Expression of pectate lyase from Colletotrichum gloeosporioides in C. magna promotes pathogenicity

To test the contribution of pectate lyase (PL) to promoting fungal pathogen icity, a pectate lyase gene (pel) from the avocado pathogen Colletotrichum gloeosporioides, isolate Cg-14, was expressed in C, magna isolate L-2.5, a pathogen of cucurbits that causes minor symptoms in watermelon seedlings an d avocado fruits. Isolate L-2.5 was transformed with pPCPH-1 containing hph -B as a selectable marker and the 4.1-kb genomic pel clone, Southern hybrid ization, with the 4.1-kb genomic pel clone or 2.13-kb hph-B cassette as pro bes, detected integration of pel in transformed C, magna isolates Cm-PL-3 a nd Cm-PL-10. Western blot (immunoblot) analysis with antibodies against Cg- 14 PL detected a single pi, secreted by L-2.5 at a molecular mass of 41.5 k Da, whereas the PL of C, gloeosporioides had a molecular mass of 39 kDa. Wh en pi, activity was measured 4 days after inoculation in pectolytic enzyme- inducing media (PEIM), transformed isolates Cm-PL-3 and Cm-PL-10 showed add itive PL activity relative to both Cg-14 and L-2.5. Transformed isolates al so showed additive maceration capabilities on avocado pericarp relative to the wild-type C, magna alone, but did not reach the maceration ability of C , gloeosporioides, However, more severe maceration and damping off develope d in watermelon seedlings inoculated with the transformed isolates compared with the two wild-type isolates, which showed no symptom development on th ese seedlings during the same period. Results clearly show the contribution of a single pel to the pathogenic abilities of C, magna and suggest that P L, is a pathogenicity factor required for the penetration and colonization of Colletotrichum species.