Construction and characterization of subtractive stage-specific expressed sequence tag (EST) libraries of the pinewood nematode Bursaphelenchus xylophilus

To establish expressed sequence tag databases of the two life stages (the dispersal and propagative stages) of pinewood nematode Bursaphelenchus xylophilus, subtractive EST libraries that were specific to the dispersal 4th larval stage (D4S) and the pine-grown propagative mixed (PGPS) stage were constructed by suppressed subtractive hybridization, and annotated by BLASTx and Gene Ontology (GO). A total of 1112 (57.7%) contigs from the D4S-cDNA library and 1215 (46.7%) contigs from the PGPS-specific cDNA libraries had matched BLASTx hits (E ≤ 10− 2), among which 913 (47.4%) and 960 (36.9%) contigs, respectively, were classified into three GO categories. A total of 14 genes were selected on the basis of stage-specific abundances and GO subcategories, and their transcription levels were analyzed by quantitative real-time PCR. We discussed the potentials of some stage-specific genes, such as sorbitol dehydrogenase, cysteine protease, venom allergen-like protein, and FMRFamide-like peptide, as diagnosis markers and novel control targets.