Abstract
The FlaA gene from Vibrio harveyi, with a short nucleotide sequence encoding the Flag marker, was cloned into the eukaryotic expression vector pcDNA3.1(+) (designated as pcFlaA). Ninety grouper (Epinephelus awoara) were separated into three equal size groups. An experimental group was immunized with pcFlaA, Control I group was immunized with the vector pcDNA3.1(+), and Control II group was immunized with PBS. The expression of pcFlaA mRNA and protein was examined using reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry. We also evaluated the immunogenicity and protective efficacy of pcFlaA against V. harveyi by measuring the lymphocyte proliferation response and serum levels of specific antibody and conducting a bacterial challenge test. We successfully transfected the fish muscle with pcFlaA. The pcFlaA mRNA and protein was expressed in the muscle cells for up to one month following injection. The proliferation response of lymphocytes in fish immunized with pcFlaA was significantly higher than in control group II. Furthermore, the immunized fish generated specific antibody. The vaccination also resulted in significantly higher survival during the bacterial challenge test.
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Caselli E, Betti M, Crossi M P, et al. 1999. DNA immunization with HIV-1 tat mutated in the trans activation domain induces humoral and cellular immune responses against wild-type Tat. J. Immunol., 162(9): 5 631–5 638.
Daly J G, Moore A R, Olivier G. 1995. A colorimetric assay for the quantification of brook trout (Salvelinus fontinalis) lymphocyte mitogenesis. Fish & Shellfish Immunology, 5: 265–273.
Das M, Chopra A K, Wood T, et al. 1998. Cloning, sequencing and expression of the flagellin core protein and other genes encoding structural proteins of the Vibrio cholerae flagellum. FEMS Micro. Lett., 165: 239–246.
Garver K A, Conway C M, Kurath G. 2006. Introduction of translation stop codons into the viral glycoprotein gene in a fish DNA vaccine eliminates induction of protective immunity. Marine Biotechnology, 8(4): 351–356.
Gómez-Chiarri M, Livingstion S K, Muro-Cacho C, et al. 1996. Introduction of foreign genes into the tissue of live fish by direct injection and particle bombardment. Diseases of Aquatic Organisms, 27: 5–12.
Hansen E, Fernandes K, Goldspink G, et al. 1991. Strong expression of foreign genes following direct injection into fish musle. FEBS Lett., 290: 73–76.
Heppell J, Davis H L. 2000. Application of DNA vaccine technology to aquaculture. Advanced Drug Delivery Reviews, 43(1): 29–43.
Heppell J, Lorenzen N, Armstrong N K, et al. 1998. Development of DNA vaccines for fish: vector design, intramuscular injection and antigen expression using viral haemorrhagic septicaemia virus genes as model. Fish & Shellfish Immunology, 8: 271–286.
Kumar S R, Ahamed V P I, Sarathi M, et al. 2008. Immunological responses of Penaeus monodon to DNA vaccine and its efficacy to protect shrimp against white spot syndrome virus(WSSV). Fish & Shellfish Immunology, 24(4): 467–478.
Kumar S R, Parameswaran V, Ahmed V P I, et al. 2007. Protective efficiency of DNA vaccination in Asian seabass (Lates calcarifer) against Vibrio angullarum. Fish & Shellfish Immunology, 23(2): 316–326.
McCarter L L, 1995. Genetic and molecular characterization of the polar flagellum of Vibrio parahaemolyticus. J. Bacteriol., 77(6): 1 595–1 609.
McGee D, Horstedt P, Milton D L. 1996. Identification and characterization of additional flagellin genes from Vibrio anguillarum. J. Bacteriol., 178(17): 5 188–5 198.
Nie J H, Wang Y C. 2007. Research updates on DNA vaccine. Chinese Pharmaceutical Affairs, 21(2): 114–118.
Qin Y X, Wang J, Su Y Q, et al. 2006. Studies on the pathogenic bacterium of ulcer disease in Epinephelus awoara. Acta Oceanologica Sinica, 25(1): 147–153.
Rosas G, Cruz-Revilla C, Fragoso G, et al. 1998. Taenia crassiceps cysticercosis: humoral immune response and protection elicited by DNA immunization. Journal of Parasitology, 84(3): 516–523.
Rout N, Kumar S, Jaganmohan S, et al. 2007. DNA vaccines encoding viral envelope proteins confer protective immunity against WSSV in black tiger shrimp. Vaccine, 25(15): 2 778–2 786.
Schulte P M, Powers D A, Schartl M. 1998. Efficient gene transfer into Xiphophorus muscle and melanoma by injection of supercoiled plasmid DNA. Mol. Mar. Biol. Biotechnol., 7: 241–247.
Yang G C, Schrank G D, Freeman B A. 1977. Purification of flagellar cores of Vibrio cholerae. J. Bacteriol, 192(2): 1 121–1 128.
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Supported by Fujian Science and Technology Innovation Foundation for Young Scientists (No. 2006F3096) and Scientific Research Foundation of Jimei University
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Qin, Y., Su, Y., Wang, S. et al. Immunogenicity and protective efficacy of Vibrio harveyi pcFlaA DNA vaccine in Epinephelus awoara . Chin. J. Ocean. Limnol. 27, 769–774 (2009). https://doi.org/10.1007/s00343-009-9215-y
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DOI: https://doi.org/10.1007/s00343-009-9215-y