Pathogen identification and fungicide screening of melon charcoal rot disease

Abstract:【Objective】Cucumis melo is one of the important economic crops of Cucurbitaceae in China. In recent years, melon root diseases are becoming more and more serious because of promotion of protected melon cultivation mode and continuous cropping. In 2018, 20 hectares of Jinxiangyu melons were planted in Ledong, Sanya, Hainan province, but the whole plants wilted suddenly and died about 20 days before the maturity period. The disease mortality rate was about 90%, the melon production losses were serious and the quality lost commodity, which caused heavy losses. The melon roots and stem bases showed brown rot and a large number of small black particles were gathered. In the study, we analyzed and clarified the cause of sudden wilt and death of melon in greenhouse and screened suitable chemical agents, and the aim was to screen out fungicides suitable for the prevention and treatment of melon charcoal rot and lay the foundation for effective prevention and control of the disease.【Methods】Melon decaying roots were collected for isolation of the pathogen by using the traditional tissue  separation method. A small piece of tissue about 4 mm × 4 mm was cut from the junction of diseased and healthy roots, sterilized and dried, and then cultured and purified on PDA, before the purified patho-gens were stored on PDA at 4 ℃. To confirm the pathogenicity, the stems and basal stem of the melon variety Jinxiangyu and pumpkin rootstocks ZGNG-12 were inoculated by stem cake inoculation meth-od, and the pathogen was reisolated from the inoculated sites. The pathogen was identified through morphological observation and molecular biology methods, and the indoor toxicity of 8 kinds of fungi- cides on the mycelium of the melon charcoal rot was determined by the growth rate method.【Results】Several isolates were obtained by tissue isolation and purification. Morphological observation revealed that the hyphae of the pathogen grew rapidly, growing radially around the surface of the medium. The colony diameter could reach 80 mm at 27 ℃ for 3 days in the dark on PDA, and it could grow up to 90 mm for 4 days. The primary hyphae were white at 24 h, the color of the colony changed from white to off-white at 48 h, and a large number of oval or round micro sclerotia appeared. The micro sclerotia aggre-gated and turned black after 72 h, and the size of the micro sclerotia was (80.3-120) μm×(65.6-120) μm. There were no pycnidia and pycnidiospores on PDA. The fragment length amplified by the fungal primers ITS1/ITS4 was 583 bp, and BLAST analysis showed that the sequence was 99% identity with the Macrophomina phaseolina (GenBank accession: FJ827625.1). The fragment length amplified by the specific primers MPKFI/MpKRI was 350 bp, and the sequence was 99% identity with charcoal rot (GenBank accession: FJ960441). Phylogenetic tree analysis based on fungal ITS and specific sequences of M. phaseolina genus showed that HN-melon isolates and M. phaseolina clustered together. Considering morphology, pathogenicity identification and disease characteristics of field plant, the results showed that the pathogen causing sudden wilt of melon in Hainan greenhouse was M. phaseolina. It was the first report on melon charcoal rot in Hainan province. The virulence test of 8 fungicides to M. phaseolina differed greatly, the best inhibitory effect was 50% prochloraz manganese salt with the EC50 at 0.684 9 mg·L^-1, and the second was pyrazomethrin 250 g· L^-1 with an EC50 at 1.612 4 mg· L^-1.【Conclusion】In this study, M. phaseolina caused melon acute wilting and death in Hainan province was identified for the first time on melon in Hainan province. 50% prochloraz manganese salt and pyraclos-trobin 250 g·L^-1 could inhibit the growth of mycelium, and the results laid the foundation for the preven-tion and control of the disease.