Suppression of Xanthomonas oryzae pv. oryzae biofilm formation by Acacia mangium methanol leaf extract

Abstract Xanthomonas oryzae pv. oryzae (Xoo), a pathogen responsible for rice bacterial leaf blight, produces biofilm to protect viable Xoo cells from antimicrobial agents. A study was conducted to determine the potency of Acacia mangium methanol (AMMH) leaf extract as a Xoo biofilm inhibitor. Four concentrations (3.13, 6.25, 9.38, and 12.5 mg/mL) of AMMH leaf extract were tested for their ability to inhibit Xoo biofilm formation on a 96-well microtiter plate. The results showed that the negative controls had the highest O.D. values from other treatments, indicating the intense formation of biofilm. This was followed by the positive control (Streptomycin sulfate, 0.2 mg/mL) and AMMH leaf extract at concentration 3.13 mg/mL, which showed no significant differences in their O.D. values (1.96 and 1.57, respectively). All other treatments at concentrations of 6.25, 9.38, and 12.5 mg/mL showed no significant differences in their O.D. values (0.91, 0.79, and 0.53, respectively). For inhibition percentages, treatment with concentration 12.5 mg/mL gave the highest result (81.25%) followed by treatment at concentrations 6.25 and 9.38 mg/mL that showed no significant differences in their inhibition percentage (67.75% and 72.23%, respectively). Concentration 3.13 mg/mL resulted in 44.49% of biofilm inhibition and the positive control resulted in 30.75% of biofilm inhibition. Confocal laser scanning microscopy (CLSM) analysis of Xoo biofilm inhibition and breakdown showed the presence of non-viable Xoo cells and changes in aggregation size due to increase in AMMH leaf extract concentration. Control slides showed the absence of Xoo dead cells.

摘要 稻黄单胞菌稻致病变种（Xanthomonas oryzae pv. oryzae, Xoo）是引发水稻细菌性叶枯病的病原菌，可产生生物膜以保护存活的Xoo细胞免受抗菌剂侵害。本研究旨在评估马占相思甲醇叶提取物（Acacia mangium methanol leaf extract, AMMH）作为Xoo生物膜抑制剂的效能。本实验选用4种浓度（3.13、6.25、9.38及12.5 mg/mL）的AMMH叶提取物，检测其在96孔微量培养板上抑制Xoo生物膜形成的能力。结果显示，阴性对照组的光密度（Optical Density, O.D.）值在所有处理组中最高，表明生物膜形成旺盛。紧随其后的是阳性对照组（硫酸链霉素，0.2 mg/mL）与3.13 mg/mL浓度的AMMH叶提取物组，二者的光密度值无显著差异（分别为1.96与1.57）。其余浓度为6.25、9.38及12.5 mg/mL的处理组，其光密度值均无显著差异（分别为0.91、0.79与0.53）。就生物膜抑制率而言，12.5 mg/mL浓度处理组的抑制率最高（81.25%），其次为6.25与9.38 mg/mL浓度处理组，二者的抑制率无显著差异（分别为67.75%与72.23%）。3.13 mg/mL浓度处理组的生物膜抑制率为44.49%，阳性对照组的抑制率为30.75%。共聚焦激光扫描显微镜（Confocal laser scanning microscopy, CLSM）对Xoo生物膜抑制与消解效果的分析显示，随着AMMH叶提取物浓度升高，样本中出现了失活的Xoo细胞，且菌体聚集尺寸发生改变。对照组玻片则未检测到死亡的Xoo细胞。