Identification, biological characteristics and fungicide screening of the pathogen causing sea buckthorn stem wilt

【Objective】From 2020 to 2024, a highly destructive disease was investigated in sea buckthorn plantations in Chaoyang City, Liaoning Province, China. The affected plants exhibited severe symptoms, including massive leaf fall, branch dieback, and eventual death. Given the significant impact of this disease on sea buckthorn production, this study was designed to identify the pathogen species causing stem wilt, determine its biological characteristics, and screen for effective and safe fungicides. The findings of this study are expected to contribute to the development of sustainable and effective management strategies for this disease.【Methods】Typical symptomatic sea buckthorn branches and roots were collected from affected plantations. The pathogen was isolated using the tissue isolation method, and the pathogenicity of the purified strains was verified by the back-joining method according to Koch’s rules. After single-spore purification, the pathogenic strains were cultured on SNA medium for 10 days. The colony morphology and color were meticulously observed and recorded. The size and morphological characteristics of conidia were measured using a light microscope to characterize the pathogen’s morphology. Molecular identification was carried out using PCR amplification with fungal universal primers ITS1/ITS4 and Fusarium-specific primers RPB2 and EF-1α. The sequencing results were analyzed using BLAST on NCBI, and phylogenetic trees were constructed using MEGA 7.0 software to confirm the identity of the pathogen. The biological characteristics of the pathogen were studied by evaluating its growth on different media, carbon sources, nitrogen sources, temperatures, light conditions, and pH levels. This comprehensive assessment aimed to understand the optimal conditions for the pathogen’s growth and sporulation. In vitro efficacy of 10 fungicides was determined using the mycelial growth rate method. Based on the results, five effective fungicides were selected for pot experiments to evaluate their control efficacy under controlled conditions. This step aimed to identify effective fungicides that could potentially be used in field applications to manage the disease effectively.【Results】 The pathogen isolation and preliminary morphological identification revealed that the strain X28 was highly pathogenic. Inoculation experiments showed that symptoms closely resembled those observed in the field. Morphological observations of the strain X28 on SNA medium revealed white colonies with abundant aerial mycelium and yellow pigment on the reverse. Macroconidia were slightly curved with three distinct septa, while microconidia were colorless, oval or ellipsoidal, and with 0–2 septa. Molecular identification confirmed the pathogen as Fusarium proliferatum. The results of biological characteristic showed that the optimal conditions for mycelial growth were PDA medium with sucrose as the carbon source and peptone as the nitrogen source, at 28 ℃, pH 9.0, and under alternating light and dark conditions (12 h light/12 h darkness). The optimal carbon source for sporulation of the pathogen was fructose, and the optimal nitrogen sources were yeast extract and peptone. Light treatment and pH had minimal effects on spore production of the pathogen. The toxicity of the ten fungicides testing on F. proliferatum showed that these fungicides exhibited inhibitory effects on the pathogen, and 98% kresoximmethyl showed the strongest inhibitory effect (EC50 = 0.070 9 mg ·L- 1 ), followed by 97% tebuconazole (EC50 = 0.261 9 mg·L-1 ). Pot experiments demonstrated that the incidence rate of inoculated plants treated with five fungicides was significantly lower than the control. The lowest incidence rates were observed with 80% tebuconazole WG and 50% kresoxim- methyl WG, at 25% and 20% , respectively. These treatments also showed the lowest disease indices (21.7% and 18.3%) and the highest control efficacies (71.6% and 76%). The findings suggest that both kresoxim-methyl and tebuconazole are highly effective in controlling F. proliferatum.【Conclusion】This study identified F. proliferatum as the pathogen causing sea buckthorn wilt in Chaoyang City, Liaoning Province. The 50% kresoxim-methyl WG demonstrated the best control efficacy among the tested fungicides. The detailed investigation of the pathogen’s biological characteristics and the screening of effective fungicides provide practical support for the field control of sea buckthorn wilt. Future research should focus on large-scale field trials to validate the long- term efficacy and safety of these fungicides. Additionally, exploring integrated management strategies, including cultural practices and biological control agents, could further enhance the control of sea buckthorn wilt. This comprehensive approach will be important for the sustainable management of sea buckthorn plantations and the mitigation of economic losses caused by this destructive disease.