Rhodosporidium Paludigenum Induces Disease Resistance And Defense-Related Mechanisms Against Decays In Citrus Fruit

In addition to physiological losses (dehydration, cold damage, etc.), latent infection and cross infection are the main factors causing fruits and vegetables decay during preharvest and postharvest proccess. Fungal pathogens exploit three main routes to penetrate the host tissue: through wounds; through natural openings, such as lenticels, stem ends, and pedicels; and by direct breaching of the host cuticle, which enables most postharvest pathogens to enter the unripe tissue and remain quiescent for months until the fruit ripens (resistance decline). Biological, physical or chemical elicitors were successfully employed to enhance host defense against various postharvest diseases, leading to a markable decrease in economic losses. In order to address the relationship between Rhodosporidium paludigenum and postharvest citrus fruits, as well as the relationship between biological elicitor and host tissues, postharvest pathology, transcriptomics, metabolomics, molecular biology and signaling pathways receptor competition /blocking technology were employed to reveal host cell changes in transcriptome and metabolic levels. Besides, the roles of ethylene and chitosan in antagonistic yeast induced citrus fruit resistance were also studied. The main findings are as follows:1. R. paludigenum was novel biocontrol yeast which could significantly inhibit the development of various postharvest fruit diseases. Meanwhile, R. paludigenum was the most effective yeast among three selected yeasts in the stimulation of the resistance to green mold of citrus fruit. Preharvest applications of R. paludigenum significantly reduced postharvest decays of mandarin orange caused by Penicillium digitatum and Penicillium italicum. The induction of resistance to Penicillium decay through the treatment with R. paludigenum is associated with significant enhancement of the activities of some defense-related enzymes, comprising β-1,3-glucanase, phenylalanine ammonia-lyase, peroxidase and polyphenoloxidase.2. The microarray data mining revealed that R. paludigenum activated transcription of genes important for plant hormones, signaling transduction, stress and defensive responses in orange peel tissue. Moreover, up-regulation of phenylalanine and tyrosine metabolism, phenylpropanoids biosynthesis, and alkaloid biosynthesis I, were observed at the transcription level. Conversely, large amounts of genes involved in starch metabolism, oligosaccharide and glycoside metabolism were markedly repressed by R. paludigenum treatment. Activation of phenylpropanoids biosynthesis pathway was correlated with the increasing production of phenolic acids and their subsequent metabolite lignin, indicating antifungal metabolites indeed contributed to biocontrol yeast enhanced fruit protection.3. Blocking ethylene perception with 1-MCP resulted in an increase in ACS1, ACS2 and A CO expression, and consequently an increase in ethylene production during mechanical wounding and resistance induction. The expression of the ethylene receptors ETR1, ETR2 and ETR4 as well as ethylene response factors (ERFs) were observed with similar responses to yeast and 1-MCP stimuli, with ETR3 mRNA accumulation being the most sensitive to yeast application while ERS1 was the least sensitive. When applied at concentrations greater than 500 ppb 1-MCP pre-fumigation significantly reduced the fruit’s natural protection and R. paludigenum induced disease resistances to P. digitatum decay, indicating that ethylene perception was involved in disease resistance induction. Moreover, the expression of the yeast up-regulated defensive genes CHI, β-1,3-glucanase, PAL and CIN that were inhibited to different degrees by the 1-MCP pre-treatment in the Ponkan mandarin.4. Treated with antagonistic yeast R. paludigenum and pathogen P. digitatum could induce 234 detectable metabolites in citrus fruit peel, and 54 substances changed significantly among them (| Fold Change|≥2). According to the responses of 54 metabolites to different microorganisms’treatment, Malic acid, L-Proline, Butanoic acid, Xylitol, D-Mannitol and Maltose were found to be specific response to antagonistic yeast stimuli in citrus peel; meanwhile, there were also nine substances showing specific responses to Penicillium infection, such as Glucaric acid, Galactonic acid, Levoglucosan and Cobaltocene. Another fourteen substances, such as Oleic acid, a-DL-Lyxopyranose, D-Ribopyranose, Isoborneol and propiophenone, shows specific responses to both microbes’ treatments at the same time. Furthermore, up-regulation of metabolic pathways, ABC transporters, Biosynthesis of secondary metabolites and phosphotransferase system were observed in metabolism level.5. Oranges treated with COS or R. paludigenum were observed having a delay in onset and progression of disease symptoms relative to wounded controls. Preharvest application of COS at different concentrations achieved similar biocontrol efficiency rates in green mold control after 4 days storage. However, the combination of COS (1%, w/v) and R. paludigenum showed a more effective decay control than any other treatments. COS (1%, w/v) alone did not negatively affect R. paludigenum growth in wounds, but severely inhibited P. digitatum spore germination than lower-dose treatments in vitro. The expression levels of the defense-related genes chitinase and phenylalanine ammonia lyase increased with decreased disease symptoms. Moreover, this phenomenon was more prominent in the integrated treatments than in the individual ones.