Identification and 16S rRNA gene diversity analysis of salt cedar(Tamarix chinensis)witches'-broom

doi:10.6048/j.issn.1001-4330.2023.10.025

Xinjiang Agricultural Sciences ›› 2023, Vol. 60 ›› Issue (10): 2551-2557.DOI: 10.6048/j.issn.1001-4330.2023.10.025

• Plant Protection·Microbes·Soil Fertilizer • Previous Articles Next Articles

Identification and 16S rRNA gene diversity analysis of salt cedar(Tamarix chinensis)witches'-broom

LI Feng¹(), LAI Ganggang¹^,², ZHAO Zhihui¹, CHEN Xiaofei¹, ZHU Tiansheng¹()

1. Key Laboratory of Integrated Pest Management (IPM) of Xinjiang Production and Construction Corps in Southern Xinjiang/ Scientific Observing and Experimental Station of Crop Pests in Alar, Ministry of Agriculture, China/ College of Agronomy, Tarim University, Aral Xinjiang 843300, China
2. Institute of Agriculture Sciences, the Fourth Agriculture Production Division of Xinjiang Production and Construction Crops, Cocodala Xinjiang 835219, China

Received:2023-01-13 Online:2023-10-20 Published:2023-11-01
Correspondence author: ZHU Tiansheng (1974-),male, native place: Pingliang, Gansu Province. professor, research field: plant pathology (E-mail)ztszky@163.com
Supported by:
National Natural Science Foundation of China(31060238);The Open Project of the XPCC's Characteristic Fruit Tree Production Engineering Laboratory(FE202001);Science and Technology R & D Program Projects of XPCC(2019DA001);Science and Technology R & D Program Projects of XPCC(2020DA003)

柽柳丛枝植原体鉴定及16S rRNA基因多样性分析

李丰¹(), 赖刚刚¹^,², 赵志惠¹, 陈小飞¹, 朱天生¹()

1.塔里木大学农学院/南疆农业有害生物综合治理兵团重点实验室/农业农村部阿拉尔作物有害生物科学观测实验站,新疆阿拉尔 843300
2.新疆生产建设兵团第四师农业科学研究所,新疆可克达拉 835219

通讯作者: 朱天生(1974-),男,甘肃平凉人,教授,硕士生导师,研究方向为植物病理,(E-mail)ztszky@163.com
作者简介:李丰(1998-),男,甘肃定西人,硕士研究生,研究方向为植物病理,(E-mail)2925008071@qq.com
基金资助:
国家自然科学基金项目(31060238);新疆生产建设兵团特色果树生产工程实验室开放课题(FE202001);新疆生产建设兵团财政科技计划项目(2019DA001);新疆生产建设兵团财政科技计划项目(2020DA003)

Abstract

Abstract:

【Objective】 To clarify the distribution, taxonomic status, morphological characteristics and genetic differentiation of 16S rRNA of salt cedar witches'-broom disease in southern Xinjiang.【Methods】 In this study, the distribution and damage of salt cedar witches'-broom(SCWB) disease in southern Xinjiang were investigated and the pathogens of SCWB was observed by transmission electron microscope, 16S rRNA gene and rp gene amplification. Meanwhile, the molecular identification of the gene was carried out to clarify its morphological characteristics and taxonomic status, and the genetic differentiation of the 16S rRNA gene of SCWB in different areas of southern Xinjiang was studied.【Results】 The average incidence of SCWB in southern Xinjiang was 9.06%, and the average disease index was 5.66. The existence of phytoplasma granules was observed in the phloem tissue of tamarisk branches. Nested PCR of 16S rRNA and rp gene bands with sizes of 1,219 bp and 1,174 bp were obtained respectively. Through homology comparison and phylogenetic tree construction, it was confirmed that the SCWB belonged to the phytoplasma disease, and belonged to the phytoplasma 16SrXXX-A subgroup in terms of taxonomic status; SCWB phytoplasma could be mainly divided into four lines, and the nucleotide similarity of 16S rRNA gene was between 96.5% and 99.8% in southern Xinjiang.【Conclusion】 In this study, a systematic study of the phytoplasma disease of SCWB is carried out, the taxonomic status of the pathogen, the distribution and damage of the disease in southern Xinjiang are clarified, and the genetic differentiation of its 16S rRNA gene is analyzed. The rp gene i amplified to increase the content of phytoplasma diseases in the 16SrXXX group, which has provided a theoretical basis for the detection and identification of the disease.

Key words: salt cedar witches'-broom; 16S rRNA; rp; transmission electron microscope; sequence analysis; diversity analysis

摘要：

【目的】 研究柽柳丛枝病在新疆南疆地区的分布危害、分类、形态特征及病原物16S rRNA基因的遗传多样性,为该病害的检测和鉴定提供理论依据。【方法】 调查新疆南疆地区柽柳丛枝病的分布及危害,采用形态学与分子生物学相结合的方法,对柽柳丛枝病病原物进行透射电镜观察、16S rRNA基因和rp基因扩增,研究其形态特征及分类地位,并对新疆南疆不同地区柽柳丛枝病病原物16S rRNA基因遗传多样性。【结果】 柽柳丛枝病在新疆南疆地区的平均发病率为9.06%,平均病情指数为5.66;在柽柳丛枝韧皮部组织中观察到有植原体颗粒的存在;16S rRNA基因和rp基因巢式PCR分别获得1 219 bp和1 174 bp大小的条带,柽柳丛枝病属于植原体病害,在分类地位上属于植原体16SrXXX-A亚组;新疆南疆柽柳丛枝植原体主要可分为4个株系,16S rRNA基因核苷酸相似度在96.5%~99.8%。【结论】 柽柳丛枝病在新疆南疆地区主要分布在阿克苏市、温宿县、巴楚县、图木舒克市、阿拉尔市、伽师县,阿拉尔市12团等地,且发生情况和危害程度均较轻,不同地区之间的发病率及病情指数存在显著差异性;柽柳丛枝病存在植原体颗粒并主要在韧皮部组织中,有球形、椭圆形和不规则形态,大小300~600 nm不等。

关键词: 柽柳丛枝植病, 16S rRNA基因, rp基因, 透射电镜, 序列分析, 多样性

CLC Number:

S432

LI Feng, LAI Ganggang, ZHAO Zhihui, CHEN Xiaofei, ZHU Tiansheng. Identification and 16S rRNA gene diversity analysis of salt cedar(Tamarix chinensis)witches'-broom[J]. Xinjiang Agricultural Sciences, 2023, 60(10): 2551-2557.

李丰, 赖刚刚, 赵志惠, 陈小飞, 朱天生. 柽柳丛枝植原体鉴定及16S rRNA基因多样性分析[J]. 新疆农业科学, 2023, 60(10): 2551-2557.

Figures/Tables 8

References 18

[1]	孙姗姗, 刘新平, 何玉惠, 等. 我国固沙植物资源化利用研究进展[J]. 世界林业研究. 2020, 33(6): 74-79.
	SUN Shanshan, LIU Xinping, HE Yuhui, et al. Research Advance in Sand-fixing Plant Resources Utilization in China[J]. World Forestry Research, 2020, 33(6):74-79.
[2]	Doi R, Shirasaki A, Sasa M. The mode of development of Japanese encephalitis virus in the mosquito Culex tritaeniorhynchus summorosus as observed by the fluorescent antibody technique[J]. The Japanese Journal of Experimental Medicine, 1967, 37(3).
[3]	耿显胜, 舒金平, 王浩杰, 等. 植原体病害的传播、流行和防治研究进展[J]. 中国农学通报. 2015, 31(25):164-170. DOI
	GENG Xiansheng, SHU Jinping, WANG Haojie, et al. Research Advance on Transmission, Epidemic and Control of Phytoplasma Disease[J]. Chinese Agricultural Science Bulletin, 2015, 31(25):164-170.
[4]	徐平洛, 翟晓巧. 植物植原体病害研究进展[J]. 河南林业科技. 2020, 40(4):20-23.
	XU Pingluo, ZHAI Xiaoqiao. Research Advances in Plant Phytoplasma Diseases[J]. Journal of Henan Forestry Science and Technology, 2020, 40(4):20-23.
[5]	Wei W, Davis R E, Lee I, et al. Computer-simulated RFLP analysis of 16S rRNA genes:identification of ten new phytoplasma groups[J]. International Journal of Systematic and Evolutionary Microbiology, 2007, 57(8):1855. DOI URL
[6]	杨毅, 姜蕾, 李世访. 植原体分类鉴定研究进展[J]. 植物检疫. 2020, 34(5):13-20.
	YANG Yi, JIANG Lei, LI Shifang. Research progress on phytoplasma identification and classification[J]. Plant Quarantine, 2020, 34(5):13-20.
[7]	刘仲健, 罗焕亮, 张景宁. 植原体病理学[M]. 北京: 中国林业出版社,1999.
	LIU Zhongjian, LUO Huanliang, ZHANG Jingning. Phytoplasma Phytopathology[M]. Beijing: China Forestry Press, 1999.
[8]	Zhao Y, Sun Q, Davis, et al. New Witches'-Broom Diseases in Northern China and Their Association with Diverse Phytoplasmas[J]. Phytopathology, 2005,(95):S117-S118.
[9]	Zhao Y, Sun Q, Wei W, et al. 'Candidatus Phytoplasma tamaricis',a novel taxon discovered in witches'-broom-diseased salt cedar (Tamarix chinensis Lour.)[J]. International Journal of Systematic Bacteriology, 2009, 59(10):2496-2504.
[10]	李成亮. 新疆六种植原体病害的检测与鉴定[D]. 石河子: 石河子大学, 2014.
	LI Chengliang. Detection and Identification of Six Phytoplasmal Diseases in Xinjiang[D]. Shihezi: Shihezi University, 2014.
[11]	赖刚刚. 南疆柳树植原体病的调查及病原鉴定[D]. 阿拉尔: 塔里木大学, 2021.
	LAI Ganggang. Investigation and identification of Willow phytoplasma diseases in Southern Xinjiang[D]. Aral: Tarim University, 2021.
[12]	韩冰. 新疆杏褪绿卷叶植原体的检测与鉴定[D]. 乌鲁木齐: 新疆农业大学, 2012.
	HAN Bing. Detection and Identification of Apricot Chlorotic Leafroll Phytoplasma in Xinjiang[D]. Urumqi: Xinjiang Agricultural University, 2012.
[13]	李正男. 西北地区常见植物植原体病害分子鉴定及株系多样性研究[D]. 杨凌: 西北农林科技大学, 2015.
	LI Zhengnan. Molecular diagnosis of phytoplasma diseases on common plants in Northwest China and diservity study of the associated phytoplasma[D]. Yangling: Northwest A&F University, 2015.
[14]	漆艳香, 谢艺贤, 张辉强, 等. 植原体DNA提取方法的改良[J]. 生物技术通报. 2004,(4): 44-46.
	QI Yanxiang, XIE Yixian, ZHANG Huiqiang, et al. A Improved Method for Extracting Phytoplasma DNA[J]. Biotechnology Bulletin, 2004,(4):44-46.
[15]	Lee I M, Gundersen-Rindal D E, Davis R E, et al. Revised classification scheme of phytoplasmas based on RFLP analyses of 16s rRNA and ribosomal protein gene sequences[J]. International Journal of Systematic Bacteriology, 1998,(48):1153-1169.
[16]	Gundersen D E, Lee I M. Ultrasensitive detection of phytoplasmas by nested-PCR assays using two universal primer pairs[J]. Phytopathologia Mediterranea. 1996, 35(3):144-151.
[17]	Kumar S, Stecher G, Li M, et al. MEGA X:Molecular Evolutionary Genetics Analysis across Computing Platforms[J]. Molecular Biology and Evolution, 2018, 35(6):1547-1549. DOI URL
[18]	Li F, Lai G G, Zhao Z H, et al. Identification of 16S rXXX-A Phytoplasma Associated with Salt Cedar (Tamarix chinensis) Witches’-Broom in Southern Xinjiang of China[J]. Plant Disease, 2021, 105(12):4146.

等级 Grade	分级标准 Standard	柽柳感病情况 Susceptibility of tamarix chinensis
0	健康	未发病
I	轻度	开始发病,不显著
Ⅱ	中度	病枝数量达总枝数量的1/3, 其余枝条正常
Ⅲ	较严重	病枝数量达总枝数量的2/3, 尚有正常枝条
IV	严重	病枝数量达总枝数量的2/3以上, 正常枝条较少

等级 Grade	分级标准 Standard	柽柳感病情况 Susceptibility of tamarix chinensis
0	健康	未发病
I	轻度	开始发病,不显著
Ⅱ	中度	病枝数量达总枝数量的1/3, 其余枝条正常
Ⅲ	较严重	病枝数量达总枝数量的2/3, 尚有正常枝条
IV	严重	病枝数量达总枝数量的2/3以上, 正常枝条较少

调查地区 Survey area	调查总数 Total number of investigations	发病总数 Total number of cases	平均发病率 Average incidence (%)	平均病情指数 Mean disease index
温宿县 Wensu	116	22	19.08^a	3.45^a
阿拉尔市12团 Aral city 12th regiment	390	72	18.20^ab	8.25^b
图木舒克市 Tumxuk	153	24	15.71^bc	5.47^b
巴楚县 Bachu	294	47	15.54^bc	3.18^b
阿拉尔市 Aral	271	36	13.40^d	4.172^c
伽师县 Jiashi	179	20	11.18^d	3.69^c
阿克苏市 Akesu	206	14	6.60^e	7.56^c
昆玉市 Kunyu	267	0	0.00^f	0.00^d
和田地区 Hetian	858	235	0.00^f	0.00^d
平均 Average			9.06	5.66

调查地区 Survey area	调查总数 Total number of investigations	发病总数 Total number of cases	平均发病率 Average incidence (%)	平均病情指数 Mean disease index
温宿县 Wensu	116	22	19.08^a	3.45^a
阿拉尔市12团 Aral city 12th regiment	390	72	18.20^ab	8.25^b
图木舒克市 Tumxuk	153	24	15.71^bc	5.47^b
巴楚县 Bachu	294	47	15.54^bc	3.18^b
阿拉尔市 Aral	271	36	13.40^d	4.172^c
伽师县 Jiashi	179	20	11.18^d	3.69^c
阿克苏市 Akesu	206	14	6.60^e	7.56^c
昆玉市 Kunyu	267	0	0.00^f	0.00^d
和田地区 Hetian	858	235	0.00^f	0.00^d
平均 Average			9.06	5.66

登录号 Login Number	OL687359	OL687355	OL687360	OL687364	OL687373	OL687370	OL687381
OL687359	100.0	2.0	2.1	2.4	2.0	2.1	3.6
OL687355	98.0	100.0	0.4	1.8	0.4	0.3	2.6
OL687360	97.9	99.6	100.0	1.7	0.3	0.2	2.7
OL687364	97.7	98.2	98.3	100.0	1.6	1.7	3.3
OL687373	98.0	99.6	99.7	98.4	100.0	0.3	2.7
OL687370	97.9	99.7	99.8	98.3	99.8	100.0	2.5
OL687381	96.5	97.5	97.3	96.8	97.4	97.6	100.0

Identification and 16S rRNA gene diversity analysis of salt cedar(Tamarix chinensis)witches'-broom

柽柳丛枝植原体鉴定及16S rRNA基因多样性分析

RichHTML

PDF

Knowledge

Abstract

Cite this article

share this article

Figures/Tables 8

References 18

Related Articles 15

Recommended Articles 0

Metrics

[1]	AN Zhe, NIU Ruichang, ZHU Xiangzhen, WANG Li, ZHANG Kaixin, LI Dongyang, JI Jichao, NIU Lin, GAO Xueke, LUO Junyu, CUI Jinjie, MA Deying. Analyze the microbial diversity of cotton aphids with different bacterial types in cotton fields [J]. Xinjiang Agricultural Sciences, 2024, 61(9): 2277-2284.
[2]	JIN Juan, Subina Xiaokelaiti, Abudoukayoumu Ayimaiti, YANG Lei, HAO Qing, FAN Dingyu. Cloning and sequence analysis of expansin gene ZjEXPA8 in Huizao [J]. Xinjiang Agricultural Sciences, 2023, 60(9): 2223-2230.
[3]	WANG Man, ZHANG Zheng, Yilidana Dilixiati, WU Bin. Cloning and bioinformatics analysis of VvGST1 from Munage tab.grapes [J]. Xinjiang Agricultural Sciences, 2023, 60(8): 1922-1930.
[4]	Nasibai Abuduwahapu, GAO Xiaojuan, Kunduziayi Abudushalamu, LI Jiangwei. Cloning and Sequence Analysis of cDNA of C3d Gene of Bactrian Camel from Xinjiang [J]. Xinjiang Agricultural Sciences, 2023, 60(2): 493-500.
[5]	WANG Zhong, FAN Zheru, ZHANG Yueqiang, LI Jianfeng, GAO Xin, SHI Jia. Cloning and Expression Analysis of Chlorophyll a Oxygenase Gene in Wheat [J]. Xinjiang Agricultural Sciences, 2021, 58(9): 1569-1576.
[6]	YANG Bo, CHE Yuhong, GUO Chunmiao, Mubareke·Ayoupu, WU Jingrong, DU Juan. Cloning and bioinformation analysis of 4CL gene in Cydonia oblonga Mill [J]. Xinjiang Agricultural Sciences, 2021, 58(6): 1055-1063.
[7]	ZHANG Guoru, TANG Yaping, YANG Tao, Patiguri Esmutola, WANG Baike, LI Ning, WANG Juan, YU Qinghui, YANG Shengbao. Agronomic Character Analysis of 272 Capsicum Resources [J]. Xinjiang Agricultural Sciences, 2021, 58(12): 2300-2311.
[8]	CHE Yuhong, YANG Bo, GUO Chunmiao, Mubareke Ayoupu, WU Jinrun, DU Juan. Cloning, Sequence Analysis and Expression of CAD Gene in Quince (Cydonia oblonga Mill) Fruit [J]. Xinjiang Agricultural Sciences, 2020, 57(5): 814-821.
[9]	ZHU Zong-cai, ZHANG Wang-bin, LI Ya-peng, YAN Hai-lin, YI Zi-bo. Study on the Cross Infection of Rot Pathogens in Different Hosts of Orchard in Xinjiang [J]. Xinjiang Agricultural Sciences, 2019, 56(6): 1112-1121.
[10]	SUN Liang, WEN Feng, LIU Feng, ZHANG Xin-yu, SUN Jie. Cloning and Sequence Analysis of 5'UTR Intron of GhFAD2-1 Gene from Cotton (Gossypium hirsutum L.) [J]. Xinjiang Agricultural Sciences, 2018, 55(1): 1-8.
[11]	ZHANG Lu-yao, LI Fei, JIAO Hai-hong, ZHAO Li, LIU Yong-hong. Genetic Evolution Analysis of F gene of PPRV in Southern Xinjiang [J]. Xinjiang Agricultural Sciences, 2017, 54(11): 2118-2129.
[12]	. Etiological Detection and Analysis of Paratuberculosis in Sheep [J]. , 2016, 53(8): 1562-1568.
[13]	WU Peng;ZHANG Xun;LIANG Tian;CHEN Xin-kai;XIAO Yuan-yuan;SHENG Jin-liang. PRRSV ORF7 Cloning and Genetic Evolution Analysis of Pigs in Northern Xinjiang [J]. , 2016, 53(11): 2149-2156.
[14]	WEI Jie;XUE Ying;MIAO Shu-kui;DOU Xiao-long;REN Xiao-bing;MA Wen-ge;HUANG Jiong. Isolation and Identification of Sheep Pox Virus Isolated from Xinjiang and Analysis of the Sequence of Structural Protein P32 [J]. , 2015, 52(4): 732-739.
[15]	SUN Xiao-xia;WANG Yu-ting;NIU Jian-xin. Identification and sequence analysis of the Apple scar skin viroid isolated from apple trees in Xinjiang Province [J]. , 2015, 52(4): 693-698.