Construction and Identification of a Recombinant Plasmid Encoding Echinococcus granulosus Oncosphere Antigen (EG95)Abstract Background: Cystic echinococcosis (CE), as a zoonotic disease cause to health threat and economic losses. Despite implemented cont
AbstractBackground: Cystic echinococcosis (CE), as a zoonotic disease cause to health threat and economic losses. Despite implemented control programs, few countries have been able to decrease or eliminate this infection. Vaccination of the intermediate host offers an additional strategy to control the parasite transmission and EG95 antigen is considered more than the others in the vaccine issue. According to the high protection induced by the EG95 recombinant vaccine, this study was designed to construct recombinant plasmid formulation of EG95 antigen.Methods: In 2015, the Echinococcus granulosus eggs were recovered from an infected dog in Parasitological laboratory of Tarbiat Modares University in Tehran, Iran. Following hatching, the oncospheres of E. granulosus were activated to increase the presence of the desired mRNA. The extracted mRNA was transcribed to the cDNA which used as template in RT-PCR. Then the EG95 gene cloned into pET28a vector and the recombinant plasmids expression was investigated in prokaryotic and eukaryotic cells.Results: The recombinant plasmid encoding EG95 antigen was successfully constructed and identified by PCR, restriction enzyme digestion and sequencing. In vitro expression of the EG95 antigen was confirmed in prokaryotic and eukaryotic systems by SDS-PAGE and western blotting analysis.Conclusion: Because of potential advantages of DNA vaccines, including ability to induce long-term immune responses, low production cost and stability in different temperatures, this study carried out to construct the EG95 gene into a vector. This recombinant vector can be evaluated in further studies as a DNA vaccine may provide new prospects for the development of a vaccine against cystic hydatid disease.
Craig PS, McManus DP, Lightowlers MW et al. Prevention and control of cystic echinococcosis. Lancet Infect Dis. 2007;7(6):385-94.
Donald P, Zhang W, Li J. Echinococcosis. Lancet. 2003;362(9392):1295-304.
Lightowlers M, Flisser A, Gauci C, Heath D, Jensen O, Rolfe R. Vaccination against cysticercosis and hydatid disease. Parasitol Today. 2000;16(5):191-6.
Eckert J, Gemmell M, Soulsby E. FAO/UNEP/WHO guidelines for surveillance, prevention and control of echinococcosis hydatidosis. 1981.
Rokni MM. Echinococcosis/hydatidosis in Iran. Iran J Parasitol. 2009;4(2):1-16.
Jabbar A, Jenkins D, Crawford S, Walduck A, Gauci C, Lightowlers M. Oncospheral penetration glands are the source of the EG95 vaccine antigen against cystic hydatid disease. Parasitology. 2011;138(01):89-99.
Lightowlers M, Jensen O, Fernandez E et al. Vaccination trials in Australia and Argentina confirm the effectiveness of the EG95 hydatid vaccine in sheep. Int J Parasitol. 1999;29(4):531-4.
Lightowlers M, Lawrence S, Gauci C et al. Vaccination against hydatidosis using a defined recombinant antigen. Parasite Immunol. 1996;18(9):457-62.
Heath D, Lightowlers M. Vaccination on hydatidology—state of the art. Arch Int Hidatid. 1997;33:14-6.
Karami A. DNA Vaccine (Genetic Immunization). Baqiyatallah University of Medical Sciences; 2014.
Pirestani M, Dalimi A, Sarvi S, Khoramabadi N. Evaluation of Immunogenicity of Novel Isoform of EG95 (EG95-5G1) From Echinococcus granulosus in BALB/C Mice. Iran J Parasitol. 2014;9(4):491-502.
Sarvi S, Dalimi A, Ghafarifar F. Molecular cloning and expression of EG95 gene of Iranian isolates of Echinococcus granulosus. Iran J Parasitol. 2012;7(2):1-7.
Zhang N-Z, Huang S-Y, Zhou D-H et al. Protective immunity against Toxoplasma gondii induced by DNA immunization with the gene encoding a novel vaccine candidate: calcium-dependent protein kinase 3. BMC Infect Dis. 2013;13(1):512.
Meng M, Zhou A, Lu G, Wang L et al. DNA prime and peptide boost immunization protocol encoding the Toxoplasma gondii GRA4 induces strong protective immunity in BALB/c mice. BMC Infect Dis. 2013;13(1):1.
Cui X, Lei T, Yang D, Hao P, Li B, Liu Q. Toxoplasma gondii immune mapped protein-1 (TgIMP1) is a novel vaccine candidate against toxoplasmosis. Vaccine. 2012;30(13):2282-7.
Robinson HL, Torres CA, editors. DNA vaccines. Seminars in Immunology; 1997: Elsevier.
Fynan EF, Webster RG, Fuller DH, Haynes JR, Santoro JC, Robinson HL. DNA vaccines: a novel approach to immunization. Int J Immunopharmacol. 1995;17(2):79-83.
Lightowlers MW, Gauci CG. Vaccines against cysticercosis and hydatidosis. Vet Parasitol. 2001;101(3):337-52.
Woollard D, Gauci C, Heath D, Lightowlers M. Epitope specificities and antibody responses to the EG95 hydatid vaccine. Parasite Immunol. 1998;20(11):535-40.
Dutton S, Fleming SB, Ueda N, Heath DD, Hibma MH, Mercer AA. Delivery of Echinococcus granulosus antigen EG95 to mice and sheep using recombinant vaccinia virus. Parasite Immunol. 2012;34(6):312-7.
Paykari H, Heath DD, Dalimi AH, Karimi GR, Motamedi GR. Experimental vaccination of sheep against hydatid cyst using EG95 recombinant vaccine. Arch Razi Institute. 2008;63(2):29-34.
Heath DD, Robinson C, Shakes T et al. Vaccination of bovines against Echinococcus granulosus (cystic echinococcosis). Vaccine. 2012;30(20):3076-81.
Osborn P, Heath D. Immunisation of lambs against Echinococcus granulosus using antigens obtained by incubation of oncospheres in vitro. Res Vet Sci. 1982.
Heath Dd, Lawrence SB. Antigenic polypeptides of Echinococcus granulosus oncospheres and definition of protective molecules. Parasite Immunol. 1996;18(7):347-57.
Chow C, Gauci CG, Cowman AF, Lightowlers MW. A gene family expressing a host-protective antigen of Echinococcus granulosus. Mol Biochem Parasitol. 2001;118(1):83-8.
Chow C, Gauci CG, Cowman AF, Lightowlers MW. Echinococcus granulosus: oncosphere-specific transcription of genes encoding a host-protective antigen. Exp Parasitol. 2004;106(3-4):183-6.
Lin RY, Ding JB, Wen H. Cloning, homological analysis and construction of Eg95 Xinjiang strain DNA vaccine. Hepatobiliary Pancreat Dis Int. 2003;2(4):545-8.
Sarvi S, Dalimi-Asl A, Ghaffarifar F, Sharifi Z. Determination and cloning of the gene encoding EG95 protein in Iranian isolate of Echinococcus granulosus. Feyz. 2011;15(3):194-9.