Yellow Leaf Disease Resistance and Melanaphis sacchari Preference in Commercial Sugarcane Cultivars.

Sugarcane yellow leaf disease (YLD) caused by sugarcane yellow leaf virus (ScYLV) is a major threat for the sugarcane industry worldwide, and the aphid Melanaphis sacchari is its main vector. Breeding programs in Brazil have provided cultivars with intermediate resistance to ScYLV, whereas the incidence of ScYLV has been underestimated partly due to the complexity of YLD symptom expression and identification. Here, we evaluated YLD symptoms in a field assay using eight sugarcane genotypes comprising six well-established commercial high-sucrose cultivars, one biomass yield cultivar, and a susceptible reference under greenhouse conditions, along with estimation of virus titer through RT-qPCR from leaf samples. Additionally, a free-choice bioassay was used to determine the number of aphids feeding on the SCYLV-infected cultivars. Most of the cultivars showed some degree of resistance to YLD, while also revealing positive RT-qPCR results for ScYLV and virus titers with non-significant correlation with YLD severity. The cultivars IACSP01-5503 and IACBIO-266 were similar in terms of aphid preference and ScYLV resistance traits, whereas the least preferred cultivar by M. sacchari, IACSP96-7569, showed intermediate symptoms but similar virus titer to the susceptible reference, SP71-6163. We conclude that current genetic resistance incorporated into sugarcane commercial cultivars does not effectively prevent the spread of ScYLV by its main aphid vector.

Sugarcane mosaic virus mediated changes in cytosine methylation pattern and differentially transcribed fragments in resistance-contrasting sugarcane genotypes.

Sugarcane mosaic virus (SCMV) is the causal agent of sugarcane mosaic disease (SMD) in Brazil; it is mainly controlled by using resistant cultivars. Studies on the changes in sugarcane transcriptome provided the first insights about the molecular basis underlying the genetic resistance to SMD; nonetheless, epigenetic modifications such as cytosine methylation is also informative, considering its roles in gene expression regulation. In our previous study, differentially transcribed fragments (DTFs) were obtained using cDNA-amplified fragment length polymorphism by comparing mock- and SCMV-inoculated plants from two sugarcane cultivars with contrasting responses to SMD. In this study, the identification of unexplored DTFs was continued while the same leaf samples were used to evaluate SCMV-mediated changes in the cytosine methylation pattern by using methylation-sensitive amplification polymorphism. This analysis revealed minor changes in cytosine methylation in response to SCMV infection, but distinct changes between the cultivars with contrasting responses to SMD, with higher hypomethylation events 24 and 72 h post-inoculation in the resistant cultivar. The differentially methylated fragments (DMFs) aligned with transcripts, putative promoters, and genomic regions, with a preponderant distribution within CpG islands. The transcripts found were associated with plant immunity and other stress responses, epigenetic changes, and transposable elements. The DTFs aligned with transcripts assigned to stress responses, epigenetic changes, photosynthesis, lipid transport, and oxidoreductases, in which the transcriptional start site is located in proximity with CpG islands and tandem repeats. Real-time quantitative polymerase chain reaction results revealed significant upregulation in the resistant cultivar of aspartyl protease and VQ protein, respectively, selected from DMF and DTF alignments, suggesting their roles in genetic resistance to SMD and supporting the influence of cytosine methylation in gene expression. Thus, we identified new candidate genes for further validation and showed that the changes in cytosine methylation may regulate important mechanisms underlying the genetic resistance to SMD.

Reference genes for gene expression studies targeting sugarcane infected with Sugarcane mosaic virus (SCMV).

OBJECTIVE: The selection of reference genes in sugarcane under Sugarcane mosaic virus (SCMV) infection has not been reported and is indispensable to get reliable reverse transcription quantitative PCR (RT-qPCR) results for validation of transcriptome analysis. In this regard, seven potential reference genes were tested by RT-qPCR and ranked according to their stability using BestKeeper, NormFinder and GeNorm algorithms, and RefFinder WEB-based software in an experiment performed with samples from two sugarcane cultivars contrasting for SCMV resistance, when mechanically inoculated with a severe SCMV strain and using mock inoculated plant controls. RESULTS: The genes Uridylate kinase (UK) and Ubiquitin-conjugating enzyme 18 (UBC18) were the most stable according to GeNorm algorithm and the Pearson correlation coefficients with the BestKeeper index. On the other hand, ribosomal protein L35-4 (RPL1), Actin (ACT) and Ubiquitin1 (UBQ1) were the least stable genes for all algorithms tested.