2-辛基-3-异噻唑啉酮抗烟草赤星病菌的生物活性及作用机制

doi:10.3864/j.issn.0578-1752.2026.13.007

Abstract

Abstract:

【Objective】The objective of this study is to clarify the inhibitory effect of 2-octyl-3-isothiazolinone, a metabolite from the co-culture of Trichoderma asperellum HG1 and Bacillus subtilis Tpb55, on Alternaria alternata (the pathogen causing tobacco brown spot) and its impact on the physiological metabolism of the pathogen, and to provide a theoretical basis for the application of this natural product in the management of plant fungal diseases.【Method】The antifungal activities of eight secondary metabolites from the co-culture against A. alternata were assessed using the mycelial growth rate method. The toxicity of 2-octyl-3-isothiazolinone was evaluated, and its EC₅₀ value was calculated. Field trials were conducted to determine its control efficacy against tobacco brown spot. Transcriptome sequencing was employed to analyze the gene expression profiles of A. alternata following treatment with 2-octyl-3-isothiazolinone. The impact on energy metabolism was assessed by measuring ATP content and ATPase activity. Propidium iodide (PI) staining and reactive oxygen species (ROS) detection were used to evaluate cell membrane integrity and oxidative stress levels, respectively.【Result】At a concentration of 10 μg·mL^-1, 2-octyl-3-isothiazolinone exhibited 100% inhibition against A. alternata, which was significantly superior to that of the positive control dimethachlon (27.60%). In the field experiment, the control efficacies at concentrations of 100 and 200 μg·mL^-1 were 80.80% and 84.33%, respectively, which were equivalent to that of the positive control (40% dimethachlon diluted 400-fold, 80.15%), and there was no toxic effect on tobacco itself. Further studies showed that 2-octyl-3-isothiazolinone exerted its antifungal effect through a dual mechanism: on the one hand, it significantly inhibited the key enzymes of glycolysis and the tricarboxylic acid cycle, blocking the energy metabolism of the pathogen; on the other hand, the expression levels of genes related to ROS scavenging in the pathogen (AALT_g9032, AALT_g9033) were significantly down-regulated, eventually leading to the massive accumulation of ROS, causing oxidative damage and destroying the integrity and biological function of the cell membrane. Further verification demonstrated that 2-octyl-3-isothiazolinone significantly reduced ATP content and ATPase activity, confirming disruption of energy metabolism. PI staining revealed concentration-dependent damage to cell membrane integrity, while DCFH-DA fluorescence staining indicated substantial intracellular ROS accumulation, triggering oxidative stress and exacerbating cellular damage. These results were consistent with the transcriptomic findings.【Conclusion】As a green pesticide with broad application prospects, 2-octyl-3-isothiazolinone interferes with the normal life activities of A. alternata through the synergistic effect of energy blockage-oxidative damage. This study provides a theoretical basis and application support for its use as a new antifungal agent to control tobacco brown spot.

Key words: 2-octyl-3-isothiazolinone, Alternaria alternata, antifungal activity, transcriptome analysis, mechanism of action

GUO FuQiang, PENG ZhiXin, LI DeRui, LIN MuSen, XU LiFeng, ZHANG ChengSheng. Biological Activity and Mechanism of 2-Octyl-3-isothiazolinone Against Alternaria alternata Causing Tobacco Brown Spot[J].Scientia Agricultura Sinica, 2026, 59(13): 2841-2852.

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References 33

[1]	周素婷, 任兵杨, 李欲轲, 乙引, 唐明. 链格孢致病机制、防治措施与开发利用研究进展[J]. 分子植物育种, 2025, 23(3): 782-791.
	Zhou S T, Ren B Y, Li Y K, Yi Y, Tang M. Research progress on pathogenic mechanism, control measures and utilization of Alternaria[J]. Molecular Plant Breeding, 2025, 23(3): 782-791. (in Chinese)
[2]	彭坤, 王士良, 刘富, 马波, 沈先奎, 吴建昌, 黄保宏, 刘露. 烟草赤星病病原菌生物学特性及致病性研究[J]. 现代农业科技, 2021(2): 78-80.
	Peng K, Wang S L, Liu F, Ma B, Shen X K, Wu J C, Huang B H, Liu L. Biological characteristics and pathogenicity of the pathogen causing tobacco brown spot[J]. Modern Agricultural Science and Technology, 2021(2): 78-80. (in Chinese)
[3]	刘丰银. 禾本科植物链格孢菌多样性研究[D]. 荆州: 长江大学, 2024.
	Liu F Y. Study on diversity of Alternaria from Gramineae plants[D]. Jingzhou: Yangtze University, 2024. (in Chinese)
[4]	刘晓慧, 杨体诗, 李自林, 和镜芳, 李学卫. 四种药剂对烟草赤星病的防治效果[J]. 中南农业科技, 2025, 46(6): 250-252.
	Liu X H, Yang T S, Li Z L, He J F, Li X W. Control effects of four pesticides on tobacco brown spot[J]. South-Central Agricultural Science and Technology, 2025, 46(6): 250-252. (in Chinese)
[5]	程曦. 烟草赤星病的生物防治研究[D]. 杨凌: 西北农林科技大学, 2024.
	Cheng X. Study on biological control of tobacco brown spot[D]. Yangling: Northwest A&F University, 2024. (in Chinese)
[6]	李行善, 王娜娜, 崔传斌, 王平平, 杨明明, 颜霞, 黄丽丽. 烟草赤星病拮抗菌株的筛选鉴定及防治效果研究[J]. 西北农林科技大学学报(自然科学版), 2025, 53(2): 91-101.
	Li X S, Wang N N, Cui C B, Wang P P, Yang M M, Yan X, Huang L L. Screening, identification and control efficacy of antagonistic strains against tobacco brown spot[J]. Journal of Northwest A&F University (Natural Science Edition), 2025, 53(2): 91-101. (in Chinese)
[7]	刘乔斐, 党军政, 朱峰, 王智慧, 丁明石, 耿伟, 李文琪, 陈璐. 农药施用方式对烟草赤星病防治效果及烟叶品质的影响[J]. 农业工程, 2025, 15(10): 69-74.
	Liu Q F, Dang J Z, Zhu F, Wang Z H, Ding M S, Geng W, Li W Q, Chen L. Effects of pesticide application methods on control efficacy of tobacco brown spot and quality of tobacco leaves[J]. Agricultural Engineering, 2025, 15(10): 69-74. (in Chinese)
[8]	周怡青, 肖友利. 活性天然产物靶标蛋白的鉴定[J]. 化学学报, 2018, 76: 177-189.
	Zhou Y Q, Xiao Y L. Target identification of bioactive natural products[J]. Acta Chimica Sinica, 2018, 76: 177-189. (in Chinese)
[9]	Sparks T C, Bryant R J. Impact of natural products on discovery of, and innovation in, crop protection compounds[J]. Pest Management Science, 2022, 78(2): 399-408.
[10]	Maglangit F, Fang Q, Kyeremeh K, Sternberg J M, Ebel R, Deng H. A co-culturing approach enables discovery and biosynthesis of a bioactive indole alkaloid metabolite[J]. Molecules, 2020, 25(2): 256.
[11]	Ebrahim W, El-Neketi M, Lewald L, Orfali R S, Lin W, Rehberg N, Kalscheuer R, Daletos G, Proksch P. Metabolites from the fungal endophyte Aspergillus austroafricanus in axenic culture and in fungal-bacterial mixed cultures[J]. Journal of Natural Products, 2016, 79(4): 914-922.
[12]	Akdemir H, Silva A, Zha J, Zagorevski D V, Koffas M A G. Production of pyranoanthocyanins using Escherichia coli co-cultures[J]. Metabolic Engineering, 2019, 55: 290-298.
[13]	Sun S, Wang H, Yan K, Lou J, Ding J, Snyder S A, Wu L, Xu J. Metabolic interactions in a bacterial co-culture accelerate phenanthrene degradation[J]. Journal of Hazardous Materials, 2021, 403: 123825.
[14]	Niu B, Wang W, Yuan Z, Sederoff R R, Sederoff H, Chiang V L, Borriss R. Microbial interactions within multiple-strain biological control agents impact soil-borne plant disease[J]. Frontiers in Microbiology, 2020, 11: 585404.
[15]	李琦, 杨一洲, 闫合. 异噻唑啉酮及其衍生物的合成及生物活性综述[J]. 应用化工, 2024, 53(7): 1658-1662, 1667.
	Li Q, Yang Y Z, Yan H. Review on synthesis and biological activity of isothiazolinone and its derivatives[J]. Applied Chemical Industry, 2024, 53(7): 1658-1662, 1667. (in Chinese)
[16]	李世昌, 张龚敏, 敖晓娟, 王浩江, 谭卓华, 王飞, 马玫, 雷祖碧, 刘煜, 杨育农. 异噻唑啉酮化合物的合成研究进展[J]. 合成材料老化与应用, 2018, 47(4): 120-123.
	Li S C, Zhang G M, Ao X J, Wang H J, Tan Z H, Wang F, Ma M, Lei Z B, Liu Y, Yang Y N. Research progress of synthesis of isothiazolinone compounds[J]. Synthetic Materials Aging and Application, 2018, 47(4): 120-123. (in Chinese)
[17]	王磊, 武绍峰, 顾学斌. 异噻唑啉酮类杀菌剂的应用研究[J]. 工业微生物, 2015, 45(5): 60-64.
	Wang L, Wu S F, Gu X B. Applied research of isothiazolinone fungicides[J]. Industrial Microorganisms, 2015, 45(5): 60-64. (in Chinese)
[18]	Li Q, Lin W, Zhang X, Wang M, Zheng Y, Wang X, Gao G, Li Y, Zhao D, Zhang C. Transcriptomics integrated with metabolomics reveal the competitive relationship between co-cultured Trichoderma asperellum HG1 and Bacillus subtilis Tpb55[J]. Microbiological Research, 2024, 280: 127598.
[19]	李清钰. 枯草芽孢杆菌与棘孢木霉共培养条件优化及互作机制解析[D]. 北京: 中国农业科学院, 2023.
	Li Q Y. Optimization of co-culture conditions and analysis of interaction mechanism between Bacillus subtilis and Trichoderma asperellum[D]. Beijing: Chinese Academy of Agricultural Sciences, 2023. (in Chinese)
[20]	Yang Q, Wang J, Zhang P, Xie S, Yuan X, Hou X, Yan N, Fang Y, Du Y. In vitro and in vivo antifungal activity and preliminary mechanism of cembratrien-diols against Botrytis cinerea[J]. Industrial Crops and Products, 2020, 154: 112745.
[21]	Sui X, Han X, Wang X, Wan J, Wen M, Zhao D, Zheng Y, Zhang C, Xu C, Wang Y. 2-Nonanol produced by Bacillus velezensis EM-1: A new biocontrol agent against tobacco brown spot[J]. Frontiers in Microbiology, 2025, 30(16): 1582372.
[22]	Jin X, Zhang M, Lu J, Duan X, Chen J, Liu Y, Chang W, Lou H. Hinokitiol chelates intracellular iron to retard fungal growth by disturbing mitochondrial respiration[J]. Journal of Advanced Research, 2021, 34: 65-77.
[23]	Kim H, Xue X. Detection of total reactive oxygen species in adherent cells by 2',7'-dichlorodihydrofluorescein diacetate staining [J]. Journal of Visualized Experiments, 2020, 160: e60682.
[24]	Bader J, Mast-Gerlach E, Popović M K, Bajpai R, Stahl U. Relevance of microbial coculture fermentations in biotechnology[J]. Journal of Applied Microbiology, 2010, 109(2): 371-387.
[25]	Wang M, Ma J, Fan L, Fu K, Yu C, Gao J, Chen J. Biological control of southern corn leaf blight by Trichoderma atroviride SG3403[J]. Biocontrol Science and Technology, 2015, 25(10): 1133-1146.
[26]	冯文. 异噻唑啉酮类化合物的合成及海洋防污性能研究[D]. 海口: 海南师范大学, 2019.
	Feng W. Study on synthesis and marine antifouling properties of isothiazolinone compounds[D]. Haikou: Hainan Normal University, 2019. (in Chinese)
[27]	马春风, 刘光明, 张广照. 环境友好海洋防污体系的研究进展[J]. 大学化学, 2016, 31(2): 1-5.
	Ma C F, Liu G M, Zhang G Z. Progress in the development of environmentally friendly marine anti-fouling systems[J]. University Chemistry, 2016, 31(2): 1-5. (in Chinese)
[28]	Xu Y Q, Huo Z Y, Bai Y, Wang Y, Wang W L, Wang H B, Gao Y J, Wang R N, Li C Y, Liu H, Huang W E, Hu H Y, Wu Y H. Isothiazolinone biocide aggravate disinfection-resistant bacterial fouling on reverse osmosis membranes[J]. Desalination, 2026, 620: 119637.
[29]	Wang S, Chen Z, Ren X, Ding C, Yang X, Tang Y, Ge R S, Chen W, Liu Y. Isothiazolinone biocides as aromatase inhibitors: Hormonal disruption, and in silico docking and SAR/QSAR analysis[J]. Environmental Research, 2025, 285(1): 122339.
[30]	Silva V, Silva C, SoareS P, Garrido E M, Borges F, Garrido J. Isothiazolinone biocides: Chemistry, biological, and toxicity profiles[J]. Molecules, 2020, 25(4): 991.
[31]	毛玉帅, 段亚冰, 周明国. 琥珀酸脱氢酶抑制剂类杀菌剂抗性研究进展[J]. 农药学学报, 2022, 24(5): 937-948.
	Mao Y S, Duan Y B, Zhou M G. Research progress of the resistance to succinate dehydrogenase inhibitors[J]. Chinese Journal of Pesticide Science, 2022, 24(5): 937-948. (in Chinese)
[32]	张灿, 高续恒, 周俞辛, 刘西莉. 线粒体呼吸链复合物III抑制剂作用机制和抗性分子机制研究进展[J]. 农药学学报, 2019, 21(增刊1): 747-758.
	Zhang C, Gao X H, Zhou Y X, Liu X L. Research progress on action mechanism and resistance molecular mechanism of mitochondrial respiratory chain complex III inhibitors[J]. Chinese Journal of Pesticide Science, 2019, 21(Suppl. 1): 747-758. (in Chinese)
[33]	Yaakoub H, Mina S, Calenda A, Bouchara J P, Papon N. Oxidative stress response pathways in fungi[J]. Cellular and Molecular Life Sciences, 2022, 79(6): 333.

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基因编号Gene ID	正向引物Forward primer (5′-3′)	反向引物Reverse primer (5′-3′)
β-Tubulin	TTCAACGAAGCCTCCAACAAC	GTCGCCGGGCTCGAGAT
AALT_g2668	CTCAGCATACTCAGGAGCGA	TAGATGCGGTTCCTGGTGTT
AALT_g583	AACACCGACATCATCACCCT	TAGGCAATGCTCTTGAACGC
AALT_g2895	GTACCTTGACTTGGGTCCGA	GGAGCATCTCGATCCTGGAA
AALT_g995	AACCTCCATCATCTGCACCA	GCGAACGACATTGAGACCAA
AALT_g5819	TGTTGCACCAATGGGAAAGG	AGACATGCGAATGGTGAAGC
AALT_g9032	ACATCACTGGCGCATCTTTC	GCAACAGTGGAGAATCGCAT

次级代谢物 Secondary metabolite	抑菌率 Inhibition rate (%)	次级代谢物 Secondary metabolite	抑菌率 Inhibition rate (%)
壬二酸Azelaic acid	13.26±0.32f	亚油酸Linoleic acid	0g
香芹酮Carvone	14.24±0.46f	棕榈油酸Palmitoleic acid	0g
10-十一碳烯酸10-Undecenoic acid	35.11±0.54d	巴龙霉素 Paromomycin	75.61±0.57b
苹果酸Malic acid	46.82±0.88c	2-辛基-3-异噻唑啉酮2-Octyl-3-isothiazolinone	100.00a
菌核净Dimethachlon	27.60±0.71e

处理Treatment	病情指数Disease index	相对防效Relative control efficacy (%)
CK (0.5% DMSO）	25.84a	—
100 µg·mL^-1 2-辛基-3-异噻唑啉酮2-Octyl-3-isothiazolinone	4.96b	80.80a
200 µg·mL^-1 2-辛基-3-异噻唑啉酮2-Octyl-3-isothiazolinone	4.05c	84.33a
40%菌核净400倍液40% Dimethachlon diluted 400 times	5.13b	80.15a

Biological Activity and Mechanism of 2-Octyl-3-isothiazolinone Against Alternaria alternata Causing Tobacco Brown Spot

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