Effect Of Chitin On The Biocontrol Activity Of Marine Yeast Rhodosporidium Paludigenum Against Postharvest Fruit Diseases And The Possible Mechanisms Involved

Postharvest decay of fruits caused by fungal disease leads to substantial economic losses world-widely. Postharvest biological control employing antagonistic yeasts has emerged as a promising alternative to synthetic fungicides considering certain progress has made in recent years. Our previous studies indicated that the marine yeast strain of Rhodosporidium paludigenum Fell&Tallman isolated from the East China Sea has great biocontrol efficacy in inhibiting various postharvest diseases of fruit. However, at present, when used alone, the yeast antagonists could not control the postharvest diseases as effectively as synthetic fungicides when used alone, hence how to effectively improve the efficacy of antagonistic yeasts is the key factor affecting commercial application.Chitin is a linear homopolymer of N-acetylglucosamine being linked by β-1,4glycosidic bonds, which as a biopesticide approved by U.S. Environmental Protection Agency (EPA) and also a food additive approved by U.S. Food and Drug Administration (FDA), has wide range of application potential in various fields. This paper mainly investigated the effect of chitin on the antagonistic activity of R. paludigenum, the influence factor and the possible mechanisms involved. Meanwhile, the proteomic analysis and comparison of R. paludigenum in response to chitin was analyzed based on isobaric tags for relative and absolute quantitation (iTRAQ).The main results were summarized as follows:(1) The antagonistic activity of R. paludigenum against a variety of postharvest pathogen was greatly enhanced when it was cultivated in nutrient yeast dextrose with added chitin (NYCB) medium compared with that cultivated in nutrient yeast dextrose (NYDB) medium. The enhancement of antagonistic activity of the yeast harvested from NYCB was greater than that harvested from NYDB amended with other substances including trehalose, glucan and chitosan.(2) The optimal induction concentration of chitin was1.0%(w/v), and the optimal cultivation time was36h. After the optimal conditions for induction culture, the biocontrol efficacy of R. paludigenum was significantly increased at each concentration. Moreover, under refrigerated conditions and preharvest treatment conditions, the biocontrol efficacy of R. paludigenum was also remarkably enhanced through cultivation in NYDB added with chitin.(3) Adding chitin and sodium chloride to NYDB could both improve the antagonistic activity of R. paludigenum to a certain extent, while there was no synergistic effect between these two substances, and chitin might play the leading role in combination induction cultivation.(4) The possible mechanisms involved that cultivated in chitin-supplement medium could significantly improve the biocontrol efficacy of R. paludigenum might include the following:①population growth in fruit wounds and population dynamics on fruit surface of R. paludigenum harvested from NYCB were considerably higher than that harvested from NYDB;②the ability of R. paludigenum produced in NYCB to induce resistance was significantly enhanced, which might be associated with stimulation of resistance related enzymes activities and Inhibition of lipid peroxidation in fruit treated with the yeast;③ antioxidant enzymes activities of R. paludigenum were stimulated by cultivating in NYCB, while malondialdehyde accumulation in the yeast cells was suppressed;④the cell-free filtrate of R. paludigenum cultivated in NYCB showed higher activities of chitinase and β-1,3-glucanase than that cultivated in NYDB.(5) The proteomic analysis and comparison of R. paludigenum in response to chitin was analyzed through iTRAQ, and the results showed:①there were587differentially expressed proteins between the yeast harvested from NYCB and NYDB, in which327up-regulated proteins and260down-regulated proteins;②ribosome proteins were up-regulated overall, mainly for40S small subunit and60S large subunit;③proteins related to glycolysis and pentose phosphate pathways were up-regulated overall including hexokinase which was the key enzyme in the glycolytic pathway and glucose-6-phosphate1-dehydrogenase which was the key enzyme in the pentose phosphate pathway;④significant changes of protein expression in multiple pathways of amino acid metabolism were occurred, among them delta-1-pyrroline-5-carboxylate dehydrogenase was up-regulated, which played an important role in catalytic formation of proline from glutamic acid metabolism;⑤9antioxidant-related proteins were significantly up-regulated including catalase A, cytochrome-c peroxidase, glutathione peroxidase, peroxiredoxin5and mitochondrial peroxiredoxin6;⑥trehalose6-P phosphatase and trehalose synthase were significantly up-regulated which related to trehalose synthesis;⑦one heat shock protein/chaperonin and one heat shock transcription factor were up-regulated while one protein of heat shock protein Hsp70family down-regulated.In conclusion, adding chitin to normal media might be an effective method to improve the antagonistic activity of marine yeast R. paludigenum, and the possible mechanisms involved were analyzed through conventional methods and iTRAQ, while deserves to be elucidated further.