杨树溃疡病拮抗链霉菌的筛选及鉴定

doi:10.11707/j.1001-7488.20210710

摘要/Abstract

摘要：

目的: 以生物防治防控杨树溃疡病发生与危害为目标，筛选能有效抑制杨树溃疡病且具有开发潜力的拮抗菌株，为杨树溃疡病的生物防治提供新的生防因子。方法: 从吉林省长白山国家级自然保护区横山保护站林下土壤样品中筛选获得1株对杨树溃疡病菌有良好抑制效果的链霉菌，编号为HS1。采用多重筛选法确定菌株HS1及其发酵液的抑菌活性，通过形态学、生理生化特征、16S rDNA序列分析等方法明确其分类地位，运用人工接种法测定菌株HS1发酵液对杨树溃疡病的预防和治疗效果。结果: 菌株HS1及其发酵液对供试植物病原菌均有抑制作用，抑菌谱广，其中对杨树溃疡病病原菌抑制作用最强，抑菌带宽达20.66 mm，发酵液抑菌圈直径达40.26 mm；16S rDNA序列比对分析发现，菌株HS1与Streptomyces scopuliridis同源性最高；人工接种试验结果表明，菌株HS1发酵液对杨树溃疡病的预防和治疗效果分别为60.87%和71.74%。结论: 生防菌株HS1可有效预防和控制杨树溃疡病，且对多种农林重要病害病原菌的抑菌作用效果明显，具有广阔的开发应用前景。

关键词: 杨树溃疡病, 链霉菌, 生物防治, 分类鉴定

Abstract:

Objective: In order to control the poplar canker disease by biological control agent,the present study aimed to screen and obtain antagonistic strains from Streptomyces that can effectively inhibit poplar canker. Method: A streptomyces strain with good biocontrol effect against the pathogen of poplar canker was obtained from the soil in the larch forest Hengshan Protection Station,Changbaishan Mountain National Nature Reserve,Jilin Province. The number was recorded as HS1. Multiple screening was used to determine the antibacterial activity of the strain and its fermentation broth; morphological,physiological and biochemical characteristics,16S rDNA sequence analysis were used to determine its taxonomic status; artificial inoculation method was used to determine the preventive and therapeutic effects of HS1 fermentation broth on poplar canker. Result: The result showed that strain HS1 and its fermentation broth had inhibitory effect on the tested plant pathogens,and the antibacterial spectrum was relatively wide. Among them,it had the strongest inhibitory effect on Botryosphaeria dothidea. The width of inhibition zone averagely was more than 20.66 mm and the diameter of the bacteriostatic circle of the fermentation broth reached 40.26 mm.The 16S rDNA sequence alignment analysis showed that the strain had the highest homology with Streptomyces scopuliridis; The result of artificial inoculation test showed that the preventive and therapeutic effects of strain HS1 fermentation broth on poplar canker were 60.87% and 71.74% respectirly. Conclusion: The screened strain HS1 and its fermentation broth could effectively prevent and inhibit the occurrence and expansion of poplar canker disease,and also have a good antibacterial effect on other pathogens of many important agricultural and forest diseases. Therefore,it has a good for development and application prospects in biocontrol of forest pathogens.

Key words: poplar canker, Streptomyces, biological control, screening and identification

中图分类号:

S432.1

陈越渠,刘庆珍,李立梅,张杨,韩姣,张永安. 杨树溃疡病拮抗链霉菌的筛选及鉴定[J]. 林业科学, 2021, 57(7): 92-100.

Yuequ Chen,Qingzhen Liu,Limei Li,Yang Zhang,Jiao Han,Yong'an Zhang. Screening and Identification of Antagonistic Streptomyces for Biocontrol of Poplar Canker[J]. Scientia Silvae Sinicae, 2021, 57(7): 92-100.

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菌株 Strains	抑菌带宽 Inhibition zone/mm	菌株Strains	抑菌带宽 Inhibition zone/mm	菌株Strains	抑菌带宽 Inhibition zone/mm
HS1	20.66±0.647 15	HS6	17.70±1.453 41	A-1-14	12.08±0.962 70
HS2	19.92±2.305 91	CX1	13.18±0.870 17	F12	14.92±0.472 86
HS3	19.96±1.833 79	CX2	13.94±1.107 61	F15	10.48±0.468 61
HS4	17.30±1.487 14	CX3	14.60±0.603 99
HS5	15.06±0.554 62	CX4	14.66±1.177 96

病原菌 Pathogens	抑菌带宽 Inhibition zone/mm	抑菌直径 Diameter of inhibition zone/mm
蓝莓枯枝病菌小新壳梭孢Neofusicoccum parvum	12.28±1.36f	20.69±1.15f
杨树烂皮病菌金黄壳囊孢菌Cytospora chrysosperma	20.30±2.36bc	40.02±0.59a
烟草赤星病菌链格孢Alternaria alternata	17.96±0.78d	32.80±0.78c
玉米大斑病菌Exserohilum turcicum	18.46±0.76cd	39.77±0.79a
甜瓜枯萎病菌尖孢镰刀菌Fusarium oxysporium	15.40±0.93e	21.28±1.17f
大豆炭疽病菌Colletotrichum truncatum	21.20±1.10b	37.61±1.55b
蓝莓溃疡病菌葡萄座腔菌Botryosphaeria dothidea	21.72±1.86b	39.84±0.67a
瓜果腐霉病菌Pythium aphanidermatum	14.94±1.07e	17.12±0.87g
水稻恶苗病菌串株镰刀菌Fusarium moniliforme	14.32±0.38e	29.15±0.59d
云杉立枯病菌尖孢镰刀菌Fusarium oxysporium	20.84±1.87b	29.90±1.05d
辣椒炭疽病菌Colletotrichum capsici	15.64±0.71e	33.70±0.44c
茄子褐纹病菌Phomopsis vexans	24.80±0.89a	36.28±0.63b
杨树溃疡病菌葡萄座腔菌Botryosphaeria dothidea	20.66±0.65b	40.26±0.51a

处理 Treatment	预防作用Prevention			治疗作用Curation
处理 Treatment	发病率 Rate of disease(%)	病情指数 Disease indexes	预防效果 Effect of Prevention (%)	发病率 Rate of disease(%)	病情指数 Disease indexes	预防效果 Effect of Prevention (%)
HS1	50.00±8.33bc	24.99±3.61b	60.87±5.66a	41.65±8.35c	18.05±2.40c	71.74±3.76a
多菌灵Carbendazim	61.31±9.64b	31.94±2.40b	50.00±3.76b	58.34±8.35bc	33.33±2.08b	47.83±3.26b
对照组CK	94.44±9.62a	63.89±3.19a	—	94.44±9.62a	63.89±3.19a	—

培养基 Medium	气生菌丝体 Aerial mycelium	基质菌丝体 Substrate mycelium	可溶性色素 Soluble pigment	丰茂度 Abundance
高氏一号琼脂 Gause No.1 agar	蚌肉白 Clam white	酪黄 Buttery yellow	-	+++
葡萄糖天门冬素 Glucose asparagine agar	荷花白 Lotus white	蚌肉白 Clam white	-	+++
蔗糖察氏 Czaper’s agar	粉白 Powder white	Ⅰ-3-1′	-	+
克氏合成一号琼脂 Carrot No.1 agar	蚌肉白 Clam white	豆汁黄 Soy sauce yellow	-	+++
马铃薯葡萄糖琼脂 Potato glucose agar	淡红灰 Light red gray	风帆黄 Sail yellow	鲛青 Cyan	+++
酵母膏麦芽膏琼脂 Yeast extract malt extract agar	Ⅳ-1-2′	豆汁黄 Soy sauce yellow	-	+++
淀粉铵琼脂 starch ammonium salt medium	Ⅰ-2-2′	莲子白 Lotus seed white	-	+++
贝奈特琼脂 Bennett’s agar	鹿角棕 Antler brown	鹅掌黄 Liriodendron yellow	-	++
葡萄糖酵母膏琼脂 Glucose yeast extract agar	荷花白 Lotus white	软木黄 Cork yellow	-	+++
甘油天冬素培养基 Glycerol asparagine agar	鲛青 Cyan	珠母灰 Pearlescent gray	Ⅶ-7-1′	++
甘油精氨酸培养基 Glycerine arginine medium	荷花白 Lotus white	粉白 Powder white	-	+++
黄豆粉蛋白胨培养基 Soybean powder peptone medium	Ⅳ-2-1′	粉白 Powder white	-	+++
燕麦粉琼脂 Oatemeal agar	Ⅲ-3-3′	香水玫瑰黄 Perfume rose yellow	-	+++
淀粉琼脂 Soluble starch agar	乳白 Milky	淡蜜黄 Light honey yellow	鹅掌黄 Liriodendron yellow	++
黄豆粉琼脂培养基 Bean agar medium	Ⅳ-3-2′	蚌肉白 Clam white	-	+++

pH	丰茂度 Abundance	NaCl(%)	丰茂度 Abundance	温度 Temperature/℃	丰茂度 Abundance
2	-	0	+	4	+
5	++	10	++	15	++
7	+++	20	++	28	+++
9	++	30	+++	37	-
12	+	40	++	50	-