玉米茎腐病生防菌哈茨木霉CCTH-2鉴定及其生物学特性

doi:10.7668/hbnxb.20192851

摘要/Abstract

摘要：

玉米茎腐病(CSR)对玉米产量和品质危害巨大。为尽快展开玉米茎腐病的有效防治,将吉林省玉米茎腐病的优势致病菌禾谷镰孢,与14株从玉米根际土壤中分离得到木霉菌株分别进行平板对峙试验。观察木霉对禾谷镰孢的抑制作用,筛选得到抑制效果较好的菌株CCTH-2和CCTH-6。复筛通过这2株木霉的发酵液、易挥发性代谢产物、难挥发性代谢产物对禾谷镰孢的抑制效果和对其他病原真菌的抑制效果,最终筛选到一株具有良好防效的木霉菌株CCTH-2;CCTH-2经过形态学和分子生物学鉴定显示,为哈茨木霉,并对其生物学特性进行初步研究。结果表明,平板对峙培养条件下CCTH-2对禾谷镰孢抑制率为82.83%,其发酵液、易挥发代谢物和难挥发代谢物对禾谷镰孢的抑制率为36.56%,46.75%,32.48%,具有显著抑制效果。生物学特性试验显示,CCTH-2菌落生长和产孢的最适培养基为PDA,最适温度为25 ℃,最适光照条件为全光照,外源添加Fe²⁺有利于CCTH-2的生长发育,并且CCTH-2具有一定的适应环境酸碱度和盐含量变化的能力。因此,可以确认哈茨木霉CCTH-2是一株有着良好防效且潜力巨大的生防菌。

关键词: 玉米茎腐病, 禾谷镰孢, 哈茨木霉CCTH-2, 生防菌株

Abstract:

Corn stalk rot is harmful to yield and quality of corn,so effective control of corn stalk rot should be carried out as soon as possible.For Fusarium graminearum,the dominant strains of corn stalk rot in Jilin Province,and fourteen Trichoderma sp.strains were isolated from corn rhizosphere soil were subjected to plate confrontation experiment respectively.By observing the inhibitory effect of Trichoderma sp.strains on Fusarium graminearum,the strains of CCTH-2 and CCTH-6 with better inhibitory effect could be screed.The inhibition of Fusarium graminearum by fermentation broth,volatile metabolites and non-volatile metabolites of these two Trichoderma sp.strains were screened again,and inhibitory effect of these two Trichoderma sp.strains on other pathogens.Finally,Trichoderma sp.strain CCTH-2 with good control effect was screened by comprehensive analysis.Morphological and molecular identification showed Trichoderma harzianum,and its biological characteristics were preliminarily studied.The results showed that the inhibition rate of CCTH-2 against Fusarium graminearum was 82.83% under culture conditions of slab face-off method and the fermentation broth,volatile metabolites and non-volatile metabolites of CCTH-2 against Fusarium graminearum was 36.56%,46.75%,32.48%,showed significant inhibition effect.The biological characteristics test showed that PDA was the optimal medium for the growth and sporulation of CCTH-2 colonies,the optimal temperature was 25 ℃,the optimal illunination condition was total illumination,exogenous addition of Fe²⁺ was beneficial to the growth and development of CCTH-2,and CCTH-2 had the ability to adapt to the changes of pH value and salt contene.Therefore,it could be confirmed that Trichoderma harzianum CCTH-2 was a biocontrol bacterium with good control effect and great potential,providing information guarantee and theoretical basis for the subsequent development and utilization of biocontrol bacteria of corn stalk rot.

Key words: Corn stalk rot, Fusarium graminearum, Trichoderma harzianum CCTH-2, Biological characteristics

佟昀铮, 于汇琳, 潘洪玉, 王迎春. 玉米茎腐病生防菌哈茨木霉CCTH-2鉴定及其生物学特性[J]. 华北农学报, 2022, 37(S1): 309-317. doi: 10.7668/hbnxb.20192851.

TONG Yunzheng, YU Huilin, PAN Hongyu, WANG Yingchun. Identification of Trichoderma harzianum CCTH-2 and Its Biological Characteristics,a Biocontrol Fungi Against Corn Stalk Rot[J]. Acta Agriculturae Boreali-Sinica, 2022, 37(S1): 309-317. doi: 10.7668/hbnxb.20192851.

http://www.hbnxb.net/CN/Y2022/V37/IS1/309

图/表 19

表1 木霉菌对禾谷镰孢的抑菌率

Tab.1 Inhibitory rate of Trichoderma spp. to Fusarium graminearum

菌株 Strains	禾谷镰孢 F. graminearum
菌株 Strains	菌落半径/cm The radius of colony	抑制率/% Inhibation rate
CCTH-1	1.10	72.58±0.60bc
CCTH-2	0.69	82.83±0.25a
CCTH-3	1.24	68.92±0.49c
CCTH-4	1.36	66.08±0.56d
CCTH-5	1.09	72.75±0.31bc
CCTH-6	0.70	82.42±0.25a
CCTH-7	0.95	76.33±0.30b
CCTH-8	1.23	69.33±0.18c
CCTH-9	1.21	69.67±0.25c
CCTH-10	1.11	71.83±0.25bc
CCTH-11	1.11	72.33±0.64bc
CCTH-12	1.14	71.42±0.27bc
CCTH-13	1.16	71.00±0.42c
CCTH-14	1.07	73.83±0.45b
CK	4.00	-

图1 禾谷镰孢菌落生长形态图(培养基正反两面) A.木霉菌CCTH-2、CCTH-6的发酵液对禾谷镰孢的抑制作用(3 d);B.木霉菌CCTH-2、CCTH-6的易挥发性代谢物对禾谷镰孢的抑制作用(5 d);C.木霉菌CCTH-2、CCTH-6的难挥发性代谢物对禾谷镰孢的抑制作用(36 h)。

Fig.1 Colony growth morphology diagram of Fusarium graminearum(Both sides of culture medium) A.Inhibition activities of fermentation broth of Trichoderma sp. CCTH-2 and CCTH-6 species against Fusarium graminearum(3 d);.B.Inhibition activities of volatile metabolites of Trichoderma sp. CCTH-2 and CCTH-6 species against Fusarium graminearum(5 d);C.Inhibition activities of non-volatile metabolites of Trichoderma sp. CCTH-2 and CCTH-6 species against Fusarium graminearum(36 h).

表2 木霉菌CCTH-2和CCTH-6对禾谷镰孢的抑制作用

Tab.2 Inhibitory activities of Trichoderma sp. CCTH-2 and CCTH-6 species against Fusarium graminearum

菌株 Strains	发酵液 Fermentation broth		易挥发性代谢产物 Volatile metabolites		难挥发性代谢产物 Non-volatile metabolites
菌株 Strains	菌落半径/cm The radius of colony	抑制率/% Inhibition rate	菌落半径/cm The radius of colony	抑制率/% Inhibition rate	菌落半径/cm The radius of colony	抑制率/% Inhibition rate
CCTH-2	2.03	36.56±1.21a	2.13	46.75±0.18a	1.06	32.48±1.59b
CCTH-6	2.18	31.87±0.92b	2.28	43.00±0.68b	0.91	42.04±1.59a
CK	3.20	-	4.00	-	1.57	-

表3 木霉菌CCTH-2和CCTH-6对其他病原菌的抑制作用

Tab.3 Inhibitory activities of Trichoderma sp.CCTH-2 and CCTH-6 species against other pathogenic fungi

菌株 Strains	对照组 Control group	木霉菌CCTH-2 Trichoderma sp. CCTH-2		木霉菌CCTH-6 Trichoderma sp. CCTH-6
菌株 Strains	菌落半径/cm The radius of colony	菌落半径/cm The radius of colony	抑制率/% Inhibition rate	菌落半径/cm The radius of colony	抑制率/% Inhibition rate
立枯丝核菌R.solani	7.50	2.27	69.73±0.86ab	2.63	64.93±0.77b
玉米大斑病菌S. turcica	3.45	1.05	69.57±0.80b	1.11	67.83±0.55b
玉米小斑病菌C. heterostrophus	3.42	1.17	65.79±1.51b	1.08	68.42±1.90b
核盘菌S. sclerotiorum	7.50	2.83	62.27±1.35b	2.57	65.73±1.78b
玉米灰斑病菌C. zeae-maydis	7.50	1.40	81.33±2.26a	1.46	80.53±0.93a
灰葡萄孢B.cinerea	6.07	1.31	78.42±1.87a	1.36	77.59±1.32a

图2 木霉菌CCTH-2和CCTH-6对其他病原菌的抑制作用

Fig.2 Inhibitory activities of Trichoderma sp.CCTH-2 and CCTH-6 species against other pathogenic fungi

图3 哈茨木霉菌形态 A.初期菌落形态;B.后期菌落形态;C.分生孢子梗;D.分生孢子。

Fig.3 Trichoderma harzianum morphologic characteristic A.Colony on PDA(initial stage);B.Colony reverse on PDA(late stage);C.Conidiophore;D.Conidium.

图4 哈茨木霉CCTH-2的系统发育树

Fig.4 Phylogenetic tree of Trichoderma harzianum CCTH-2

图5 不同的培养基上哈茨木霉CCTH-2的菌落形态

Fig.5 Colony morphology of Trichoderma harzianum CCTH-2 on different media

图6 不同的培养基对哈茨木霉CCTH-2菌丝生长和产孢的影响

Fig.6 Effects of different media on mycelial growth and sporulation of Trichoderma harzianum CCTH-2

图7 不同的温度条件下哈茨木霉CCTH-2的菌落形态

Fig.7 Colony morphology of Trichoderma harzianum CCTH-2 under different temperature conditions

图8 不同温度对哈茨木霉CCTH-2菌丝生长和产孢的影响

Fig.8 Effects of different temperatures on mycelial growth and sporulation of Trichoderma harzianum CCTH-2

图9 不同的光照类型下哈茨木霉CCTH-2的菌落形态

Fig.9 Colony morphology of Trichoderma harzianum CCTH-2 under different light types

图10 不同光照类型对哈茨木霉CCTH-2菌丝生长和产孢的影响

Fig.10 Effects of different light types on mycelial growth and sporulation of Trichoderma harzianum CCTH-2

图11 不同微量元素下哈茨木霉CCTH-2的菌落形态

Fig.11 Colony morphology of Trichoderma harzianum CCTH-2 under different trace elements

图12 不同微量元素对哈茨木霉CCTH-2菌丝生长和产孢的影响

Fig.12 Effects of different trace elements on mycelial growth and sporulation of Trichoderma harzianum CCTH-2

图13 不同pH值条件下哈茨木霉CCTH-2的菌落形态

Fig.13 Colony morphology of Trichoderma harzianum CCTH-2 under different pH conditions

图14 不同pH值对哈茨木霉CCTH-2菌丝生长和产孢的影响

Fig.14 Effects of different pH values on mycelial growth and sporulation of Trichoderma harzianum CCTH-2

图15 不同的含盐量条件下哈茨木霉CCTH-2的菌落形态

Fig.15 Colony morphology of T.harzianum CCTH-2 under different salt content conditions

图16 不同的含盐量对哈茨木霉CCTH-2菌丝生长和产孢的影响

Fig.16 Effects of different salt content on mycelial growth and sporulation of Trichoderma harzianum CCTH-2

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