WO2023115830A1 - Complex microbial agent for promoting growth of soybeans and improving population abundance of probiotic microorganisms - Google Patents

Complex microbial agent for promoting growth of soybeans and improving population abundance of probiotic microorganisms Download PDF

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WO2023115830A1
WO2023115830A1 PCT/CN2022/096584 CN2022096584W WO2023115830A1 WO 2023115830 A1 WO2023115830 A1 WO 2023115830A1 CN 2022096584 W CN2022096584 W CN 2022096584W WO 2023115830 A1 WO2023115830 A1 WO 2023115830A1
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bacterial agent
soybean
increased
bacteria
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庄家尧
毛国豪
张文韬
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南京林业大学
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    • C—CHEMISTRY; METALLURGY
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    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
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    • C05G3/00—Mixtures of one or more fertilisers with additives not having a specially fertilising activity

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  • Microbial fertilizers are more and more widely used in agriculture due to their high-efficiency and green features, and microbial agents as their raw materials have been widely valued and researched by agricultural producers.
  • compound inoculum has been vigorously promoted and applied due to its advantages of rich variety, complete fertility, and remarkable effect. Therefore, the screening and development of high-quality composite bacterial agents are inseparable from improving the soybean production in my country and realizing the self-reliance of my country's soybean industry.
  • the volume ratio of Bacillus highlander Y1 and Pseudomonas H1 in the composite bacterial agent are equal and the concentration is not less than 10 9 CFU/ml.
  • a phylogenetic tree will be constructed for the strains with high similarity (see Figure 4 for details), and the analysis of the developmental tree shows that the Y1 strain belongs to Bacillus altitudinis of the genus Bacillus, and it is named Bacillus altitudinis (Bacillus altitudinis) Y1.
  • the concentration of available phosphorus and alkaline hydrolyzed nitrogen in group H treated with compound bacterial agent was 3.49mg/kg and 486.00mg/kg respectively, compared with blank group C, the available phosphorus content increased significantly by 66.96% (P ⁇ 0.05), and the concentration of alkaline hydrolyzed Nitrogen increased by 29.14%, and the pH decreased from 7.13 to 6.73.
  • the soil was acidified to a certain extent; compared with the single bacterial agent J group and Y group, the available phosphorus content increased by 22.03% and 32.20% respectively (P ⁇ 0.05), and the alkali-hydrolyzed nitrogen was respectively Increased by 4.59% and 10.54%, the pH decreased to a certain extent.
  • the present invention discloses a composite bacterial agent that promotes the growth of soybeans and increases the abundance of probiotic microbial populations. Effects, especially to promote aboveground biomass and seedling height of soybean plants; increase the relative abundance of Rhizobium and Devosbacteriaceae bacteria in soybean roots, so that the symbiotic nitrogen fixation ability has been effectively exerted.
  • Figure 1 shows the three bacterial agents in the YMB medium test tube, the left is the Y1 fermentation broth, the middle is the H1 fermentation broth, and the right is the H1 and Y1 composite strain fermentation broth.
  • Fig. 2 is a colony diagram of Bacillus alboteria Y1 on the YMA medium plate.
  • Bacillus altitudinis Bacillus altitudinis (Bacillus altitudinis) Y1 was isolated and screened from wild soybean root nodules in our laboratory, and was preserved by the China Center for Type Culture Collection (CCTCC).
  • the address of the preservation unit is Wuhan, China, Wuhan University, and the preservation number is CCTCC NO: M 20211043, the deposit date is September 2, 2021.
  • composition and content of YMB medium are: mannitol 10g, magnesium sulfate heptahydrate 0.2g, sodium chloride 0.1g, yeast powder 3g, dipotassium hydrogen phosphate 0.25g, potassium dihydrogen phosphate 0.25g, calcium carbonate 3g , 1000ml deionized water, pH 7.2-7.4.
  • control group was treated with equal volumes of 100-fold diluted Y1 fermentation broth, 100-fold diluted H1 fermentation broth, and sterile water.
  • the present invention has carried out four treatments to soybean plants, including blank control group C, Y1 bacterial agent group Y group, H1 bacterial agent J group, and compound bacterial agent H group. Compared with the sterile blank control group, the root length of the underground part of soybean was significantly increased in the three treatments of adding bacteria.
  • the top 5 species with the highest abundance at the level of known bacterial genus are Geobacillus (0.00%-61.22%), Actinomadura , 0.00% ⁇ 11.02%), Bacillus (2.66% ⁇ 11.95%), Luteimonas (Luteimonas, 0.31% ⁇ 5.97%) are the main known dominant flora, in each treatment
  • the relative abundance is 11.91%-52.31%.
  • the relative abundance of Actinomadura after compound microbial agent H was higher than that of blank group C, single bacterial agent group J, and Y group. Actinomadura has an antibacterial effect and is beneficial for the production of antibiotic drugs.
  • the present invention involves two groups of treatment groups: sterile control group c and compound microbial agent H group.
  • the collected and arranged two groups of soybean rhizosphere samples are sent to Guangzhou Kidio Co., Ltd. for detection, and the roots are analyzed by high-throughput sequencing.
  • Microbial diversity exploring the composition of microbial communities in most roots.
  • the top 7 species with the highest abundance at the level of bacterial families are Xanthomonadaceae, Burkholderia Burkholderiaceae, Sphingomonadaceae, Methylophilaceae, Devosiaceae, Pseudomonadaceae, Rhizobiaceae.

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Abstract

A complex microbial agent for promoting the growth of soybeans and improving the population abundance of probiotic microorganisms, prepared by mixing a Pseudomonas sp. H1 fermentation broth and a Bacillus altitudinis Y1 fermentation broth. When the complex microbial agent is applied to soybean planting, the growth of soybean plants can be promoted, the population abundance of the probiotic microorganisms in soil can be improved, and the complex microbial agent has an improvement effect on the soil.

Description

一种对大豆促生和提高益生微生物种群丰度的复合菌剂A compound microbial agent for promoting soybean growth and increasing the abundance of probiotic microbial populations 技术领域technical field
本发明涉及一种对大豆促生和提高益生微生物种群丰度的复合菌剂，属于微生物技术领域。The invention relates to a composite microbial agent for promoting the growth of soybeans and increasing the abundance of probiotic microbial populations, and belongs to the technical field of microorganisms.
背景技术Background technique
大豆作为我国主要的食用油料来源，在食用、饲用市场需求巨大。数据显示，中国内地市场大豆进口数量连续五年增加，2020年我国进口大豆数量更是首次超过一亿吨。此外，市场上大豆品质的良莠不齐也是一大问题，如何提高我国自身的大豆产量和品质已然成为限制农业发展的一大问题。As the main source of edible oil in my country, soybean has a huge demand in the food and feed market. Data show that the number of soybean imports in the Chinese mainland market has increased for five consecutive years. In 2020, the number of soybeans imported by my country will exceed 100 million tons for the first time. In addition, the uneven quality of soybeans in the market is also a major problem. How to improve the yield and quality of soybeans in my country has become a major problem restricting agricultural development.
农业发展离不开化学肥料的施用，但是近年来化肥的不科学使用带来了水体污染、重金属富集、土壤盐渍化等许多问题。调查显示传统化肥的使用并没有带来预期中作物产量的增加，反而一些农田甚至出现土壤肥力低下、作物生长差的现象。为此，广大农业生产者迫切需要一种可以替代传统化肥的新型肥料的出现。Agricultural development is inseparable from the application of chemical fertilizers, but the unscientific use of chemical fertilizers in recent years has brought about many problems such as water pollution, heavy metal enrichment, and soil salinization. Surveys have shown that the use of traditional chemical fertilizers has not brought about the expected increase in crop yields. On the contrary, some farmlands have even suffered from low soil fertility and poor crop growth. For this reason, numerous agricultural producers urgently need the appearance of a kind of novel fertilizer that can replace traditional chemical fertilizer.
微生物肥料以其高效绿色特点在农业应用上愈加广泛，作为其原料的微生物菌剂得到了农业生产者的普遍重视与研究。作为微生物菌剂中的一个分支，复合菌剂因其种类丰富、肥力齐全、效果显著等优点获得大力推广和应用。因此，提高我国大豆产量、实现我国大豆产业的自力更生，离不开优质复合菌剂的筛选和开发。Microbial fertilizers are more and more widely used in agriculture due to their high-efficiency and green features, and microbial agents as their raw materials have been widely valued and researched by agricultural producers. As a branch of microbial inoculum, compound inoculum has been vigorously promoted and applied due to its advantages of rich variety, complete fertility, and remarkable effect. Therefore, the screening and development of high-quality composite bacterial agents are inseparable from improving the soybean production in my country and realizing the self-reliance of my country's soybean industry.
因此，能否从自然界中筛选出促进大豆生长的菌株并制成复合菌剂成为解决我国大豆产业发展问题的一大方案，同时也是研究微生物肥料工作者亟需解决的问题。Therefore, whether the strains that can promote soybean growth can be screened from nature and made into a compound bacterial agent has become a major solution to the development of my country's soybean industry, and it is also an urgent problem for researchers who study microbial fertilizers.
发明内容Contents of the invention
[发明目的][Purpose of the invention]
本发明旨在从自然界中筛选出对大豆促生的菌株并制成一种促进大豆生长的菌剂，为提高我国自身大豆产量和品质提供解决方案。The present invention aims to screen out the soybean growth-promoting bacterial strain from nature and make a soybean growth-promoting bacterial agent, so as to provide a solution for improving the yield and quality of my country's own soybean.
[技术方案][Technical solutions]
为解决上述技术问题，本发明公开了一种复合菌剂，该复合菌剂中的菌株由保藏编号为CCTCC NO:M 20211043的高地芽孢杆菌(Bacillus altitudinis)Y1和为保藏编号为CCTCC NO:M 20211042的假单胞菌(Pseudomonas sp.)H1组成。In order to solve the above-mentioned technical problems, the present invention discloses a composite microbial agent. The bacterial strain in the composite microbial agent consists of Bacillus altitudinis (Bacillus altitudinis) Y1 with a preservation number of CCTCC NO:M 20211043 and Bacillus altitudinis with a preservation number of CCTCC NO:M 20211042 Pseudomonas sp. H1 composition.
本发明所述高地芽孢杆菌(Bacillus altitudinis)Y1和假单胞菌(Pseudomonas sp.)H1已知都具有促进豆科植物生长的功能。Both Bacillus altitudinis Y1 and Pseudomonas sp. H1 described in the present invention have the function of promoting the growth of leguminous plants.
复合菌剂中高地芽孢杆菌Y1菌液和假单胞菌H1菌液体积比相等且浓度均不低于10 9CFU/ml。 The volume ratio of Bacillus highlander Y1 and Pseudomonas H1 in the composite bacterial agent are equal and the concentration is not less than 10 9 CFU/ml.
本发明所述高地芽孢杆菌(Bacillus altitudinis)Y1是从来源于湖南省株洲市攸县野生大豆根 瘤中分离得到的，在南京林业大学水土保持实验室-80摄氏度冰箱内保存，保藏于中国典型培养物保藏中心，地址：中国，武汉，武汉大学。保藏编号为CCTCC NO:M 20211043，保藏日期为2021年9月2日。The Bacillus altitudinis Y1 of the present invention is isolated from wild soybean root nodules in You County, Zhuzhou City, Hunan Province, stored in a refrigerator at -80 degrees Celsius in the Soil and Water Conservation Laboratory of Nanjing Forestry University, and preserved in China's typical culture Object Preservation Center, Address: Wuhan University, Wuhan, China. The deposit number is CCTCC NO:M 20211043, and the deposit date is September 2, 2021.
高地芽孢杆菌(Bacillus altitudinis)Y1菌株经测序分析，其16S rDNA序列如SEQ ID NO.2所示，将测序得到的序列在Genbank中进行核酸序列比对，结果显示该菌株为：Bacillus altitudinis(高地芽孢杆菌)，其与Bacillus pumilus(NR-043242)的同源性最近，核酸序列相似度高达99％。将与其相似度高的菌株构建***进化树(具体可见附图4)，经发育树分析显示Y1菌株属于芽孢杆菌属的高地芽孢杆菌，将其命名为高地芽孢杆菌(Bacillus altitudinis)Y1。Bacillus altitudinis (Bacillus altitudinis) Y1 strain is sequenced and analyzed, and its 16S rDNA sequence is shown in SEQ ID NO.2, and the sequence obtained by sequencing is carried out nucleic acid sequence alignment in Genbank, and the result shows that this bacterial strain is: Bacillus altitudinis (Highland Bacillus), which has the closest homology with Bacillus pumilus (NR-043242), with a nucleic acid sequence similarity as high as 99%. A phylogenetic tree will be constructed for the strains with high similarity (see Figure 4 for details), and the analysis of the developmental tree shows that the Y1 strain belongs to Bacillus altitudinis of the genus Bacillus, and it is named Bacillus altitudinis (Bacillus altitudinis) Y1.
本发明所述的假单胞菌(Pseudomonas sp.)H1是从来源于湖南省株洲市攸县野生大豆根瘤中分离得到的，在南京林业大学水土保持实验室-80摄氏度冰箱内保存，保藏于中国典型培养物保藏中心，地址：中国，武汉，武汉大学。保藏编号为CCTCC NO:M 20211042，保藏日期为2021年9月2日。Pseudomonas sp. H1 of the present invention is isolated from wild soybean root nodules in You County, Zhuzhou City, Hunan Province, and is stored in a -80 degree Celsius refrigerator in the Soil and Water Conservation Laboratory of Nanjing Forestry University. China Type Culture Collection Center, Address: Wuhan University, Wuhan, China. The deposit number is CCTCC NO:M 20211042, and the deposit date is September 2, 2021.
假单胞菌(Pseudomonas sp.)H1菌株经测序分析，其16S rDNA序列如SEQ ID NO.1所示，将测序得到的序列在Genbank中进行核酸序列比对，结果显示，与Pseudomonas sp(MG491596.1)和Pseudomonas chlororaphis(MT078671.1)的同源性较近，为99％。将与其相似度高的菌株构建***进化树(具体可见附图5)，结果显示菌株属于假单胞菌属，将其命名为假单胞菌(Pseudomonas sp.)H1。Pseudomonas sp. (Pseudomonas sp.) H1 strain is sequenced and analyzed, and its 16S rDNA sequence is shown in SEQ ID NO.1, and the sequence obtained by sequencing is compared with the nucleic acid sequence in Genbank. .1) and Pseudomonas chlororaphis (MT078671.1) have a close homology of 99%. A phylogenetic tree was constructed for strains with high similarity (see Figure 5 for details), and the results showed that the strain belonged to the genus Pseudomonas, and it was named Pseudomonas sp. H1.
本发明还提供了一种复合菌剂在促进大豆植株生长中的应用，使用高地芽孢杆菌Y1菌剂和假单胞菌H1菌剂混合的复合菌剂作为菌肥施用，植株为大豆植株。高地芽孢杆菌Y1保藏编号为CCTCC NO:M 20211043；假单胞菌H1的保藏编号为CCTCC NO:M 20211042。The present invention also provides an application of a composite bacterial agent in promoting the growth of soybean plants, using a composite bacterial agent mixed with highlanderia Y1 bacterial agent and Pseudomonas H1 bacterial agent as bacterial fertilizer, and the plants are soybean plants. The preservation number of Bacillus highlander Y1 is CCTCC NO:M 20211043; the preservation number of Pseudomonas H1 is CCTCC NO:M 20211042.
本发明还提供了一种提高益生微生物种群相对丰度的方法，使用高地芽孢杆菌Y1和假单胞菌H1混合的复合菌剂作为菌肥施用。高地芽孢杆菌Y1保藏编号为CCTCC NO:M 20211043；假单胞菌H1的保藏编号为CCTCC NO:M 20211042。The present invention also provides a method for increasing the relative abundance of probiotic microbial populations, using a composite microbial agent mixed with highlanderia Y1 and pseudomonas H1 as bacterial fertilizer. The preservation number of Bacillus highlander Y1 is CCTCC NO:M 20211043; the preservation number of Pseudomonas H1 is CCTCC NO:M 20211042.
进一步地，本发明的复合菌剂按照以下步骤进行制备并施入大豆盆栽中:Further, the composite microbial agent of the present invention is prepared according to the following steps and applied in soybean potted plants:
1)从保存完好的斜面中分别挑取一假单胞菌H1、高地芽孢杆菌Y1的菌体接种在不同的营养琼脂固体培养基中，25℃活化48h。1) Pseudomonas H1 and Bacillus highlander Y1 were picked from the well-preserved slant and inoculated in different nutrient agar solid media, and activated at 25°C for 48 hours.
2)将活化后的假单胞菌H1、高地芽孢杆菌Y1菌株分别用接种环挑取一环菌泥加入到不同LB液体培养基中，25℃，200r/min恒温震荡24h制备种子液。2) Pick a loop of sludge from the activated Pseudomonas H1 and Bacillus highlander Y1 strains with an inoculation loop, respectively, and add them to different LB liquid culture medium, and shake at 25°C and 200r/min for 24h to prepare seed liquid.
3)将两种种子液按5％的接种量接种在不同YMB培养基中进行扩增培养，25℃，200r/min摇床振荡培养2-3d通过加无菌水稀释或继续发酵保证菌液OD600为0.8-1.2，即得两种发酵菌液。3) Inoculate the two seed liquids with 5% inoculum amount in different YMB medium for expansion culture, 25°C, 200r/min shaker culture for 2-3d, dilute with sterile water or continue to ferment to ensure the bacterial liquid When the OD600 is 0.8-1.2, two kinds of fermentation broths are obtained.
4)将上述单独培养的两种发酵菌液按1：1等体积比混匀，用紫外分光光度计测量OD600，通过加无菌水稀释或继续发酵保证菌液OD600值在0.8～1.2范围内即得复合菌株发酵液，然后密封储存在4℃冰箱中备用；将上述的发酵液用无菌水稀释100倍加入大豆盆栽根部土壤中，每盆6mL。4) Mix the above-mentioned two kinds of fermented bacteria cultured separately according to the equal volume ratio of 1:1, measure the OD600 with an ultraviolet spectrophotometer, and ensure that the OD600 value of the bacterial liquid is in the range of 0.8 to 1.2 by diluting with sterile water or continuing to ferment The fermented broth of the compound strain was obtained, and then sealed and stored in a refrigerator at 4°C for later use; the above-mentioned fermented broth was diluted 100 times with sterile water and added to the root soil of soybean pots, 6 mL per pot.
5)对照采用等体积的稀释100倍的Y1发酵菌液、稀释100倍的H1发酵菌液、无菌水三种处理。5) As a control, the same volumes of 100-fold diluted Y1 fermentation broth, 100-fold diluted H1 fermentation broth, and sterile water were used for three treatments.
有益效果Beneficial effect
本发明提供了一种可促进大豆生长的复合菌剂，将此复合菌剂使用到大豆植株中，能有效增加大豆地上部分生物量，促进大豆生长发育，发展前景可期。具体为：The invention provides a composite bacterial agent capable of promoting soybean growth. Using the composite bacterial agent in soybean plants can effectively increase the biomass of the aboveground part of soybean, promote the growth and development of soybean, and has a promising development prospect. Specifically:
在大豆生长指标上on soybean growth indicators
大豆幼苗经H复合菌剂处理后，地上部分生物量平均为2.20g；株高平均为32.87cm；茎粗平均为4.12mm，与空白对照组相比地上生物量增加60.58％、株高增加34.99％、茎粗增加72.38％；相较于H1菌剂处理组地上生物量增加51.72％、株高增加20.31％、茎粗增加10.75％，相较于Y1菌剂处理组地上生物量增加22.22％、株高增加9.02％、茎粗增加12.56％。After the soybean seedlings were treated with H compound bacterial agent, the average aboveground biomass was 2.20g; the average plant height was 32.87cm; the average stem diameter was 4.12mm. Compared with the blank control group, the aboveground biomass increased by 60.58%, and the plant height increased by 34.99% %, stem diameter increased by 72.38%; compared with the H1 bacterial agent treatment group, the aboveground biomass increased by 51.72%, the plant height increased by 20.31%, and the stem diameter increased by 10.75%. Compared with the Y1 bacterial agent treatment group, the aboveground biomass increased by 22.22%, The plant height increased by 9.02%, and the stem diameter increased by 12.56%.
在大豆根际土壤理化性质上On the physical and chemical properties of soybean rhizosphere soil
复合菌剂处理H组的盆栽有效磷及碱解氮浓度分别为3.49mg/kg和486.00mg/kg，相较于空白组C组其有效磷含量显著增加66.96％(P＜0.05)，碱解氮增加29.14％，pH由7.13降低至6.73土壤有一定程度的酸化；相较于单一菌剂J组、Y组其有效磷含量分别增加22.03％、32.20％(P＜0.05)，碱解氮分别增加4.59％、10.54％，pH均有一定程度的下降。The concentration of available phosphorus and alkaline hydrolyzed nitrogen in group H treated with compound bacterial agent was 3.49mg/kg and 486.00mg/kg respectively, compared with blank group C, the available phosphorus content increased significantly by 66.96% (P<0.05), and the concentration of alkaline hydrolyzed Nitrogen increased by 29.14%, and the pH decreased from 7.13 to 6.73. The soil was acidified to a certain extent; compared with the single bacterial agent J group and Y group, the available phosphorus content increased by 22.03% and 32.20% respectively (P<0.05), and the alkali-hydrolyzed nitrogen was respectively Increased by 4.59% and 10.54%, the pH decreased to a certain extent.
在大豆根部土壤微生物种群丰度上On the abundance of soybean root soil microbial population
在门水平上at door level
(1)与空白对照组相比，在经复合菌剂处理的大豆根部土壤微生物种群里，变形菌门细菌含量由61.22％减少到38.18％，而放线菌门细菌由10.54％增加到30.39％。(1) Compared with the blank control group, the content of Proteobacteria decreased from 61.22% to 38.18%, while Actinomycetes increased from 10.54% to 30.39%. .
(2)与H1菌剂组、Y1菌剂组相比，经复合菌剂H组处理的大豆根部土壤微生物种群中，厚壁菌门细菌含量达到9.27％，显著低于H1菌剂组37.6％和Y1菌剂组60.96％比例；放线菌门细菌含量达到30.39％，显著高于H1组7.53％、Y1组0的比例；蓝藻菌门细菌含量为0.67％，低于H1菌剂组12.86％和Y1菌剂组7.88％比例；拟杆菌门细菌含量为12.92％，高于H1菌剂组4.56％和Y1菌剂组0.87％比例。(2) Compared with the H1 bacterial agent group and the Y1 bacterial agent group, in the soybean root soil microbial population treated with the compound bacterial agent H group, the content of Firmicutes reached 9.27%, which was significantly lower than the 37.6% of the H1 bacterial agent group Compared with Y1 bacterial agent group 60.96%; Actinomycetes bacterial content reached 30.39%, significantly higher than H1 group 7.53%, Y1 group 0; Cyanobacteria bacterial content was 0.67%, lower than H1 bacterial agent group 12.86% Compared with the ratio of 7.88% in the Y1 bacterial agent group; the Bacteroidetes bacteria content was 12.92%, which was higher than the 4.56% in the H1 microbial agent group and the 0.87% ratio in the Y1 microbial agent group.
在属水平上at the genus level
(1)与空白对照组相比，在复合菌剂处理的大豆根部土壤微生物种群中，马杜拉放线菌属细菌由3.29％增加到11.02％，卵黄杆菌属细菌由1.63％增加到5.97％。(1) Compared with the blank control group, in the soybean root soil microbial population treated with the compound bacterial agent, the bacteria of the genus Madura increased from 3.29% to 11.02%, and the bacteria of the genus Oyolk increased from 1.63% to 5.97% .
(2)与H1菌剂组、在Y1菌剂组相比，复合菌剂组H组处理的大豆根部土壤微生物种群中，马杜拉放线菌属细菌含量为11.02％，高于H1菌剂组1％和Y1菌剂组0的比例，地芽孢杆菌属细菌含量为0，低于H1菌剂组16.13％和Y1菌剂组40.05％的比例。(2) Compared with the H1 bacterial agent group and the Y1 bacterial agent group, the content of Actinomyces madurai bacteria in the soybean root soil microbial population treated by the compound microbial agent group H group was 11.02%, which was higher than that of the H1 bacterial agent group The proportion of group 1% and Y1 bacterial agent group 0, the content of Geobacillus bacteria was 0, which was lower than the ratio of H1 microbial agent group 16.13% and Y1 microbial agent group 40.05%.
在大豆根部微生物种群丰度上On the abundance of microbial populations in soybean roots
经复合菌剂处理的大豆根部微生物种群中，根瘤菌科细菌含量为18.88％，高于空白对照组 5.63％含量；伯克氏菌科细菌含量为11.29％，低于空白对照组27.15％含量；小单胞菌科细菌含量为0，低于空白对照组30.15％含量。In the soybean root microbial population treated with the compound bacterial agent, the content of Rhizobiaceae bacteria was 18.88%, which was 5.63% higher than that of the blank control group; the content of Burkholderiaceae bacteria was 11.29%, which was lower than the content of 27.15% of the blank control group; The micromonas bacteria content was 0, which was 30.15% lower than that of the blank control group.
经由上述的技术方案可知，本发明公开提供了一种对大豆促生和提高益生微生物种群丰度的复合菌剂，取得的技术效果为本发明提供的复合菌剂施用后对大豆植株的促生作用，尤其是促进大豆植株地上生物量、苗高；提高了大豆根部根瘤菌科(Rhizobium)和德沃斯氏菌科细菌的相对丰度，使共生固氮能力得到了有效发挥。It can be seen from the above-mentioned technical scheme that the present invention discloses a composite bacterial agent that promotes the growth of soybeans and increases the abundance of probiotic microbial populations. Effects, especially to promote aboveground biomass and seedling height of soybean plants; increase the relative abundance of Rhizobium and Devosbacteriaceae bacteria in soybean roots, so that the symbiotic nitrogen fixation ability has been effectively exerted.
附图说明Description of drawings
为了更清楚地说明本发明实施例或现有技术中的技术方案，下面将对本发明所需要使用的附图作简单地介绍。In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following will briefly introduce the accompanying drawings required by the present invention.
图1为YMB培养基试管中的三种菌剂，左为Y1发酵菌液，中间为H1发酵菌液，右为H1和Y1复合菌株发酵液。Figure 1 shows the three bacterial agents in the YMB medium test tube, the left is the Y1 fermentation broth, the middle is the H1 fermentation broth, and the right is the H1 and Y1 composite strain fermentation broth.
图2为YMA培养基平板上的高地芽孢杆菌Y1菌落图。Fig. 2 is a colony diagram of Bacillus alboteria Y1 on the YMA medium plate.
图3为YMA培养基平板上的假单胞菌H1菌落图。Fig. 3 is a colony diagram of Pseudomonas H1 on a YMA medium plate.
图4为本发明提供的高地地孢杆菌Y1菌株进化树图。Fig. 4 is a phylogenetic tree diagram of Geobacillus highlander Y1 strain provided by the present invention.
图5为本发明提供的假单胞菌H1菌株进化树图。Fig. 5 is a phylogenetic tree diagram of the Pseudomonas H1 strain provided by the present invention.
图6为本发明提供的在NA培养基平板上假单胞菌H1和高地地孢杆菌Y1菌株拮抗实验结果图。Fig. 6 is a graph showing the results of antagonism experiments on Pseudomonas H1 and Geobacillus highlander Y1 strains on the NA medium plate provided by the present invention.
图7为本发明提供的复合菌剂H组处理的盆栽有效磷、碱解氮浓度和pH变化示意图。Fig. 7 is a schematic diagram of changes in the concentration of available phosphorus, alkaline nitrogen and pH of potted plants treated with the compound bacterial agent H group provided by the present invention.
图8为本发明提供的对照组和经复合菌剂H组处理的盆栽土中，在门水平上微生物群落组成示意图。Fig. 8 is a schematic diagram of the microbial community composition at the phylum level in the control group provided by the present invention and the potting soil treated with the compound microbial agent H group.
图9为本发明提供的对照组和经复合菌剂H组处理的盆栽土中，在属水平上微生物群落组成示意图。Fig. 9 is a schematic diagram of the microbial community composition at the genus level in the control group provided by the present invention and the potting soil treated with the compound bacterial agent H group.
图10为本发明提供的对照组和复合菌剂H组处理的大豆根部中，在科水平上微生物群落组成示意图。Fig. 10 is a schematic diagram of the microbial community composition at the family level in soybean roots treated with the control group and the compound microbial agent H group provided by the present invention.
图11为本发明提供的对照组和复合菌剂H组处理的盆栽大豆生长情况对比图。Fig. 11 is a comparison chart of the growth of potted soybeans treated by the control group and the compound bacterial agent H group provided by the present invention.
具体实施方式Detailed ways
下面将结合本发明实施例中的附图，对本发明实施例中的技术方案进行清楚、完整地描述，显然，所描述的实施例仅是本发明一部分实施例，而不是全部的实施例。基于本发明中的实施例，本领域普通技术人员在没有做出创造性劳动前提下所获得的所有其他实施例，都属于本发明保护的范围。The following will clearly and completely describe the technical solutions in the embodiments of the present invention with reference to the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only some of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by persons of ordinary skill in the art without making creative efforts belong to the protection scope of the present invention.
下述实施例中的各试剂为市售产品，而配制菌剂的两种菌体假单胞菌(Pseudomonas sp.) H1、高地芽孢杆菌(Bacillus altitudinis)Y1均为本实验室分离所得。假单胞菌(Pseudomonas sp.)H1由本实验室从野生大豆根瘤中分离、筛选所得，由中国典型培养物保藏中心(CCTCC)保藏，保藏单位地址为中国武汉，武汉大学，保藏编号为CCTCC NO:M 20211042，保藏日期为2021年9月2日。高地芽孢杆菌(Bacillus altitudinis)Y1由本实验室从野生大豆根瘤中分离、筛选所得，由中国典型培养物保藏中心(CCTCC)保藏，保藏单位地址为中国武汉，武汉大学,保藏编号为保藏编号为CCTCC NO:M 20211043，保藏日期为2021年9月2日。The reagents in the following examples are commercially available products, and the two bacteria Pseudomonas sp. H1 and Bacillus altitudinis Y1 used to prepare the bacterial preparations were all isolated in our laboratory. Pseudomonas sp. H1 was isolated and screened from wild soybean root nodules in our laboratory, and was preserved by China Center for Type Culture Collection (CCTCC). The address of the preservation unit is Wuhan University, Wuhan, China, and the preservation number is CCTCC NO : M 20211042, the deposit date is September 2, 2021. Bacillus altitudinis (Bacillus altitudinis) Y1 was isolated and screened from wild soybean root nodules in our laboratory, and was preserved by the China Center for Type Culture Collection (CCTCC). The address of the preservation unit is Wuhan, China, Wuhan University, and the preservation number is CCTCC NO: M 20211043, the deposit date is September 2, 2021.
实施例1Example 1
两种菌株拮抗性探究Antagonism study of two strains
1培养基1 medium
(1)营养琼脂固体培养基的组成和含量为：蛋白胨10g、牛肉浸粉3g、氯化钠5g、琼脂15g、1000ml去离子水，pH 7.2-7.4。(1) The composition and content of nutrient agar solid medium are: peptone 10g, beef extract powder 3g, sodium chloride 5g, agar 15g, 1000ml deionized water, pH 7.2-7.4.
(2)LB液体培养基的组成和配比方法为：容器内加入去离子水950ml、胰蛋白胨10g、酵母浸粉5g、氯化钠5g，摇动容器直至溶质溶解，用5mol/L氢氧化钠调节溶液pH至7.4，定容至1L。(2) The composition and proportioning method of LB liquid medium is as follows: add 950ml of deionized water, 10g of tryptone, 5g of yeast extract powder, and 5g of sodium chloride into the container, shake the container until the solute dissolves, and add 5mol/L sodium hydroxide Adjust the pH of the solution to 7.4, and dilute to 1L.
2拮抗实验步骤2 Antagonism experiment steps
假单胞菌H1、高地芽孢杆菌Y1分别用LB液体培养基中活化后，30℃、180r/min，摇床培养24h，取H1菌液5μL点解在在营养琼脂固体培养基平板中心位置，将5μLY1点接在营养琼脂固体培养基平板的4个边缘位置，30℃培养2-3d。Pseudomonas H1 and Bacillus highlander Y1 were respectively activated in LB liquid medium, cultured on a shaking table at 30°C and 180r/min for 24 hours, and 5 μL of H1 bacterial solution was spot-dissolved on the center of the nutrient agar solid medium plate. Spot 5 μL Y1 on the 4 edges of the nutrient agar solid medium plate, and culture at 30°C for 2-3 days.
结果如图6显示，假单胞菌菌株H1与高地芽孢杆菌菌株Y1没有产生明显的拮抗圈，说明相互之间不拮抗。The results are shown in Figure 6, Pseudomonas strain H1 and Bacillus highlander strain Y1 did not produce obvious antagonistic circles, indicating that they were not antagonistic to each other.
实施例2Example 2
复合菌剂促生大豆的能力Ability of Compound Bacteria to Promote Growth of Soybean
1制备单一菌种菌剂和复合菌剂1 Preparation of single strain bacterial agent and composite bacterial agent
1.1培养基1.1 Medium
(1)营养琼脂固体培养基的组成和含量为：蛋白胨10g、牛肉浸粉3g、氯化钠5g、琼脂15g、1000ml去离子水，pH 7.2-7.4。(1) The composition and content of nutrient agar solid medium are: peptone 10g, beef extract powder 3g, sodium chloride 5g, agar 15g, 1000ml deionized water, pH 7.2-7.4.
(2)LB液体培养基的组成和配比方法为：容器内加入去离子水950ml、胰蛋白胨10g、酵母浸粉5g、氯化钠5g，摇动容器直至溶质溶解，用5mol/L氢氧化钠调节溶液pH至7.4，定容至1L。(2) The composition and proportioning method of LB liquid medium is as follows: add 950ml of deionized water, 10g of tryptone, 5g of yeast extract powder, and 5g of sodium chloride into the container, shake the container until the solute dissolves, and add 5mol/L sodium hydroxide Adjust the pH of the solution to 7.4, and dilute to 1L.
(3)YMB培养基的组成和含量为：甘露醇10g、七水硫酸镁0.2g、氯化钠0.1g、酵母粉3g、磷酸氢二钾0.25g、磷酸二氢钾0.25g、碳酸钙3g、1000ml去离子水，pH 7.2-7.4。(3) The composition and content of YMB medium are: mannitol 10g, magnesium sulfate heptahydrate 0.2g, sodium chloride 0.1g, yeast powder 3g, dipotassium hydrogen phosphate 0.25g, potassium dihydrogen phosphate 0.25g, calcium carbonate 3g , 1000ml deionized water, pH 7.2-7.4.
1.2制备菌剂的步骤1.2 Steps for preparing bacterial agent
1)从保存完好的斜面中分别挑取一假单胞菌H1、高地芽孢杆菌Y1的菌体接种在营养琼脂固体培养基中，25℃活化48h。1) Pseudomonas H1 and Bacillus highlander Y1 were respectively picked from the well-preserved slant and inoculated in nutrient agar solid medium, and activated at 25°C for 48 hours.
2)将活化后的假单胞菌H1、高地芽孢杆菌Y1菌株分别用接种环挑取一环菌泥加入到LB液体培养基中，25℃，200r/min恒温震荡24h制备种子液。2) Pick a loop of sludge from the activated Pseudomonas H1 and Bacillus highlander Y1 strains with an inoculation loop, respectively, and add them to LB liquid medium, and shake at 25°C and 200r/min for 24 hours to prepare seed liquid.
3)将种子液按3％的接种量接种在YMB培养基中进行扩增培养，25℃，200r/min摇床振荡培养2-3d通过加无菌水稀释或继续发酵至发酵液OD600为0.8～1.2，即得两种发酵菌液。3) Inoculate the seed solution with 3% inoculation amount in YMB medium for expansion culture, 25°C, 200r/min shaker culture for 2-3d, add sterile water to dilute or continue to ferment until the OD600 of the fermentation liquid is 0.8 ~1.2, two kinds of fermentation broths were obtained.
4)将上述单独培养的两种发酵菌液按1：1混合后用紫外分光光度计测量OD600，通过加无菌水稀释或继续发酵保证菌液OD600值在0.8～1.2范围内即得复合菌株发酵液，然后密封储存在4℃冰箱中备用；将上述的发酵液用无菌水稀释100倍即得到复合菌剂。4) Mix the two kinds of fermented bacterial liquids cultured separately above at a ratio of 1:1, measure the OD600 with a UV spectrophotometer, dilute with sterile water or continue fermentation to ensure that the OD600 value of the bacterial liquid is within the range of 0.8 to 1.2 to obtain a composite strain The fermented liquid is then sealed and stored in a refrigerator at 4°C for later use; the above-mentioned fermented liquid is diluted 100 times with sterile water to obtain a composite bacterial agent.
5)对照组采用等体积的稀释100倍的Y1发酵菌液、稀释100倍的H1发酵菌液、无菌水三种处理。5) The control group was treated with equal volumes of 100-fold diluted Y1 fermentation broth, 100-fold diluted H1 fermentation broth, and sterile water.
2大豆幼苗种植2 Planting of soybean seedlings
本发明的试验对象为盆栽大豆。首先用次氯酸钠溶液对种子灭菌，然后用浸种法进行催芽。待幼芽长出后，挑取长势较好的幼芽栽植于花盆中，盆栽所用的土壤为江苏兴农基质科技有限公司所生产的育苗基质土。每盆栽入3株幼芽，放置在简易大棚。在长出第一片真叶时，每盆植物根部各接种6mL复合菌剂，以不接种和接种6mL单菌种H1、Y1菌剂为对照。每三天喷洒一次营养液保持湿润。待生长一个月后进行间苗，每盆保留一株健壮的幼苗(每盆长势一致)，保持三天喷施一次营养液。45天后，测得其地上部分与地下部分生长情况。The test object of the present invention is potted soybean. First sterilize the seeds with sodium hypochlorite solution, and then use the seed soaking method to germinate. After the sprouts grow out, pick the sprouts with better growth and plant them in flowerpots. The soil used for potting is the seedling-raising substrate soil produced by Jiangsu Xingnong Substrate Technology Co., Ltd. Three young shoots are planted in each pot and placed in a simple greenhouse. When the first true leaf grows, the root of each potted plant is inoculated with 6mL compound bacterial agent, and the control is not inoculated and inoculated with 6mL single strain H1, Y1 bacterial agent. Spray a nutrient solution every three days to keep it moist. After one month of growth, the seedlings were thinned, and a strong seedling was kept in each pot (the growth of each pot was the same), and the nutrient solution was sprayed once every three days. After 45 days, the growth conditions of its above-ground and underground parts were measured.
盆栽指标的测定和方法Determination and method of pot index
对于大豆植株地上部分：使用游标卡尺、皮尺测定幼苗的地径和苗高；使用根系扫描仪测量叶面积(每盆植株选取上中下位叶片共计10片用于测量叶面积)。For the aerial parts of soybean plants: use vernier calipers and tape measure to measure the ground diameter and seedling height of the seedlings; use a root scanner to measure the leaf area (a total of 10 upper, middle and lower leaves are selected for each potted plant to measure the leaf area).
对于大豆植株地下部分：使用根系扫描仪测量测量根系形态；测定植株的地下生物量。For the underground parts of soybean plants: measure root morphology using a root scanner; determine the belowground biomass of the plants.
对于大豆盆栽土：采用mettlertoledo ph计测定pH(水土比为5：1)；采用酸溶-钼锑抗比色法测定土壤有效磷；采用碱解扩散法测定土壤的碱解氮。For soybean potting soil: Mettlertoledo pH meter was used to measure pH (water-to-soil ratio was 5:1); soil available phosphorus was determined by acid-soluble-molybdenum-antimony anti-colorimetric method; alkali-hydrolyzed nitrogen in soil was measured by alkali-lysis diffusion method.
结果表明：使用复合菌剂H能显著增加土壤中有效磷和碱解氮含量，同时pH下降土壤有一定程度的酸化。The results showed that the use of compound microbial agent H could significantly increase the content of available phosphorus and alkaline nitrogen in the soil, and at the same time, the pH decreased and the soil was acidified to a certain extent.
对大豆植株地上部分生长的影响Effects on the Growth of the Aboveground Parts of Soybean Plants
注：P＜0.05。Note: P<0.05.
表1所示，本发明对大豆植株进行了四个处理，包括空白对照组c、Y1菌剂Y组、H1菌剂J组、复合菌剂H组。与无菌空白对照组相比，添加菌剂的三种处理中，大豆地上部分各生长指标均显著提高。其中，Y、J、H菌剂组相较于无菌剂组地上部分生物量分别增加31.39％、5.84％、60.58％，株高分别增加23.82％、12.20％、34.99％，茎粗分别增加53.14％、55.65％、72.38％。As shown in Table 1, the present invention has carried out four treatments to soybean plants, including blank control group c, Y1 bacterial agent Y group, H1 bacterial agent J group, and compound bacterial agent H group. Compared with the sterile blank control group, in the three treatments of adding bacteria agents, the growth indexes of soybean aerial parts were significantly improved. Among them, compared with the sterile agent group, the above-ground biomass of the Y, J, and H bacterial agent groups increased by 31.39%, 5.84%, and 60.58%, respectively, the plant height increased by 23.82%, 12.20%, and 34.99%, and the stem diameter increased by 53.14%. %, 55.65%, 72.38%.
从表1可知，与添加单一菌株的J、Y组相比，复合菌剂组H组中大豆的促生效果最显著，其地上生物量、株高、茎粗分别为2.2g、32.87cm、4.12mm，相较于J处理组地上生物量增加51.72％、株高增加20.31％、茎粗增加10.75％，相较于Y处理组地上生物量增加22.22％、株高增加9.02％、茎粗增加12.56％。It can be seen from Table 1 that compared with groups J and Y added with a single strain, the growth-promoting effect of soybeans in group H of the compound bacterial agent group was the most significant, and its aboveground biomass, plant height, and stem diameter were 2.2 g, 32.87 cm, and 32.87 cm, respectively. 4.12mm, compared with the J treatment group, the aboveground biomass increased by 51.72%, the plant height increased by 20.31%, and the stem diameter increased by 10.75%. Compared with the Y treatment group, the aboveground biomass increased by 22.22%, the plant height increased by 9.02%, and the stem diameter increased 12.56%.
表1不同菌剂处理后大豆植株地上部分生长情况比较Table 1 Comparison of the growth of the above-ground parts of soybean plants after different bacterial agents
Figure PCTCN2022096584-appb-000001
Figure PCTCN2022096584-appb-000001
对大豆植株地下部分根长的影响Effects on Root Length of Underground Parts of Soybean Plants
注：P＜0.05。Note: P<0.05.
表2所示，本发明对大豆植株进行了四个处理，包括空白对照组C、Y1菌剂组Y组、H1菌剂J组、复合菌剂H组。与无菌空白对照组相比，添加菌剂的三种处理中，大豆地下部分根长显著提高。As shown in Table 2, the present invention has carried out four treatments to soybean plants, including blank control group C, Y1 bacterial agent group Y group, H1 bacterial agent J group, and compound bacterial agent H group. Compared with the sterile blank control group, the root length of the underground part of soybean was significantly increased in the three treatments of adding bacteria.
从表2可知，与添加单一菌株的J、Y菌剂组相比，复合菌剂组H组中大豆的促生效果最显著，根长为9.15cm，相比于J组增加24.83％，相较于Y组增加9.84％。It can be seen from Table 2 that compared with the J and Y bacterial agent groups added with a single strain, the growth-promoting effect of soybean in the compound bacterial agent group H group was the most significant, and the root length was 9.15cm, which was 24.83% higher than that of the J group. Compared with group Y, it increased by 9.84%.
表2不同菌剂处理后大豆植株地下部分根长比较Table 2 Comparison of the root lengths of the underground parts of soybean plants after different microbial treatments
Figure PCTCN2022096584-appb-000002
Figure PCTCN2022096584-appb-000002
对盆栽土壤理化性质的影响Effects on Physical and Chemical Properties of Potted Soil
由表3和图7可知，复合菌剂处理H组的盆栽有效磷及碱解氮浓度分别为3.49mg/kg和 486.00mg/kg，相较于空白组C组其有效磷含量显著增加66.96％(P＜0.05)，碱解氮增加29.14％，土壤有一定程度的酸化pH由7.13降低至6.73；相较于单一菌剂J组、Y组其有效磷含量分别增加22.03％、32.20％(P＜0.05)，碱解氮分别增加4.59％、10.54％，pH均有一定程度的下降。It can be seen from Table 3 and Figure 7 that the concentration of available phosphorus and alkali-hydrolyzed nitrogen in group H treated with compound bacteria agent was 3.49mg/kg and 486.00mg/kg respectively, and the available phosphorus content in group C significantly increased by 66.96% compared with blank group C (P<0.05), the alkaline hydrolysis nitrogen increased by 29.14%, and the pH of the soil acidified to a certain extent decreased from 7.13 to 6.73; compared with the single bacterial agent J group and Y group, the available phosphorus content increased by 22.03% and 32.20% respectively (P <0.05), the alkaline hydrolysis nitrogen increased by 4.59% and 10.54%, respectively, and the pH decreased to a certain extent.
表3不同菌剂处理后大豆盆栽土壤理化性质比较Table 3 Comparison of physical and chemical properties of soybean potting soil after different bacterial agents
Figure PCTCN2022096584-appb-000003
Figure PCTCN2022096584-appb-000003
实施例3Example 3
探究复合菌剂对大豆根际土壤微生物群落组成的影响。To explore the effect of compound bacterial agent on the microbial community composition of soybean rhizosphere soil.
本发明涉及四组处理组：无菌对照组C组、H1菌剂组J组、Y1菌剂Y组、复合菌剂H组，将收集整理好的四组大豆根际土样送至广州基迪奥有限公司进行测序，通过高通量测序的方法分析盆栽土中微生物多样性，探究盆栽土的微生物群落组成。The present invention involves four groups of treatment groups: aseptic control group C, H1 bacterial agent group J, Y1 bacterial agent Y group, and compound bacterial agent H group. The collected four groups of soybean rhizosphere soil samples were sent to Guangzhou Base Dior Co., Ltd. conducts sequencing, analyzes the microbial diversity in potting soil through high-throughput sequencing, and explores the microbial community composition of potting soil.
微生物门水平分析Microbial phylum level analysis
由附图8和表4结果分析各处理在细菌门水平上最大丰度排名前5的物种，变形菌门(Proteobacteria，26.96％～61.22％)、放线菌门(Actinobacteria，0.01％～30.39％)、厚壁菌门(Firmicutes，7.20％～60.96％)、拟杆菌门(Bacteroidetes，0.87％～12.92％)、蓝藻菌门(Cyanobacteria，0.67％～12.86％)是主要的优势菌群，在各处理中的相对丰度为88.22％～96.82％。施加复合菌剂H后Proteobacteria相对丰度比空白组C组降低但是高于单一菌剂组J、Y组。同时H组Actinobacteria相对丰度均高于C、J、Y组，Actinobacteria是可以产生抗生素的一类细菌，能促使土壤中的动物和植物遗骸腐烂。同时，Firmicutes相对丰度比J、Y组降低，比C组提高，Firmicutes产生的可逆性孢子，可以抵抗极端环境，Firmicutes包括芽孢杆菌属，丰度显著提升可能与本实验施加高地芽孢杆菌Y1有关。对比J、Y组，H组Proteobacteria、Actinobacteria、Bacteroidetes相对丰度均提高，Firmicutes、Cyanobacteria相对丰度均降低。对比空白组C组，H组Proteobacteria相对丰度降低，Actinobacteria、Firmicutes、Bacteroidetes相对丰度均提高。By accompanying drawing 8 and table 4 result analysis each handles the species of the top 5 maximum abundances on the bacterial phylum level, Proteobacteria (Proteobacteria, 26.96%～61.22%), Actinobacteria (Actinobacteria, 0.01%～30.39%) ), Firmicutes (7.20%-60.96%), Bacteroidetes (0.87%-12.92%), and Cyanobacteria (0.67%-12.86%) are the main dominant flora. The relative abundance in the treatment ranged from 88.22% to 96.82%. The relative abundance of Proteobacteria after compound bacterial agent H was lower than that of blank group C but higher than that of single bacterial agent group J and Y. At the same time, the relative abundance of Actinobacteria in group H was higher than that in groups C, J, and Y. Actinobacteria is a type of bacteria that can produce antibiotics and can promote the decay of animal and plant remains in the soil. At the same time, the relative abundance of Firmicutes was lower than that of J and Y groups, and higher than that of C group. The reversible spores produced by Firmicutes can resist extreme environments. Firmicutes include Bacillus, and the significant increase in abundance may be related to the application of Bacillus uplander Y1. . Compared with groups J and Y, the relative abundance of Proteobacteria, Actinobacteria, and Bacteroidetes in group H increased, while the relative abundance of Firmicutes and Cyanobacteria decreased. Compared with the blank group C, the relative abundance of Proteobacteria in group H decreased, while the relative abundance of Actinobacteria, Firmicutes, and Bacteroidetes increased.
表4门水平上大豆根际土壤主要微生物群落组成比较Table 4 Comparison of main microbial communities in soybean rhizosphere soil at phylum level
Figure PCTCN2022096584-appb-000004
Figure PCTCN2022096584-appb-000004
Figure PCTCN2022096584-appb-000005
Figure PCTCN2022096584-appb-000005
微生物属水平分析Microbial genus level analysis
如附图9和表5所示，在已知的细菌属水平上最大丰度排名前5的物种，地芽孢杆菌属(Geobacillus，0.00％～61.22％)、马杜拉放线菌属(Actinomadura，0.00％～11.02％)、芽孢杆菌属(Bacillus，2.66％～11.95％)、藤黄色单胞菌属(Luteimonas，0.31％～5.97％)是主要已知的优势菌群，在各处理中的相对丰度为11.91％～52.31％。施加复合菌剂H后Actinomadura相对丰度比空白组C组、单一菌剂组J、Y组均提高。Actinomadura具有抗菌作用，有利于生产抗生素药物。同时H组Bacillus相比于空白组C组提高，相比于菌剂组J、Y组降低。此外，H组Luteimonas均高于C、J、Y组。Luteimonas代谢产物具有抗菌作用，对大肠杆菌、金黄色葡萄球菌具有抑制作用。对比菌剂组J、Y组，H组Actinomadura、Luteimonas相对丰度均提高，Bacillus、Geobacillu相对丰度均降低。对比空白组C组，H组Actinomadura、Bacillus、Luteimonas相对丰度均提高。As shown in Figure 9 and Table 5, the top 5 species with the highest abundance at the level of known bacterial genus are Geobacillus (0.00%-61.22%), Actinomadura , 0.00%～11.02%), Bacillus (2.66%～11.95%), Luteimonas (Luteimonas, 0.31%～5.97%) are the main known dominant flora, in each treatment The relative abundance is 11.91%-52.31%. The relative abundance of Actinomadura after compound microbial agent H was higher than that of blank group C, single bacterial agent group J, and Y group. Actinomadura has an antibacterial effect and is beneficial for the production of antibiotic drugs. At the same time, Bacillus in group H increased compared with blank group C, and decreased compared with bacterial agent groups J and Y. In addition, Luteimonas in group H were higher than those in groups C, J, and Y. The metabolites of Luteimonas have antibacterial effects and have inhibitory effects on Escherichia coli and Staphylococcus aureus. Compared with bacterial agent groups J and Y, the relative abundances of Actinomadura and Luteimonas in H group increased, while the relative abundances of Bacillus and Geobacillu decreased. Compared with the blank group C, the relative abundance of Actinomadura, Bacillus, and Luteimonas in the H group increased.
表5属水平上大豆根际土壤主要微生物群落组成比较Table 5 Comparison of main microbial communities in soybean rhizosphere soil at the genus level
Figure PCTCN2022096584-appb-000006
Figure PCTCN2022096584-appb-000006
实施例4Example 4
探究菌株对大豆根部微生物群落组成的影响。To investigate the effect of bacterial strains on the composition of soybean root microbial community.
将收集整理好的大豆根际土样送至广州基迪奥有限公司进行测序，通过高通量测序的方法分析大豆植株中微生物多样性，探究大豆根部的微生物群落组成。The collected soybean rhizosphere soil samples were sent to Guangzhou Gideo Co., Ltd. for sequencing, and high-throughput sequencing was used to analyze the microbial diversity in soybean plants and explore the composition of microbial communities in soybean roots.
微生物科水平分析Microbiology level analysis
本发明涉及两组处理组：无菌对照组c、复合菌剂H组，将收集整理好的两组大豆根际样本送至广州基迪奥有限公司进行检测，通过高通量测序的方法分析根部微生物多样性，探究大都根部的微生物群落组成。由附图10和表6可以看出，经复合菌剂处理的大豆根部微生物种群中，在细菌科水平上最大丰度排名前7的物种为黄单胞菌科(Xanthomonadaceae)、伯克氏菌科(Burkholderiaceae)、鞘脂单胞菌科(Sphingomonadaceae)、嗜甲基菌科(Methylophilaceae)、德沃斯氏菌科(Devosiaceae)、假单胞菌科(Pseudomonadaceae)、根瘤菌科(Rhizobiaceae)。相较于空白组C组，H组在前7种主要菌的相对丰度 上均提高。Rhizobiaceae菌种具有固氮作用，复合菌剂组H组Rhizobiaceae相对丰度为18.88％，高于空白无菌对照组5.63％相对丰度。Devosiaceae也是一种固氮细菌，复合菌剂组H组Devosiaceae相对丰度为9.58％，高于空白无菌对照组1.17％相对丰度。伯克氏菌科细菌相对丰度为11.29％，低于空白对照组27.15％相对丰度。Pseudomonadaceae是一种可以促进植物生长的共生菌，复合菌剂组H组Pseudomonadaceae相对丰度为5.16％，高于空白无菌对照组2.29％相对丰度。Sphingomonadaceae是一种可以促进植物抗重金属胁迫酶活性的菌种，同时有利于植物降解多环芳烃、农药。复合菌剂组H组Sphingomonadaceae相对丰度为8.12％，高于空白无菌对照组3.00％相对丰度。Xanthomonadaceae是一种植物病原细菌，一些物种能够激活植物内生菌群的抑病功能，同时一些种具有生产黄原胶的潜力，后者具有使微生物能够吸附到生物表面，保护自身免受干燥和疏水分子的影响，浓缩营养并固定有毒元素的价值。相比于空白无菌对照组4.63％相对丰度，复合菌剂组H组Xanthomonadaceae相对丰度为14.11％。The present invention involves two groups of treatment groups: sterile control group c and compound microbial agent H group. The collected and arranged two groups of soybean rhizosphere samples are sent to Guangzhou Kidio Co., Ltd. for detection, and the roots are analyzed by high-throughput sequencing. Microbial diversity, exploring the composition of microbial communities in most roots. As can be seen from Figure 10 and Table 6, among the soybean root microbial populations treated with compound bacterial agents, the top 7 species with the highest abundance at the level of bacterial families are Xanthomonadaceae, Burkholderia Burkholderiaceae, Sphingomonadaceae, Methylophilaceae, Devosiaceae, Pseudomonadaceae, Rhizobiaceae. Compared with the blank group C, the relative abundance of the first seven main bacteria in group H increased. Rhizobiaceae strains have nitrogen fixation, and the relative abundance of Rhizobiaceae in group H of the compound bacterial agent group was 18.88%, which was higher than the relative abundance of 5.63% in the blank sterile control group. Devosiaceae is also a kind of nitrogen-fixing bacteria. The relative abundance of Devosiaceae in group H of the compound bacterial agent group was 9.58%, which was higher than the relative abundance of 1.17% in the blank sterile control group. The relative abundance of Burkholderiaceae was 11.29%, which was lower than the 27.15% relative abundance of the blank control group. Pseudomonadaceae is a symbiotic bacterium that can promote plant growth. The relative abundance of Pseudomonadaceae in group H in the compound bacterial agent group was 5.16%, which was higher than that in the blank sterile control group (2.29%). Sphingomonadaceae is a strain that can promote the activity of plant anti-heavy metal stress enzymes, and at the same time help plants degrade polycyclic aromatic hydrocarbons and pesticides. The relative abundance of Sphingomonadaceae in group H of compound bacterial agent group was 8.12%, which was 3.00% higher than that of blank sterile control group. Xanthomonadaceae is a phytopathogenic bacterium, some species are able to activate the disease-suppressive function of plant endophytic flora, while some species have the potential to produce xanthan gum, which has the ability to allow microorganisms to adsorb to biological surfaces and protect themselves from desiccation and The influence of hydrophobic molecules, concentrates nutrients and fixes the value of toxic elements. Compared with the 4.63% relative abundance of the blank sterile control group, the relative abundance of Xanthomonadaceae in group H of the compound bacterial agent group was 14.11%.
表6科水平上大豆根部微生物群落组成比较Table 6 Comparison of soybean root microbial community composition at the family level
Figure PCTCN2022096584-appb-000007
Figure PCTCN2022096584-appb-000007

Claims (10)

  1. 一种复合菌剂，其特征在于，该复合菌剂中的菌株由保藏号为CCTCC NO：M 20211043的高地芽孢杆菌(Bacillus altitudinis)Y1与保藏号为CCTCC NO：M 20211042的假单胞菌(Pseudomonas sp.)H1混合组成。A composite bacterial agent, characterized in that the bacterial strain in the composite bacterial agent consists of Bacillus altitudinis Y1 with a preservation number of CCTCC NO: M 20211043 and Pseudomonas with a preservation number of CCTCC NO: M 20211042 ( Pseudomonas sp.) H1 mixed composition.
  2. 权利要求1所述的复合菌剂，其特征在于，由浓度不低于10 9个/ml的所述的高地芽孢杆菌Y1发酵液与浓度不低于10 9CFU/ml的所述的假单胞菌H1发酵液等体积比混合而成；两种发酵液的浓度相同。 The composite bacterial agent according to claim 1, characterized in that, the concentration of the Bacillus highlander Y1 fermentation broth is not lower than 10 9 /ml and the pseudo-sheet of the concentration is not lower than 10 9 CFU/ml Bacteria H1 fermentation broth is mixed in equal volume ratio; the concentration of the two fermentation broths is the same.
  3. 一种权利要求1所述的复合菌剂的制备方法，其特征在于，所述的复合菌剂按照以下步骤进行制备：A preparation method of the composite bacterial agent according to claim 1, characterized in that, the composite bacterial agent is prepared according to the following steps:
    1)从保存完好的斜面上分别挑取假单胞菌H1、高地芽孢杆菌Y1的菌体接种在营养琼脂固体培养基上，25℃活化48h；1) Pseudomonas H1 and Bacillus highlander Y1 were picked from the well-preserved slant and inoculated on the nutrient agar solid medium, and activated at 25°C for 48 hours;
    2)将活化后的假单胞菌H1、高地芽孢杆菌Y1菌株分别用接种环挑取一环菌泥加入到两个LB液体培养基中，25℃，200r/min恒温震荡24h制备种子液；2) Pick up a ring of sludge from the activated Pseudomonas H1 and Bacillus highlander Y1 strains with an inoculation loop, respectively, and add them to two LB liquid mediums, and shake at 25°C and 200r/min for 24 hours to prepare seed liquids;
    3)将两种种子液按5％的接种量接种在YMB培养基中进行扩增培养，25℃，200r/min摇床振荡培养2-3d通过稀释或继续发酵至发酵液OD 600为0.8-1.2，即得两种发酵菌液； 3) Inoculate the two seed liquids in YMB medium with 5% inoculum amount for expansion culture, 25°C, 200r/min shaker shaking culture for 2-3d by diluting or continuing to ferment until the OD600 of the fermentation liquid is 0.8- 1.2, to obtain two kinds of fermentation liquid;
    4)将上述单独培养的两种发酵菌液按1∶1混合后用紫外分光光度计测量OD 600，通过稀释或继续发酵保证菌液OD 600值在0.8～1.2范围内，即得复合菌株发酵液，然后密封储存在4℃冰箱中备用； 4) Mix the above-mentioned two kinds of fermented bacterial liquids cultured separately at a ratio of 1:1, measure the OD 600 with a UV spectrophotometer, and ensure that the OD 600 value of the bacterial liquid is within the range of 0.8 to 1.2 through dilution or continuous fermentation, and then the composite bacterial strain is fermented. solution, and then sealed and stored in a 4°C refrigerator for later use;
    5)将步骤4)中复合菌株发酵液用无菌水稀释100倍即得到复合菌剂。5) Dilute the complex bacterial strain fermentation liquid in step 4) with sterile water 100 times to obtain the complex microbial agent.
  4. 根据权利要求3所述的制备方法，其特征在于，步骤1)中，所述营养琼脂固体培养基的组成和含量为：蛋白胨10g、牛肉浸粉3g、氯化钠5g、琼脂15g、1000ml去离子水，pH 7.2-7.4。The preparation method according to claim 3, characterized in that, in step 1), the composition and content of the nutrient agar solid medium are: peptone 10g, beef extract powder 3g, sodium chloride 5g, agar 15g, 1000ml Ionized water, pH 7.2-7.4.
  5. 根据权利要求3所述的制备方法，其特征在于，步骤2)中，所述LB液体培养基的组成和配比方法为：容器内加入去离子水950ml、胰蛋白胨10g、酵母浸粉5g、氯化钠5g，摇动容器直至溶质溶解，用5mol/L氢氧化钠调节溶液pH至7.4，定容至1L。The preparation method according to claim 3, characterized in that, in step 2), the composition and proportioning method of the LB liquid medium is: add 950ml of deionized water, 10g of tryptone, 5g of yeast extract powder, Sodium chloride 5g, shake the container until the solute is dissolved, adjust the pH of the solution to 7.4 with 5mol/L sodium hydroxide, and dilute to 1L.
  6. 根据权利要求3所述的制备方法，其特征在于，步骤3)中，所述YMB培养基的组成和含量为：甘露醇10g、七水硫酸镁0.2g、氯化钠0.1g、酵母粉3g、磷酸氢二钾0.25g、磷酸二氢钾0.25g、碳酸钙3g、1000ml去离子水，pH 7.2-7.4。The preparation method according to claim 3, wherein, in step 3), the composition and content of the YMB medium are: mannitol 10g, magnesium sulfate heptahydrate 0.2g, sodium chloride 0.1g, yeast powder 3g , Dipotassium hydrogen phosphate 0.25g, potassium dihydrogen phosphate 0.25g, calcium carbonate 3g, 1000ml deionized water, pH 7.2-7.4.
  7. 权利要求1或2所述的复合菌剂在大豆植株促生和提高大豆根部益生微生物相对丰度方面的应用。The application of the compound bacterial agent described in claim 1 or 2 in promoting the growth of soybean plants and improving the relative abundance of probiotic microorganisms in soybean roots.
  8. 根据权利要求7所述的应用，其特征在于：将所述复合菌剂按每盆6mL的接种量接种到大豆植株根部土壤中进行处理。The application according to claim 7, characterized in that: the composite microbial agent is inoculated into the soil at the root of soybean plants with an inoculum amount of 6 mL per pot for treatment.
  9. 如权利要求8所述的应用，其特征在于：经所述复合菌剂处理后，大豆根部益生微生物种群相对丰度提高，根瘤菌科、德沃斯氏菌科这两种固氮菌种群丰度显著增加，大豆根部的根瘤菌科细菌相对丰度由5.63％增加到18.88％，德沃斯氏菌科细菌相对丰度由1.17提高到9.58％；促进植物生长的共生菌假单胞菌科细菌相对丰度由2.29％提高到5.16％；促进植物抗重金属胁迫酶活性的鞘脂单胞菌科细菌相对丰度由3.00％增加到8.12％；有利于植物浓缩营养并固定有毒元素的黄单胞菌科细菌相对丰度由4.63％增加至14.11％。The application according to claim 8, characterized in that: after being treated with the compound bacterial agent, the relative abundance of probiotic microbial populations in soybean roots increases, and the two nitrogen-fixing bacteria populations of Rhizobiaceae and Devosellaceae are abundant. The relative abundance of Rhizobiaceae bacteria in soybean roots increased from 5.63% to 18.88%, and the relative abundance of De Vossiaceae bacteria increased from 1.17 to 9.58%; the symbiotic bacteria Pseudomonas family The relative abundance of bacteria increased from 2.29% to 5.16%; the relative abundance of Sphingomonas bacteria that promoted the enzyme activity of heavy metal stress resistance in plants increased from 3.00% to 8.12%; The relative abundance of Mycobacteriaceae increased from 4.63% to 14.11%.
  10. 如权利要求8所述的应用，其特征在于：经所述复合菌剂处理的大豆，由于益生微生物种群丰度增加，对植物根系和地上部分具有促生作用；地上部分生物量平均为2.20g，显著增加60.58％；株高平均为32.87cm，显著增加34.99％；茎粗平均为4.12mm，显著增加72.38％；The application according to claim 8, characterized in that: the soybeans treated with the compound bacterial agent have a growth-promoting effect on plant roots and aerial parts due to the increase in the abundance of probiotic microbial populations; the average biomass of the aerial parts is 2.20g , a significant increase of 60.58%; the average plant height was 32.87cm, a significant increase of 34.99%; the average stem diameter was 4.12mm, a significant increase of 72.38%;
    根际土壤有效磷含量显著增加66.96％，碱解氮增加29.14％，pH由7.13降低至6.73。The available phosphorus content in the rhizosphere soil increased significantly by 66.96%, the alkaline nitrogen increased by 29.14%, and the pH decreased from 7.13 to 6.73.
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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN115960765A (en) * 2022-11-02 2023-04-14 云南大学 Flavobacterium bacteria and application thereof in low-temperature composting

Families Citing this family (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114015629B (en) * 2021-12-20 2022-04-29 南京林业大学 Complex microbial inoculum for promoting growth of soybeans and improving abundance of probiotic microbial populations

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20200187507A1 (en) * 2018-12-06 2020-06-18 Auburn University Inoculants including plant growth producing rhizobacteria (pgpr) and insectides for promoting plant growth and health
CN111849815A (en) * 2020-07-21 2020-10-30 广西民族大学 Plant growth promoting rhizosphere strain Gxun-20 and application thereof in plant growth promotion
CN112210510A (en) * 2020-09-18 2021-01-12 南京林业大学 Phosphate-solubilizing growth-promoting bacterium and salt-tolerant arthrobacter X-1 for promoting growth of nodule and improving abundance of probiotic microorganism population and application thereof
CN113667732A (en) * 2021-09-04 2021-11-19 山东农业大学 Plant-mediated-based screening method for microbial composite strains
CN114015629A (en) * 2021-12-20 2022-02-08 南京林业大学 Complex microbial inoculum for promoting growth of soybeans and improving abundance of probiotic microbial populations

Family Cites Families (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2015180983A1 (en) * 2014-05-27 2015-12-03 Basf Se Mixtures comprising nitrogen-fixing bacteria and biopesticides and chemical pesticides
WO2016034353A1 (en) * 2014-09-03 2016-03-10 Basf Se Pesticidally active mixtures
CN108624528B (en) * 2018-05-14 2021-01-29 华中农业大学 Composite microbial inoculum with growth promoting and yield increasing effects on leguminous plants and application thereof
CN111955484B (en) * 2020-08-25 2021-04-30 南京林业大学 Penicillium simplicissimum NL-Z1 strain for promoting growth of nodule and improving abundance of probiotic microorganism population
CN112608868B (en) * 2020-12-23 2022-10-25 上海交通大学 Bacillus altitudinis and application thereof
CN112662589B (en) * 2021-01-19 2022-08-12 南京农业大学 Development and application of plant rhizosphere biofilm co-colonization type multifunctional complex microbial inoculum

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20200187507A1 (en) * 2018-12-06 2020-06-18 Auburn University Inoculants including plant growth producing rhizobacteria (pgpr) and insectides for promoting plant growth and health
CN111849815A (en) * 2020-07-21 2020-10-30 广西民族大学 Plant growth promoting rhizosphere strain Gxun-20 and application thereof in plant growth promotion
CN112210510A (en) * 2020-09-18 2021-01-12 南京林业大学 Phosphate-solubilizing growth-promoting bacterium and salt-tolerant arthrobacter X-1 for promoting growth of nodule and improving abundance of probiotic microorganism population and application thereof
CN113667732A (en) * 2021-09-04 2021-11-19 山东农业大学 Plant-mediated-based screening method for microbial composite strains
CN114015629A (en) * 2021-12-20 2022-02-08 南京林业大学 Complex microbial inoculum for promoting growth of soybeans and improving abundance of probiotic microbial populations

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
LIU KE, MCINROY JOHN A., HU CHIA-HUI, KLOEPPER JOSEPH W.: "Mixtures of Plant-Growth-Promoting Rhizobacteria Enhance Biological Control of Multiple Plant Diseases and Plant-Growth Promotion in the Presence of Pathogens", PLANT DISEASE, THE AMERICAN PHYTOPATHOLOGICAL SOCIETY, US, vol. 102, no. 1, 1 January 2018 (2018-01-01), US , pages 67 - 72, XP093074730, ISSN: 0191-2917, DOI: 10.1094/PDIS-04-17-0478-RE *
XIANG NI, LAWRENCE KATHY S., KLOEPPER JOSEPH W., DONALD PATRICIA A., MCINROY JOHN A.: "Biological control of Heterodera glycines by spore-forming plant growth-promoting rhizobacteria (PGPR) on soybean", PLOS ONE, vol. 12, no. 7, pages e0181201, XP093074728, DOI: 10.1371/journal.pone.0181201 *
XUE XIAO-YUN ，, FENG RUI-HUA, GUAN DA-WEI, LI JUN, CAO FENG-MING: "Screening and Analysis for Efficient Co-inoculation System of Soybean Rhizobia and Plant Growth-promoting Rhizobacteria", DADOU KEXUE - SOYBEAN SCIENCE., GAI KAN BIANJIBU, HA ERBIN., CN, vol. 30, no. 4, 31 August 2011 (2011-08-31), CN , pages 613 - 620, XP093074725, ISSN: 1000-9841 *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN115960765A (en) * 2022-11-02 2023-04-14 云南大学 Flavobacterium bacteria and application thereof in low-temperature composting
CN115960765B (en) * 2022-11-02 2024-04-26 云南大学 Flavobacterium strain and application thereof in low-temperature composting

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