for aMP vaccine prepared under iron restricted condition. All the vaccine groups had shown a significant difference from the control group at all stages of study. On homologous challenging OMP vaccine under iron restricted media on 42nd day PV afforded 60 per cent protection. On 60th day PV afforded 50 per cent protection. Though the aMP vaccine prepared under iron restricted condition was found to provide high antibody titre, the protection percentage afforded by it in comparison with ordinary bacterin and aMP vaccine prepared under iron sufficient media was low. Hence, the aMP vaccine prepared under iron restricted condition need to be subjected to further investigation.