Abstract:

The Enterocytozoon hepatopenaei(EHP) from shrimp is a very important pathogenic microorganism in aquaculture. In present, the common method for EHP detection is PCR reaction which use ribosomal 18S gene as a target. The problem with this method is nonspecific amplification because of high conservatism of 18S gene. In this study, the spore wall protein 2 (EHPSWP2) was cloned with silico cloning method. The length of the cDNA of the SWP2 was 1 019 bp containing an open reading frame of 687 bp encoding a protein with 229 amino acids with predicted molecular weight of 26.32 ku, and 5’ UTR is of 120 bp and 3’ UTR is of 179 bp. The amino acids sequences and phylogenetic tree analysis showed that there is a close relationship among EHP, Enterocytozoon bieneusi and Hepatospora eriocheir. The recombinant expression vector pET15b-SWP2 was constructed for SWP2 protein expression, the molecular weight was 28 ku and the monoclonal antibodies against SWP2 protein were produced. The methods of PCR and western blot method were applied for EHP detection with Macrobrachium rosenbergii samples. This study provided a technical guarantee for the diagnosis of EHP disease by farmers, and laid the foundation for SWP2 protein research.

Key words: Enterocytozoon hepatopenaei, SWP2, protein expression, monoclonal antibody, disease detection