殺菌劑在木材中的擴散性及對褐根病菌防治效果評估

Abstract

褐根病 (brown root rot disease) 為熱帶及亞熱帶地區樹木的重要真菌性病害，對臺灣都市行道樹及綠化樹種皆構成威脅。褐根病菌 (Phellinus noxius, syn. Pyrrhoderma noxium) 主要侵染根部與莖基部組織，造成樹木生長衰弱，枝葉黃化及小葉化，並使根系喪失水分與養分運輸及支持功能，增加樹木倒伏及死亡風險。現行藥劑防治篩選方式，多以含藥培養基進行，能否準確反映複雜木材環境下的殺菌效果值得探討。由於樹木受褐根病菌侵染後失去輸導功能，而心材本身亦不具運輸能力，因此藥劑在木材內的擴散性，決定其能否有效到達罹病處並發揮藥效。本研究使用邊長 5 毫米之正榕 (Ficus macrocarpa) 立方體木塊開發簡便之含藥木塊測定法，並選擇分屬於 5 種作用機制 [Fungicide Resistance Action Committee (FRAC) 3、7、9、11及12] 之 9 種殺菌劑，評估在正榕木塊與MA+4培養基中的抑菌效力。殺菌劑預防性處理木塊的試驗中，得克利 (tebuconazole) 對褐根病菌之抑制效果最優異 (EC₅₀ = 0.131 mg kg-1)，治療性處理則以依普座 (epoxiconazole) 最佳 (EC₅₀ = 0.256 mg kg-1)。培養基與木塊測定法之 EC₅₀ 呈顯著正相關 (r = 0.77–0.79)，木塊對培養基之 EC₅₀ 比值介於 3.6 至 197.5，顯示木材基質顯著降低殺菌劑效力。為釐清殺菌劑在木材內的擴散性，將殺菌劑 (1.2 mL 75 g L-1 有效成分) 注射於正榕段木，7天後以電鑽於注射孔上方、下方及右側 2 cm、4 cm 處分別鑽取木屑樣本，透過液相層析串聯質譜儀 (liquid chromatography-tandem mass spectrometry, LC/MS-MS) 定量藥劑濃度後，發現在注射點上方及下方 2 cm 處，百克敏 (pyraclostrobin)、得克利、依普座等藥劑平均濃度達68.1–132.0 mg kg-1，而右方 2 cm 處僅 0.3–1.8 mg kg-1，顯示殺菌劑主要沿縱向傳輸，橫向擴散極為有限。滅普寧 (mepronil)、賽普洛 (cyprodinil) 及護汰寧 (fludioxonil) 擴散性極差，不排除與其劑型為水懸劑或水分散性粒劑相關。進一步以 4 種殺菌劑在溫室進行垂榕 (Ficus benjamina) 苗防治試驗，分別以土壤澆灌與莖部注射方式施用殺菌劑，並以褐根病菌麥粒接種源進行枝條創傷接種。結果發現，得克利土壤澆灌組之葉片黃化萎凋率 (15.6%) 在接種後 7 天比對照組 (40.3%) 顯著降低，但效果未能持續至 14 天。整體而言，所有處理組皆未呈現顯著防治效果，顯示未來需建立更適合褐根病藥劑篩選之植體測定系統。本研究突破傳統培養基篩選之侷限，釐清殺菌劑於木材內的擴散特性與基質效應，可作為開發木材腐朽病害新穎藥劑防治配方或施用技術之依據。

Brown root rot disease (BRRD) is an important fungal disease of trees in tropical and subtropical regions, posing a serious threat to urban street trees and ornamental trees in Taiwan. Phellinus noxius (syn. Pyrrhoderma noxium), the causal agent of BRRD, primarily infects root and stem base tissues, causing growth decline, foliar chlorosis, small leaves, and loss of water and nutrient transport and support functions in root systems, thereby increasing the risk of tree toppling and mortality. Current fungicide screening methods predominantly employ fungicide-amended culture media; however, whether such methods accurately reflect fungicidal efficacy in the complex wood environment remains questionable. Since trees lose their transport function upon infection by P. noxius, and heartwood inherently lacks transport capacity, the diffusion of fungicides within wood determines whether they can effectively reach infected sites and exert their efficacy. In this study, a simple fungicide-amended wood block assay was developed using 5-mm cubes of Ficus macrocarpa and MA+4 selective medium. Nine fungicides, representing five modes of action [Fungicide Resistance Action Committee (FRAC) codes 3, 7, 9, 11, and 12], were selected to evaluate their inhibitory efficacy on this method and culture medium assay. For the wood block assay, tebuconazole exhibited the most superior inhibitory effect against P. noxius (EC₅₀ = 0.131 mg kg⁻¹) in preventive treatment trials, while epoxiconazole performed best in curative treatment (EC₅₀ = 0.256 mg kg⁻¹). The EC₅₀ values from culture medium and wood block assays showed significant positive correlation (r = 0.77–0.79), with wood-to-medium EC₅₀ ratios ranging from 3.6 to 197.5, suggesting that the wood matrix significantly attenuates fungicide efficacy. To elucidate fungicide diffusion efficiency within wood, fungicides (1.2 mL at 75 g L⁻¹ active ingredient) were injected into F. macrocarpa wood logs, and wood powder samples were collected by drilling at 2 cm and 4 cm above, below, and to the right of the injection point, respectively, after 7 days. Fungicide concentrations were quantified using liquid chromatography-tandem mass spectrometry (LC-MS/MS). At 2 cm above and below the injection point, average concentrations of pyraclostrobin, tebuconazole, and epoxiconazole reached 68.1–132.0 mg kg⁻¹, whereas concentrations at 2 cm to the right were only 0.3–1.8 mg kg⁻¹, indicating that fungicides primarily transport longitudinally with extremely limited lateral diffusion. Mepronil, cyprodinil, and fludioxonil showed very poor diffusion, possibly related to their formulations as suspension concentrates or water-dispersible granules. Greenhouse seedling trials were conducted using Ficus benjamina. Branches were wound-inoculated with P. noxius-colonized grain inoculum, and four fungicides were applied via soil drench or trunk injection. The leaf yellowing and wilting rate of the tebuconazole soil drench treatment (15.6%) was significantly lower than the control (40.3%) at 7 days post-inoculation (dpi); however, this effect did not persist to day 14. Overall, none of the treatments exhibited significant disease control efficacy, indicating the need to establish a more suitable in planta evaluation system for BRRD fungicide screening. This study addresses the limitations of conventional culture-medium-based screening by elucidating fungicide diffusion characteristics and matrix effects in wood, thereby providing a basis for the development of novel formulations or application strategies for the chemical control of wood decay diseases.