Results And Effect As Well As Mechanism Of A New Model Genetic Vaccine Against Somatostatin In The Swine

In this study, the immune efficacy and influence factors of a novel somatostatin DNA vaccine-Attenuated Mouse Typhus Salmonella strains CS022(pGM-CSF/SS) immunized against piglets with oral administration and nasal feeding were investigated, using methods of genetic immunization, ELISA, and paraffin section. Mice were administered attenuated Cholera Swine Salmonella strains C500(pGS/2SS-M4GFP-asd) through intramuscular injection to detect the temporal and spatial patterns of the exogenous gene expression in various tissues and organs, which will contribute to immunization program optimization of novel growth-promoting DNA vaccine, elevation of their growth-promoting effects on the piglets and to accelerate its clinical application.1. Immune responses and Growth-promoting effect of piglets to the novelsomatostatin DNA vaccine by oral vaccination. So as to study the immune responses and the growth-promoting effect of the novel somatostatin DNA vaccine by oral way. a total of40three-ways crossing commercial piglets at9weeks of age were randomly divided into4groups, and each group was evenly composed of5males and5females. T1, T2and T3groups were administered with attenuated Salmonella strain CS022(pGM-CSF/SS) somatostatin DNA vaccine orally with low doses (5×108CFU/each), moderate doses (5×109CFU/each), and high doses (5×1010CFU/each) respectively. Control group C1was administered with5ml PBS solution. A booster was performed with the same doses and same treatment at1week after primary immunization. Body weight gains of the piglets were measured three times (before immunization, after immunization4and8weeks). The blood samples were collected two times at former vena cava (before immunization and after immunization5weeks). IgG antibody levels in plasma was detected using indirect ELISA and SS concentration in plasma was detected using radioimmunoassay.The results showed that the piglets had normal behavior after vaccination. Plasma SS antibodies were detected in all experimental groups, T3group had the highest level. Compared with C2group, T2group did not have significant difference (P>0.05) but both T2and T3had very significant difference (P<0.01). The immunoreactive pigs occurred in each experimental group, and the ratio of T1and T2group were10.0%and T3group was40.0%. Compared with those of pre-immunization and the Cl group, plasma SS concentration declined significantly for each group (p<0.01), but there was no significant difference among experimental groups (P>0.05).T3group pigs average daily gain increased by20.69%compared with Cl group pigs (P<0.05), it had significant difference, and increasing by27.59%compared with T1group pigs (P<0.01) which it had significantly difference after immunization0-4weeks, but it increased only by13.79%compared with T2that had no significant difference. There was no significant difference (P>0.05) among all groups though T1group pigs average daily gain increased by11.11%compared with C1control group after immunization5-8weeks, and also T3group pigs average daily gain increased only by10.00%compared with C1control group after immunization0-8weeks.These results demonstrated that the vaccine could induce the immune response in piglets and the immune effects were significant. There was positive correlation-dependent among antibody levels and the ratio of antibody postive piglets and average daily gain and dose amount.2. Growth-promoting effect of novel somatostatin DNA vaccine by nasal injectionSo as to study the growth-promoting effect of the novel somatostatin DNA vaccine by nasal injection, a total of75three-ways crossing commercial castrated male piglets with the same genetic background and weight21.0±2.0Kg, were divided into four groups. There were20pigs for each group of T4, T5and T6, and15for C2group. The pigs were administered vaccine3times at intervals of4weeks through nasal injection, with low dose (2×108CFU/each), moderate dose (2×109CFU/each) and high doses (2×1010CFU/each) respectively. C2group was treated with PBS solution with a dose of2ml. Body weight gains of the piglets were measured twice (before immunization and after immunization12weeks). The results showed that T4(low dose) group pigs average daily gain increased by11.11%compared with C2control group after immunization, T5(middle dose) and T6(high dose) group increased by6.17%and1.23%respectively. There was no significant difference (P>0.05) between groups. Average daily gain declined with the increasing dose of vaccine, which implied that immune efficacy had negative correlation with vaccine dose. Nasal immunization needed vaccine doses far less than oral immunization for the same daily gain. 3. The temporal and spatial patterns of the target gene and the SS fusion protein of immunized mice through intramuscular injection with novel somatostatin DNA vaccineSo as to study the temporal and spatial patterns of the target gene and the SS fusion protein of immunized mice through intramuscular injection with novel somatostatin DNA vaccine, a total of78Kunming mice at7weeks old were randomly divided into13groups, and each group was evenly composed of3males and3females. Longitudinal muscle injection with dose of150μL×109CFU/each at two hind quadriceps in mice was implemented. The immunized groups A1-A6were inoculated attenuated Salmonella choleraesuis C500(pGS/2SS-M4GFP-asd) somatostatin DNA vaccine, and negative control group B1-B6were inoculated with plasmid-free C500bacteria vaccine, and the control group C3inoculated with150μL PBS saline. Six mice of each group were killed after Ethyl ether smoked dizzy in turn with ether A1-A6and B1-B6at immunization24h,48h,96h,144h,192h and240h, and C3group mice was killed by the same way at immunization24h. Paraffin sections were made after collecting their heart, liver, spleen, lung, kidney and hindlimb skeletal muscle, and to deal with by Immunohistochemical staining method.The results showed that SS fusion protein had been expressed in turn with at macrophage of spleen, liver, muscle with injection site, and its expression reaches the maximum all of them at immunization144h. It occurred firstly at muscle tissue (immunization24h), the second at spleen (immunization48h), the last at liver (immunization144h). Muscle tissue had sustained more time than spleen and liver, and had more quantity until immunization240h. Liver had the shortest time of continuous expression, and there was no SS fusion protein at immunization240h.It demonstrated that it was viable to immune livestock the way of intramuscular injection with novel somatostatin DNA vaccine. It was short time that the exogenous plasmid DNA and purpose protein had continuous expression in mice by intramuscular injection.