| Polylactic-co-glyconlicacid （PLGA） microparticle vaccines containing GBS whole cell and supernatant after ultrasonication were prepared by double emulsion-solvent evaporation method. The GBS was isolated from sick body of tilapia in tilapia farms. We studied the parameters of microspheres preparation for oral, detected the embedding and release characteristics of the microspheres and investigated the immune protective effects of the microparticle oral vaccine on Nile tilapia（Oreochromisniloticus）. The main contents and results are as follows:1. Parameters of microspheres preparation containing GBS and it’s characteristicsThe influences of the concentration of PLGA, emulsifler concentration and the volume of outer water phase on mean diameter of the microparticles containing supernatant by Control variable method were analyzed. The results indicate that with the increase of PLGA concentration, emulsifier concentration and volume of outer water phase, the mean diameter of microparticles increased.2.5%of PLGA,0.1%of emulsifier concentration and20mL of outer water phase were used for preparation of microparticles containing supernatant. The preparation parameters were similar for GBS whole cell microparticles except that the emulsifier concentration was0.2%. We observed the surface morphology of the microspheres with scanning electron microscope （SEM）, and analyzed the encapsulation efficiency, drug loading and drug release performance of microspheres in vitro by BCA. The mean diameter of GBS whole cell, supernatant microparticles are9.4μm and3.9μm respectively. All the microparticles prepared are spherical in shape. The encapsulation efficiency of GBS whole cell and supernatant microparticles are68.07%and63.49%respectively; the drug loading are5.49x108/mg and3.58%respectively; The cumulative rate of drug-release over28days are64.2%and82.8%respectively. Taken together, the encapsulation efficiency, drug loading and cumulative rate of drug-release, as well as the microparticle diameter are suitable for oral immunization. This result will facilitate evaluation of efficiency of the oral vaccine against GBS in tilapia.2.Immune protective effects of the microparticle containing GBS oral vaccine on Nile tilapiaPLGA microparticles containing GBS whole cell, supernatant after ultrasonicationand PBS were administrate to Nile tilapia orally, and formalin killed Streptococcus agalactiaeas positive control group by intragastric and intraperitoneal（IP） injection. The Percentage of phagocytes（PP）, Phagocytic index（PI）, lysozymeactivities, superoxide dismutase（SOD） activities and antibody level(OD450nm) are detectedposted7,14,21,28d. Than tilapia were challenged by artificial infection with Streptococcus agalactiae, calculating relative percent survivals （RPS）. The result is that PP, PI, lysozymeactivities, SOD activities of experimental groups are higher than negative control group, lower than positive control group. Antibody level of experimental groups has the similar result, but control groups go down after14d and experimental groups keep high level all the time. The RPS of GBS whole celland supernatant after ultrasonicationgroups are57.63%and34.40%respectively, positive control group is94.53%. The results suggest that PLGA microparticles, as adjuvants, encapsulated GBS are effective to protect Nile tilapia against GBS infection abviously, and they can increase the immune protection time by controlled-release effect, but the RPS was lower than positive control group. We can carry out further study on concentration of microparticle for improving the immune protective effects of the microparticle containing GBS oral vaccine on tilapia. |
