| Effects of Aeromonas hydrophila whole-cell oral vaccine with or withoutβ-glucan adjuvant on the immunological function and disease resistance of Chinese mitten crab Eriocheir sinensis, preparation of microcapsules of A. hydrophila and effects on immuno- logical function of E. sinensis were studiesd in the paper. The results are summarized as follows:Experiment one: The purpose of this study is to determine the variance of immune function of E. sinensis after being fed with the diets added lyophilized powder of A. hydrophila whole-cell vaccine, hydrophila whole-cell vaccine andβ-glucan (1:1) dosed at 0, 1 3 or 6 g/kg dry diet for 1 week, respectively. The total haemocyte count (THC), different haemocyte count (DHC), the phagocytic activities, respiratory burst (release of superoxide anion) activities of haemocytes, the phenoloxidase (PO), nitric oxide synthase (NOS) activities and total antioxidative competencein (T-AOC) in serum, the peroxidase (POD), superoxide dismutase (SOD), acid phosphatase (ACP), alkaline phosphatase (ALP) and lysozyme (LSZ) activities in serum or hepatopancreas were examined at 0, 1, 2, and 3 weeks after last administration with the diets of test groups. Crab were challenged at 1.2×107 colony forming units (cfu) /kg crab weight with a virulent strain of Aeromonas hydrophila (CL99920) of E. sinensis at 2 weeks after last administration with the diets of test groups, and mortalities were recorded over a 10-day period. The results were as follows: (1) The THC and HC, the phagocytic activities, respiratory burst activities of haemocytes, the PO, POD, SOD, ACP, ALP and LSZ activities in serum, the POD, ACP, ALP and LSZ activities in hepatopancreas of E. sinensis fed with the A. hydrophila whole-cell vaccine by certain dose were significantly higher than control group at many sampling time points (P<0.05). However, no significant differences were observed in the SGC and GC, the T-AOC, NOS activities in serum, and the of SOD activities in hepatopancreas of E. sinensis fed with the A. hydrophila whole-cell vaccine compared with control group (P>0.05). Furthermore, the crabs fed with the A. hydrophila whole-cell vaccine were the more resistant. The best immune protection rate was 57.14% at the group of 3 g/kg dry diet. (2) The THC, the phagocytic activities, respiratory burst activities of haemocytes, the PO, SOD, ACP, ALP, LSZ, and NOS activities in serum, the POD, SOD, ACP and LSZ activities in hepatopancreas of E. sinensis fed with the A. hydrophila whole-cell vaccine withβ-glucan adjuvant by certain dose were significantly higher than control group at many sampling time points (P<0.05). However, no significant differences were observed in the DHC, the POD activities, T-AOC in serum, and the of ALP activities in hepatopancreas of E. sinensis fed with the A. hydrophila whole-cell vaccine withβ-glucan adjuvant compared with control group (P>0.05). Furthermore, the crabs fed with the A. hydrophila whole-cell vaccine withβ-glucan adjuvant were the more resistant. The best immune protection rate was 57.14% at the group of 3 g/kg dry diet.Experiment two: The sodium alginate which has good biology consistent was used as shell-material and the A. hydrophila whole-cell vaccine was used as core-material, then preparing microcapsulation with emulsification method. The optimal technological conditions of making the microcapsule were determined through single factor test and orthogonal experiment of L9 (34). The results were as follows: the concentration of sodium alginate was 2%, the emulsification time was 15 min, the stirring speed was 800 rpm, the proportion of A. hydrophila whole-cell vaccine and sodium alginate was 1:8. This method can prepare microcapsules which have good enteric solubility and acid resistance, embedding rate was 71.34%.Experiment three: The purpose of this study is to determine the variance of immune function of E. sinensis after being fed with the diets added lyophilized powder of A. hydrophila microencapsulated vaccine dosed at 0, 1 3 or 6 g/kg dry diet for 1 week, respectively. The THC, DHC, the phagocytic activities, respiratory burst activities of haemocytes, the PO, NOS activities and T-AOC in serum, the POD, SOD, ACP, ALP and LSZ activities in serum or hepatopancreas were examined at 0, 1, 2, and 3 weeks after last administration with the diets of test groups. Crab were challenged at 1.2×107 colony forming units (cfu) /kg crab weight with a virulent strain of A. hydrophila (CL99920) of E. sinensis at 2 weeks after last administration with the diets of test groups, and mortalities were recorded over a 10-day period. The results were as follows: The THC, the phagocytic activities, respiratory burst activities of haemocytes, the PO, POD, SOD, ACP, ALP, LSZ, and NOS activities in serum, the POD, SOD, ALP and LSZ activities in hepatopancreas of E. sinensis fed with the A. hydrophila microencapsulated vaccine by certain dose were significantly higher than control group at many sampling time points (P<0.05). However, no significant differences were observed in the DHC, the T-AOC in serum, and the of ACP activities in hepatopancreas of E. sinensis fed with the A. hydrophila microencapsulated vaccine compared with control group (P>0.05). Furthermore, the crabs fed with the A. hydrophila microencapsulated vaccine were the more resistant. The best immune protection rate was 71.43% at the group of 3 g/kg dry diet.
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