Exogenous Melatonin Induces Disease Resistance Against Pathogen Infection In Cherry Tomato Fruit During Postharvest Storage

Cherry tomato fruit(Lycopersicon esculentum)is widely consumed due to its delicious taste,rich nutrition and low calories.However,tomato fruit is highly susceptible to Botrytis cinerea during postharvest storage,which can not only causes gray mold but also results in significant economic losses.Melatonin,which belongs to a highly conserved and small tryptophan molecule,and emerges by plant evolution,is widely distributed in the plant kingdom and plays pleiotropic roles in fruit,such as development,ripening,senescence and various biotic and abiotic stress.However,the impact and regulatory mechanisms of melatonin on improving quality and attenuating fungal infection in cherry tomato fruit are still unclear.In this study,we investigated systemically the potential effects of pre-harvest and post-harvest melatonin treatments on fruit ripening,fruit quality and disease resistance in cherry tomato fruit.The results showed that:(1)Melatonin treatment with low concentrations(0.05 and 0.1 mM)can delays significantly the color development(green turned red),the degradation of firmness,and reduces the decay incidence and weight loss,enhances total soluble solids content(TSS)of cherry tomato fruit,which suggests that melatonin may delays the ripening process and promots the qulitity of cherry tomato fruit during postharvest storage.Additionally,melatonin treatment increased the content of reduced glutathione(GSH)and ascorbic acid(ASA),promoted the activities of relative enzymes in the ascorbate-glutathione(GSH-ASA)cycle,and enhanced the total antioxide capacity(T-AOC)of tomato fruit,which may contributes to reducing the content of the relative membrane permeability and malondialdehyde(MDA),and delaying the senescence in cherry tomato fruit.In addation,exogenous melatonin treatment enhanced significantly the content of endogenous melatonin by trigerring the genes expression of melatonin biosynthesis tryptophan decarboxylase(SlTDC)and N-acetylserotonin methyltransferase(SlASMT)in cherry tomato fruit during postharvest storage,which may relates to its effects to regulating fruit ripening and senescence of cherry tomato.(2)Exogenous melatonin with low concentrations(0.05 and 0.1 mM)promoted the mycelial growth of Botrytis cinerea,but inhibited significantly the disease diameter of tomato fruit inoculated with Botrytis cinerea,which may implies that melatonin can inhibited the infection of Botrytis cinerea by inducing the disease resistance of cherry tomato fruit furing postahravest storage.Moreover,melatonin treatment induced the reactive oxygen species(ROS)burst,promoted the endogenous salicylic acid(SA),but inhibited significantly the content of nitric oxide(NO),and did not influence the content of jasmonic acid(JA),which may suggests that melatonin inducing disease resistance in tomato fruit may depends on the ROS and SA signaling.At the same time,melatonin treatment enhanced the enzymatic activities of NAPDH oxidase,chitinase(CHT),?-1,3-glucanase(GLU)and peroxidase(POD)and promoted the phenylpropanoid pathway by promoting activities of phenylalanine ammonia-lyase(PAL),4-coumarate-coenzyme A ligase(4CL)and cinnamate-4-hydroxylase(C4H)and accompanied the higher contents of flavonoids,total phenols and lignin in fruit,which can be conducive to enhancing disease resistance against pathogenic fungi such as B.cinerea in cherry tomato fruit during postharvest storage.(3)In order to further reveal whether the melatonin inducing disease resistance mainly dependent on the endogenous SA or not,the paclobutrazol(PAC),which can inhibits the SA biosynthesis in plant,was selected to treatment the postharvested tomato fruit.The results indicated that melatonin treatment increased the activity and expression of SlNADPH oxidase,induced early ROS burst,as well as enhanced the enzymatic activities of superoxide dismutase(SOD),ascorbate peroxidase(APX),and POD,thus it can maintains ROS levels.However,compared with the melatonin treatment,compound treatment(melatonin combined with PAC)did not make significantly differences for the enzymatic activities of NADPH oxidase,SOD and POD,but inhibited the early burst of ROS throught regulating the activity of catalase(CAT).Moreover,melatonin treatment can reduces the content of NO by inhibiting the activity of nitric oxide synthase(NOS),as well as the compound treatment further decreased the cotent of NO and the activity of NOS.Those results can indicate that melatonin-induced disease resistance depended on NADPH-dependent ROS signaling but not NO signaling.Exogenous melatonin treatment enhanced significantly the content of the endogenous melatonin by trigerring the genes expression of melatonin biosynthesis including SlTDC,serotonin N-acetyltransferase(SlSNAT)and SlASMT.However,compared with the melatonin treatment,compound treatment did not cause significantly effects on the content of endogenous melatonin in fruit,which seemed to indicate that the synthesis of melatonin in fruit may be dependent on ROS signaling,and independent on NO signaling.Melatonin treatment enhanced the accumulation of SA by promoting the enzymatic activities of PAL and benzoic acid 2-hydroxylase(BA2H),increasing the gene expression of SlPAL5 in cherry tomato fruit.Furthermore,melatonin treatment upregulated markedly the genes expression of SA signaling pathway and pathogenesis-related(PR)proteins,such as SlWRKY70,SlPAD4,SlNPR1,SlTGA5,SlPR1 and SlPR2,as well as boosted the activities of chitinase(CHI)and?-1,3-glucanase(GLU).Melatonin treatment inhibited effectively the disease diameter of cherry tomato fruit inoculated with Botrytis cinerea and postharvest decay of fruit during postharvest storage.However,compared with the melatonin treatment,compound treatment significant decreased the content of SA by inhibiting the enzymatic activity of BA2H and the gene expression of SlPAL5,inhibited significantly the vital genes expression of SA signaling pathway,reduced the enzymatic activities of the CHT and GLU,and accompanied the serious disease in tomato fruit.Those results indicated that metaonin inducing disease resistance is mainly dependent on SA signaling,and related to activiting systemic acquire resistance(SAR)in tomato fruit during storage.(4)Pre-harvest spraying with melatonin one time at different special phase during fruit development of tomato fruit increased the content of TSS,but decreased the content of titrable acids(TA)and the rate of weight loss,as well as maintainted the firmness of fruit,delayed the the color development(green turned red)and inhibited effectively the disease diameter of cherry tomato fruit inoculated with Botrytis cinerea.The respiration rate and decay incidence of fruit were enhanced by melatonin-treated during the earlier period of the fruit developing(young stage and early fruit swelling),however,the opposite results of respiration rate and decay incidence were found in melatonin-treated during the later period of the fruit developing(late fruit enlargement and pre-harvest two day).Interestingly,compared with other treatments,melatonin-treated at the late fruit enlargement reduced significantly the rotting rate of tomato fruit during storage.At the same time,pre-harvest melatonin-treated is better than postharvest melatonin-treated on improving the quality and reducing decay incidence in tomato fruit during storage.Therefore,in order to revealing the mechanisms of melatonin decreasing the disease infection by pre-harvest spraying treatment,we researched the vital resistance signals of fruit during postharvest storage in this chapter.The results showed that melatonin treatment enhanced the capacity of total antioxide(T-AOC),and maintained the integrity of cell membrane,thus it can delays fruit senescence.Preharvest treatment with melatonin promoted the accumulation of endogenous melatonin by trigerring the genes expression of melatonin biosynthesis,including SlSNAT and SlASMT,induced ROS burst by enhancing the activity of NADPH oxidase,enhenced the content of endogenous SA by trigerring the gene expression of SlICS,as well as up-regulated the vital genes expression of SlICS,SlWRKY70,SlPAD4,SlNPR,SlTGA2.2,SlTGA5,SlPR1,SlPR2,SlPR5,boosted of activity of some protection enzymes such as CHI and GLU,and reduced significantly the content of NO during the whole storage,which can suggests that melatonin enhancing the disease resistance is mainly by inducing the ROS brust,enhancing the SA signaling,and then activating systemic acquire resisance in tomato fruit during postharvest storage.In addition,pre-harvest treatment with melatonin enhanced the activities of PAL,C4H and 4CL,and accompanied the higher contents of flavonoids,total phenols and lignin in tomato fruit.It indcated that pre-harvest treatment with melatonin can enhances the phenylpropanoid pathway that contribute to enhancing resistance in tomato against fungus during postharvest storage.(5)Transcriptomic date of pre-harvest treatment with melatonin(1mM)during the late fruit enlargement of cherry tomato fruit suggested that melatonin treatment altered significantly the genes expression of mainly related to disease resistance pathways,including plant hormone signal transduction,plant-pathogen interaction,MAPK signaling pathway-plant,phenylpropanoid biosynthesis and flavonoid biosynthesis.Moreover,melatonin regulating ROS and NO signals may relate with Ca²⁺-dependent inducing hypersensitive response(HR),which can enhances cell wall reinforcement and promotes the disease resistance in tomato fruit.The genes expression of relation with SA signaling pathway,including SlNPR1,SlTGA7 and SlPR1,were up-regulated by melatonin-treated in cherry tomato fruit.At the same time,the most significantly differential genes of phenylpropanoid biosynthesis pathway were enriched in lignin biosynthesis pathway.In conclusion,the results indicated that exogenous melatonin treatment can delays the ripening and senescence of cherry tomato fruit by adjusting the fruit respiratory rate,promoting the ascorbic acid-glutathione(ASA-GSH)cycle,enhancing the activity of antioxidant enzymes,maintaining the integrity of cell membrane during postharvest storage.Exogenous melatonin treatment can enhances the disease resistance of cherry tomato fruit by inducing the burst of ROS,enhancing the content of SA,up-regulating the genes expressions of SA signaling pathway,and promoting the accumulation of pathogenesis-related proteins(PR1,PR2,PR5,CHT),activating phenylpropanoid biosynthesis pathway and enhancing the accumulation of antimicrobial substances(including the flavonoids,total phenols and lignin).