解磷真菌JL-7的筛选及其作用于低品位磷矿的肥效研究

李小军; 吉俐; 张景宁; 盛定红; 谢承卫

doi:10.19303/j.issn.1008-0384.2022.009.017

解磷真菌JL-7的筛选及其作用于低品位磷矿的肥效研究

doi: 10.19303/j.issn.1008-0384.2022.009.017

贵州大学化学与化工学院，贵州　贵阳　550025

基金项目: 贵州省科学技术基金项目（20171028）；贵州省科技厅科技支撑计划项目（黔科合[2016]2808号）；中国烟草总公司贵州省公司科技项目（201708）

详细信息

作者简介:
李小军（1998−），男，硕士研究生，主要从事微生物肥料研究（E-mail：846736603@qq.com）

通讯作者:
谢承卫（1964−），男，教授，主要从事微生物、烟草肥料及各类矿产资源的应用研究工作（E-mail：cwxie@gzu.edu.cn）

中图分类号: S 154
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出版历程
- 收稿日期: 2022-03-05
- 录用日期: 2022-03-05
- 修回日期: 2022-07-22
- 网络出版日期: 2022-10-05
- 刊出日期: 2022-09-30

Efficiency Improvement of Low-grade Phosphorus Fertilizer by Phosphate-solubilizing Fungus JL-7

School of Chemistry and Chemical Engineering, Guizhou University, Guiyang, Guizhou　550025, China

摘要

摘要: 目的针对我国低品位磷矿资源利用率低、耕地土壤有效磷含量低以及传统化肥的大量使用造成的土壤易固化等一系列问题，筛选可解离低品位磷矿的高效菌株，为低品位磷矿的资源利用提供途径。方法以砂培法从贵州铜仁烟草种植基地土壤中筛选出一株新型解磷真菌，将其命名为JL-7，经过生理生化试验和分子生物学鉴定该菌为烟曲霉菌（Aspergillus fumigatiaffinis）。采用单因素和正交试验对该菌的解磷性能进行优化，在最佳优化条件下制备微生物菌肥并通过烟草盆栽试验进行肥效验证。结果从烟草根际土壤中筛选的真菌JL-7在优化条件（接菌量1×10⁵ cfu·mL⁻¹、初始pH 6、解离时间8 d、解离温度26 ℃）下对低品位磷矿的最高解磷量达到967.4 mg·kg⁻¹，真菌解离液中pH可降低至2.9左右。以高效解磷菌株JL-7制备菌肥，通过烟草盆栽种植试验，该菌肥对烟草（云烟87）的茎围、株高、最大叶面积分别提升44.60%、57.29%、62.90%。种植后对土壤进行分析，结果表明，该菌肥对土壤中的有效磷、速效钾、碱解氮含量分别提升48.5%、3.7%、9.1%。结论菌株JL-7解磷效果优异且性能稳定，具有制备新型微生物肥料的潜力，具备一定的推广及应用价值。
- 解磷真菌 /
- 低品位磷矿 /
- 条件优化 /
- 微生物肥料
Abstract: Objective A fungal strain highly efficient in dissociating low-grade phosphorus ore for fertilization was isolated to determine the applicability. Method Soil samples at Guizhou Tongren Tobacco Planting Base were screened using the sand culture method for fungi that exhibited capability to solubilize phosphate. The selected isolate was identified by physiological, biochemical, and molecular biological tests. Conditions for optimal phosphate solubilization were determined by single factor and orthogonal experiments. Effect of the fungal addition on fertilization was verified by a pot test conducted on tobacco seedlings. Result The selected isolate was code-named JL-7 and, subsequently, identified to be a strain of Aspergillus fumigatiaffinis. The optimized conditions to maximize the phosphate-solubilization of JL-7 applied an inoculation at the rate of 1×10⁵ cfu·mL⁻¹ with an initial pH of 6 to incubate at 26 ℃ for 8 d. The process dissolved a low-grade phosphorus ore material up to 967.4 mg·kg⁻¹ with the resulting solution reduced to approximately pH 2.9. In the pot experiment, the Yunyan 87 tobacco seedlings grown on a medium with the addition of a JL-7-inoculated phosphorus fertilizer had the stem girth, plant height, and maximum leaf area increased by 44.60%, 57.29%, and 62.90%, respectively, over control. Meanwhile, the pot soil increased 48.5% on available phosphorus, 3.7% on available potassium, and 9.1% on alkali-hydrolysable nitrogen after the planting as well. Conclusion The identified strongly phosphate-solubilizing Fungus JL-7 displayed a significant and consistent ability of dissolving phosphate. It was considered a potential candidate to be widely promoted as a microbial fertilization enhancer.
- Phosphate-dissolving fungus /
- low-grade phosphate ore /
- condition optimization /
- microbial fertilization enhancer

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图 1 JL-7的生长变化过程及显微图

a：初生菌丝生长情况；b：5天后菌落生长情况；c、d：9天后菌落生长情况和平板背面观察到的菌落外观；e、f：光学显微镜下放大1 000倍观察到的真菌菌丝和分生孢子梗。

Figure 1. Growth and change process and micrograph of JL-7

a: Growth of primary mycelium; b: growth of colony after 5 d; c and d: growth of colony after 9 d and rear view of colony on plate; e and f: fungal mycelium and conidiophore under optical microscope (1 000×).

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图 2 JL-7的系统发育树

Figure 2. Phylogenetic tree of JL-7

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图 3 菌株JL-7的生长曲线及真菌解离液中pH的变化曲线

Figure 3. Growth curve of JL-7 and pH changes of phosphae-dissociation solution

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图 4 不同因素对真菌JL-7解磷量的影响

Figure 4. Effect of various factors on phosphate solubilization of JL-7

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图 5 菌肥对烟草生长促进效果

图中不同小写字母表示差异显著(P＜0.05)。

Figure 5. Effect of fungal fertilization enhancer on tobacco growth

Data with different lowercase letters indicate significant differences (P＜0.05).

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表 1 JL-7的生理生化试验结果

Table 1. Physiological and biochemical test results on JL-7

项目 Item	结果 Result	项目 Item	结果 Result	项目 Item	结果 Result
D-阿拉伯糖	−	D-核糖	+	麦芽三糖	+
L-阿拉伯糖	+	D-棉子糖	+	N-乙酰-半乳糖胺	+
D-纤维二糖	+	L-鼠李糖	+	N-乙酰-葡萄糖胺	+
L-山梨糖	−	D-甘露糖	+	N-乙酰-甘露糖胺	+
D-果糖	+	D-松三糖	+	D-葡萄糖胺	+
L-岩藻糖	+	D-蜜二糖	+	葡糖醛酰胺	−
D-半乳糖	−	D-塔格糖	+	丙酸胺	+
+表示阳性反应；−表示阴性反应。 + means positive reaction; − means negative reaction.

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表 2 单因素设计的L₉(3)⁴正交试验表

Table 2. L₉(3)⁴ orthogonal experiment design with single factor experiment

因素 Level	接菌量 Inoculation amount/(cfu·mL⁻¹)	初始pH Initial pH	解离时间 Dissociation time/d	解离温度 Dissociation temperature/ ℃
1	1×10⁴	5	6	26
2	1×10⁵	6	7	28
3	1×10⁶	7	8	30

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表 3 正交试验对菌株JL-7解磷条件的优化结果

Table 3. Orthogonal optimized conditions for JL-7 phosphate solubilization

因素 Level	接菌量 Inoculation amount/(cfu·mL⁻¹)	初始pH Initial pH	解离时间 Dissociation time/d	解离温度 Dissociation temperature/ ℃	有效磷含量 Available phosphorus/(mg·kg⁻¹)
1	1×10⁴	5	6	26	779. 8
2	1×10⁴	6	7	28	811.3
3	1×10⁴	7	8	30	793.1
4	1×10⁵	5	7	30	852.5
5	1×10⁵	6	8	26	872.9
6	1×10⁵	7	6	28	828.8
7	1×10⁶	5	8	28	837.6
8	1×10⁶	6	6	30	812.1
9	1×10⁶	7	7	26	803.6
K1	794.733	823.300	806.900	818.767
K2	851.400	832.100	822.467	825.900
K3	817.767	805.500	834.533	819.233
极差 R	56.667	23.600	27.633	7.133

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表 4 菌肥对土壤肥力的影响

Table 4. Effect of fungal fertilization enhancer on soil fertility （mg·kg⁻¹）

处理方式 Processing method	碱解氮 Alkali-hydrolyzable nitrogen	有效磷 Available phosphorus	速效钾 Available potassium
CK（对照）	107.47±2.09 c	58.87±2.56 c	139.20±1.06 b
A（施用A菌肥）	124.43±1.94 a	73.10±2.59 b	144.45±2.17 a
B（施用B菌肥）	117.17±2.65 b	87.41±0.97 a	144.42±1.88 a
同列数据后不同小写字母表示差异显著(P＜0.05)。 Data with different letters on the same column indicate significant differences (P＜0.05).

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