Identification, Biological characteristics and fungicide selection of the pathogen causing Yuelu melon Fusarium fruit rot in Guangdong

【Objective】The Yuelu melon is a premium melon variety. After being introduced and cultivated in limited quantities in Hainan, Fujian, and Guangdong provinces, it has demonstrated significant economic value. In 2025, a Fusarium fruit rot disease affecting Yuelu melon was discovered in Guangzhou, with an incidence rate ranging from 5% to 8%. The disease altered the aroma, flavor, and quality of the melon, thereby diminishing its nutritional and commercial value. During cultivation, symptoms were observed exclusively within the fruit, with no visible external manifestations, which significantly increased the challenges associated with disease detection and management. The study aimed to identi-fy the pathogen and elucidate its suitable growth conditions, including temperature, pH, light, as well as carbon and nitrogen sources. In addition, the toxicity of various fungicides to the pathogen was evaluated to assess their efficacy, thereby providing insights for effective disease control.【Methods】From April to June 2025, a survey was conducted to assess the incidence and severity of fruit rot in Yuelu melon at the Xiangfeng Special Fruit Industrial Park in Zengcheng District, Guangzhou City. Field symptoms were documented photographically. Diseased tissue samples were collected and, following surface disinfection, they were cultured on potato dextrose agar (PDA) at 26 ℃ for 3 days using the tissue isolation method. Mycelia from the margins of developing colonies were then transferred to PDA plates. Purified strains were obtained through single-spore isolation and stored at 4 ℃ for subsequent use. Pathogenicity was confirmed through two inoculation methods. The first involved mycelia inoculation: The mycelia surfaces of the tested strains were adhered to the puncture site of the melon fruits, with blank agar discs used as controls. The second method employed spore suspension inoculation: 100 µL of spore suspension (5×106 spores · mL- 1 ) was injected into the fruit from the tail end. All treated fruits were incubated at 26 ℃. Once softening or clear symptoms developed, the pathogens were isolated and purified again for verification in accordance with Koch’s postulates. The strains were inoculated on potato dextrose agar (PDA), synthetic nutrient-poor agar (SNA), and carnation leaf agar (CLA) in the dark at 26 ℃ for 10 days. Mycelia, conidiophores, conidia attachment patterns, conidia and chlamydospores were observed under an optical microscope, and the size of conidiophores and conidia were measured (n=80). Genomic DNA was extracted from the tested strains, and the calmodulin (CAM), translation elongation factor 1-alpha (EF1-α), and the second largest subunit of RNA polymerase Ⅱ (RPB2) gene regions were amplified via PCR. The products were sequenced by Shanghai Sangon Biotech Co., Ltd. The resulting sequences were submitted to the NCBI database to obtain accession numbers. Concurrently, BLAST analyses were performed on NCBI to identify and retrieve sequences of closely related species from GenBank. Sequence alignment was conducted using Mega 7.0 software, followed by manual splicing of the aligned datasets. A phylogenetic tree was constructed using the concatenated sequences of CAM, EF1-α, and RPB2 through the neighbor-joining method implemented in MEGA 7.0 to confirm species identification. Furthermore, the tested strains were cultured in an incubator under varying growth conditions, including temperature, pH, light intensity, carbon sources, and nitrogen sources, to determine the optimal culture environment for mycelia growth and spore production. Subsequently, to evaluate the sensitivity of the pathogen to various fungicides, eight commercial fungicides were tested. Mycelia discs were inoculated onto potato dextrose agar (PDA) plates supplemented with each fungicide at designated concentrations and incubated at 26 ℃. Colony diameters were measured after 5 days of incubation.【Results】The external appearance of diseased fruits showed no visible symptoms in the field. However, their tail ends felt soft to the press, and internal examination revealed brown, watersoaked rot, occasionally accompanied by a thin layer of white mold on the diseased areas. A total of 27 Fusarium spp. isolates with similar morphological characteristics were obtained. Pathogenicity tests revealed that artificial inoculation with strains YLG-4, YLG-13, and YLG-21 produced symptoms consistent with those observed under field conditions. On PDA medium, the colonies were initially white and later turned light yellow with cotton-like and abundant aerial mycelia. Conidiogenous cells were phialidic, 2.72 to 49.69 µm (n=80) on SNA medium and 3.63 to 52.65 µm (n=80) on CLA medium. Macroconidia were falcate, slightly curved, tapering apically with 1 to 5 septa, (17.45-46.78) µm×(2.62-4.93) µm (n=80) on SNA medium, (15.84-44.27) µm×(2.71-4.95) µm (n=80) on CLA medium, borne false heads from monophialides and polyphialides. Chlamydospores were spherical on both media. The CAM, EF1- α, and RPB2 gene fragments of strains YLG-4, YLG-13, and YLG-21 were amplified and sequenced, producing fragments approximately 550 bp, 840 bp, and 600 bp in length, respectively. BLAST analysis against the GenBank database showed that the nucleotide sequences of the CAM, EF1-α and RPB2 regions from these strains shared 99.29% to 100% identity with corresponding sequences of Fusarium pernambucanum strains CBS 791.70, CBS 132194, CBS 132894, and CBS 133024. A phylogenetic tree was constructed based on the concatenated sequences of CAM, EF1-α, and RPB2 using the neighborjoining method, which demonstrated that YLG-4, YLG-13, and YLG-21 form a cluster with F. pernambucanum. Integrating morphological characteristics and molecular evidence, strains YLG-4, YLG-13, and YLG-21 were identified as F. pernambucanum. The optimal temperature range for mycelia growth of strain YLG-13 was 24-30 ℃, with the most rapid colony expansion observed at 26-28 ℃ and the highest conidia production occurring at 30 ℃. Mycelia growth was favorable within a pH range of 5- 10, with an optimum at pH 7-9. Light promoted mycelia development. Carbon sources such as glucose and soluble starch, as well as organic nitrogen sources including tryptone, beef extract powder, and yeast extract, were found to enhance both mycelia growth and spore production in YLG-13. Among the eight fungicides tested, all exhibited varying degrees of inhibition on mycelia growth. Fludioxonil showed the strongest inhibitory effect, with an EC50 value of 0.014 7 mg · L- 1 , followed by prochloraz, which had an EC50 value of 0.637 3 mg ·L-1 .【Conclusion】In this study, the causal agent of fruit rot in Yuelu melon was identified as Fusarium pernambucanum through pathogenicity tests, morphological characterization, and multi-locus phylogenetic analysis based on the CAM, EF1-α, and RPB2 gene regions. The pathogen showed adaptation to relatively high temperatures and neutral to alkaline conditions. Fungicide assays revealed that fludioxonil and prochloraz exhibited the strongest antifungal activity, suggesting their potential as effective agents for disease control. These findings offer a theoretical foundation for diagnosing and managing this disease.