Shi Mingming, Li Huixia, Guo Wei, Luo Ning, Chen Jinghuan, Liu Yonggang, Liu Rui, Mao Zhenchuan
College of Plant Protection, Gansu Agricultural University/Biocontrol Engineering Laboratory of Crop Diseases and Pests of Gansu Province, Lanzhou, Gansu, China.
Institute of Plant Protection, Gansu Academy of Agricultural Sciences, Lanzhou, Gansu, China.
Front Plant Sci. 2025 Aug 15;16:1586118. doi: 10.3389/fpls.2025.1586118. eCollection 2025.
root rot, caused by , has caused severe damage to the industry. Due to the unclear pathogenic mechanisms of on , the effective implementation of control measures has been greatly restricted.
An efficient protoplast preparation and genetic transformation system was established for FO-1, enabling real-time tracking of fungal colonization in . Single-factor experiments were conducted to determine optimal conditions, followed by response surface methodology to further optimize enzymatic parameters. PEG-mediated transformation was performed to generate GFP-tagged strains for infection tracking.
Single-factor experiments identified the optimal conditions as 12-hour-old mycelia treated with 0.7 M NaCl and 20 mg/mL driselase at 28°C and 180 rpm for 4 h. Response surface methodology optimized parameters to 188.24 rpm, 4.51 h, and 27.5°C, yielding 1.44 × 10 CFU/mL protoplasts, representing a 30-fold improvement over single-factor optimization. PEG-mediated transformation produced 11 GFP-tagged strains, with FO-GFP-7 retaining wild-type morphology, growth rate, and pathogenicity. Microscopic observation revealed infection dynamics: conidia aggregated at the rhizome by 2 days post-inoculation (dpi), followed by phloem colonization at 4 dpi and vascular invasion at 6 dpi. Wound inoculation at the rhizome accelerated infection, consistent with field disease patterns linked to soil microfauna-induced injuries.
This study provides a robust platform for investigating F. oxysporum pathogenicity in and offers guidance on protective measures to maintain rhizome integrity during cultivation.
由[病原菌名称未给出]引起的根腐病已对[行业名称未给出]造成严重损害。由于[病原菌名称未给出]对[宿主名称未给出]的致病机制尚不清楚，控制措施的有效实施受到了极大限制。
为[菌株名称未给出]FO - 1建立了高效的原生质体制备和遗传转化系统，能够实时追踪真菌在[宿主名称未给出]中的定殖情况。进行单因素实验以确定最佳条件，随后采用响应面法进一步优化酶解参数。通过聚乙二醇（PEG）介导的转化生成用于感染追踪的绿色荧光蛋白（GFP）标记菌株。
单因素实验确定最佳条件为用0.7 M氯化钠和20 mg/mL裂解酶处理12小时龄的菌丝体，在28°C和180 rpm条件下处理4小时。响应面法将参数优化为188.24 rpm、4.51小时和27.5°C，产生1.44×10 CFU/mL原生质体，比单因素优化提高了30倍。PEG介导的转化产生了11个GFP标记菌株，其中FO - GFP - 7保留了野生型形态、生长速率和致病性。显微镜观察揭示了感染动态：接种后2天（dpi）分生孢子在根茎处聚集，随后在4 dpi时韧皮部定殖，6 dpi时维管束入侵。根茎处伤口接种加速了感染，这与与土壤小型动物引起的损伤相关的田间病害模式一致。
本研究为研究尖孢镰刀菌在[宿主名称未给出]中的致病性提供了一个强大的平台，并为在栽培过程中维持根茎完整性的保护措施提供了指导。
