Comprehensive Mining And Preliminary Analysis Of Effector Candidates In Penicillium Expansum(Apple Blue Mold Decay)

Blue mold decay,caused by species of Penicillium expansum Link,is one of the most important postharvest diseases of apples worldwide.Besides causing economic losses,P.expansum has also potential public health significance since it produces patulin.This mycotoxin is teratogenic,carcinogenic and immunotoxic,and has potential harm to human and animal health.During the past decade,there have been many reports on controlling P.expansum caused blue mold decay,infection process and pathogenic mechanism of P.expansum are far from well known.With the aim to mine and identify the candidate virulence effectors,differentially expressed genes and secreted proteins of P.expansum grown in the host plant tissues derived medium were analyzed in this work via transcriptomic and proteomic approaches,which might deepen our understanding of the fungi pathogenic mechanisms of P.expansum and provide new strategies for the control of apple blue mold decay.The main results and conclusions are as follows:1.As compared with the fungi grown in Czapek-Dox Medium（control）,a total of 3786 genes at 24 h of incubation and 4840 genes at 72 h with increased expression levels（p≤0.05）were identified in P.expansum grown in apple juice medium（host plant tissue derived medium）by RNA-Seq analysis.Among the up-regulated genes from GO terms,there were 23 groups at the biological level,15 groups at the cellular level and 14 groups at the molecular level.Cells,binding and catalytic activity were the most abundant groups within the GO terms.A total of 22 groups were assigned KOG terms for the up-regulated genes,there were a large number of up-regulated genes involved in lipid transport,metabolism,biosynthesis of secondary metabolites and general functional prediction.Among them,550 genes at 24 h and 727 genes at72 h were identified as genes encoding secreted proteins.A total of 13 differentially expressed genes were randomly selected for qRT-PCR verification,the results show that the gene expression abundance changes were substantially consistent with RNA-Seq analysis.2.HPLC-MS/MS was used to analyse the secreted proteins of P.expansumgrown in apple juice medium,and 166 secreted proteins at 24 h and 272 secreted proteins at 72 h were identified,respectively.In addition to a number of hypothetical proteins,these identified proteins with known function are mainly related to carbohydrates metabolism and secondary metabolism.3.Based on the RNA-Seq and secretome data,PEX1₀69590（Lipase,class 3）,PEX1₀31490（Peptidase aspartic,catalytic）and PEX1₀48740（Peptidase S10,serine carboxypeptidase）were selected as candidate effectors for further characterization.With the presence of specific enzyme inhibitor,blue mod decay caused by P.expansum and lesion diameter were significantly reduced,suggesting these genes might be effector candidates and play a key role in the pathogenicity of P.expansum.