Isolation and identification of endophytic bacteria from soybean nodule and their effects on soybean plant

doi:10.3969/j.issn.1004-1524.20221120

Acta Agriculturae Zhejiangensis ›› 2023, Vol. 35 ›› Issue (7): 1532-1541.DOI: 10.3969/j.issn.1004-1524.20221120

• Crop Science • Previous Articles Next Articles

Isolation and identification of endophytic bacteria from soybean nodule and their effects on soybean plant

SUN Xiujuan(), XU Weihui, WANG Zhigang()

College of Life Science and Agriculture and Forestry, Qiqihar University, Qiqihar 161006, Heilongjiang, China

Received:2022-08-02 Online:2023-07-25 Published:2023-08-17
Contact: WANG Zhigang

Abstract

Abstract:

To study the growth-promoting effect of endophytic bacteria in soybean root nodules on soybean growth, soybean root nodules were used as experimental materials to isolate and screen nodule endophytic bacteria, and their ability to secrete plant hormones and their promotion effect on soybean plant height, stem diameter, dry weight, fresh weight, and root growth were measured. The results showed that a total of 17 endophytic bacteria were screened out from root nodules. After physiological and biochemical analysis and 16S rDNA identification, 10 strains belonged to Bacillus spp., 3 strains belonged to Priestia spp, and the remaining 4 strains belonged to Enterobacteriaceae, Klebsiella, Acinetobacter, and Agrobacterium. Through hemolysis test and 16S rDNA identification, 10 strains with strong growth-promoting ability were selected. Among them, the IAA secretion of Klebsiella S8 was highest at 147.55 mg·L^-1; the gibberellins of Agrobacterium S10 had the highest secretion at 39.34 mg·L^-1; the siderophores activity of Bacillus S3 was the highest at 70.57%. The plant height, root length, dry fresh weight and stem diameter of soybean treated by these 10 strains were significantly higher than those of the control. In conclusion, the endophytic bacteria screened in this experiment had strong growth-promoting effect on soybean growth.

Key words: root nodule, endophytic bacterium, soybean, hemolysis test

CLC Number:

SUN Xiujuan, XU Weihui, WANG Zhigang. Isolation and identification of endophytic bacteria from soybean nodule and their effects on soybean plant[J]. Acta Agriculturae Zhejiangensis, 2023, 35(7): 1532-1541.

Figures/Tables 5

Fig.1 Phylogenetic tree of endophytic bacteria based on 16S rDNA sequences

Table 1 Physiological and biochemical characteristics of endophytic bacteria

菌株编号 Strain No.	淀粉水解试验 Starch hydrolysis test	油脂水解试验 Oil hydrolysis test	乳糖发酵试验 Lactose fermentation test	葡萄糖发酵试验 Glucose fermentation test
S1	+	-	+	+
S2	+	-	+	+
S3	+	-	+	+
S4	-	+	-	+
S5	+	-	+	-
S6	+	-	-	-
S7	-	+	+	+
S8	-	+	+	+
S9	-	+	-	+
S10	-	+	+	-

Fig.2 Ability of the tested strains to secrete IAA, gibberellin and siderophores Data on the bars marked without the same lowercase letter indicated significant differences at P<0.05. The same as below.

Fig.3 Growth promoting effect of endophytic bacteria on soybean seed The three seedling growth plots are one treatment, from left to right are CK, diluted 10 times, 20 times and 30 times.

Fig.4 Effect of endophytic bacteria on growth promotion of soybean plants and comprehensive ability evaluation F, Evaluation of comprehensive ability.

References 35

[1]	杨红旗, 郝仰坤. 我国大豆产业回顾、现状与发展对策[J]. 广东农业科学, 2010, 37(1): 188-191.
	YANG H Q, HAO Y K. Historical review, current situation and development countermeasure of Chinese soybean[J]. Guangdong Agricultural Sciences, 2010, 37(1): 188-191. (in Chinese with English abstract)
[2]	黄国东, 宋清晖, 王晓慧, 等. 含枯草芽孢杆菌肥料对大豆叶片光合作用及土壤酶活性的影响[J]. 土壤与作物, 2021, 10(1): 99-107.
	HUANG G D, SONG Q H, WANG X H, et al. Effects of fertilizers containing Bacillus subtilis on photosynthesis of soybean leaves and soil enzyme activities[J]. Soils and Crops, 2021, 10(1): 99-107. (in Chinese with English abstract)
[3]	柏琼芝, 肖石江, 王晓瑞, 等. 化肥减量配施生物有机肥对秋马铃薯产量的影响[J]. 土壤与作物, 2019, 8(2): 158-165.
	BAI Q Z, XIAO S J, WANG X R, et al. Effect of reduced chemical fertilizer plus biological fertilizer on autumn potato yield[J]. Soils and Crops, 2019, 8(2): 158-165. (in Chinese with English abstract)
[4]	刘江苇, 刘颖, 徐婷, 等. 水稻内生菌研究进展及展望[J]. 生命科学研究, 2021, 25(3): 232-239.
	LIU J W, LIU Y, XU T, et al. Advances and prospects of rice endophytes[J]. Life Science Research, 2021, 25(3): 232-239. (in Chinese with English abstract)
[5]	VARDHARAJULA S, SKZ A, KRISHNA PRASAD VURUKONDA S S, et al. Plant growth promoting endophytes and their interaction with plants to alleviate abiotic stress[J]. Current Biotechnology, 2017, 6(3): 252-263.
[6]	PEIX A, RAMÍREZ-BAHENA M H, VELÁZQUEZ E, et al. Bacterial associations with legumes[J]. Critical Reviews in Plant Sciences, 2015, 34(1/2/3): 17-42.
[7]	ZHAO L F, XU Y J, SUN R, et al. Identification and characterization of the endophytic plant growth prompter Bacillus Cereus strain mq23 isolated from Sophora Alopecuroides root nodules[J]. Brazilian Journal of Microbiology, 2011, 42(2): 567-575.
[8]	ZHAO L F, DENG Z S, YANG W Q, et al. Diverse rhizobia associated with Sophora alopecuroides grown in different regions of Loess Plateau in China[J]. Systematic and Applied Microbiology, 2010, 33(8): 468-477.
[9]	赵龙飞. 我国大豆根瘤内生菌资源多样性研究[J]. 广东农业科学, 2014, 41(9): 15-19.
	ZHAO L F. Research on resource diversity of soybean [Glycine max(L.) Merr] nodule endophytes in China[J]. Guangdong Agricultural Sciences, 2014, 41(9): 15-19. (in Chinese with English abstract)
[10]	钟宇舟, 余秀梅, 陈强, 等. 四川盆地大豆根瘤内生细菌的分离鉴定及促生效果[J]. 应用与环境生物学报, 2017, 23(1): 46-53.
	ZHONG Y Z, YU X M, CHEN Q, et al. Isolation, identification and plant growth promotion ability evaluation of the endophytic bacteria isolated from soybean root nodule in Sichuan Basin[J]. Chinese Journal of Applied and Environmental Biology, 2017, 23(1): 46-53. (in Chinese with English abstract)
[11]	赵龙飞, 徐亚军, 曹冬建, 等. 溶磷性大豆根瘤内生菌的筛选、抗性及系统发育和促生[J]. 生态学报, 2015, 35(13): 4425-4435.
	ZHAO L F, XU Y J, CAO D J, et al. Screening, resistance, phylogeny and growth promoting of phosphorus solubilizing bacteria isolated from soybean root nodules[J]. Acta Ecologica Sinica, 2015, 35(13): 4425-4435. (in Chinese with English abstract)
[12]	FERCHICHI N, TOUKABRI W, BOULARESS M, et al. Isolation, identification and plant growth promotion ability of endophytic bacteria associated with lupine root nodule grown in Tunisian soil[J]. Archives of Microbiology, 2019, 201(10): 1333-1349.
[13]	黄秀梨, 辛明秀. 微生物学实验指导[M]. 2版. 北京: 高等教育出版社, 2008.
[14]	李培根, 要雅倩, 宋吉祥, 等. 马铃薯根际产IAA芽孢杆菌的分离鉴定及促生效果研究[J]. 生物技术通报, 2020, 36(9): 109-116.
	LI P G, YAO Y Q, SONG J X, et al. Isolation and identification of IAA-producing Bacillus sp on potato rhizosphere and its growth-promoting effect[J]. Biotechnology Bulletin, 2020, 36(9): 109-116. (in Chinese with English abstract)
[15]	徐伟慧, 吕智航, 史一然, 等. 西瓜复合根际促生菌剂构建与促生效应研究[J]. 浙江农业学报, 2018, 30(5): 778-786.
	XU W H, LYU Z H, SHI Y R, et al. Establishment of complex growth-promoting rhizobacteria for watermelon and promoting effect on watermelon roots[J]. Acta Agriculturae Zhejiangensis, 2018, 30(5): 778-786. (in Chinese with English abstract)
[16]	徐伟慧, 刘泽平, 符春敏, 等. 根际芽孢杆菌对水稻根系的促生效应[J]. 河南农业科学, 2018, 47(4): 59-63.
	XU W H, LIU Z P, FU C M, et al. Promoting effect of Bacillus rhizobacteria on rice root[J]. Journal of Henan Agricultural Sciences, 2018, 47(4): 59-63. (in Chinese with English abstract)
[17]	陈峥, 刘波, 邓文琼, 等. 14株芽孢杆菌菌株的分子鉴定及其促生机理分析[J]. 热带作物学报, 2021, 42(3): 789-799.
	CHEN Z, LIU B, DENG W Q, et al. Molecular identification and growth promoting mechanisms of 14 Bacillus strains[J]. Chinese Journal of Tropical Crops, 2021, 42(3): 789-799. (in Chinese with English abstract)
[18]	薛晓昀, 冯瑞华, 关大伟, 等. 大豆根瘤菌与促生菌复合系筛选及机理研究[J]. 大豆科学, 2011, 30(4): 613-620.
	XUE X Y, FENG R H, GUAN D W, et al. Screening and analysis for efficient co-inoculation system of soybean rhizobia and plant growth-promoting rhizobacteria[J]. Soybean Science, 2011, 30(4): 613-620. (in Chinese with English abstract)
[19]	李晶, 蔡少丽, 林金新. 高效紫云英根瘤菌的选育[J]. 南方农业, 2018, 12(18): 184-185.
	LI J, CAI S L, LIN J X. Breeding of rhizobia isolated from Chinese milk vetch with high efficiency[J]. South China Agriculture, 2018, 12(18): 184-185. (in Chinese)
[20]	邱并生. 根瘤内生细菌[J]. 微生物学通报, 2013, 40(3): 544.
	QIU B S. Nodule endophytic bacteria[J]. Microbiology China, 2013, 40(3): 544. (in Chinese)
[21]	张淑婷, 周利华, 赵文卓. 植物内生细菌对植物健康的作用[J]. 植物医生, 2020, 33(1): 6-11.
	ZHANG S T, ZHOU L H, ZHAO W Z. The role of plant endophytes in plant health[J]. Plant Doctor, 2020, 33(1): 6-11. (in Chinese with English abstract)
[22]	刘杰, 汪恩涛, 陈文新. 豆科植物根瘤内生细菌的发现及其研究进展[J]. 微生物学报, 2011, 51(8): 1001-1006.
	LIU J, WANG E T, CHEN W X. Discovery and research progress of endophytic bacteria in the root nodules of legumes: a review[J]. Acta Microbiologica Sinica, 2011, 51(8): 1001-1006. (in Chinese with English abstract)
[23]	张翠绵, 贾楠, 马佳, 等. 番茄根际多功能益生芽孢杆菌的筛选与鉴定[J]. 河北农业科学, 2019, 23(4): 47-52.
	ZHANG C M, JIA N, MA J, et al. Screening and identification of multifunctional beneficial Bacillus in tomato rhizosphere soil[J]. Journal of Hebei Agricultural Sciences, 2019, 23(4): 47-52. (in Chinese with English abstract)
[24]	MILJAKOVIĆ D, MARINKOVIĆ J, TAMINDŽIĆ G, et al. Effect of plant growth promoting Bacillus spp. on germination and seedling growth of soybean[J]. Legume Research-an International Journal, 2022, 45(4): 487-491.
[25]	李梦娇, 彭晟, 徐绍忠, 等. 克雷伯氏菌在农业与环境治理上的应用[J]. 生物技术进展, 2014, 4(6): 415-420.
	LI M J, PENG S, XU S Z, et al. Application of Klebsiella spp. in agriculture and environmental management[J]. Current Biotechnology, 2014, 4(6): 415-420. (in Chinese with English abstract)
[26]	韩梅, 罗培宇, 肖亦农, 等. 玉米内生固氮菌的分离鉴定及其促生长作用研究[J]. 沈阳农业大学学报, 2010, 41(1): 94-97.
	HAN M, LUO P Y, XIAO Y N, et al. Isolation of endophytic N2-fixing bacteria of corn rice and their function for promoting growth[J]. Journal of Shenyang Agricultural University, 2010, 41(1): 94-97. (in Chinese with English abstract)
[27]	GARCÍA-FRAILE P, CARRO L, ROBLEDO M, et al. Rhizobium promotes non-legumes growth and quality in several production steps: towards a biofertilization of edible raw vegetables healthy for humans[J]. PLoS One, 2012, 7(5): e38122.
[28]	赵晓妍, 曹越, 董芮萌, 等. 一株野生大豆内生细菌YDX14菌株的分离、鉴定及促生效应研究[J]. 大豆科学, 2021, 40(2): 224-231.
	ZHAO X Y, CAO Y, DONG R M, et al. Isolation, identification and growth promoting effect of an endophytic bacteria YDX14 in wild soybean[J]. Soybean Science, 2021, 40(2): 224-231. (in Chinese with English abstract)
[29]	丁文珺, 王珊珊, 任婧祺, 等. 植物内生菌研究进展[J]. 生物技术进展, 2015, 5(6): 425-428.
	DING W J, WANG S S, REN J Q, et al. Progress on plant endophyte[J]. Current Biotechnology, 2015, 5(6): 425-428. (in Chinese with English abstract)
[30]	田振祥. 大豆内生细菌的分离及其促生作用研究[D]. 哈尔滨: 东北农业大学, 2021: 22-23.
	TIAN Z X. Isolation and growth promoting effect of endophytic bacteria from soybean[D]. Harbin: Northeast Agricultural University, 2021: 22-23. (in Chinese with English abstract)
[31]	周怡, 毛亮, 张婷婷, 等. 大豆内生芽孢杆菌的分离和促生菌株的筛选及鉴定[J]. 大豆科学, 2009, 28(3): 502-506.
	ZHOU Y, MAO L, ZHANG T T, et al. Isolation of endophytic Bacillus from soybean seed and screening and identification of promoting strain[J]. Soybean Science, 2009, 28(3): 502-506. (in Chinese with English abstract)
[32]	衡楠楠, 陈远学, 徐开未, 等. 大豆根瘤菌SCAUs8的接种效果、促生性及系统发育研究[J]. 微生物学通报, 2016, 43(8): 1708-1714.
	HENG N N, CHEN Y X, XU K W, et al. Field assessment of symbiotic efficiency, growth-promoting ability and phylogeny of soybean rhizobial strain SCAUs8[J]. Microbiology China, 2016, 43(8): 1708-1714. (in Chinese with English abstract)
[33]	丁玮. 白花泡桐幼林根瘤内生细菌的分离及鉴定[D]. 南宁: 广西大学, 2019: 36-37.
	DING W. Isolation and idntification of endophytic bacteria in the root nodule of young period Paulownia fortunei[D]. Nanning: Guangxi University, 2019: 36-37. (in Chinese with English abstract)
[34]	ZHANG D N, XU H L, GAO J Y, et al. Endophytic Bacillus altitudinis strain uses different novelty molecular pathways to enhance plant growth[J]. Frontiers in Microbiology, 2021, 12: 692313.
[35]	何建清, 张格杰. 植物根际促生菌肥代替部分化肥对黑青稞生长、产量和品质的影响[J]. 河南农业科学, 2022, 51(7): 93-101.
	HE J Q, ZHANG G J. Effects of plant growth promoting rhizobacteria (PGPR) fertilizer instead of part of chemical fertilizer on growth, yield and quality of black highland barley[J]. Journal of Henan Agricultural Sciences, 2022, 51(7): 93-101. (in Chinese with English abstract)

Isolation and identification of endophytic bacteria from soybean nodule and their effects on soybean plant

RichHTML

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

Figures/Tables 5

References 35

Related Articles 15

Recommended Articles

Metrics

Comments

菌株编号 Strain No.	淀粉水解试验 Starch hydrolysis test	油脂水解试验 Oil hydrolysis test	乳糖发酵试验 Lactose fermentation test	葡萄糖发酵试验 Glucose fermentation test
S1	+	-	+	+
S2	+	-	+	+
S3	+	-	+	+
S4	-	+	-	+
S5	+	-	+	-
S6	+	-	-	-
S7	-	+	+	+
S8	-	+	+	+
S9	-	+	-	+
S10	-	+	+	-

菌株编号 Strain No.	淀粉水解试验 Starch hydrolysis test	油脂水解试验 Oil hydrolysis test	乳糖发酵试验 Lactose fermentation test	葡萄糖发酵试验 Glucose fermentation test
S1	+	-	+	+
S2	+	-	+	+
S3	+	-	+	+
S4	-	+	-	+
S5	+	-	+	-
S6	+	-	-	-
S7	-	+	+	+
S8	-	+	+	+
S9	-	+	-	+
S10	-	+	+	-

[1]	BU Yuanpeng, LIU Na, ZHANG Guwen, FENG Zhijuan, WANG Bin, GONG Yaming, XU Linying. Diversity evaluation of agronomic traits and construction of core collection and taste quality evaluation system in vegetable soybean germplasm resources [J]. Acta Agriculturae Zhejiangensis, 2023, 35(6): 1307-1314.
[2]	YANG Songhua, SHI Guiyang, WANG Jingqin, CHEN Zhu. Effects of soybean root exudates on insoluble phosphorus in soil under low phosphorus stress [J]. Acta Agriculturae Zhejiangensis, 2023, 35(6): 1396-1406.
[3]	TAN Shuxia, ZHAO Taodi, YANG Hao, NING Kejun, LIU Li, HE Qingyuan, HUANG Shoucheng, SHU Yingjie. Effects of shading on agronomic characters, yield and nitrogen metabolism of 10 vegetable soybean varieties [J]. Acta Agriculturae Zhejiangensis, 2023, 35(4): 729-735.
[4]	ZHANG Meng, SHE Bao, YANG Yuying, HUANG Linsheng, ZHU Mengqi. Study on extraction method of soybean planting areas based on unmanned aerial vehicle RGB image [J]. Acta Agriculturae Zhejiangensis, 2023, 35(4): 952-961.
[5]	LI Wenchen, LIU Xin, QI Zezheng, YU Lu, WANG Fang. Bioinformatics of Huipizhi Black soybean GmPUB24 and expression under Heterodera glycines infection [J]. Acta Agriculturae Zhejiangensis, 2022, 34(6): 1124-1132.
[6]	YANG Xinxia, TANG Mansheng, ZHANG Bin. Identification of soybean PP2C family genes and transcriptome analysis in response to salt stress [J]. Acta Agriculturae Zhejiangensis, 2022, 34(2): 207-220.
[7]	LIU Na, FAN Qiaochu, ZHOU Jia, SONG Yajing, ZHANG Guwen, FENG Zhijuan, BO Yuanpeng, WANG Bin, GONG Yaming. Identification and control of anthracnose in vegetable soybean [J]. Acta Agriculturae Zhejiangensis, 2022, 34(12): 2682-2688.
[8]	HE Xiu, XU Meiyu, XIN Weigang, ZHANG Qilin, WANG Feng, LIN Lianbing. Effects of soybean meal addition and fermentation time on nutritional quality and bacterial diversity of Pennisetum purpureum silage [J]. Acta Agriculturae Zhejiangensis, 2022, 34(10): 2160-2171.
[9]	YANG Xinxia, ZHANG Bin. Identification of soybean LAZ1 gene family and functional analysis of GmLAZ1-9 [J]. Acta Agriculturae Zhejiangensis, 2021, 33(4): 586-594.
[10]	ZHANG Weimei, ZHANG Guwen, FENG Zhijuan, LIU Na, WANG Bin, BU Yuanpeng. Research progress on genetic and regulatory mechanisms of sucrose in vegetable soybean seeds [J]. Acta Agriculturae Zhejiangensis, 2021, 33(12): 2446-2456.
[11]	YANG Ju, DENG Junliang, XIA Jiangying, SONG Tianhao, PANG Lianfeng, REN Zhihua. Effect of vitamin A on barrier function damage of piglet intestinal epithelial cells induced by soybean 7S globulin [J]. Acta Agriculturae Zhejiangensis, 2021, 33(11): 2026-2033.
[12]	XU Yue, XU Yaxin, HUANG Xingjun, WU Shu, CHEN Guodong, WU Quanzhong, ZHAI Yunlong. Effects of inoculation method of rhizobia on dry matter accumulation and yield of multiple soybean [J]. Acta Agriculturae Zhejiangensis, 2021, 33(10): 1808-1816.
[13]	HE Jiaqi, ZHAI Ying, ZHANG Jun, QIU Shuang, LI Mingyang, ZHAO Yan, ZHANG Meijuan, MA Tianyi. Cloning and expression analysis of GmDof1.5 in soybean under abiotic stress [J]. Acta Agriculturae Zhejiangensis, 2021, 33(1): 1-7.
[14]	YANG Yiwei, XIAO Hua, CHEN Hu, XIAO Niejia, GUO Cheng. Structural characteristics of soil mite communities under different modes of rose-based agroforestry in Karst area [J]. Acta Agriculturae Zhejiangensis, 2021, 33(1): 112-121.
[15]	ZHANG Guwen, SHEN Li, ZHENG Huazhang, LIU Na, FENG Zhijuan, GONG Yaming. Research advances of zinc finger protein transcription factor Di19 in regulation of soybean responding to drought stress [J]. , 2020, 32(2): 373-382.

菌株编号 Strain No.	淀粉水解试验 Starch hydrolysis test	油脂水解试验 Oil hydrolysis test	乳糖发酵试验 Lactose fermentation test	葡萄糖发酵试验 Glucose fermentation test
S1	+	-	+	+
S2	+	-	+	+
S3	+	-	+	+
S4	-	+	-	+
S5	+	-	+	-
S6	+	-	-	-
S7	-	+	+	+
S8	-	+	+	+
S9	-	+	-	+
S10	-	+	+	-

菌株编号 Strain No.	淀粉水解试验 Starch hydrolysis test	油脂水解试验 Oil hydrolysis test	乳糖发酵试验 Lactose fermentation test	葡萄糖发酵试验 Glucose fermentation test
S1	+	-	+	+
S2	+	-	+	+
S3	+	-	+	+
S4	-	+	-	+
S5	+	-	+	-
S6	+	-	-	-
S7	-	+	+	+
S8	-	+	+	+
S9	-	+	-	+
S10	-	+	+	-

菌株编号 Strain No.	淀粉水解试验 Starch hydrolysis test	油脂水解试验 Oil hydrolysis test	乳糖发酵试验 Lactose fermentation test	葡萄糖发酵试验 Glucose fermentation test
S1	+	-	+	+
S2	+	-	+	+
S3	+	-	+	+
S4	-	+	-	+
S5	+	-	+	-
S6	+	-	-	-
S7	-	+	+	+
S8	-	+	+	+
S9	-	+	-	+
S10	-	+	+	-

菌株编号 Strain No.	淀粉水解试验 Starch hydrolysis test	油脂水解试验 Oil hydrolysis test	乳糖发酵试验 Lactose fermentation test	葡萄糖发酵试验 Glucose fermentation test
S1	+	-	+	+
S2	+	-	+	+
S3	+	-	+	+
S4	-	+	-	+
S5	+	-	+	-
S6	+	-	-	-
S7	-	+	+	+
S8	-	+	+	+
S9	-	+	-	+
S10	-	+	+	-