A survey on achieving recombinant DNA vaccine (live delivery) against Streptococcusis in rainbow trout (Oncorhynchus mykiss) immunization

The main purpose of this study was to survey on achieving recombinant DNA vaccine (live delivery) against Streptococcusis for rainbow trout (Oncorhynchus mykiss) Immunization. Initially, a total of ۵۱۵ samples were collected from the head kidney of diseased fish (weighing ۵۰_۲۰۰g) in ۷۲ farms in ۸ provinces. Approximately, ۴۰% (۲۰۶ samples) of specimens were infected with Streptococcus species. Then isolated ۱۷۲ DNA samples were and consequently, five pathogenic species have been identified, including Streptococcus iniae, Streptococcus faecium, Streptococcus agalactiae, Streptococcus dysgalatiae, and Streptococcus uberis. The enzyme phosphoglucomutase (PGM) has recently been discovered to play an important role in polysaccharide capsule production and virulence in S. iniae. Therefore, was initially isolated S. iniae and cloned the phosphoglucomutase gene. Then, the PGM gene was amplified successfully and cloned in pTZ۵۷R cloning vector. The recombinant plasmid was subcloned into the pETD uet-l expression vector by restriction enzymes and confirmed by PCR. Meanwhile, for amplifying simA and cpsD genes were used universal primers pNZ۸۱۴۸ and special for simA and cpsD genes. The recombinant bacteria Lactococcus lactis (NZ۹۰۰۰) was used to transform the plasmid into L. lactis. Vaccination was performed by bath and injection (peritoneal) methods. The efficiency of g۲ was better than g۱in these two methods and in all of the groups. The detection of anti S. iniae antibody and determination of IgM level was carried out by using ELISA. The results revealed that there was a significant (p<۰.۰۵) difference between the level of IgM in both two methods and experiment groups compared to the control group. The results of the challenge of vaccinated fish with S. iniae showed that fish RPS in all groups were more than ۵۰ percent while in the control group was ۲۱.۴۳ percent. The highest fish RPS belonged to group ۵(۶۱.۲۵ percent) and statistical analyses revealed that significant (p<۰.۰۵) difference between fish vaccinated RPS, compare to the control group.

کلیدواژه ها:

Recombinant DNA vaccine ، Rainbow trout ، Streptococcusis ، Phosphoglucomutase