Molecular Characterization of Three B-1,4-Endoglucanase Genes in Pratylenchus loosi and Functional Analysis of Pl-eng-2 Gene

Abstract

Pratylenchus loosi is an important root-lesion nematode that causes damage to tea plantations in Iran and all over the world. The present study reports on the characterization and evolution of three ß-1,4-endoglucanase genes: Pl-eng-2, Pl-eng-3 and Pl-eng-4. The gene structure of Pl-eng-2 was fully determined with the predicted signal peptide and devoid of the linker domain and carbohydrate-binding domain, while Pl-eng-3 and Pl-eng-4 were only partially sequenced. The transcription of Pl-eng-2 was localized in the secretory esophageal glands of all life stages, but it was upregulated in male and female stages. The exon/intron structures of Pl-eng-2, Pl-eng-3 and Pl-eng-4 confirmed that they resulted from gene duplication followed by sequence and gene structure diversification with loss of the linker domain and carbohydrate-binding domain during evolution. A phylogenetic analysis further confirmed that nematode endoglucanases resulted from the horizontal gene transfer of a bacterial gene, as Pl-eng-3 showed sister relationships with the CelB cellulase of Bacillus subtilis. Silencing Pl-eng-2 by in vitro RNA interference produced a 60% decrease of the transcript level. The reproductive ability of silenced P. loosi showed a 35% reduction of eggs and larval stages compared to untreated nematodes, suggesting that this gene is involved in the early steps of invasion.