Comparative analysis of the sensitivity to distinct antimicrobials among Penicillium spp. causing fruit postharvest decay

Abstract

The postharvest fungal pathogensPenicillium digitatum, P. italicumand P. expansumare an in-creasing problem for the Mediterranean orchards and fruit industry. This study was designed to gain knowl-edge on factors affecting susceptibility of Penicilliumspp. to antimicrobial peptides (AMPs) as new antifungal compounds for plant protection. The previously characterized PAF26 is a novel penetratin-type AMP with ac-tivity against phytopathogenic fungi. Comparative analyses were conducted on the sensitivity of Penicilliumspp. to PAF26, to the cytolytic peptide melittin and to other antimicrobials. The research included microscopic observations, chitin quantification, virulence assays on citrus and apple fruits, and molecular phylogenetic re-lationships within Penicillium isolates from citrus fruit. Virulence analysis and phylogenetic reconstruction confirmed the host specificity and monophyletic origin for P. digitatum, contrary to the closely-related species P. expansumand P. italicum. A parallelism was found between sensitivity to PAF26 of Penicilliumisolates and to the chitin dye calcofluor white (CFW). No such correlation was found between sensitivity to PAF26 and to the membrane perturbing compound SDS or the oxidizing agent H2O2. Microscopy studies showed that mycelium and conidia from the PAF26-sensitive fungi were also prone to CFW staining, but no direct correlation with the mycelial chitin content was found. The data are consistent with the fact that fungal cell walls influence the outcome of the interaction of AMPs with fungi, and that PAF26 is more active towards Penicilliumcitrus fruit pathogens. In this context, CFW could help both to elucidate AMPs mode of action and in studies of the mechanisms of virulence and host specificity within Penicillium spp.