PGPR微胶囊菌剂研发及其对玉米的促生效果评价

刘文佳; 刘天一; 张俐敏; 徐畅; 莫继先

PGPR微胶囊菌剂研发及其对玉米的促生效果评价

齐齐哈尔大学生命科学与农林学院，黑龙江　齐齐哈尔　161006

基金项目: 黑龙江省省属高等学校基本科研业务费项目（135409420）；黑龙江省大学生创新创业训练计划项目（S202310232017）

详细信息

作者简介:
刘文佳（1999 —），女，硕士研究生，主要从事资源环境微生物学研究，E-mail：1740203063@qq.com

通讯作者:
莫继先（1982 —），男，博士，教授，主要从事资源环境微生物学研究，E-mail：mojixian8208@sina.com

中图分类号: S541
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出版历程
- 收稿日期: 2023-12-22
- 修回日期: 2024-02-07
- 网络出版日期: 2024-03-28

Microencapsulated Rhizosphere Bacteria for Promoting Growth of Corn Plants

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

摘要

摘要: 目的为了提高植物根际促生菌（Plant growth-promoting rhizobacteria, PGPR）对作物促生作用的稳定性，研制出以PGPR复合菌液为原料的微胶囊菌剂，探究其对玉米的促生效果。方法从玉米根际土壤中分离得到2株PGPR菌株并制备成复合液体菌液。分别用海藻酸钠（SA）和CaCl₂作为包埋剂和交联剂，对复合菌液进行包埋以制备微胶囊菌剂。以复合微胶囊菌剂制备的难易程度和菌株包埋率、增殖前活菌数及增殖后活菌数为评价指标，确定微胶囊菌剂制备的最优条件。通过分析微胶囊菌剂在不同聚乙二醇含量、温度、pH、存储时间以及盐含量下的菌株生长及其对玉米生长和根系形态的影响确定PGPR微胶囊菌剂的特性。结果分离得到了2株PGPR菌株，具有多种植物促生功能。使用1%SA-3%CaCl₂制备的微胶囊菌剂稳定性较好，包埋率达91.27%，增殖后的微胶囊活菌数达到8.73×10⁹ cfu·g⁻¹，增殖7.79倍。微胶囊菌剂在不同干旱、温度、pH、存储时间和盐浓度条件下对PGPR菌株具有较好的保护作用。PGPR微胶囊菌剂对玉米有显著的促生作用，施用微胶囊菌剂使玉米株高、苗干重和根干重分别增加91.83%、81.82%和29.57%。同时微胶囊菌剂也显著提高了玉米总根长、根表面积、根平均直径、根体积和根尖数。结论探明了一种PGPR微胶囊菌剂的制备方法，施用该PGPR微胶囊剂能够显著促进玉米生长、改善玉米根系形态，对玉米的促生效果优于液体菌剂。
- 玉米 /
- 根际促生菌 /
- 促生效果 /
- 微胶囊菌剂
Abstract: Objective Microencapsulation was applied to improve the stability of PGPRs for promoting corn growth. Method Rhizosphere bacteria that promote corn growth were isolated. The bacteria in broth were encapsulated using sodium alginate (SA) and CaCl₂. Based on operation efficiency and PGPR survival rate, optimal conditions for the encapsulation were determined. Properties of the microencapsulated agent were analyzed for field applications with respect to the bacterial proliferation under varied artificial dehydration created by using polyethylene glycol (PEG) and salt, temperatures, pHs, and storage times as well as the growth and root morphology of the treated corn plants. Result Two PGPR strains capable of promoting corn growth were isolated. The optimized process using 1% SA and 3% CaCl₂ yielded a high encapsulation rate of 91.27% with a viable bacteria count of 8.73×10⁹ cfu·g⁻¹ on 7.79x proliferation. Significant protection against the imposed adverse conditions was observed. In comparison with the virgin PGPRs broth, the encapsulation also significantly enhanced the growth of the treated corn plants with a 91.83% increase on plant height, 81.82% on seedling dry weight, and 29.57% on root dry weight over control. In addition, the total length, surface area, diameter, volume, and tip number of the roots of the corn plants grown with the encapsulated PGPRs were significantly increased as well. Conclusion Microencapsulation significantly improved the stability as well as the root morphology and growth promoting effect of PGPRs on the corn plants.
- Corn /
- rhizosphere bacteria /
- growth-promoting effect /
- microencapsulated

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图 1 菌株P1和K5的菌间拮抗试验

Figure 1. Interbacterial antagonism between P1 and K5

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图 2 菌株 P1（a）和K5（b）的显微形态

Figure 2. Microscopic morphology of P1 (a) and K5 (b)

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图 3 菌株P1的系统进化树

Figure 3. Phylogenetic tree of P1

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图 4 菌株K5的系统进化树

Figure 4. Phylogenetic tree of K5

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图 5 菌株P1和K5的生长曲线

Figure 5. Growth curves of P1 and K5

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图 6 SA1%-CaCl₂3%微胶囊颗粒形态

Figure 6. Morphology of 1% SA-3% CaCl₂ microcapsules

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图 7 不同环境条件对两种剂型菌株生长的影响

＊表示两组之间差异显著（P＜0.05）

Figure 7. Effect of environmental conditions on proliferation of PGPRs in two different dosages

＊indicates significant difference (P＜0.05).

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图 8 不同剂型PGPR对玉米生长的影响

a：株高；b：苗干重；c：根干重；不同小写字母表示不同组别间差异显著（P＜0.05）。

Figure 8. Effect of PGPR dosage on growth of corn plants

a: plant height; b: dry weight; c: root dry weight; data with different letters indicate significant differences (P＜0.05).

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图 9 不同剂型PGPR对玉米根系形态的影响

a：根长；b：根总表面积；c：根平均直径；d：根尖数；e：根体积；不同字母表示不同组别间差异显著（P＜0.05）

Figure 9. Effect of PGPR dosage on root morphology of corn plants

a: root length; b: total root surface area; c: average root diameter; d: number of root tips; e: root volume; data with different letters indicate significant differences (P＜0.05).

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表 1 玉米根际土壤中筛选的具有促生特性的菌株及促生特性分析

Table 1. Strains and properties of corn growth-promoting bacteria from rhizosphere soill

菌株 Strains	解磷量 Phosphate solubilizing/ (μg·mL⁻¹)	解钾量 Potassium releasing/ (μg·mL⁻¹)	固氮量 Nitrogen fixation/ (μg·mL⁻¹)	吲哚乙酸 Indoleacetic acid/ (μg·mL⁻¹)	铁载体活性 Siderophore activity/%	ACC含量 ACC content/ (μg·mL⁻¹)
P1	33.50±0.37a	—	1.44±0.16b	78.06±0.51a	55.72±0.67a	0.19±0.07e
P2	31.49±0.51b	—	—	65.74±1.61b	44.19±0.94d	0.30±0.07b
P3	31.34±0.58b	—	—	—	49.12±0.35b	0.27±0.05c
P4	30.85±0.59b	—	—	—	46.47±0.27c	0.22±0.14d
P5	30.12±0.03c	—	—	46.23±3.25d	40.48±0.44e	0.14±0.07f
P6	31.13±0.04b	—	—	—	39.65±0.38e	0.20±0.06e
N1	—	—	1.96±0.35a	—	33.5±0.527f	0.27±0.11c
N2	—	—	1.27±0.21b	—	—	0.14±0.02f
N3	—	—	1.23±0.10b	—	—	0.22±0.03d
K1	—	1.22±0.09b	—	55.63±0.54c	31.52±0.39g	0.14±0.02f
K2	—	1.17±0.07b	—	—	—	0.30±0.06b
K3	—	1.26±0.07b	—	—	29.34±0.35g	0.25±0.13cd
K4	—	1.35±0.28b	—	—	—	0.18±0.07e
K5	—	2.26±0.07a	—	63.43±2.9b	50.46±0.71b	0.34±0.05a
同列数据后不同字母表示差异显著（P＜0.05）；—表示无此项促生功能。 Data with different letters on same column indicate significant differences (P＜0.05); —indicate no growth-promoting function.

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表 2 菌株P1和K5的生理生化特性

Table 2. Physiological and biochemical characteristics of P1 and K5

试验项目 Items		淀粉水解实验 Starch hydrolysis	蔗糖发酵实验 Sucrose fermentation	乳糖发酵实验 Lactose fermentation	葡萄糖发酵实验 Glucose fermentation	甲基红实验 Methyl red experiment	柠檬酸盐实验 Citrate experiment	吲哚实验 Indole experiment	革兰氏染色实验 Gram staining experiment	触酶实验 Catalase experiment	伏-普实验 Volt-P experiment
菌株 Strains	P1	−	+	−	+	−	−	+	+	+	+
菌株 Strains	K5	−	−	+	−	−	−	+	+	+	−
+：阳性；−：阴性； +: positive; −: negative.

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表 3 SA-CaCl₂不同质量浓度配比下微胶囊的操作性和成球性

Table 3. Microencapsulation of PGPRs with varied SA-CaCl₂ combinations

配比组合 Varied combination		操作性 Operation	成球性 Sphericity
1%SA	2%CaCl₂	易 Easy	较好，拖尾，无黏连 Better, trailing, no sticking
	3%CaCl₂	易 Easy	较好，拖尾，少量黏连 Better, trailing, few sticks
	4%CaCl₂	易 Easy	较好，拖尾，少量黏连 Better, trailing, few sticks
2%SA	2%CaCl₂	易 Easy	较好，拖尾，无黏连 Better, trailing, no sticking
	3%CaCl₂	易 Easy	较好，拖尾，少量黏连 Better, trailing, few sticks
	4%CaCl₂	易 Easy	较好，拖尾，少量黏连 Better, trailing, few sticks
3%S	2%CaCl₂	难 Difficult	较差，无拖尾，少量破碎 Bad, no tailing, few fragile
	3%CaCl₂	难 Difficult	较差，拖尾，少量黏连 Bad, trailing, few sticks
	4%CaCl₂	难 Difficult	差，拖尾，少量破碎，易黏连 Poor, trailing, few fragile, sticks

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表 4 微胶囊菌剂颗粒的理化性质

Table 4. Physicochemical properties of microcapsules

配比组合 Varied combination	直径 Diameter/mm	机械强度 Mechanical Strength/g	增殖前胶囊活菌数 Number of viable cells before proliferation/ (×10⁹ cfu·g⁻¹)	包埋率 Embedding rate/%	增殖后胶囊活菌数 Number of viable cells in after proliferation/ (×10⁹ cfu·g⁻¹)	增殖倍数 Multiplication ratio
SA1%-CaCl₂2%	3.14±0.13b	3.29±0.16b	1.01±0.11b	85.60±2.25b	5.68±0.37b	5.62
SA1%-CaCl₂3%	3.11±0.07c	3.21±0.13c	1.12±0.03c	91.27±1.05c	8.73±0.50c	7.79
SA2%-CaCl₂2%	3.22±0.04a	3.35±0.03a	1.20±0.03a	81.17±1.91a	6.91±0.49a	5.76
同列数据后不同字母表示差异显著（P＜0.05） Data with different letters on same column indicate significant differences (P＜0.05).

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