张晟恺2,
焦艳阳2,
林煜2,
吴蕙明2,
秦贤金2,
刘亚洲2,
陈军2,
林文雄2,,
1.安徽农业大学资源与环境学院 合肥 230031
2.福建农林大学农业生态研究所/福建省农业生态过程与安全监控重点实验室 福州 350002
基金项目: 国家自然科学基金项目U1205021
国家自然科学基金项目82003884
国家自然科学基金项目81573530
中国博士后科学基金项目2019M650150
详细信息
作者简介:吴红淼, 主要研究方向为根际生态学过程与调控。E-mail: wuhongmiao@ahau.edu.cn
通讯作者:林文雄, 主要研究方向为植物生理与分子生态学、农业生态学。E-mail: lwx@fafu.edu.cn
中图分类号:S181计量
文章访问数:108
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被引次数:0
出版历程
收稿日期:2020-12-06
录用日期:2021-02-17
刊出日期:2021-08-01
Effects of microbial fertilizer application on the quality and pharmacological activities of radix pseudostellariae under continuous monoculture regimes
WU Hongmiao1, 2,,ZHANG Shengkai2,
JIAO Yanyang2,
LIN Yu2,
WU Huiming2,
QIN Xianjin2,
LIU Yazhou2,
CHEN Jun2,
LIN Wenxiong2,,
1. College of Resources and Environment, Anhui Agricultural University, Hefei 230031, China
2. Institute of Agroecology, Fujian Agriculture and Forestry University/Fujian Provincial Key Laboratory of Agroecological Processing and Safety Monitoring, Fuzhou 350002, China
Funds: the National Natural Science Foundation of ChinaU1205021
the National Natural Science Foundation of China82003884
the National Natural Science Foundation of China81573530
the China Postdoctoral Science Foundation2019M650150
More Information
Corresponding author:LIN Wenxiong, E-mail: lwx@fafu.edu.cn
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摘要
摘要:连作障碍的发生与发展对药用植物的产量和品质构成了严重威胁，探寻有效的连作障碍消减策略尤为重要。本研究经连续4年的田间定位试验，分析了微生物菌肥在减缓太子参连作障碍中的作用；并采用qRT-PCR和HPLC-MS技术分析菌肥改良对太子参根际关键微生物和太子参主效成分的影响；结合药理试验评估了不同处理太子参的功效差异。结果表明，田间菌肥改良重茬太子参效果连续4年均较为显著，重茬地经菌肥改良后较不改良重茬产量分别增长68.28%、111.58%、257.54%和133.23%。菌肥改良能显著增加重茬太子参根际土壤中有益假单胞菌属丰度、减少致病尖孢镰刀菌丰度，也增加太子参中总多糖和环肽B含量；菌肥改良后太子参中氨基酸种类和含量与重茬1年和标准品组无显著差异，且8种氨基酸含量与正茬无显著差异。药理试验结果表明，太子参可以缓解环磷酰胺对小鼠造成的伤害，菌肥改良太子参对小鼠脾脏指数、肝脏重量、附睾脂肪重量、全血白细胞和红细胞含量的影响与正茬太子参无显著差异，且在血小板恢复上与标准品太子参一致。此外，菌肥改良太子参组总抗氧化能力（T-AOC）最强，超氧化物歧化酶（SOD）活性也显著高于正茬和标准品太子参；且与正茬太子参相比，菌肥处理组能显著提高小鼠肝脏组织中免疫因子IL-2和IFN-r mRNA表达水平；而在免疫因子TNF-α表达水平上，菌肥处理组与正茬无显著差异。总体而言，功能微生物菌肥能有效减缓太子参连作障碍问题，改善重茬太子参质量和药理作用。
关键词:太子参/
连作障碍/
菌肥/
根际调控/
药理作用
Abstract:The continuous monoculture problem commonly causes huge economic losses in modern agricultural production. The occurrence and development of continuous cropping obstacles pose a serious threat to the yield and quality of Chinese medicinal plants; therefore, it is important to explore effective strategies to alleviate the continuous cropping obstacles. In this study, we evaluated the effects of microbial fertilizer application on the continuous cropping obstacles and pharmacological activities of radix pseudostellariae (Pseudostellaria heterophylla). The effects of microbial fertilizer application on the yields of continuous monocultured radix pseudostellariae for 4 years under field localization experiments were investigated. Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) and high-performance liquid chromatography-mass spectrometry (HPLC-MS) were used to analyze the effects on the rhizosphere soil microorganisms and the main effective components of radix pseudostellariae. We also used pharmacological tests to analyze the efficiency of radix pseudostellariae under different treatments. Our results showed that microbial fertilizer significantly improved the growth of continuous monocultured radix pseudostellariae, and the yield increased by 68.28%, 111.58%, 257.54%, and 133.23% in 4 years, respectively. Microbial fertilizer application significantly increased the abundance of beneficial Pseudomonas spp., decreased pathogenic Fusarium oxysporum in the rhizosphere soil, and increased the contents of total polysaccharides and heterophyllin B in radix pseudostellariae as compared with those of the consecutive monoculture treatments. The types and contents of amino acids in radix pseudostellariae under microbial fertilizer application were similar to those in the 1-year monoculture and standard radix pseudostellariae. Eight types of amino acids in the microbial fertilizer amendment and the newly planted radix pseudostellariae were not significantly different. Pharmacological tests showed that radix pseudostellariae alleviated the damage caused by cyclophosphamide in mice. There were no significant differences in the spleen index, liver weight, epididymal fat weight, and white or red blood cell content of mice treated with plants of the microbial fertilizer amended and newly planted radix pseudostellariae. The effect of radix pseudostellariae on platelet recovery of mice under microbial fertilizer amendment was consistent with that of standard radix pseudostellariae. Moreover, the total antioxidant capacity of radix pseudostellariae under microbial fertilizer amendment was stronger, and the superoxide dismutase content was significantly higher than that of newly planted and standard radix pseudostellariae. Compared with newly planted radix pseudostellariae, the microbial fertilizer amendment significantly increased the expression of immune factors IL-2 and IFN-r in the liver tissue of mice. There was no significant difference in the expression of TNF-α between the microbial fertilizer amendment and newly planted radix pseudostellariae. In conclusion, microbial fertilizer amendment effectively alleviated the continuous cropping obstacle and improved the quality and pharmacological activities of radix pseudostellariae. Our results provide guidance for studying alleviation strategies in other Chinese medicinal plants under consecutive monoculture regimes and for evaluating the quality and pharmacological activities of modified medicinal plants.
Key words:Radix pseudostellariae/
Continuous cropping obstacle/
Microbial fertilizer/
Rhizosphere management/
Pharmacological activities
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图1菌肥改良后太子参产量变化
不同小写字母表示同一年份不同处理间差异显著(P < 0.05)。
Figure1.Effects of microbial fertilizer on the yields of radix pseudostellariae
Different lowercase letters mean significant differences among treatments in the same year at P < 0.05 level.
下载: 全尺寸图片幻灯片
图2菌肥改良后太子参根际微生物丰度变化
FY: 正茬; SY: 重茬1年; TY: 重茬2年; BIO: 重茬+菌肥改良。不同小写字母表示不同处理间差异显著(P < 0.05)。
Figure2.Effects of microbial fertilizer on populations of microorganisms in radix pseudostellariae rhizosphere soil
FY, SY and TY indicate treatments of newly planted soil, one-year monocultured soil and two-year monocultured soil, respectively. BIO indicates the treatment of application of microbial fertilizer on three-year monocultured soil. Different lowercase letters mean significant differences among treatments at P < 0.05 level.
下载: 全尺寸图片幻灯片
图3菌肥改良后太子参中总多糖和环肽B含量变化
FY: 正茬; SY: 重茬1年; BIO: 重茬+菌肥改良; ST: 太子参标准品。不同小写字母表示不同处理间差异显著(P < 0.05)。
Figure3.Effects of microbial fertilizer on contents of total polysaccharide and heterophyllin B in radix pseudostellariae
FY and SY indicate the treatments of newly planted soil and one-year monocultured soil, respectively. BIO indicates the treatment of application of microbial fertilizer on three-year monocultured soil. ST indicates the standard radix pseudostellariae. Different lowercase letters mean significant differences among treatments at P < 0.05 level.
下载: 全尺寸图片幻灯片
图4菌肥改良的太子参对小鼠体重的动态影响
CK: 未灌胃太子参和注射生理盐水; CTX: 未灌胃太子参和注射环磷酰胺; CTX+FY: 灌胃正茬太子参和注射环磷酰胺; CTX+SY: 灌胃重茬太子参和注射环磷酰胺; CTX+BIO: 灌胃菌肥改良太子参和注射环磷酰胺; CTX+ST: 灌胃标准品太子参和注射环磷酰胺。
Figure4.Effect of radix pseudostellariae under different treatments of application of microbial fertilizer on weight of mice
CK: no-radix pseudostellariae treatment and physiological saline injection; CTX: no-radix pseudostellariae treatment and cyclophosphamide injection; CTX+FY: newly planted radix pseudostellariae treatment and cyclophosphamide injection; CTX+SY: one-year monoculture radix pseudostellariae treatment and cyclophosphamide injection; CTX+BIO: microbial fertilizer amendment radix pseudostellariae treatment and cyclophosphamide injection; CTX+ST: standard radix pseudostellariae treatment and cyclophosphamide injection.
下载: 全尺寸图片幻灯片
图5菌肥改良的太子参对小鼠生理生化指标的影响
CK: 未灌胃太子参和注射生理盐水; CTX: 未灌胃太子参和注射环磷酰胺; CTX+FY: 灌胃正茬太子参和注射环磷酰胺; CTX+SY: 灌胃重茬太子参和注射环磷酰胺; CTX+BIO: 灌胃菌肥改良太子参和注射环磷酰胺; CTX+ST: 灌胃标准品太子参和注射环磷酰胺。不同小写字母表示不同处理间差异显著(P < 0.05)。
Figure5.Effect of radix pseudostellariae under different treatments of application of microbial fertilizer on physiological and biochemical indexes of mice
CK: no-radix pseudostellariae treatment and physiological saline injection; CTX: no-radix pseudostellariae treatment and cyclophosphamide injection; CTX+FY: newly planted radix pseudostellariae treatment and cyclophosphamide injection; CTX+SY: one-year monoculture radix pseudostellariae treatment and cyclophosphamide injection; CTX+BIO: microbial fertilizer amendment radix pseudostellariae treatment and cyclophosphamide injection; CTX+ST: standard radix pseudostellariae treatment and cyclophosphamide injection. Different lowercase letters mean significant differences among treatments at P < 0.05 level.
下载: 全尺寸图片幻灯片
图6菌肥改良的太子参对小鼠肝脏免疫因子表达量的影响
CTX: 未灌胃太子参和注射环磷酰胺; CTX+FY: 灌胃正茬太子参和注射环磷酰胺; CTX+SY: 灌胃重茬太子参和注射环磷酰胺; CTX+BIO: 灌胃菌肥改良太子参和注射环磷酰胺; CTX+ST: 灌胃标准品太子参和注射环磷酰胺。不同小写字母表示不同处理间差异显著(P < 0.05)。
Figure6.Effect of radix pseudostellariae under different treatments of application of microbial fertilizer on the expression of liver immune factors in mice
CTX: no-radix pseudostellariae treatment and cyclophosphamide injection; CTX+FY: newly planted radix pseudostellariae treatment and cyclophosphamide injection; CTX+SY: one-year monoculture radix pseudostellariae treatment and cyclophosphamide injection; CTX+BIO: microbial fertilizer amendment radix pseudostellariae treatment and cyclophosphamide injection; CTX+ST: standard radix pseudostellariae treatment and cyclophosphamide injection. Different lowercase letters mean significant differences among treatments at P < 0.05 level.
下载: 全尺寸图片幻灯片表1土壤特异微生物菌群和小鼠肝脏免疫因子qRT-PCR分析引物序列及退火温度
Table1.Taxon-specific and liver immune factors primer sets and their annealing temperatures for quantitative PCR
| 微生物类群/基因名称 Target group/Gene name | 引物名称 Primer name | 序列(5’-3’) Sequence (5’-3’) | 退火温度 Annealing temperature (℃) |
| 假单胞菌属 Pseudomonas spp. | Ps-for [14] | TTAGCTCCACCTCGCGGC | 64.0 |
| Ps-rev | GGTCTGAGAGGATGATCAGT | ||
| 尖孢镰刀菌 Fusarium oxysporum | ITS1F [15] | CTTGGTCATTTAGAGGA AGTAA | 60.4 |
| AFP308R | CGAATTAACGCGAGTCCCAA | ||
| ACTB | ACTB F [16-17] | GAAGATCAAGATCATTGCTCCT | 60.8 |
| ACTB R | TACTCCTGCTTGCTGATCCA | ||
| IL-2 | IL-2 F [16-17] | CTGCGGCATGTTCTGGATTTG | 60.5 |
| IL-2 R | TTGAGGGCTTGTTGAGATGATGC | ||
| IFN-γ | IFN-γ F [16-17] | AATGMCGCTACACACTGCA | 58.3 |
| IFN-γ R | TGAAGAAGGTAGTMTCAGG | ||
| TNF-α | TNF-α F [16-17] | ATGAGCACAGAAAGCATGATCCGC | 63.0 |
| TNF-α R | AAAGTAGACCTGCCCGGACTC |
下载: 导出CSV表2太子参环肽B检测液相洗脱条件
Table2.Elution conditions of the HPLC used to detect the heterophyllin B of radix pseudostellariae
| 时间 Time (min) | A相 Mobile phase A (%) | B相 Mobile phase B (%) | 流速 Rate (μL·min–1) |
| 0 | 95 | 5 | 300 |
| 2 | 70 | 30 | 300 |
| 12 | 60 | 40 | 300 |
| 15 | 55 | 45 | 300 |
| 16 | 50 | 50 | 300 |
| 20 | 40 | 60 | 300 |
| 22 | 30 | 70 | 300 |
| 22.1 | 95 | 5 | 300 |
| 26 | 95 | 5 | 300 |
| 流动相A: 水; 流动相B: 乙腈。Mobile phase A: water; mobile phase B: acetonitrile. | |||
下载: 导出CSV表3菌肥改良后太子参中氨基酸各组分含量变化
Table3.Effects of microbial fertilizer on contents of amino acids in radix pseudostellariae
| 氨基酸Amino acid | FY | SY | BIO | ST |
| 谷氨酸Glutamic acid | 13.27±2.36aA | 10.42±2.13aB | 12.61±3.75aA | 10.30±3.17aA |
| 丝氨酸Serine | 3.93±1.75aDE | 2.28±0.44aD | 2.91±1.31aBC | 2.25±0.91aC |
| 组氨酸Histidine | 5.99±0.83aC | 4.31±0.89aC | 5.57±1.45aB | 4.67±1.13aB |
| 甘氨酸Glycine | 0.29±0.08aG | 0.17±0.05aF | 0.23±0.09aC | 0.17±0.06aC |
| 精氨酸Arginine | 11.44±2.52aB | 12.35±2.02aA | 13.58±4.16aA | 11.61±2.67aA |
| 苏氨酸Threonine | 3.70±1.32aDE | 2.00±0.53aDE | 2.32±0.75aC | 2.03±0.50aC |
| 酪氨酸Tyrosine | 2.20±0.63aF | 0.90±0.28bDEF | 1.30±0.35bC | 1.15±0.30bC |
| 缬氨酸Valine | 2.00±1.18aF | 0.24±0.20bF | 0.69±0.46bC | 0.41±0.25bC |
| 蛋氨酸Methionine | 0.58±0.17aG | 0.47±0.16aEF | 0.64±0.14aC | 0.56±0.13aC |
| 异亮氨酸Isoleucine | 2.27±1.32aF | 0.47±0.18bEF | 0.81±0.32bC | 0.62±0.23bC |
| 亮氨酸Leucine | 3.95±1.09aDE | 1.64±0.45bDEF | 2.39±0.71bC | 2.00±0.53bC |
| 苯丙氨酸Phenylalanine | 3.07±0.94aEF | 1.28±0.35bDEF | 2.06±0.59abC | 1.73±0.39bC |
| 赖氨酸Lysine | 4.47±0.77aD | 1.86±0.56bDE | 2.89±0.93bBC | 2.23±0.63bC |
| FY: 正茬; SY: 重茬1年; BIO: 重茬+菌肥改良; ST: 太子参标准品。同行不同小写字母表示不同处理间差异显著(P < 0.05), 同列不同大写字母表示同一处理不同组分间差异显著(P < 0.01)。FY and SY indicate the treatments of newly planted soil and one-year monocultured soil, respectively. BIO indicates the treatments of application of microbial fertilizer on three-year monocultured soil. ST indicates the standard radix pseudostellariae. Different lowercase letters in the same line mean significant differences among treatments at P < 0.05 level. Different capital letters in the same column mean significant differences among amino acids at P < 0.01 level. | ||||
下载: 导出CSV参考文献
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