哈茨木霉LTR-2与产脲节杆菌DnL1-1协同对小麦茎基腐病的防治效果与机理

doi:10.3969/j.issn.1004-1524.2023.06.16

浙江农业学报 ›› 2023, Vol. 35 ›› Issue (6): 1385-1395.DOI: 10.3969/j.issn.1004-1524.2023.06.16

哈茨木霉LTR-2与产脲节杆菌DnL1-1协同对小麦茎基腐病的防治效果与机理

杨凯(), 陈凯, 李红梅, 赵忠娟, 扈进冬, 李纪顺^*(), 杨合同

齐鲁工业大学(山东省科学院)生态研究所,山东省应用微生物重点实验室,山东济南 250103

收稿日期:2022-07-18 出版日期:2023-06-25 发布日期:2023-07-04
通讯作者: *李纪顺,E-mail: yewu2@sdas.org
作者简介:杨凯(1988—),女,山东德州人,博士,助理研究员,研究方向为环境微生物。E-mail: yangkai1988@qlu.edu.cn
基金资助:
山东省自然科学基金(ZR2022MC215);校(院)科教产融合试点工程(基础研究类)(2022PX099);校(院)科教产融合试点工程(国际合作项目)(2022GH011);山东省重点研发计划(重大科技创新工程)(2020CXGC010805);山东省重点研发计划(重大科技创新工程)(2020CXGC010803-02);山东省自然科学基金青年基金(ZR2020QC044)

Biocontrol efficacy and action mechanism of Trichoderma harzianum LTR-2 and Arthrobacter ureafaciens DnL1-1 against crown rot of wheat

YANG Kai(), CHEN Kai, LI Hongmei, ZHAO Zhongjuan, HU Jindong, LI Jishun^*(), YANG Hetong

Shandong Provincial Key Laboratory of Applied Microbiology, Ecology Institute, Qilu University of Technology (Shandong Academy of Sciences), Jinan 250103, China

Received:2022-07-18 Online:2023-06-25 Published:2023-07-04

摘要/Abstract

摘要：

为明确哈茨木霉(Trichoderma harzianum)LTR-2与产脲节杆菌(Arthrobacter ureafaciens)DnL1-1协同对小麦茎基腐病菌的防治效果,以假禾谷镰孢(Fusarium pseudograminearum)为指示菌对2种菌进行对峙培养和盆栽防效等研究;通过菌丝生长、孢子萌发等指标,探讨LTR-2与DnL1-1对病原菌的抑菌机制。结果表明,随着假禾谷镰孢接种浓度逐渐降低,小麦茎基腐病病情指数逐渐降低,各处理对小麦茎基腐病防效差异显著(P<0.05),其中LTR-2+DnL1-1联合接种对小麦茎基腐病的防效最好,对假禾谷镰孢20倍、50倍和100倍稀释液的防效分别为34.7%、50.3%和87.2%。平板对峙试验中,LTR-2+DnL1-1协同与LTR-2单独对假禾谷镰孢的抑菌率均可达100%,2种处理没有显著差异;对峙试验显微结果显示,LTR-2+DnL1-1协同与LTR-2单独均能使假禾谷镰孢菌丝形态发生明显改变,原生质浓缩,隔膜增多和菌丝断裂;LTR-2+DnL1-1共培养与LTR-2单独培养的发酵滤液处理对假禾谷镰孢的菌丝形态均有明显影响,使菌丝膨大畸变,后期只产生分生孢子,几乎不产生厚垣孢子,且显著抑制其分生孢子的萌发。综上所述,DnL1-1和LTR-2对小麦茎基腐病的防效具有协同增效作用,以上结果为两者的联合应用提供了科学依据。

关键词: 哈茨木霉, 产脲节杆菌, 小麦茎基腐病, 生物防治

Abstract:

To assess the synergism control of Trichoderma harzianum LTR-2 and Arthrobacter ureafaciens DnL1-1 against wheat crown rot. The antimicrobial activities of LTR-2 and DnL1-1 against pathogenic fungus Fusarium pseudograminearum were examined using confrontation culture experiment and disease-control pot experiment. Then the inhibitory mechanism of LTR-2 and DnL1-1 against F. pseudograminearum was studied by measuring mycelial growth and spore germination. The results showed that with the decrease of F. pseudograminearum inoculation concentration, the disease index of wheat crown rot gradually decreased, and the control effects of various treatments on wheat crown rot were significantly (P<0.05) different. Among these treatments, the combined inoculation of LTR-2 and DnL1-1 showed the best control effects on wheat crown rot with the values of 34.7%, 50.3% and 87.2%, respectively. In plate confrontation test, the inhibitory rates of LTR-2+DnL1-1 and LTR-2 alone against F. pseudograminearum were both 100%, and there was no significant difference between the two treatments. The microscopic results of confrontation test showed that the morphology of F. pseudograminearum mycelia changed significantly with protoplast condensation, septum increase and mycelium fracture under the effect of LTR-2 alone and LTR-2 and DnL1-1 synergy. The fermentation filtrates of the LTR-2 alone culture and combined culture of LTR-2 and DnL1-1 both had a significant effect on the mycelia morphology of F. pseudograminearum, resulting in mycelial expansion and distortion, and affected the sporulation type, so that only produced conidia, but almost no chlamydospore. In addition, the germination of conidia was inhibited significantly. In conclusion, DnL1-1 had a synergistic effect on the control effect of LTR-2 against wheat crown rot, and the above results afforded scientific basis for the combined application of the two strains.

Key words: Trichoderma harzianum, Arthrobacter ureafaciens, wheat crown rot, biological control

中图分类号:

S435.121

杨凯, 陈凯, 李红梅, 赵忠娟, 扈进冬, 李纪顺, 杨合同. 哈茨木霉LTR-2与产脲节杆菌DnL1-1协同对小麦茎基腐病的防治效果与机理[J]. 浙江农业学报, 2023, 35(6): 1385-1395.

YANG Kai, CHEN Kai, LI Hongmei, ZHAO Zhongjuan, HU Jindong, LI Jishun, YANG Hetong. Biocontrol efficacy and action mechanism of Trichoderma harzianum LTR-2 and Arthrobacter ureafaciens DnL1-1 against crown rot of wheat[J]. Acta Agriculturae Zhejiangensis, 2023, 35(6): 1385-1395.

图/表 9

表1 LTR-2与DnL1-1对小麦茎基腐病病情指数的影响

Table 1 Effect of LTR-2 and DnL1-1 on disease index of wheat crown rot

处理 Treatment	病情指数Disease index
处理 Treatment	100倍稀释液 Dilution ratio 100×	50倍稀释液 Dilution ratio 50×	20倍稀释液 Dilution ratio 20×
CK	25.8±6.3 a	62.0±7.5 a	79.2±5.2 a
酷拉斯 Kuras	3.3±1.4 c	27.5±7.5 b	51.7±1.4 b
LTR-2+DnL1-1	3.3±3.8 c	30.8±8.8 b	51.7±1.4 b
LTR-2	15.8±6.3 b	42.5±2.5 ab	53.3±7.6 b
DnL1-1	27.5±5.0 a	56.7±11.8 a	71.7±8.0 a

图1 LTR-2与DnL1-1对小麦茎基腐病的防治效果柱上无相同小写字母表示差异显著(P<0.05)。

Fig.1 Control effect of LTR-2 and DnL1-1 on wheat crown rot Data on the bars marked without the same lowercase letter indicated significant differences at P<0.05.

表2 假禾谷镰孢20倍稀释接种条件下不同处理对小麦植株生长的影响

Table 2 Effects of different treatments on wheat growth under 20 times dilution inoculation of Fusarium pseudograminearum

处理 Treatment	株高 Plant height/cm	苗鲜重 Fresh weight of seedling/g	根鲜重 Fresh weight of root/g	根冠比 Root shoot ratio/%
CK	23.28±0.81 ab	4.14±0.13 b	1.81±0.30 b	0.44±0.09 b
酷拉斯Kuras	23.84±1.16 ab	5.33±0.56 a	2.78±0.73 a	0.53±0.17 a
LTR-2+DnL1-1	24.42±0.76 a	4.32±0.22 b	2.89±0.30 a	0.67±0.08 a
LTR-2	23.29±0.70 ab	4.54±0.18 b	2.93±0.25 a	0.65±0.07 a
DnL1-1	22.85±1.12 bc	4.14±0.44 b	2.41±0.93 a	0.57±0.19 a

图2 LTR-2与DnL1-1对假禾谷镰孢菌的抑制效果 A~D分别为对照、DnL1-1、LTR-2、DnL1-1+LTR-2与假禾谷镰孢菌对峙培养7 d。

Fig.2 Inhibitory effect of LTR-2 and DnL1-1 on Fusarium pseudograminearum A-D represent the control, DnL1-1, LTR-2 and DnL1-1+LTR-2 confront cultured with Fusarium pseudograminearum for 7 days, respectively.

图3 LTR-2与DnL1-1对假禾谷镰孢菌丝生长的影响 A,空白对照;B,DnL1-1与假禾谷镰孢对峙培养;C、D,LTR-2与假禾谷镰孢对峙培养;E、F、G,LTR-2+DnL1-1与假禾谷镰孢对峙培养。图中黑色箭头所指为LTR-2以缠绕、穿插、紧贴等方式寄生于假禾谷镰孢处,或假禾谷镰孢菌丝原生质浓缩、隔膜增多、细胞变短或菌丝断裂处。

Fig.3 Effects of LTR-2 and DnL1-1 on mycelial growth of Fusarium pseudograminearum A, Control; B, DnL1-1 confront cultured with Fusarium pseudograminearum; C and D, LTR-2 confront cultured with Fusarium pseudograminearum; E, F and G, DnL1-1+LTR-2 confront cultured with Fusarium pseudograminearum. The black arrows in the figure represented that Trichoderma parasitize Fusarium pseudograminearum in ways of twining, penetrating and clinging, and lead to Fusarium pseudograminearum mycelium protoplasm shrinkage, septum increased, cell became shorter or mycelium broke.

图4 不同处理发酵滤液对假禾谷镰孢菌丝生长的影响 A、B分别为培养2、5 d的结果,自左向右依次为加入无菌水的对照组,加入DnL1-1发酵滤液、LTR-2发酵滤液、DnL1-1+LTR-2共培养发酵滤液的处理组,上图为正面,下图为反面。

Fig.4 Inhibition of different fermentation filtrates on growth of Fusarium pseudograminearum mycelia A and B represent the results of cultured for 2 and 5 days, respectively. From left to right are the sterile water control group, DnL1-1 fermentation filtrate treatment group, LTR-2 fermentation filtrate treatment group and DnL1-1+LTR-2 combined fermentation filtrate treatment group. The top is the front side, and the bottom is the back side.

图5 LTR-2与DnL1-1发酵滤液对假禾谷镰孢液体培养中菌丝生长的影响 A,对照(空白发酵滤液+假禾谷镰孢);B,DnL1-1发酵滤液+假禾谷镰孢;C,LTR-2发酵滤液+假禾谷镰孢;D,DnL1-1+LTR-2发酵滤液+假禾谷镰孢。图中黑色箭头所指为菌丝膨大、扭曲或溶断处。

Fig.5 Effects of LTR-2 and DnL1-1 fermentation filtrate on mycelia growth of Fusarium pseudograminearum in liquid culture A, B, C and D represent the sterile water, DnL1-1 fermentation filtrate, LTR-2 fermentation filtrate and DnL1-1+LTR-2 fermentation filtrate cultured with Fusarium pseudograminearum, respectively. The black arrows in the figure refer to the area where the hyphae expand, twist or fuse.

图6 LTR-2与DnL1-1发酵滤液对假禾谷镰孢液体培养中产孢的影响 A、B,对照(空白发酵滤液+假禾谷镰孢);C,DnL1-1发酵滤液+假禾谷镰孢;D,LTR-2发酵滤液+假禾谷镰孢;E,DnL1-1+LTR-2发酵滤液+假禾谷镰孢。图中黑色箭头所指为不同孢子类型,包括厚垣孢子、分生孢子。

Fig.6 Effect of LTR-2 and DnL1-1 fermentation filtrate on sporulation of Fusarium pseudograminearum in liquid culture A, B, C, D and E represent the sterile water, DnL1-1 fermentation filtrate, LTR-2 fermentation filtrate and DnL1-1+LTR-2 fermentation filtrate cultured with Fusarium pseudograminearum, respectively. The black arrows in the figure indicate different spore types, including chlamydospores and conidia.

图7 LTR-2与DnL1-1发酵滤液对假禾谷镰孢分生孢子萌发的影响

Fig.7 Effects of LTR-2 and DnL1-1 fermentation filtrate on conidial germination of Fusarium pseudograminearum

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哈茨木霉LTR-2与产脲节杆菌DnL1-1协同对小麦茎基腐病的防治效果与机理

Biocontrol efficacy and action mechanism of Trichoderma harzianum LTR-2 and Arthrobacter ureafaciens DnL1-1 against crown rot of wheat

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