Submitted to: International Society for Horticultural Science Meeting
Publication Type: Abstract only
Publication Acceptance Date: 6/1/2004
Publication Date: 6/30/2004
Citation: Droby, S., Bar-Shimon, M., Yehuda, H., Cohen, L., Daus, A., Goldway, M., Wisniewski, M.E. 2004. Involvement of lytic enzymes in the mode of action of the yeast candida oleophila used to control postharvest diseases. Proceedings of the 5th International Postharvest Symposium, Verona, Italy, June 6-11, 2004. Interpretive Summary:
Technical Abstract: Considerable research effort has been devoted in recent years to identifying microbial antagonists that effectively control postharvest diseases of fruit and vegetables. Among these, the yeast antagonist Candida oleophila was developed as a commercial product under the trade name AspireTM. Although the biocontrol activity of C. oleophila was demonstrated on a variety of commodities in commercial applications, its mode of action has not been fully elucidated. The current study was aimed at characterizing the ability of the yeast C. oleophila to produce a variety of lytic enzymes as well as defining their role in its mode of action. C. oleophila is capable of producing and secreting various cell wall degrading enzymes including exo-B-1,3-glucanase, chitinase, and protease in the growth medium as well as fruit surface wounds. Production of these enzymes was influenced by the presence of cell-wall fragments of Penicillium digitatum. Exo-B-1,3-glucanse (CoEXG1) and chitinase (CoCH1) genes were cloned and fully characterized. A transformation system for C. oleophila was developed to study the role of lytic enzymes in the biocontrol activity. Initially, the role of exo-glucanse in the biocontrol activity of C. oleophila was tested by using CoEXG1-knockouts and double-CoEXG1 over-producing transformants. Exo-glucanase activity was completely lost in the CoEXG1-knockouts, whereas, at least two-fold activity was detected in the over-producing transformants. CoEXG1-knockouts transformants showed lower inhibitory activity against Penicillium digitatum in in vitro and in vivo tests. C. oleophila was found, also, to produce and secrete a ribonuclease (RNase) into the growth medium. The T2-RNase gene in C. oleophila was amplified using PCR with degenerative primers that were derived from relatively homologous sequences in T2 RNase of C. albicans, cloned and sequenced. In in vitro assay, spore germination of P. digitatum, P. italicum and P. expansum was readily inhibited by yeast ribonuclease.