Identification, phylogenetic analysis and expression profile of an anionic insect defensin gene, with antibacterial activity, from bacterial-challenged cotton leafworm, Spodoptera littoralis

BACKGROUND: Defensins are a well known family of cationic antibacterial peptides (AMPs) isolated from fungi, plants, insects, mussels, birds, and various mammals. They are predominantly active against gram (+) bacteria, and a few of them are also active against gram (-) bacteria and fungi. All insect defensins belonging to the invertebrate class have a consensus motif, C-X(5-16)-C-X(3)-C-X(9-10)-C-X(4-7)-CX(1)-C. Only seven AMPs have already been found in different lepidopteran species. No report was published on the isolation of defensin from the Egyptian cotton leafworm, Spodoptera littoralis. RESULTS: An anionic defensin, termed SpliDef, was isolated from the haemolymph of the cotton leafworm, S. littoralis, after bacterial challenge using differential display technique. Based on sequence analyses of the data, specific primers for full length and mature peptide of defensin were designed and successfully amplified 471 and 150 bp amplicons. The integration of the results revealed that the 471 bp-PCR product has one open reading frame (orf) of 303 bp long, including both start codon (AUG) and stop codon (UGA). The deduced peptide consists of a 23-residues signal peptide, a 27-residues propeptide and a 50-residues mature peptide with the conserved six-cysteine motif of insect defensins. Both haemolymph and expressed protein exhibited antibacterial activities comparable to positive control. The RT-qPCR indicated that it was more than 41-folds up-regulated at 48 h p.i. CONCLUSION: Our results highlight an important immune role of the defensin gene in Spodoptera littoralis by cooperating with other AMPs to control bacterial infection.