ABSTRACT
In order to detect Infectious hematopoietic necrosis virus with immunological methods, the surface glycoprotein of a recent IHNV-Sn isolated from farmed rainbow trout ( Oncorhynchus mykiss ) in China was amplified and cloned into pET27b(+) vector (designated as pET27b-G ). The expression of recombinant plasmid pET27b-G in E. coli BL21(DE3) was induced and determined by SDS-PAGE analysis. The predicted molecular weight of glycoprotein protein was approximately 55 kD and was confirmed in this study. The inclusion body of glycoprotein was treated with urea at different urea concentrations, and dialyzed into PBS buffer. Purified glycoprotein with high concentration was obtained after dialyzed in the PBS buffer. Antisera against glycoprotein were produced from immunized rabbits. The prepared antisera could react specifically with both the recombinant glycoprotein and natural glycoprotein of the IHNV-Sn isolated in the test of indirect ELISA, and the titer against the recombinant glycoprotein was 1:20,000. IFA showed that the antisera can recognize the glycoprotein located on the surface of IHNV-Sn and IHNV reference strain. These results indicated that the expressed glycoprotein was immunogenical and antigenical and could be functional as the natural IHNV glycoprotein. These results established a foundation for further study on vaccine and rapid diagnosis of IHNV.
Subject(s)
Animals , Female , Rabbits , Antibodies, Viral , Allergy and Immunology , Escherichia coli , Genetics , Metabolism , Fish Diseases , Allergy and Immunology , Virology , Gene Expression , Glycoproteins , Genetics , Allergy and Immunology , Infectious hematopoietic necrosis virus , Genetics , Allergy and Immunology , Neutralization Tests , Oncorhynchus mykiss , Rhabdoviridae Infections , Allergy and Immunology , Virology , Viral Proteins , Genetics , Allergy and ImmunologyABSTRACT
Infectious hematopoietic necrosis [IHN], viral hemorrhagic septicemia [VHS] and infectious pancreatic necrosis [IPN] as important viral diseases of salmonids, especially rainbow trout can be led to mass mortality among cultured fish. The aim of the present study was to show if IHN, IPN and VHS viruses can be detected in Iranian and imported rainbow trout eyed eggs and to compare their abundances among the hatcheries of 3 regions [Farsan, Koohrang and Lordegan] in Chaharmahal va Bakhtyari Province. In each area, three hatcheries were selected, 20 eyed eggs were randomly sampled from each farm. The samples were transferred into the sterile tubes and identified by reverse transcriptase -PCR. Meanwhile, physicochemical factors were recorded for each fish. While total infection rate in eggs was 23.3%, the level of infection for IPN, IHN and VHS was 12.5%, 10% and 0.83%, respectively. In this respect, maximum and minimum frequency were observed in Farsan [19.15%] and Koohrang [0.83%]. Comparison of the infected of eggs, based on their origin, showed that the infection rates in Iranian and imported eggs were 20% and 3.33%, respectively. The results revealed that rainbow trout eggs should be considered as major source for transmission of aquatic viruses. Hence, molecular identification of above mentioned viruses in rainbow trout eggs should be done