Heterografting has long been used to enhance the chilling tolerance of temperature-sensitive crops, including watermelon, whose mechanism is known to involve bidirectional long-distance mRNA movements. Despite several studies reporting on mobile mRNA (mb-mRNA) profiles in plants, accurate identification of mb-mRNAs is challenging owing to an array of technical problems. Here, we developed a bioinformatical pipeline that took most of the known technical concerns into consideration and is considered to be a universal tool for mb-mRNA detection in heterografts. By applying this pipeline to a commercial watermelon–bottle gourd heterografting system, we detected 130 and 1144 mb-mRNAs upwardly and 167 and 1051 mb-mRNAs downwardly transmitted under normal and chilling-stress conditions, respectively. Quantitative real-time PCR indicated a high accuracy rate (88.2%) of mb-mRNA prediction with our pipeline. We further revealed that the mobility of mRNAs was not associated with their abundance. Functional annotation and classification implied that scions may convey the stress signal to the rootstock, subsequently triggering energy metabolism reprogramming and abscisic acid-mediated stress responses by upward movement of effective mRNAs, ultimately leading to enhanced chilling tolerance. This study provides a universal tool for mb-mRNA detection in plant heterografting systems and novel insights into heterografting advantages under chilling stress.

Grafting technology can improve the yield and quality of crops. In this study, we investigated the effects of grafting on watermelon using transcriptomic and metabolomic analysis. A total of 216 differentially accumulated metabolites (DAMs) were identified between pumpkin-grafted watermelon and self-grafted watermelon. KO (Kyoto Encyclopedia of Genes and Genomes Orthology) analysis revealed that the DAMs were mainly enriched in the flavone and flavonol biosynthesis pathway. In addition, high levels of phloretin and citric acid were found in pumpkin-grafted watermelon, which contributes to fruit quality. Meanwhile, compounds such as olivetol and ferulaldehyde, which confer a bitter taste, were downregulated in pumpkin-grafted watermelon. The transcriptome data indicated that the differentially expressed genes (DEGs) identified in the pulp were enriched in fructose and mannose metabolism, biosynthesis of secondary metabolites, and flavone and flavonol biosynthesis pathways. Moreover, genes related to the microtubule, cell wall, and fiber were highly expressed in the stem of pumpkin-grafted watermelon, suggesting that grafting could change the structure of the stem and improve the quality of watermelon fruit. Our study provides a comprehensive picture of the transcriptional and metabolic profile of watermelon induced by grafting, which furthers our understanding of the molecular mechanisms involved in improving watermelon fruit quality by grafting.

Alternative oxidase (AOX) has been documented to mitigate the oxidative stress caused by abiotic stresses. However, it remains unknown how AOX regulates the antioxidant system and photosynthesis under waterlogging. To address this issue, we used two watermelon (Citrullus lanatus L.) cultivars (waterlogging tolerant cultivar ‘YL’ and sensitive cultivar ‘Zaojia8424’) as materials and the AOX inhibitor salicylhydroxamic acid (SHAM) to investigate the effects of AOX on photosynthesis and reactive oxygen species metabolism under waterlogging. We found that waterlogging decreased leaf photosynthesis and quantum yield of photosynthesis in watermelon, and the waterlogging tolerant cultivar ‘YL’ showed higher expression level of ClaAOX than the sensitive cultivar ‘Zaojia8424’. Net photosynthesis rate was higher in ‘YL’ than ‘Zaojia8424’. Moreover, waterlogging induced photoinhibition in ‘Zaojia8424’ but not in ‘YL’. Meanwhile, waterlogging promoted the accumulation of superoxide and peroxide hydrogen, and triggered oxidative damage. ‘YL’ suffered from less severe oxidative damage due to increased contents of ascorbate, a higher ratio of reduced glutathione (GSH) to oxidized glutathione (GSSG), a higher activity of ascorbate peroxidase (APX) and catalase (CAT), and enhanced levels of CAT and APX expression, relative to ‘Zaojia8424’. However, the alleviation of photosynthesis and oxidative damage, increased content of ascorbate and higher GSH/GSSG ratio were abolished by SHAM. Our results suggested that photosynthetic electronic transfer and glutathione-ascorbate cycle are involved in waterlogging tolerance mediated by the AOX pathway in watermelon.

Cucumber green mottle mosaic virus (CGMMV) belongs to the Tobamovirus genus and is an important quarantine virus of cucurbit crops. Seedborne transmission is one of the principal modes for CGMMV spread, and effective early detection is helpful to prevent the occurrence of the disease. Quantitative real-time reverse-transcription PCR (RT-qPCR) is a sensitive and rapid method for detecting CGMMV nucleic acids, but it cannot distinguish between infectious and noninfectious viruses. In the present work, a propidium monoazide (PMA) assisted RT-qPCR method (PMA-RT-qPCR) was developed to rapidly distinguish infectious and inactive CGMMV. PMA is a photoactive dye that can selectively react with viral RNA released or inside inactive CGMMV virions but not viral RNA inside active virions. The formation of PMA-RNA conjugates prevents PCR amplification, leaving only infectious virions to be amplified. The primer pair cp3-1F/cp3-1R was designed based on the coat protein (cp) gene for specific amplification of CGMMV RNA by RT-qPCR. The detection limit of the RT-qPCR assay was 1.57 × 10² copies·μL⁻¹. PMA at 120 μmol·L⁻¹ was suitable for the selective quantification of infectious CGMMV virions. Under optimal conditions, RT-qPCR detection of heat-inactivated CGMMV resulted in Ct value differences larger than 16 between PMA-treated and non-PMA-treated groups, while Ct differences less than 0.23 were observed in the detection of infectious CGMMV. For naturally contaminated watermelon leaf, fruit and seedlot samples, infectious CGMMV were quantified in 13 out of the 22 samples, with infestation levels of 10²~10⁵ copies·g⁻¹. Application of this assay enabled the selective detection of infectious CGMMV and facilitated the monitoring of the viral pathogen in watermelon seeds and tissues, which could be useful for avoiding the potential risks of primary inoculum sources.

In September 2014, a root rot and vine decline was observed on melon (Cucumis melo L.) growing in several greenhouses with 85% disease incidence in Zhejiang Province in eastern mainland China. This pathogen had been previously separated from melon in a small area test field in Gansu Province in mainland China (He and Bai 2010). The diseased melons exhibited chlorosis of crown leaves and collapse of vines just prior to harvest. The roots of affected plants exhibited necrotic lesions and loss of secondary and tertiary roots. Brown discolorations of vascular tissues was apparent but rarely extended more than a few centimeters in above-ground plant parts. Numerous perithecia in the root cortex were visible to the naked eye several days after plant death. Small pieces of symptomatic roots were surface sterilized for 1 min with 1% sodium hypochlorite, rinsed in sterile water, and then placed on potato dextrose agar (PDA) medium amended with streptomycin sulphate (25 mg/l). A pale brown colony with white aerial mycelia developed after incubation at 25°C in the dark for 7 days. Black, spherical perithecia, 400 μm in diameter, were present 20 days after incubation on PDA. Each ascus contained a single spherical ascospore, 26-30 μm in diameter, typical of Monosporascus cannonballus Pollack & Uecker (Pollack and Uecker 1974). DNA from this isolate was extracted and the internal transcribed spacer (ITS) region was amplified using the primers ITS1 and ITS4 (White et al. 1990), sequenced, and submitted to GenBank (Accession No. KT826570). BLAST indicated that the 564-bp sequence shared 99% identity with M. cannonballus in C. melo (Accession No. AM167937.1). Pathogenicity tests were performed on melon (cv. Yellow 9818) in a temperature-controlled growth chamber. Five 7-day-old mycelial plugs (5 mm in diameter) from colony periphery were inoculated to 100 ml sand-wheat grain medium (3 Vboiled bulge wheat grain: 1 Vsand) and kept at 25°C in the dark for 15 days. The medium overgrown with mycelial was used as inoculum. Five melon seeds were sown in sterilized potting mixture(1 Vsand: 1 Vpeat moss) and inoculum was added to reach a soil concentration of 5%. Five melons grown in noninoculated containers were used as controls. The plants inoculated showed root necrosis and cotyledon wilt after 10 days with an 12-h photoperiod at 30°C/20°C while the controls were symptomless. M. cannonballus was reisolated from symptomatic roots confirming Koch's postulates. This disease has been reported on watermelon and/or melon in 19 countries around the world including Egypt, India, Italy, Japan, Korea, Mexico, Spain, Taiwan and the United States (Cohen et al. 2012; Martyn and Miller 1996). To our knowledge, this is the first report of M. cannonballus on melon in eastern mainland China, extending the known range of the pathogen in China. Due to the wide distribution of melon cultivation and the loose transport policy to plant materials, there is a potential risk for disease spread throughout the country. So grafting of melon onto Cucurbita spp. more tolerance to the pathogen would be an effective disease management. References: Cohen, R., et al. 2012. Hortic. Rev. 39:77-120. He, S. Q., and Bai, B. 2010. Plant Pro. (In Chinese) 36:116-119. Martyn, R. D., and Miller, M. E. 1996. Plant Dis. 80:716-725. Pollack, F. G., and Uecker, F. A. 1974. Mycol. 66:346-349. White, T. J., et al. 1990. Pages 315 in: PCR Protocols: A Guide to Methods and Applications. M. A. Innis et al., eds. Academic Press, San Diego, CA.

Melon (Cucumis melo L.) is a member of the Cucurbitaceae family, an important economical and horticultural crop, which is widely grown in China. In May 2020, fruit rot disease with water-soaked les...