Pathogen Identification and Its Compound Formulations’ Control Efficacy against Solidago canadensis

doi:10.13560/j.cnki.biotech.bull.1985.2025-1218

Abstract

Abstract:

Objective This study aimed to isolate pathogenic fungi with high efficacy against Canadian goldenrod (Solidago canadensis L.), develop a combined formulation integrating the fungal agent with a chemical herbicide, and evaluate its control efficacy in pot experiments. Methods Pathogenic fungi from diseased S. canadensis leaves and roots were isolated, screened, and identified via morphological and molecular methods. Furthermore, chemical herbicides compatible with the fungi were screened, and a compound wettable powder was developed. The control efficacy of this formulation was then tested in pot experiments. Results The isolated pathogen was identified as Diaporthe sojae strain SC02. The compound wettable powder formulation consisted of the following components: 3% Tween 80, 1% sodium lignosulfonate, 0.9% fulvic acid, 0.5 μL/mL 20% fluroxypyr, 7% kaolin, and 40% SC02 strain spores (1×10⁹ CFU/mL). Pot experiment results showed that at concentrations ranging from 0.83 to 1.25 g/L, the compound formulation achieved a plant control efficacy of 70%–90% and a fresh weight control efficacy of 63%–73% against S. canadensis, with no adverse effects observed on surrounding plants. Conclusion The wettable powder formulated with SC02 meets national quality standards, exhibits nearly 90% control efficacy against S. canadensis plants, and has a low recurrence rate, demonstrating satisfactory weed control performance.

Key words: Solidago canadensis, Diaporthe sojae, combined wettable powder, biological control

JIN Wen-tao, LI Ying-xin, YANG Shao-hong, LIU Ying-gao, ZHAI Sen, JIANG Ping, SONG Lei, LI Shu-jiang. Pathogen Identification and Its Compound Formulations’ Control Efficacy against Solidago canadensis[J]. Biotechnology Bulletin, doi: 10.13560/j.cnki.biotech.bull.1985.2025-1218.

Figures/Tables 13

Table 1 Primers for RT-qPCR

基因名称 Gene name	引物名称 Primer name	序列 Sequence （5′-3′）
ITS	ITS1	TCCCTAGCTGAACCTCCCC
ITS	ITS4	TCCTCCGCTTATTGATATCC
CAL	CAL-F	GAGTTCAAGGAGGCCTTCTCCC
CAL	CAL-R	CATCTTCTGGCCATCATGG
EF-1α	B.d-F	CATCGAGAAGTTCGAGAAGG
EF-1α	B.d-R	TACTTGAAGGAACCCTTACC
HIS	H3F	AGGTCCACTGGTGGCAAG
HIS	H3R	GCGGGCGAGCTGGATGTCCTT

Table 2 Screening of the optimal ratio of three adjuvants

序号 Serial number	实验因素 Experimental factor
序号 Serial number	润湿剂 Wetting agent （%）	分散剂 Dispersant （%）	紫外保护剂 UV protectant （%）
1	1	1	0.5
2	5	3	0.5
3	3	5	0.5
4	5	1	0.7
5	3	3	0.7
6	1	5	0.9
7	3	1	0.9
8	1	3	0.9
9	5	5	0.9

Table 3 Isolation results of pathogenic fungi

编号 No.	叶片分离株数 Number of leaf isolates	叶片分离率 Leaf separation rate （%）	根部分离株数 Number of root isolates	根部分离率 Root separation rate （%）
SC01	7	7	0	0
SC02	43	43	69	69
SC03	4	4	0	0
SC04	5	5	12	12
SC05	10	10	0	0
SC06	17	17	0	0
SC07	14	14	19	19

Table 4 Pathogenicity of screened strains against S. canadensis and its non-host plants

编号 No.	叶部病级 Leaf disease grade			根部病级 Root disease grade
编号 No.	加拿大一枝黄花 S. canadensis	香樟 C. camphora	楠木 P. zhennan	加拿大一枝黄花 S. canadensis	香樟 C. camphora	楠木 P. zhennan
SC01	1	2	1	0	0	0
SC02	5	1	0	5	0	0
SC03	1	0	1	0	0	0
SC04	2	2	2	0	0	1
SC05	2	3	3	0	0	0
SC06	1	4	5	0	0	0
SC07	4	5	4	1	0	0

Fig. 1 Leaf and root inoculation diagramA-C: Leaf sterile agar block control (C. camphora, P. zhennan, S. canadensis, the same below); D-F: leaf inoculation with SC02; G-I: leaf inoculation with SC06; J-L: leaf inoculation with SC07; M-O: root inoculation with SC02

Fig. 2 Identification results of SC02 strainA: Front view of the colony; B: back view of the colony; C: sporulation medium; D: ascus shell; E: ascus; F: α-conidia; G: β-conidia; H: phylogenetic tree of strains

Fig. 3 Screening results of compound herbicidesA-H: Single-dose test. CK, 20% clopyralid, 25% carfentrazone, 41% glyphosate isopropylamine salt, 48% triclopyr butoxyethyl ester, 10% quizalofop-p-ethyl, 20% flumioxazin, 20% dimethachlor aqueous solution group, respectively. I-P: Screening plate for recommended concentrations of chemical herbicides. CK, 20% clopyralid, 25% carfentrazone SL, 41% glyphosate isopropylamine salt, 48% triclopyr butoxyethyl ester EC, 10% quizalofop-p-ethyl, 24% flumioxazin, 20% dimethachlor aqueous solution group, respectively. Q-T: Screening plate for optimal concentrations of chemical herbicides. 20% clopyralid, 25% carfentrazone SL, 41% glyphosate isopropylamine salt, 24% flumioxazin group, respectively. U: Bar chart of screening results for compound herbicides against S. canadensis. The recommended concentration refers to the dilution concentration officially recommended for the purchased herbicide. The optimal concentration indicates the dilution of the recommended concentration that supports the optimal growth of the pathogen. Different lowercase letters indicate significant differences among different treatments at the same time (P0.001), the same below

Table 5 Screening results of individual chemical herbicide

化学除草剂 Chemical herbicides	推荐浓度 Recommended concentration	死亡率 Mortality rate （%）	死亡时间 Time of death （d）	死亡症状 Symptoms of death
20%氯氟吡氧乙酸	1 μL/mL	100	4	全株死亡，茎秆叶片全部萎蔫枯死
25%环嗪酮SL	0.02 g/mL	100	8	全株死亡，但有少量叶片茎秆泛绿，短时间未完全失活
41%草甘膦异丙铵盐	0.02 g/mL	100	6	全株死亡，但部分茎秆短时间未失活
48%三氯吡氧乙酸丁氧基乙酯EC	10 μL/mL	90	9	大部分植株枯死，但有少量植株茎秆存活
10%精草胺膦胺盐	10 μL/mL	100	8	全株死亡，茎秆叶片全部萎蔫枯死
24%唏草酮	1 μL/mL	100	5	全株死亡，茎秆叶片全部萎蔫枯死
20%二甲四氯水剂	5 μg/mL	100	4	全株死亡，茎秆叶片全部萎蔫枯死

Fig. 4 Screening results for carriers and three adjuvantsA-E: Carrier (CK, diatomaceous earth, kaolin, bentonite, silica gel) screening results. F-I: Wetting agent (CK, Tween 20, Tween 80, PEG 2000) screening results. J-Q: Ultraviolet protection agent (CK, UV-treated CK, humic acid, UV-treated humic acid, sodium alginate, UV-treated sodium alginate, dextrin, UV-treated dextrin) screening results. R-U: Dispersant (CK, sodium lignosulfonate, calcium lignosulfonate, methylene bis-naphthalenesulfonate) screening results. V-Y: Screening results for carrier, wetting agent, ultraviolet protection agent, and dispersant

Table 6 Determination of wetting time of different wetting agents

润湿剂 Wetting agents	（润湿时间 Wetting time （±SE）（s））
润湿剂 Wetting agents	1%	3%	5%
吐温20 Tween 20	32±0.58b	20±0.52b	11±1.88b
吐温80 Tween 80	24±0.68c	14±1.28c	10±0.75b
PEG2000	54±0.78a	45±1.05a	36±0.62a

Fig. 5 Screening results of the optimal ratio of three additives

Table 7 Quality test results of wettable powder mixture of D. sojae SC02

指标 Index	国家标准值 National standards	测量值 Measured value
润湿时间 Wetting time （s）	≤180	36±1
悬浮率 Suspensibility （%）	≥70	74
含孢量 Spore content （CFU/g）	-	1.65×10⁹
pH	5.0-8.0	6.5
细度 Fineness （%）	≥90	92
起泡性 Frothiness （mL）	≤10 mL	7 mL
干燥减量 Drying reduction （%）	≤6	5.65
贮存稳定性 Storage stability （%）	≥80	≥81.5

Fig. 6 Results of the pot experimentsA-J: CK1 distilled water, CK2 clopyralid, 20, 10, 5, 2.5, 1.25, 0.83, 0.63, 0.5 g/L dilution concentration treatment effect. K-O: Plant height, leaf necrosis rate, fresh weight control effect, root necrosis rate, and plant control effect results in potted control trials

References 41

[1]	鞠瑞亭, 李慧, 石正人, 等. 近十年中国生物入侵研究进展 [J]. 生物多样性, 2012, 20(5): 581-611.
	Ju RT, Li H, Shi ZR, et al. Progress of biological invasions research in China over the last decade [J]. Biodivers Sci, 2012, 20(5): 581-611.
[2]	Theoharides KA, Dukes JS. Plant invasion across space and time: factors affecting nonindigenous species success during four stages of invasion [J]. New Phytol, 2007, 176(2): 256-273.
[3]	杨昌洪, 肖晓华, 刘春, 等. 噬菌核霉可湿性粉剂防治油菜菌核病试验示范 [J]. 南方农业, 2016, 10(19): 19-21.
	Yang CH, Xiao XH, Liu C, et al. Experimental demonstration on control of Sclerotinia sclerotiorum with wettable powder [J]. South China Agric, 2016, 10(19): 19-21.
[4]	陈娥. 顶孢霉可湿性粉剂和微粉剂的药效评价及田间应用技术研究 [D]. 兰州: 甘肃农业大学, 2016.
	Chen E. Evaluation and field application technology of WP and MDP of Acremonium hansfordii [D]. Lanzhou: Gansu Agricultural University, 2016.
[5]	王均伟, 黄晓慧, 杨然迪, 等. 草甘膦与新型原卟啉原氧化酶抑制剂X18002复配减量应用研究 [J]. 农药学学报, 2023, 25(4): 817-828.
	Wang JW, Huang XH, Yang RD, et al. Study on usage reduction of glyphosate mixed with a novel protoporphyrinogen oxidase inhibitor-X18002 [J]. Chin J Pestic Sci, 2023, 25(4): 817-828.
[6]	何丽烂, 区焯林, 王芳, 等. 尖孢镰刀菌与草甘膦复配对空心莲子草防御酶的影响 [J]. 广东农业科学, 2011, 38(22): 78-79, 86.
	He LL, Ou ZL, Wang F, et al. Influence of Fusarium oxysporum mixed with Glyphosate on Alternanthera philoxeroides defense enzymes [J]. Guangdong Agric Sci, 2011, 38(22): 78-79, 86.
[7]	李鑫, 郭冉, 杨娟, 等. 黏质沙雷氏菌Ha1培养条件优化及其与化学除草剂复配研究 [J]. 杂草学报, 2020, 38(2): 21-32.
	Li X, Guo R, Yang J, et al. Optimization of culture conditions of Serratia marcescens Ha1 and its compounding with herbicides [J]. J Weed Sci, 2020, 38(2): 21-32.
[8]	商美妮. 木霉固体发酵培养条件优化及含木霉生物基质育苗效应研究 [D]. 南京: 南京农业大学, 2020.
	Shang MN. Study on optimization of solid fermentation culture conditions of Trichoderma and the effect of biological substrate containing Trichoderma on seedling growth [D]. Nanjing: Nanjing Agricultural University, 2020.
[9]	翟森, 靳文涛, 柳丽杰, 等. 龙泉山森林公园入侵植物加拿大一枝黄花的调查及风险性分析 [J]. 安徽农业科学, 2025, 53(9): 118-122.
	Zhai S, Jin WT, Liu LJ, et al. Investigation and risk analysis of invasive plant Solidago canadensis in longquanshan forest park [J]. J Anhui Agric Sci, 2025, 53(9): 118-122.
[10]	王前程, 张迎迎, 戴陶宇, 等. 拟康宁木霉T-51菌株对番茄枯萎病的生物防治及其机理研究 [J]. 西北植物学报, 2022, 42(6): 974-982.
	Wang QC, Zhang YY, Dai TY, et al. Study on biological control effect and mechanism of Trichoderma koningiopsis T-51 on tomato Fusarium wilt [J]. Acta Bot Boreali Occidentalia Sin, 2022, 42(6): 974-982.
[11]	袁树忠, 何平平, 廉吉衷, 等. 一种镰刀菌对空心莲子草的致病力与寄主专一性测定 [J]. 中国生物防治, 2006(2): 137-141.
	Yuan SZ, He PP, Lian JZ, et al. Host range and pathogenicity of a Fusarium species to Alternanthera philoxeroides [J]. Chin J Biol Control, 2006(2): 137-141.
[12]	李思, 尚晓静, 张富美, 等. 蓝莓根部内生木霉分离鉴定及其拮抗作用 [J]. 核农学报, 2022, 36(8): 1569-1578.
	Li S, Shang XJ, Zhang FM, et al. Diversity of endophytic Trichoderma in the roots of blueberries and its antagonistic effects [J]. J Nucl Agric Sci, 2022, 36(8): 1569-1578.
[13]	司璐, 吴彤, 甄锦程, 等. 大兴安岭凋落物中可培养真菌分离、鉴定及活性筛选 [J]. 中国农学通报, 2022, 38(26): 118-123.
	Si L, Wu T, Zhen JC, et al. Isolation, identification and activity screening of culturable fungi from litter in the greater Hinggan Mountains [J]. Chin Agric Sci Bull, 2022, 38(26): 118-123.
[14]	闫佳佳, 万璐, 尤梦瑶, 等. 产香兰素内生真菌GP20的鉴定及发酵条件研究 [J]. 中国调味品, 2022, 47(11): 44-49.
	Yan JJ, Wan L, You MY, et al. Study on identification and fermentation conditions of vanillin-producing endophytic fungus GP20 [J]. China Condiment, 2022, 47(11): 44-49.
[15]	巨亚雯, 陈亚丽, 徐蓬, 等. 植物生长调节剂与磺酰脲类除草剂混用对防除麦田杂草的协同增效作用 [J]. 杂草学报, 2022, 40(4): 43-49.
	Ju YW, Chen YL, Xu P, et al. Synergistic interaction of plant growth regulators mixed with sulfonylurea herbicides on weeding in wheat fields [J]. J Weed Sci, 2022, 40(4): 43-49.
[16]	张英财, 农向群, 张泽华, 等. 18种化学农药与绿僵菌相容性研究 [J]. 中国生物防治学报, 2012, 28(2): 186-191.
	Zhang YC, Nong XQ, Zhang ZH, et al. Compatibility of eighteen chemical pesticides with Metarhizium anisopliae [J]. Chin J Biol Control, 2012, 28(2): 186-191.
[17]	朱文珊. 长枝木霉拌种对芸豆生长及根际土壤酶活性的影响 [D]. 大庆: 黑龙江八一农垦大学, 2022.
	Zhu WS. Effects of seed dressing in Trichoderma longibrachiatum on kidney bean growth and enzyme activities in rhizosphere soil [D]. Daqing: Heilongjiang Bayi Agricultural University, 2022.
[18]	何祖传, 周健, 张洪泉, 等. 化学除草剂对加拿大一枝黄花的防除效果试验 [J]. 现代农业科技, 2020(10): 82, 88.
	He ZC, Zhou J, Zhang HQ, et al. Effect of chemical herbicide on Solidago canadensis L [J]. Mod Agric Sci Technol, 2020(10): 82, 88.
[19]	韩敏, 张海艳, 洪爱梅, 等. 几种除草剂对加拿大一枝黄花的防除效果评价 [J]. 世界农药, 2024, 46(10): 56-62.
	Han M, Zhang HY, Hong AM, et al. Evaluation of the control effects of several herbicides on Solidago canadensis [J]. World Pestic, 2024, 46(10): 56-62.
[20]	王博, 李嘉, 马莉莉, 等. 异丙甲草胺乳油不同用量防治玉米田禾本科杂草田间药效试验 [J]. 云南农业科技, 2021(5): 36-38.
	Wang B, Li J, Ma LL, et al. Field experiments of metolachlor EC with different amounts on grassy weeds in corn field [J]. Yunnan Agric Sci Technol, 2021(5): 36-38.
[21]	咸洪泉, 李雅华. 木霉菌株Td31代谢产物对立枯丝核菌的拮抗作用 [J]. 沈阳农业大学学报, 2009, 40(5): 566-570.
	Xian HQ, Li YH. The antagonism of metabolite by Trichoderma.Td31 to Rhizoctonia solani [J]. J Shenyang Agric Univ, 2009, 40(5): 566-570.
[22]	孙文. 短短芽胞杆菌ch2-22发酵动力学与水分散粒剂的研究 [D]. 天津: 河北工业大学, 2017.
	Sun W. Study on fermentation kinetics of Brevibacillus brevis Ch2-22 and preparation of water dispresible granule [D]. Tianjin: Hebei University of Technology, 2017.
[23]	陈慧. 绛红褐链霉菌颗粒剂的研究 [D]. 雅安: 四川农业大学, 2015.
	Chen H. Reseach of granule formulation for Streptomyces purpeofuscus [D]. Yaan: Sichuan Agricultural University, 2015.
[24]	古丽君, 徐秉良, 张德瑞, 等. 深绿木霉病原菌制剂加工工艺研究: 中国植物保护学会2009年学术年会[C], 武汉, 2009.
	Gu LJ, Xu BL, Zhang DR, et al. Study on Processing Technology of Trichoderma atroviride Pathogen Preparation[C]. Proceedings of the 2009 Annual Conference of the China Society of Plant Protection, Wuhan, 2009.
[25]	戴启星. 长枝木霉T6菌株可湿性粉剂研制及复配菌剂协同增效作用研究 [D]. 兰州: 甘肃农业大学, 2019.
	Dai QX. Development of Trichoderma longibrachiatum T6 wettable powder and synergistic compounding agents [D]. Lanzhou: Gansu Agricultural University, 2019.
[26]	邹益泽. 草茎点霉水分散粒剂的研发及其对鸭跖草防除效果研究 [D]. 沈阳: 沈阳农业大学, 2019.
	Zou YZ. Development of Phoma herbarum WG and its control effect against Commelina communis L. [D]. Shenyang: Shenyang Agricultural University, 2019.
[27]	王建锋, 陆宁海. 木霉菌在农业中的研究与应用前景 [J]. 安徽农学通报, 2008, 14(14): 41-43, 136.
	Wang JF, Lu NH. Rogress in agriculture research with Trichoderma [J]. Anhui Agric Sci Bull, 2008, 14(14): 41-43, 136.
[28]	叶素丹, 冯明光. 生防真菌耐旱特性的生理生化基础及其利用 [J]. 应用生态学报, 2004, 15(12): 2383-2387.
	Ye SD, Feng MG. Physiological and biochemical bases of xero-tolerance of fungal biocontrol agents and their application [J]. Chin J Appl Ecol, 2004, 15(12): 2383-2387.
[29]	杨肖. 金龟子绿僵菌和小檗碱复配及其可湿性粉剂和水分散粒剂的制备 [D]. 北京: 北京化工大学, 2021.
	Yang X. Preparation of Metarhizium anisopliae and berberine and their wettable powder and water dispersible granuie [D]. Beijing: Beijing University of Chemical Technology, 2021.
[30]	农药可湿性粉剂润湿性测定方法: [S]. 2001.
	Test method for wettability of pesticide wettable powders: [S]. 2001.
[31]	农药可湿性粉剂悬浮率剂测定方法: [S]. 1993.
	Test method for suspensibility of pesticide wettable powders: [S]. 1993.
[32]	冯月. 木霉菌株产酶培养及其特定底物酶解过程研究 [D]. 北京: 北京林业大学, 2012.
	Feng Y. Studies on cellulase-produced fermentation of Trichoderma strains and enzymatic hydrolysis of the specific substrate [D]. Beijing: Beijing Forestry University, 2012.
[33]	农药pH值的测定方法: [S]. 1993.
	Test method for pH value of pesticides: [S]. 1993.
[34]	国家技术监督局. 农药粉剂、可湿性粉剂细度测定方法: [S]. 北京: 中国标准出版社, 2004.
	State Bureau of Quality and Technical Supervision of the People's Republic of China. Seive test for dustable and wettable powders of pesticides: [S]. Beijing: Standards Press of China, 2004.
[35]	国家质量监督检验检疫总局, 中国国家标准化管理委员会. 农药持久起泡性测定方法: [S]. 北京: 中国标准出版社, 2012.
	General Administration of Quality Supervision, Inspection and Quarantine of the People's Republic of China, Standardization Administration of the People's Republic of China. Testing method of persistent foaming for pesticides: [S]. Beijing: Standards Press of China, 2012.
[36]	农药干燥减量的测定方法: [S]. 2013.
	Test method for drying reduction of pesticides: [S]. 2013.
[37]	周欣欣, 张宏军, 张佳, 等. 除草剂田间药效试验申请应注意的几个问题 [J]. 农药科学与管理, 2012, 33(7): 8-12.
	Zhou XX, Zhang HJ, Zhang J, et al. Several problems that should be paid attention to in the application of herbicide field efficacy test [J]. Pestic Sci Adm, 2012, 33(7): 8-12.
[38]	常悦, 李玉静, 李周旭, 等. 基于Maxent和ArcGIS加拿大一枝黄花适生区预测 [J]. 农学学报, 2024, 14(3): 40-47.
	Chang Y, Li YJ, Li ZX, et al. Prediction of suitable growing areas of Solidago canadensis L. based on maxent and ArcGIS [J]. J Agric, 2024, 14(3): 40-47.
[39]	叶梅荣, 刘庆安, 马纪忠, 等. 41%草甘膦水剂对加拿大一枝黄花的防除效果 [J]. 江苏农业科学, 2010, 38(2): 130-132.
	Ye MR, Liu QA, Ma JZ, et al. Controlling effects of 41% glyphosate aqueous solution on Solidago canadensis L [J]. Jiangsu Agric Sci, 2010, 38(2): 130-132.
[40]	李玲燕. 生物除草剂齐整小核菌SC64颗粒剂的研究 [D]. 南京: 南京农业大学, 2015.
	Li LY. A study on solid granule of bioherbicide Sclerotium rolfsii strain SC64 [D]. Nanjing: Nanjing Agricultural University, 2015.
[41]	曹晶晶, 谷涛, 张自常, 等. 防除不同生境下加拿大一枝黄花的除草剂研究 [J]. 杂草学报, 2024, 42(4): 82-90.
	Cao JJ, Gu T, Zhang ZC, et al. Research on herbicides for controlling Solidago canadensis L. in different habitats [J]. J Weed Sci, 2024, 42(4): 82-90.