Plutella xylostella is the major cosmopolitan pest of brassica and other crucifer crops, the larval midgut of which is a dynamic organ that interfaces with a divers earray of physiological and toxicological processes. The draft sequence of the P.xylostella genome was recently released, but its annotation remains challenging because of the low coverage of this branch of life.Peptide sequencing by computational assignment of tandem mass spectra to a database of putative protein sequences provides an independent approach to confirm or refute protein prediction, which has been termed proteogenomics. In this study we carried out an in-depth proteogenomic analysis using shotgun HPLC-ESI-MS/MS approach with a multi-algorithme pipeline to complement genome annotation in the P.xylostella larval midgut.