新疆拜城县马铃薯疮痂病病原菌的分离鉴定及生物学特性分析

doi:10.6048/j.issn.1001-4330.2023.09.021

新疆农业科学 ›› 2023, Vol. 60 ›› Issue (9): 2258-2265.DOI: 10.6048/j.issn.1001-4330.2023.09.021

新疆拜城县马铃薯疮痂病病原菌的分离鉴定及生物学特性分析

徐李娟¹^,²(), 陈勇³, 王则玉³, 王博², 艾尼江·尔斯满², 郭瑞¹, 李克梅¹(), 宋素琴²^,⁴()

1.新疆农业大学农学院,乌鲁木齐 830052
2.新疆农业科学院微生物应用研究所/新疆特殊环境微生物重点实验室,乌鲁木齐 830091
3.新疆农业科学院土壤肥料与农业节水研究所,乌鲁木齐 830091
4.新疆绿洲农业病虫害治理与植保资源利用自治区高校重点实验室/石河子大学农学院,新疆石河子 832003

收稿日期:2022-12-15 出版日期:2023-09-20 发布日期:2023-09-19
通信作者: 李克梅 (1972-),女,江苏如皋人,教授,硕士生导师,研究方向为牧草病害及防治,(E-mail)likemei@xjau.edu.cn；宋素琴(1976-),女,山西长治人,研究员,研究方向为微生物活性产物及作物病害,(E-mail)suqin_song@163.com
作者简介:徐李娟(1997-),女,河南鹤壁人,硕士研究生,研究方向为马铃薯病原学及生物防治, (E-mail)2462300072@qq.com
基金资助:
新疆维吾尔自治区重点研发项目任务专项“新疆马铃薯绿色高产高效栽培技术研究与集成应用”(2022B02044-2);新疆农业科学院科技创新重点培育专项“马铃薯疮痂病病原菌致病毒素thaxtomins产量差异分析及其对新疆农田杂草的除草性能研究”(xjkcpy-202203);农业部西北荒漠绿洲作物有害生物综合治理重点实验室开放基金课题“新疆马铃薯疮痂病病原菌Steptomyces spp.多样性及其致病力分化研究”(KFJJ202006)

Isolation, identification and biological characteristics of potato scab pathogen in Baicheng, Xinjiang

XU Lijuan¹^,²(), CHEN Yong³, WANG Zeyu³, WANG Bo², Ainijiang Ersiman², GUO Rui¹, LI Kemei¹(), SONG Suqin²^,⁴()

1. College of Agriculture, Xinjiang Agricultural University, Urumqi 830052,China
2. Institute of Microbiology Application, Xinjiang Academy of Agricultural Sciences/Xinjiang key Laboratory of Special Environmental Microorganisms, Urumqi 830091,China
3. Institute of soil,Fertilizer and Agricultural Water Conservation,Xinjiang Academy of Agricultural Aciences,Urumqi 830091,China
4. College of Agriculture,Shihezi University/Key Laboratory at the Universities of Xinjiang Uygur Autonomous Region for Oasis Agricultural Pest Management and Plant Protection Resource Utilization,Shihezi Xinjiang 832003,China

Received:2022-12-15 Online:2023-09-20 Published:2023-09-19
Correspondence author: SONG Suqin(1976-),female, Changzhi County, Shanxi Province, associate researcher, major in microbial active products and crop diseases,(E-mail)suqin_song@163.com; LI Kemei (1972-),female,professor, major in grass disease and control,(E-mail)likemei@xjau.edu.cn
Supported by:
Research and integrated application of green high-yield and high-efficiency cultivation technology of potato in Xinjiang(2022B02044-2);Key S & T Innovation Incubation Project of Xinjiang Academy of Agricultural Sciences "Analysis on Thaxtomins Yield of Pathogenic Toxin of Potato Pathogen and Weeds in Xinjiang"(xjkcpy-202203);Open Fund Sponsored by the Key Laboratory of Comprehensive Treatment of Northwest Desert Oasis of the Ministry of Agriculture and Rural Affaris "Study on Steptomyces spp. Diversity and Pathogenicity Differentiation of Potato Scab Pathogen in Xinjiang"(KFJJ202006)

摘要/Abstract

摘要：

【目的】研究新疆阿克苏地区拜城县马铃薯疮痂病病原菌的种类。【方法】采集新疆阿克苏地区拜城县3个不同种植区的马铃薯疮痂病块茎及薯表土壤,采用组织分离法和稀释涂布分离法,并通过致病性测定、形态学、分子生物学及生理生化特性对病原菌进行鉴定。【结果】在分离出的72株放线菌中,菌株K6含致病基因txtAB、 tomA、 nec1且对萝卜幼苗有较强致病性;经鉴定为同一种链霉菌属Streptomyces bobili,为新疆首次报道的病原菌。所有供试碳源、氮源均可作为单一碳源、氮源被菌株S.bobili K6利用,在酪氨酸琼脂培养基上有黑色素产生,对6%NaCl、0.1%苯酚、20 μg/mL链霉素敏感,在pH4的条件下不能正常生长。【结论】新疆拜城县马铃薯疮痂病病原菌为S.bobili,包含3种致病基因txtAB、 tomA、 nec1,且致病性较强。

关键词: 马铃薯疮痂病病原菌; 链霉菌; 分离鉴定; 生物学特性

Abstract:

【Objective】 To clarify the species of potato common scab pathogenic bacteria in Baicheng County, Aksu Prefecture, Xinjiang.【Methods】 Potato common scab tubers and potato surface soil were collected from three different growing areas in Baicheng County, Aksu Prefecture, Xinjiang, and the pathogenic bacteria were identified by tissue isolation method and dilution coating isolation method, and by pathogenicity determination, morphology, molecular biology and physiological and biochemical characteristics.【Results】 Among the 72 isolated actinomycetes, six strains contained the pathogenic genes txtAB, tomA and nec1 and were strongly pathogenic to radish seedlings; they were identified as the same Streptomyces bobili, which was the first reported pathogen in Xinjiang.All the tested carbon and nitrogen sources were available as single carbon and nitrogen sources by strain S.bobili K6, with melanin production on complex agar medium, sensitive to 6% NaCl, 0.1% phenol, 20 μg/mL streptomycin, and not growing any more at pH 4 normally.【Conclusion】 The pathogenic of potato common scab is S.bobili in Baicheng County of Xinjiang, which contains three pathogenic genes txtAB, tomA, and nec1, and belongs to highly pathogenicity.

Key words: potato common scab disease in Baicheng of Xinjiang; Streptomyces; isolation and identification; biological characteristics

中图分类号:

S436.32

徐李娟, 陈勇, 王则玉, 王博, 艾尼江·尔斯满, 郭瑞, 李克梅, 宋素琴. 新疆拜城县马铃薯疮痂病病原菌的分离鉴定及生物学特性分析[J]. 新疆农业科学, 2023, 60(9): 2258-2265.

XU Lijuan, CHEN Yong, WANG Zeyu, WANG Bo, Ainijiang Ersiman, GUO Rui, LI Kemei, SONG Suqin. Isolation, identification and biological characteristics of potato scab pathogen in Baicheng, Xinjiang[J]. Xinjiang Agricultural Sciences, 2023, 60(9): 2258-2265.

图/表 10

参考文献 32

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体系成分 System composition	txtAB 反应体系 txtAB reaction system	nec1 反应体系 nec1 reaction system	tomA 反应体系 tomA reaction system
2×Es Taq Master Mix	20 μL	20 μL	20 μL
上游引物 Upstream primer	0.6 μL	0.4 μL	0.4 μL
下游引物 Downstream primer	0.6 μL	0.4 μL	0.4 μL
ddH₂O	12.8 μL	12.8 μL	12.8 μL
DNA模板 DNA templates	1 μL	1 μL	1 μL

体系成分 System composition	txtAB 反应体系 txtAB reaction system	nec1 反应体系 nec1 reaction system	tomA 反应体系 tomA reaction system
2×Es Taq Master Mix	20 μL	20 μL	20 μL
上游引物 Upstream primer	0.6 μL	0.4 μL	0.4 μL
下游引物 Downstream primer	0.6 μL	0.4 μL	0.4 μL
ddH₂O	12.8 μL	12.8 μL	12.8 μL
DNA模板 DNA templates	1 μL	1 μL	1 μL

扩增基因 Amplified gene	引物 Primer	引物序列 Primer sequence	片段大小 Fragment size(bp)	参考文献 Reference
16S rRNA	PA PH	5'- TTTGATCCTGGCTCAG -3' 5'- AAGGAGGTGATCCAGCCGCA -3'	1 500	^[30]
txtAB	Txt AB1 Txt AB2	5'-CCACCAGGACCTGCTCTTC-3' 5'-TCGAGTGGACCTCACAGATG-3'	385	^[6]
nec1	Nf Nr	5'-ATGAGCGCGAACGGAAGCCCCGGA-3' 5'-GCAGGTCGTCACGAAGGATCG-3'	700	^[4]
tomA	Tom3 Tom4	5'-GAGGCGTTGGTGGAGTTCTA-3' 5'-TTGGGGTTGTACTCCTCGTC-3'	392	^[6]

扩增基因 Amplified gene	引物 Primer	引物序列 Primer sequence	片段大小 Fragment size(bp)	参考文献 Reference
16S rRNA	PA PH	5'- TTTGATCCTGGCTCAG -3' 5'- AAGGAGGTGATCCAGCCGCA -3'	1 500	^[30]
txtAB	Txt AB1 Txt AB2	5'-CCACCAGGACCTGCTCTTC-3' 5'-TCGAGTGGACCTCACAGATG-3'	385	^[6]
nec1	Nf Nr	5'-ATGAGCGCGAACGGAAGCCCCGGA-3' 5'-GCAGGTCGTCACGAAGGATCG-3'	700	^[4]
tomA	Tom3 Tom4	5'-GAGGCGTTGGTGGAGTTCTA-3' 5'-TTGGGGTTGTACTCCTCGTC-3'	392	^[6]

条件 Condition	txtAB (385 bp)	nec1 (700 bp)	tomA (392 bp)
预变性 Pre-denaturation	95℃ 3 min	95℃ 3 min	95℃ 3 min
变性 Degeneration	95℃ 30 s	95℃ 30 s	95℃ 30 s
退火 Annealing	51℃ 30 s	62℃ 30 s	57℃ 30 s
延伸 Elongation	72℃ 30 s	72℃ 30 s	72℃ 30 s
再延伸 Re-extension	72℃ 7 min	72℃ 7 min	72℃ 7 min
循环 Cycle	30次	30次	30次

新疆拜城县马铃薯疮痂病病原菌的分离鉴定及生物学特性分析

Isolation, identification and biological characteristics of potato scab pathogen in Baicheng, Xinjiang

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图/表 10

参考文献 32

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Metrics

指标特征 Index characteristics	K6	指标特征 Index characteristics	K6
D-葡萄糖 D-glucose	+	1% NaCl	+
D-甘露醇 D-mannitol	+	3% NaCl	+
棉子糖 Raffinose	+	6% NaCl	-
α-乳糖 α-lactose	+	pH=4	+
D-果糖 D-fructose	+	pH=5	+
海藻糖 Alginic sugar	+	pH=6	+
山梨醇 Glucitol	+	pH=7	+
L-阿拉伯糖 L-arabinose	+	pH=8	+
肌醇 Inositol	+	pH=9	+
甲硫氨酸 Methionine	+	淀粉水解 Amylohydrolysis	+
酵母粉 Yeast extract	+	H₂S产生 H₂S production	+
硫酸铵 Ammonium sulfate	+	苯酚 Phenol (0.1%)	-
组氨酸 Histidine	+	青霉素 Penicillin(10 U/mL)	+
黑色素 Nigrosin	+	链霉素 Streptomycin(20 μg/mL)	-

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