A Study On The Biocontrol Efficacy Of Pichia Anomala Induced With Chitosan Against Blue Mold Disease Of Table Grapes Caused By Penicillium Expansum

Blue mold diseases caused by Penicillium expansum was earlier known as apple postharvest disease.Recently,numerous reports showed that P.expansum is the main cause of grapes deterioration.In the last five decades,chemical fungicides such as sulfur dioxide(SO₂)were used to control the postharvest diseases of table grapes.However,due to consumer's interest in organic consumption and environmental sustainability induced searches for alternative control methods become crucial.Among the available methods,bio-control methods are gaining more interest in their environmental friendliness,consumer health benefits,and sustainability.Antagonistic microorganisms such as yeasts are increasingly reported for their ability to control postharvest diseases of numerous fruits and vegetables.This is because yeasts present more appealing characteristics like their do not to produce mycotoxins,their non-allergenic effect,and benefits to animal&human health.Yeast can grow on a wide range of nutrients and can also survive in low water activity and low oxygen environment.The ascomycetous yeast Pichia anomala belongs to the non-saccaromyces wine yeasts and contributes to wine aroma by the production of volatile compounds.It has the ability to grow under low p H,low oxygen,and high osmotic pressure.The yeast can also use a broad range of nitrogen and phosphor sources,which makes it a potential agent to decrease environmental pollution by organic residues from agriculture.Previous studies reported that P.anomala has a high antagonist effect against Penicillium roqueforti,P.carneum,and P.paneum.P.anomala is also reported as a bio-control agent during the storage of high-moisture feed grain under airtight conditions.However,it's potential has never been tested in grapes affected by P.expansum.Biological control is a safe and effective method,and protecting fruits from postharvest diseases with biocontrol yeasts has become a hot spot in academic research.Compared with chemical fungicides,however,the biocontrol effect of yeast preparation is weak,so how to improve the effect is an urgent problem to be solved.One of the key measures to improve the biocontrol effect is to induce antagonistic yeast with elicitor.Chitosan is a natural,safe,non-toxic,and biodegradable food additive.This study aimed to analyse the bio-control efficacy of P.anomala induced with chitosan against blue mold disease of table grapes caused by P.expansum,moreover,the study determines the mechanisms of blue mold disease control efficacy in enhanced disease resistance of table grapes induced by P.anomala cultured with chitosan from both physiological and molecular levels.The summary of main research outputs are the following:(1)Table grapes induced by P.anomala cultured with chitosan(1%w/v)can significantly increased the population of the yeast both in the fruit surface,wound and in the media.Moreover,it up-regulated the yeast enzymatic activities like?-1,3-glucanase,malondialdehyde,and catalase.P.anomala cultured with chitosan(1%w/v).P.anomala induced with chitosan(1%w/v)significantly reduced the disease incidence,lesion diameter and natural decay of grapes without affecting the fruit quality compared with the control.Furthermore,the yeast inhibited the spore germination and hyphae length of P.expansum after 10 h incubation in the PDB media.The result confirmed that P.anomala has direct antagonism characteristics against P.expansum.(2)Table grapes induced by P.anomala induced with chitosan(1%w/v)up-regulated disease defence related enzymatic and non-enzymatic activities,mainly includes:polyphenol oxidase(PPO),catalase(CAT),peroxidase(POD)and phenylalanine ammonia-lyase(PAL),chitinase(CHI),ascorbate peroxidase(APX)and chitinase(CHI),and the expression levels of genes coding these corresponding enzyme;enhanced the total phenol and flavonoid contents of table grapes;decreased the formation of malondialdehyde(MDA)and hydrogen peroxide(H₂O₂)which are responsible for fruits deterioration.In addition,the scanning electron microscope(SEM)concisely illustrated that high denser the yeast can directly inhibit the growth of P.expansum mycelia.(3)The gene expression levels of table grapes treated by P.anomala induced with chitosan(1%w/v)were assessed by transcriptome technology;compared to untreated table grapes,table grapes induced by P.anomala induced with chitosan(1%w/v)were up-regulated the gene expression of the plant-pathogen interaction pathway,plant MAPK signaling pathway,plant hormone signal transduction pathway,the pathway of glutathione metabolism,the pathway of phenylaline,tryosine,and trptopham biosynthesis,and the pathway of flavonoid biosynthesis in table grapes.These help the fruit to synthesize disease resistance substances,regulate the reactive oxygen species(ROS),and enhance the fruit cell wall,and enrich the hormone signal transduction during the pathogen attack,thereby enhancing the disease resistance of table grapes.(4)The final study determined the physico-chemical quality parameters(including water loss,titratable acidity,soluble solid content,hardness,ascorbic acid content)and sensory qualities(including the appearance of the grape,the color of the pedicle,the degree of expansion,the taste,etc.)after table grapes treated by P.anomala induced with chitosan(1%w/v).The result showed that the combination of cold storage and treatment with the yeast better retained the fruit physico-chemical quality parameters and make the table grapes more acceptable with wider sensory attributes even after long storage of 30 days as compared to the non-treated berries.