Mechanisms Of Apple Resistance Enhancement Toward Valsa Canker Disease Mediated By Potassium Application

Apple Valsa canker is a devastating disease in apple production,known as the "apple tree cancer",it has seriously hampered the development of the apple industry in China.Previous laboratory studies found that the potassium content of the apple tree effects canker disease severity.This study aims to determine how potassium improves the tree body resistance.Since apple is a perennial plant with seasonal variation in growth rate,large differences in nutritional level among trees,studying the relationship between nutrition level and disease resistance is difficult to carry out.In this study,hydroponic culture and callus culture were taken advantage to explore the effect of plant potassium content on Valsa canker resistance level;Through qRT-PCR,the expression pattern of genes related to phenolic synthesis and pathogenesis-associated proteins in callus tissues with different potassium levels were determined.This study aims to clarify the role of potassium in the interaction between apple Valsa canker and apple,and to provide a theoretical basis for controlling apple tree rottenness with tree nutrition.Main results are listed as follows:1.A Gala tissue culture system was established and callus with different contents of potassium were obtained,which laid a foundation for the study of Valsa canker resistance mechanisms.First culture medium is: MS + agar 8 g + Sucrose 30 g + 0.5mg / L6-BA + 0.5mg / L IBA + 0.1 mg / LIAA + PVP 4 g;subculture medium is: MS + agar 8 g + sucrose 30 g + 0.5 mg /L6-BA+0.2 mg / LIBA.Rooting medium was: 1 / 2MS + sucrose agar 8 g + 15 g + 0.3mg / LIBA + 1.0mg / LIAA.Callus induction medium is: MS + sucrose agar 8 g + 30 g + 2.0 mg / L6-BA + 2.0 mg / L 2,4-D + 0.4 mg / LNAA.The callus potassium level on MS medium was set as 1 k,based on which 2 K,1/2 K,and 0 k levels were set up.Callus was cultured on such medium for one month to obtain callus tissues with different potassium levels.2.Seedlings of root stock M26 and T33 with different potassium contents were obtained with hydroponic culture method: The potassium level of 1/4 hoagland nutrient solution was set as 1 k,based on which 2 K,1/2 K,and 0 k levels were set up.Apple seedlings were cultivated for 20 weeks to obtain hydroponic root stock with different potassium levels.3.Inoculation experiment was performed with annual hydroponic apple root stocks M26 and T337 containing different levels of potassium,in which M26 showed better resistance at the same level of potassium: M26 stock was immune toward Valsa canker when the leaf K content reached 1.5%,while T337 was still highly susceptible when the leaf potassium content reached 1.5%.T337 showed an immune response,when the leaf potassium content reached 2.3%.4.Inoculation experiment was performed with callus containing different levels of potassium,in which a positive correlation was found between callus potassium content and resistance level at the tested range.Callus with high potassium content exhibited tiny lesion at 48 h post inoculation whereas callus with low potassium content showed water-soaking and rot symptoms.These results showed that callus tissue could be used for studying the resistance mechanism of apple toward Valsa canker disease.5.In callus infection system,potassium level significantly affects the gene expression of phenolics synthesis pathway 4-p-coumaric acyl CoA ligase,flavonoid glycosyltransferase,chalcone isomerase,phenylalanine ammonia lyase,chalcone synthase,and two flavanones 3-hydroxylase,the gene expression at 2K were 2.51 times,6.67 times,2.04 times,4.35 times,4.17 times,2.56 times of 0K respectively,all being significantly different.Moreover,the expression of pathogenesis related protein gene such as ?-1,3-glucanase,chitinase gene,thaumatin like protein gene first increased and then decreased during Valsa canker infection,regardless of the callus content level.On the other hand,the fold-change dynamics and peak expression time point differ among K treatments.At all assessed time points,the relative gene expression of ?-1,3-glucanase,chitinase,thaumatin like protein were significantly higher in high potassium treatment than that of low potassium treatment.These results indicate that potassium increases apple Valsa canker resistance through effecting gene expression of phenolic metabolism related genes,pathogenesis related genes and fungal cell wall degrading enzymes.