3株芽孢杆菌在水稻根际定殖促生及其在土壤中的存活
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摘要
植物根际促生菌(PGPR)可分泌植物生长激素,促进土壤养分循环,是生物肥料重要的种质资源。PGPR吸附于作物根系是形成定殖与功能的必要条件,抗性药物标记是研究功能菌株定殖促生能力的一种手段。利用解钾功能菌(Bacillus aryabhattai LZP01,简称LZP01)、解磷功能菌(Bacillus pumilus LZP02,简称LZP02)和溶磷功能菌(Bacillus megaterium LZP03,简称LZP03)这3株高效的水稻根际促生菌,通过抗生素药物利福平梯度筛选分别得到标记菌株K-LZP01、K-LZP02和K-LZP03,并且研究标记菌株的功能保留情况,进而检测标记菌株在水稻根际与土壤中定殖存活及促生能力。结果表明,水稻幼根浸入菌液后吸附量随浸入时间的增加而增大,其中K-LZP01在10min后吸附基本稳定;K-LZP01在50 min时吸附量达到最大;K-LZP03在5 min时吸附量达到最大,最大吸附量为12×10~7CFU·g~(-1),之后趋于动态稳定。将水稻幼苗浸入不同浓度菌悬液中,幼根吸附量随菌悬液浓度的增大而增大。标记菌株在接种于土壤中10 d后定殖量趋于稳定,且定殖能力较强。距离水稻根际越近标记菌株含量越高,表明菌株定殖竞争力越强。在灭菌土与未灭菌土条件下,3株菌株均具有促进水稻根系生长的能力。
Plant growth-promoting rhizobacteria(PGPR)can secrete plant growth hormone and promote soil nutrient circulation,which is an important germplasm resource of biological fertilizers. Adsorption of PGPR to crop roots is a necessary condition for colonization and function,and drug resistance marker is a mean of studying the ability of functional strains to colonize and promote growth. The purpose of the experiment is to label the strain by the antibiotic drug rifampicin,and to study the retention of the function of the strain,and then to determine the colonization survival ability of the strain in the rhizosphere of rice. The results show that the adsorption of rice roots to strain increased with the immersion time,and the adsorption content of K-LZP01 was basically stable after dipping for 10 min. K-LZP02 reached the maximum adsorption capacity at 50 min. The maximum adsorption capacity of K-LZP03 was 12×10~7 CFU·g~(-1) at 5 min,and then became stable. When the rice seedlings were dipped in different concentrations of bacterial suspension,the adsorption capacity of radicle increased with increasing of bacterial suspension concentration. After 10 days of inoculation in soil,the colonization quantity of the labeled strains tended to be stable and the colonization ability is strong. The results found that the closer the distance to the rice rhizosphere,the higher the strain content,which indicate that the strains had good colonization competitiveness. In both sterilized and unsterilized soils,the three strains could all promote the growth of rice. In this study,researches on the resistance marker of PGPR strain as well as its colonization and growth promotion in rice rhizosphere were carried out,and the results are important to interpret the mechanism of PGPR and guide the production practice.
Plant growth-promoting rhizobacteria(PGPR)can secrete plant growth hormone and promote soil nutrient circulation,which is an important germplasm resource of biological fertilizers. Adsorption of PGPR to crop roots is a necessary condition for colonization and function,and drug resistance marker is a mean of studying the ability of functional strains to colonize and promote growth. The purpose of the experiment is to label the strain by the antibiotic drug rifampicin,and to study the retention of the function of the strain,and then to determine the colonization survival ability of the strain in the rhizosphere of rice. The results show that the adsorption of rice roots to strain increased with the immersion time,and the adsorption content of K-LZP01 was basically stable after dipping for 10 min. K-LZP02 reached the maximum adsorption capacity at 50 min. The maximum adsorption capacity of K-LZP03 was 12×10~7 CFU·g~(-1) at 5 min,and then became stable. When the rice seedlings were dipped in different concentrations of bacterial suspension,the adsorption capacity of radicle increased with increasing of bacterial suspension concentration. After 10 days of inoculation in soil,the colonization quantity of the labeled strains tended to be stable and the colonization ability is strong. The results found that the closer the distance to the rice rhizosphere,the higher the strain content,which indicate that the strains had good colonization competitiveness. In both sterilized and unsterilized soils,the three strains could all promote the growth of rice. In this study,researches on the resistance marker of PGPR strain as well as its colonization and growth promotion in rice rhizosphere were carried out,and the results are important to interpret the mechanism of PGPR and guide the production practice.
引文
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[20]王海霞,陈园,王超,等.组合抗生素抗性选育多杀菌素高产菌株[J].粮油食品科技,2017,25(4):70-75.[WANG Hai-xia,CHEN Yuan,WANG Chao,et al. Breeding of Spinosad High-Pro-ducing Strains by Combinatorial Antibiotic Resistance[J]. Sci-ence and Technology of Cereals,Oils and Foods,2017,25(4):70-75.]
[21]车建美,蓝江林,刘波.转绿色荧光蛋白基因的青枯雷尔氏菌生物学特性[J].中国农业科学,2008,41(11):3626-3635.[CHE Jian-mei,LAN Jiang-lin,LIU Bo. GFP Tagging Ralstonia-solanacearum With gfp/luxAB Mini-Tn5[J]. Scientia Agricultura Sinica,2008,41(11):3626-3635.]
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[23]刘庆丰,熊国如,毛自朝,等.枯草芽胞杆菌XF-1的根围定殖能力分析[J].植物保护学报,2012,39(5):425-430.[LIU Qing-feng,XIONG Guo-ru,MAO Zi-chao,et al. Analyses for the Colonization Ability of Bacillus subtilis XF-1 in the Rhizosphere[J]. Acta Phytophylacica Sinica,2012,39(5):425-430.]
[24] CHEN C,BAUSKE E M,MUSSON G,et al. Biological Control of Fusarium Wilt on Cotton by Use of Endophytic Bacteria[J]. Bio-logical Control,1995,5(1):83-91.
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[26] THEODULOZ C,VEGA A,SALAZAR M,et al. Expression of a Bacillus thuringiensisδ-Endotoxin cry1Ab Gene in Bacillus subtil-is and Bacillus licheniformis Strains That Naturally Colonize the Phylloplane of Tomato Plants(Lycopersicon esculentum,Mills)[J]. Journal of Applied Microbiology,2003,94(3):375-381.
[27]余国运,唐文华,陈延熙.增产菌在小麦叶面定殖的研究[J].生物防治通报,1992,8(2):83-86.[YU Guo-yun,TANG Wen-hua,CHEN Yan-xi. Colonization of“Yield-Increasing Bacte-ria”on the Phylloplane of Wheat[J]. Chinese Journal of Biologi-cal Control,1992,8(2):83-86.]
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[2]付景,杨建昌.中国水稻栽培理论与技术发展的回顾与展[J].作物杂志,2010(5):1-4.[FU Jing,YANG Jian-chang. Retro-spect and Prospect for the Development of Theory and Technolo-gy of Rice Cultivation in China[J]. Crops,2010(5):1-4.]
[3]曹丹,白林燕,冯建中,等.东北三省水稻种植面积时空变化监测与分析[J].江苏农业科学,2018,46(10):260-265.[CAO Dan,BAI Lin-yan,FENG Jian-zhong,et al. Monitoring and Analysis of Temporal and Spatial Variation of Rice Planting Area in Northeastern China[J]. Jiangsu Agricultural Sciences,2018,46(10):260-265.]
[4] WIEHE W,H?FLICH G. Survival of Plant Growth Promoting Rhizosphere Bacteria in the Rhizosphere of Different Crops and Migration to Non-Inoculated Plants Under Field Conditions in North-East Germany[J]. Microbiological Research,1995,150(2):201-206.
[5]甄静,李冠杰,王继雯,等.固定化菌株在土壤中的繁殖释放及性能分析[J].中国农学通报,2016,32(9):155-158.[ZHEN Jing,LI Guan-jie,WANG Ji-wen,et al. Immobilized Strain Propagation and Release and Its Performance[J]. Chinese Agricultural Science Bulletin,2016,32(9):155-158.]
[6]孙真,郑亮,邱浩斌.植物根际促生细菌定殖研究进展[J].生物技术通报,2017,33(2):8-15.[SUN Zhen,ZHENG Liang,QIU Hao-bin. Research Advances on Colonization of Plant Growth-Promoting Rhizobacteria[J]. Biotechnology Bulletin,2017,33(2):8-15.]
[7]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 Wei-hui,LIU Ze-ping,FU Chun-min,et al. Promoting Effect of Bacillus Rhizobacteria on Rice Root[J]. Journal of Henan Agricultural Sciences,2018,47(4):59-63.]
[8]王志刚,胡云龙,徐伟慧,等.鞘氨醇单胞菌菌株CL01的分离鉴定及其对连作西瓜的促生效应[J].农业生物技术学报,2015,23(10):1360-1367.[WANG Zhi-gang,HU Yun-long,XU Wei-hui,et al. Isolation and Identification of Sphingomonas sp.CL01 and Its Promoting Effects on Watermelon(Citrullus lana-tus)in Continuous Cropping Soil[J]. Journal of Agricultural Bio-technology,2015,23(10):1360-1367.]
[9]余贤美,侯长明,王海荣,等.枯草芽孢杆菌Bs-15在枣树体内和土壤中的定殖及其对土壤微生物多样性的影响[J].中国生物防治学报,2014,30(4):497-502.[YU Xian-mei,HOU Chang-ming,WANG Hai-rong,et al. Colonization of Bacillus sub-tilis Bs-15 in Jujube Plant and Soil and Its Influence on the Micro-bial Diversity in the Soil[J]. Chinese Journal of Biological Con-trol,2014,30(4):497-502.]
[10]刘泽平,王志刚,徐伟慧,等.水稻根际促生菌的筛选鉴定及促生能力分析[J].农业资源与环境学报,2018,35(2):119-125.[LIU Ze-ping,WANG Zhi-gang,XU Wei-hui,et al. Screen,Identification and Analysis on the Growth-Promoting Ability for the Rice Growth-Promoting Rhizobacteria[J]. Journal of Agricul-tural Resources and Environment,2018,35(2):119-125.]
[11] JI S H,GURURANI M A,CHUN S C. Isolation and Characteriza-tion of Plant Growth Promoting Endophytic Diazotrophic Bacteria From Korean Rice Cultivars?[J]. Microbiological Research,2014,169(1):83-98.
[12] CASTANHEIRA N L,DOURADO A C,PAIS I,et al. Coloniza-tion and Beneficial Effects on Annual Ryegrass by Mixed Inocula-tion With Plant Growth Promoting Bacteria[J]. Microbiological Research,2017,198:47-55.
[13]吴蔼民,顾本康,傅正擎,等.内生菌73a在不同抗性品种棉花体内的定殖和消长动态研究[J].植物病理学报,2001,31(4):289-294.[WU Ai-min,GU Ben-kang,FU Zheng-qing,et al.Studies on the Population Fluctuation of Entophytic Bacteria 73a in Cotton Plant[J]. Acta Phytopathologica Sinica,2001,31(4):289-294.]
[14] NIMNOI P,PONGSILP N,RUANPANUN P. Monitoring the Effi-ciency of Streptomyces galilaeus Strain KPS-C004 Against Root Knot Disease and the Promotion of Plant Growth in the Plant-Par-asitic Nematode Infested Soils[J]. Biological Control,2017,114:158-166.
[15] SANTOS A R S,ETTO R M,FURMAM R W,et al. Labeled Azo-spirillum brasilense Wild Type and Excretion-Ammonium Strains in Association With Barley Roots[J]. Plant Physiology and Bio-chemistry,2017,118:422-426.
[16]王志远,吴兴兴,吴毅歆,等.解淀粉芽孢杆菌B9601-Y2抗性基因标记及其在作物根部的定殖能力[J].华中农业大学学报,2012,31(3):313-319.[WANG Zhi-yuan,WU Xing-xing,WU Yi-xin,et al. Resistance Genes Labeling and Colonization Ability of a Biocontrol Agent B9601-Y2 of Bacillus amylolIquefa-ciens in Crop Rhizospheres[J]. Journal of Huazhong Agricultural University,2012,31(3):313-319.]
[17] GLANDORF D C,VERHEGGEN P,JANSEN T,et al. Effect of Genetically Modified Pseudomonas putida WCS358r on the Fun-gal Rhizosphere Microflora of Field-Grown Wheat[J]. Applied and Environmental Microbiology,2001,67(8):3371-3378.
[18] OMER Z S,TOMBOLINI R,GERHARDSON B. Plant Coloniza-tion by Pink-Pigmented Facultative Methylotrophic Bacteria(PPFMs)[J]. FEMS Microbiology Ecology,2010,47(3):319-326.
[19] BRETON Y L,MOHAPATRA N,HALDENWANG W G. In Vivo Random Mutagenesis of Bacillus subtilis by Use of TnYLB-1,a mariner-based Transposon[J]. Applied and Environmental Micro-biology,2006,72(1):327-333.
[20]王海霞,陈园,王超,等.组合抗生素抗性选育多杀菌素高产菌株[J].粮油食品科技,2017,25(4):70-75.[WANG Hai-xia,CHEN Yuan,WANG Chao,et al. Breeding of Spinosad High-Pro-ducing Strains by Combinatorial Antibiotic Resistance[J]. Sci-ence and Technology of Cereals,Oils and Foods,2017,25(4):70-75.]
[21]车建美,蓝江林,刘波.转绿色荧光蛋白基因的青枯雷尔氏菌生物学特性[J].中国农业科学,2008,41(11):3626-3635.[CHE Jian-mei,LAN Jiang-lin,LIU Bo. GFP Tagging Ralstonia-solanacearum With gfp/luxAB Mini-Tn5[J]. Scientia Agricultura Sinica,2008,41(11):3626-3635.]
[22] GUERRERO-MOLINA M F,WINIK B C,PEDRAZA R O. More Than Rhizosphere Colonization of Strawberry Plants by Azospiril-lum brasilense[J]. Applied Soil Ecology,2012,61:205-212.
[23]刘庆丰,熊国如,毛自朝,等.枯草芽胞杆菌XF-1的根围定殖能力分析[J].植物保护学报,2012,39(5):425-430.[LIU Qing-feng,XIONG Guo-ru,MAO Zi-chao,et al. Analyses for the Colonization Ability of Bacillus subtilis XF-1 in the Rhizosphere[J]. Acta Phytophylacica Sinica,2012,39(5):425-430.]
[24] CHEN C,BAUSKE E M,MUSSON G,et al. Biological Control of Fusarium Wilt on Cotton by Use of Endophytic Bacteria[J]. Bio-logical Control,1995,5(1):83-91.
[25]齐飞飞,夏觅真,唐欣昀,等. lux AB基因标记的K2116L菌株在棉花根际中的定殖[J].生态学杂志,2008,27(2):192-196.[QI Fei-fei,XIA Mi-zhen,TANG Xin-yun,et al. Colonization of lux AB Gene-Marked Bacillus K2116 Strain in Cotton Rhizosphere[J]. Chinese Journal of Ecology,2008,27(2):192-196.]
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