Study On Fermentation Process Of Haemophilus Parasuis (HPS) And And The Development Of Its Vaccine

Haemophilus parasuis(HPS)is a commensal bacterium that attaches to the upper respiratory tract of pigs and is an opportunistic organism that invades the body of the pig when the body’s immunity declines.After infected,HPS mainly causes multiple serositis and arthritis in pigs.HPS is a potential threat to the pig industry,once it breaks out,it will cause huge economic losses.Therefore,HPS has also received close attention.For the prevention of the disease,HPS is mainly immuned by injecting the inactivated HPS vaccines which have good immune protection.Based on the isolated HPS strains,the identification and the serotypes of the isolated HPS strains were done by different laboratory methods.The manufacturing process and the bacterial antigen culture conditions of the identified HPS strains were compared and optimized in the shake flask and the bioreactor.The bacterial antigens were cultured in the optimized conditions and the inactivated vaccine of HPS was prepared.And the safety and the efficacy of HPS vaccines were evaluated in this paper.The isolations of RPBS0904-11,RPBS0905-3 and RPBS0912-1 strains were isolated exactly by using of colony and morphological observation,nicotinamide adenine dinucleotide(NAD)-dependent assay,biochemical identification,PCR identification,and there were confirmed to be belongs to HPS.The identified HPS strains were serotyped by using of PCR identification and agar diffusion test(KRG),that is,the strain RPBS0904-11 was identified as HPS type 4,the strain RPBS0905-3 was HPS type 5 and strain RPBS0912-1 was HPS 12 type.Based on the culture of the identified HPS strains in shake flasks,the optimized culture conditions such as the animal serum type,the adding amount of coenzyme(NAD),and the pH environment of the medium used in the dominant culture conditions were determined;The optimized culture conditions such as the type of feed,feed rate and feeding method required under the dominant culture conditions in the bioreactor were determined.Based on the culturing HPS bacterial antigen by using of the optimized process in the bioreactor,the HPS trivalent(4,5,12)inactivated vaccine was prepared.The pre-made vaccine has a good immune effect according to the safety test by using of multiple immunization doses in piglets to evaluate the safety of the vaccine,according to the clinical manifestations and pathological observation in the vaccinated piglets,and according to the vaccine efficacy test by immunizing guinea pigs with an immunization dose and the antibody titer in guinea pigs is much higher than the effective antibody level.There is no obvious difference between the two vaccines prepared in the bioreactor process and in the shake flask process respectively,compare to the culture process in the bioreactor and in the shake flask including vaccine endotoxin content,vaccine bacterial protein content and vaccine immune effect.The bioreactor culture process vaccine can completely replace the shake flask process vaccine.