Unraveling the Molecular Interaction between Pseudomonas syringae pv. actinidiae (Psa) and the Kiwifruit Plant through RNAseq Approach

Bacterial canker, caused by Pseudomonas syringae pv. actinidiae (Psa), is the most important disease of kiwifruit, threatening kiwifruit production around the world. However, little is known about the molecular interaction between Psa and kiwifruit plants. To elucidate early molecular plant response during the different phases of the infection, gene expression in healthy or inoculated plants of Actinidia chinensis was analyzed 3, 24 and 48 h after experimental inoculation. In addition, to understand the molecular mechanism underlying the increased plant resistance observed after the exogenous application of the systemic acquired resistance (SAR) inducer acibenzolar-S-methyl (ASM), gene expression was analyzed in healthy or inoculated ASM pre-treated plants. Transcriptome analysis of A. chinensis, was performed using an Illumina GAIIx sequencer, generating 221 million 75 bp long paired end reads. The raw reads were de novo assembled into 63,943 contigs by Trinity software with an average size of 1,078 bp and N50 of 1,614 bp. Annotation of the de novo transcript contigs was done using Blast2GO software, BLASTX was performed against the NCBI non-redundant protein database with an E-value cut-off of 1e(-5). Mapped records were also aligned to protein databases such as Swiss-Prot, KEGG and COG, in order to retrieve proteins with the highest sequence similarity, to give a putative functional annotation. Finally, InterProScan was used to identify additional gene ontology (GO) annotations. Reads were mapped to the contigs using the CLC software to identify differentially expressed genes (DEGs), statistical analysis was done using the method described in the R package DESeq. A preliminary analysis of DEGs was conducted on sequences obtained 24 h after infection. A total of 349 DEGs were identified in response to infection in non-ASM pre-treated plants. When ASM pre-treated plants were considered, 637 DEGs were identified, 213 of them were in common with the 349 DEGs modulated by infection in not ASM pre-treated plants. The ASM treatment by itself was able to modulated 974 DEGs, 70 of them were further modulated by infection in ASM pre-treated plants. These results suggest that the molecular response to Psa is strongly enhanced in ASM pre-treated plants.