RESUMO
In an attempt to control the proliferation of the pathogenic bacterium Vibrio parahaemolyticus in gilthead seabream (Sparus aurata), the immunostimulant effect of lysate and ToxA from this bacterium was evaluated. Fish were intraperitoneally injected twice (first injection, day 1 of the experiment; second injection, day 7) and sampled after one week (on days 8 and 15). Afterwards, all fish specimens were experimentally infected with V. parahaemolyticus and mortality was recovered for 1 week. Fish injected with lysate, ToxA and phosphate buffer saline (control) showed 100%, 50% and 0% survival, respectively, when challenged with the pathogen. Skin mucus immune parameters and immune-related gene expression in skin and spleen were also evaluated. The results showed that mucus immune parameters were enhanced in the lysate and ToxA groups compared with the values obtained for fish from the control group. Expression of IL-1ß, TNF-α, C3 and IgM genes was significantly up-regulated in the lysate and ToxA groups, principally after infection with the bacterium. Interestingly, TLR5 gene expression increased in fish immunized with lysate. The most prominent histological characteristic in gut from infected fish was the presence of a great number of intraepithelial leucocytes as well as inflammation of the submucosa, while severe hydropic degeneration and hemosiderosis were detected in liver from infected fish. Injection of lysate or ToxA had a protective effect against the deleterious consequences of subsequent infection with V. parahaemolyticus in gut and liver. The findings underline the potential of lysate and ToxA as potent preventive antigens against this kind of vibriosis.
Assuntos
Adjuvantes Imunológicos/farmacologia , Toxinas Bacterianas/imunologia , Vacinas Bacterianas/imunologia , Doenças dos Peixes/prevenção & controle , Dourada , Vibrioses/veterinária , Vibrio parahaemolyticus/imunologia , Adjuvantes Imunológicos/administração & dosagem , Animais , Toxinas Bacterianas/administração & dosagem , Vacinas Bacterianas/administração & dosagem , Doenças dos Peixes/imunologia , Doenças dos Peixes/microbiologia , Expressão Gênica , Imunidade Inata/efeitos dos fármacos , Injeções Intraperitoneais/veterinária , Muco/imunologia , Pele/imunologia , Vibrioses/imunologia , Vibrioses/microbiologia , Vibrioses/prevenção & controleRESUMO
Immunogenicity of ToxA and Vibrio parahaemolyticus lysate was evaluated in a double immunostimulation scheme in Pacific red snapper after V. parahaemolyticus infection. Three groups of Pacific red snapper were intraperitonealy (i.p.) injected with phosphate-buffered saline (PBS group), ToxA of V. parahaemolyticus (ToxA-Vp group) or V. parahaemolyticus lysate (lysate-Vp group) (first injection, day 1; second injection, day 7). Fish were subsequently infected with live V. parahaemolyticus. Humoral immune parameters in skin mucus and serum were evaluated on days 1, 7, 8 and 14 days post-immunostimulation and 7 days post-infection. Moreover expression of immune-related genes was quantified by real time PCR in head-kidney leukocytes, spleen, liver, and intestine. The ToxA-Vp-treated group showed a higher anti-protease and catalase activity in skin mucus when compared with the PBS group. Measurements of SOD and CAT activities showed an increment in both activities a day after the second boost with ToxA-Vp or lysate-Vp. Interestingly, IgM levels in mucus and transcripts were enhanced followed the ToxA-Vp treatment even after challenge. Furthermore, IL-1ß was strongly expressed in all analyzed cell or tissues followed ToxA-Vp or Vp-lysate treatments. Finally, SOD and CAT gene expression was up-regulated in fish immunostimulated with either treatment ToxA-Vp or lysate-Vp, mainly after infection in head-kidney leukocytes and intestine. This is the first study where the effects of ToxA from V. parahaemolyticus in the immune system of Pacific red snapper was evaluated. These results suggest that ToxA-Vp would positively affect humoral immune response and up-regulate expression of genes involved in the immune system function; and could help in the control of V. parahaemolyticus infection in Pacific red snapper Lutjanus peru, an economic important fish in Mexico.