We obtained a full-length cDNA clone for the Mx gene of barramundi (Lates calcarifer), using RACE (rapid amplification of cDNA ends) polymerase chain reaction (PCR) amplification of RNA extracted from a barramundi brain cell line cBB. The Mx cDNA of 2.2kb contains an open reading frame (ORF) of 1875 nucleotides encoding a protein of 624 amino acids. The predicted barramundi Mx protein is 71.4 kDa and contains a tripartite guanosinetriphosphate (GTP)-binding motif at the amino terminal and a leucine zipper at the carboxyl terminal, characteristic of all known Mx proteins. Poly I:C-transfection induced the expression of Mx gene in cBB cells, and the induction level at 28 degrees C was higher than that at 20 degrees C. Moreover, Mx gene expression was also induced by viral infection, including fish nodavirus, birnavirus, and iridovirus. Among these, nodavirus was a stronger inducer than the other two viruses. Using an antiviral activity assay, we revealed that poly I:C-transfected cBB cells had antiviral activity against fish nodavirus and birnavirus, but not iridovirus. Furthermore, the replication of nodavirus and birnavirus could be restored after the expression of Mx gene was down-regulated by siRNA. Therefore, these results indicated that the expression of barramundi Mx gene was able to inhibit the proliferation of fish nodavirus and birnavirus.