Detection of a chitinase-like protein in the roots of Douglas-fir trees infected with Armillaria ostoyae and Phellinus weirii

Protein was extracted from root bark of 11- and 25-year-old interior Dougla s-fir (Pseudotsuga menziesii (Mirb.) France) trees that were naturally infe cted with Armillaria ostoyae (Romagnesi) Herink. The proteins were separate d by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Root bark tissue adjacent to infected areas had a significantly higher prot ein concentration than healthy tissue (P < 0.05), whereas the protein conce ntration of infected tissue was consistently lower (P < 0.05) than that of healthy tissue. The SDS;PAGE profiles of healthy, infected, and adjacent-to -infected root bark tissues revealed significant differences in concentrati ons of a 29.3-kDa protein. The N-terminal amino acid sequence of the 29.3-k Da protein displayed significant homology (P = 0.013) to a basic endochitin ase. Use of a polyclonal antibody raised against the 29.3-kDa putative endo chitinase-like protein (ECP) indicated differences in the quantities of ECP in healthy roots compared with roots infected with A, ostoyae in 11- and 2 5-year-old interior Douglas-fir trees. The antibody was also used to screen for the presence of the 29.3-kDa protein in roots of 24-year-old coastal D ouglas-fir (Pseudotsuga menziesii var. menziesii) trees that were artificia lly inoculated with and colonized by Phellinus weirii (Murr.) Gilbn. The am ount of ECP was elevated in root bark of coastal Douglas-fir in response to P. weirii infection, although in lower quantities relative to those found in the A. ostoyae-interior Douglas-fir pathosystem. The sequence homology o f the ECP with a basic chitinase, together with its increased synthesis in response to two fungal pathogens, indicate a possible role for this protein in the defense of Douglas-fr against fungal pathogens.