Full-length transcriptome and RNA-Seq analyses reveal the resistance mechanism of sesame in response to Corynespora cassiicola.

BACKGROUND: Corynespora leaf spot is a common leaf disease occurring in sesame, and the disease causes leaf yellowing and even shedding, which affects the growth quality of sesame. At present, the mechanism of sesame resistance to this disease is still unclear. Understanding the resistance mechanism of sesame to Corynespora leaf spot is highly important for the control of infection. In this study, the leaves of the sesame resistant variety (R) and the sesame susceptible variety (S) were collected at 0-48 hpi for transcriptome sequencing, and used a combined third-generation long-read and next-generation short-read technology approach to identify some key genes and main pathways related to resistance. RESULTS: The gene expression levels of the two sesame varieties were significantly different at 0, 6, 12, 24, 36 and 48 hpi, indicating that the up-regulation of differentially expressed genes in the R might enhanced the resistance. Moreover, combined with the phenotypic observations of sesame leaves inoculated at different time points, we found that 12 hpi was the key time point leading to the resistance difference between the two sesame varieties at the molecular level. The WGCNA identified two modules significantly associated with disease resistance, and screened out 10 key genes that were highly expressed in R but low expressed in S, which belonged to transcription factors (WRKY, AP2/ERF-ERF, and NAC types) and protein kinases (RLK-Pelle_DLSV, RLK-Pelle_SD-2b, and RLK-Pelle_WAK types). These genes could be the key response factors in the response of sesame to infection by Corynespora cassiicola. GO and KEGG enrichment analysis showed that specific modules could be enriched, which manifested as enrichment in biologically important pathways, such as plant signalling hormone transduction, plant-pathogen interaction, carbon metabolism, phenylpropanoid biosynthesis, glutathione metabolism, MAPK and other stress-related pathways. CONCLUSIONS: This study provides an important resource of genes contributing to disease resistance and will deepen our understanding of the regulation of disease resistance, paving the way for further molecular breeding of sesame.

First Report of Root Rot Caused by Pythium myriotylum on Sesame in China.

Sesame (Sesamum indicum L.) is a very important oilseed crop and cultivated on 11.7 million hectares, producing 6.02 million tons of seeds with an average seed yield of 512 kg ha-1 in the world (Yadav et al. 2022). In June of 2021, diseased roots were observed on sesame in the villages of Mada and Hanba, Xiangcheng city (114.88°N, 33.13°E), Henan province, China. The diseased plants appeared stunted and wilted at the seedling stage. Approximately 7.1% to 17.7% of plants were affected in two fields, 0.6 ha in total, and disease severity in each affected plant ranged from 50% to 80%. Twenty-four disease plants were collected to confirm the pathogen. The diseased roots were cut into small fragments (2 to 5 mm long), surface sterilized with 75% ethanol (for 1 min), and 10% sodium hypochlorite (for 1 min) and then rinsed in sterilized water three times (1 min each rinse). The fragments were blotted dry and transferred to a potato dextrose agar (PDA) medium (potato 200 g/L, glucose 20 g/L, agar 18 g/L) amended with streptomycin (50 µg/mL). After incubation at 28°C for 24 h, white mycelium grew out from plant fragments. Then, a total of seven morphologically similar strains were transferred onto fresh V8 agar by hyphal tip transfer (Rollins 2003). By light microscope observations, the sporangia were filamentous or digitated, and undifferentiated or inflated lobulate. The oospores were mostly aplerotic, globose or subglobose in shape, and 20.4 to 42.6 µm in diameter (n = 90, n: Total number of oospores measured). Furthermore, antheridia were bulbous-like or clavate-like and were observed attached to the surface of the oospores. The zoospores were abundant and ranged from 8.5 to 14.2 µm in diameter. The morphology characteristics of all strains were consistent with those of Pythium myriotylum (Watanabe et al. 2007). Genomic DNA was extracted from the representative strain 20210628 using the CTAB method (Wangsomboondee et al. 2002). The complete internal transcribed spacer (ITS) and cytochrome oxidase subunit I gene (COI, COX1) can be valid and useful barcodes for accurate identification of many oomycetes (Robideau et al. 2011). The ITS and COI were amplified with the primers ITS1/ITS4 (Riit et al. 2016) and primers OomCox-Levup/OomCox-Levlo (Robideau et al. 2011), respectively. The nucleotide sequences obtained were deposited in the GenBank database under the accession numbers OM230138.2 (ITS) and ON500503.1 (COI). GenBank BLAST search identified the sequences as P. myriotylum ITS and COI sequences (e.g., HQ237488.1 and MK510848.1, respectively) with 100% coverage and 100% identity. To evaluate the pathogenicity, sesame seeds (cultivar: Jinzhi No.3) were planted in 12-cm-diameter plastic pots containing a mixture of sterilized soil, vermiculite and peat mossat a ratio of 3:1:1. Oospores were collected following the procedure of Raftoyannis et al. (2006) with minor modifications. Three-leaf stage sesame roots were soaked with 5 mL of oospore suspension at 1 × 106/mL of the 20210628 strain, and the control plants were inoculated with sterilized water. All plants were maintained in a greenhouse (28±2°C, > 80% R. H.). The experiment was repeated twice with three replications. The plants inoculated with P. myriotylum showed the water soak symptom on the stem base 7 days after inoculation, while control plants were symptomless. Three weeks after inoculation, the plants showed root tissue necrosis, root rot, and dwarfing symptoms that were similar to those observed on sesame plants in the field, while control plants remained healthy. P. myriotylum was re-isolated from the inoculated plants and the morphology was the same as the original strain 20210628. These results suggest that P. myriotylum is the causal agent of sesame root rot. Previous studies have revealed that P. myriotylum can cause root rot in peanuts (Yu et al. 2019), chili pepper (Hyder et al. 2018), green bean (Serrano et al. 2008) and aerial blight of tomato (Roberts et al. 1999). To the best of our knowledge, this is the first report of P. myriotylum causing root rot on sesame. This pathogen can infect plant roots and develop rapidly if no effective control measures are implemented. Once the disease breaks out in a large area, the yield of sesame will be seriously threatened. The results provide important implications for the prevention and management of this disease.