Transcriptomic approaches to identify defense mechanism in sugarcane to Colletotrichum falcatum causing red rot

Abstract

Sugarcane (Saccharum officinarum L.) is an important commercial crop cultivated worldwide for sugar and in recent years, it also serves as an important resource of biomass, bioethanol and bioplastic. Red rot caused by the fungal pathogen Colletotrichum falcatum Went (Teleomorph: Glomerella tucumanensis [Speg.] Arx and Muller) is a devastating threat for sugarcane cultivation in India. The disease progression is characterized by reddish lesions along the stalk tissue and the lethal effects of the disease leave the sugarcane stalk vulnerable for further sugar recovery. Inspite of the disease’s impact, locating any specific trait in sugarcane is difficult because of its complex polyploidy. To understand and elucidate the initial defense mechanisms in a resistant variety of sugarcane after pathogen inoculation, two suppression subtractive hybridization (SSH) cDNA libraries were constructed with the cDNA of resistant variety at 12 and 36 h. At the end, 141 ESTs specifically upregulated in red rot resistant variety after pathogen challenge was obtained. Annotation of the ESTs revealed that they belong to the categories pertaining to recognition and signal transduction, defense and programmed cell death, oxidative stress, redox maintenance, secondary metabolite biosynthesis, energy and photosynthesis, membrane trafficking and transport, cell/nuclear structure, nucleic acid metabolism, protein metabolism and processing, ubiquitin-26S proteasome system, carbohydrate metabolism, lipid metabolism and unknown categories. Based on the transcripts profile, a hypothetical pathway was generated and it clearly revealed that recognition and signal transduction as the first level of host defense which further activates defense gene expression and causes a global change in sugarcane transcriptome at the molecular level. To further confirm the expression level of defense-associated ESTs, the gene expression of 24 of them were studied in a quantitative real-time PCR (qRT-PCR) at 6, 12, 24, 36, 48 and 72 h after C. falcatum inoculation with 25SrRNA as the endogenous control. In all the ESTs studied, the expression level was found to be high in 72 h after C. falcatum. Further, the sugarcane gene expression at 72 h after C. falcatum was validated by Affymetrix GeneChip SugarCane Genome Array. Genes belonging to the functional categories: signal transduction, defense, energy and metabolism were upregulated. This is the first detailed work on red rot resistance in sugarcane at the molecular level. This study also established that C. falcatum infection causes a global change in sugarcane at the molecular and basal level which finally leads to defense against the fungal pathogen.