Journal of Agricultural Science and Technology ›› 2024, Vol. 26 ›› Issue (7): 147-155.DOI: 10.13304/j.nykjdb.2023.0651
• ANIMAL AND PLANT HEALTH • Previous Articles
Yongjun XIE(), Xiaozhuo PAN, Fuhui CHEN, Kaibo YIN, Jiayue JIN, Yibing WANG(
)
Received:
2023-08-30
Accepted:
2024-02-06
Online:
2024-07-15
Published:
2024-07-12
Contact:
Yibing WANG
谢勇俊(), 潘小卓, 陈福慧, 尹凯波, 金嘉悦, 王一兵(
)
通讯作者:
王一兵
作者简介:
谢勇俊 E-mail:jun146ban@163.com;
基金资助:
CLC Number:
Yongjun XIE, Xiaozhuo PAN, Fuhui CHEN, Kaibo YIN, Jiayue JIN, Yibing WANG. Screening, Identification and Biocontrol of Bacteria Degrading Ginseng Phenolic Acid Autotoxic Substances[J]. Journal of Agricultural Science and Technology, 2024, 26(7): 147-155.
谢勇俊, 潘小卓, 陈福慧, 尹凯波, 金嘉悦, 王一兵. 人参酚酸类自毒物质降解菌的筛选鉴定及生防研究[J]. 中国农业科技导报, 2024, 26(7): 147-155.
培养基 Culture medium | 组分 Component |
---|---|
筛选培养基 Screening medium | 1 L培养基含(NH2)SO₄ 2 g、CaCl₂ 0.1 g、NaH2PO4 0.5 g、K2HPO4 0.5 g、MgSO4·7H2O 0.2 g、碳源(水杨酸、没食子酸、邻苯二甲酸、对羟基苯甲酸、丁香酸、香草酸、肉桂酸、魏酸)500 mg,1 000 mL蒸馏水,pH 7.2,固体培养基加入20 g琼脂粉 1 L medium contains 2 g (NH2)SO₄, 0.1 g CaCl₂, 0.5 g NaH2PO4, 0.5 g K2HPO4, 0.2 g MgSO4·7H2O, 500 mg carbon source (salicylic acid, gallic acid, phthalic acid, p-hydroxybenzoic acid, butyric acid, vanillic acid, cinnamic acid and ferulic acid), 1 000 mL water,and pH 7.2, the solid medium contains 20 g agar powder |
Hugh-Leifson 培养基 Hugh-Leifson medium | 1 L培养基含葡萄糖10 g,蛋白胨2 g,磷酸氢二钾0.3 g,氯化钠5 g,琼脂3~5 g, 质量浓度为1% 溴麝香草酚蓝3 mL 1 L medium contains 10 g glucose, 2 g peptone, 0.3 g dipotassium hydrogen phosphate, 5 g sodium chloride, 3~5 g agar and 3 mL 1% bromomuscohol blue by mass |
明胶培养基 Gelatin medium | 1 L培养基含蛋白胨5 g,牛肉膏3 g,明胶120 g,pH 6.8~7.0 1 L medium contains 5 g peptone, 3 g beef paste, 120 g gelatin, pH 6.8~7.0 |
硝酸盐培养基 Nitrate medium | 1 L培养基含KNO3(不含NO2-)0.2 g,蛋白胨5 g,pH 7.4 1 L medium contains 0.2 g KNO3 (NO2- free), 5 g peptone, pH 7.4 |
尿素培养基 Urea medium | 1 L培养基含尿素20.0 g,Na2HPO4 9.5 g,酵母浸膏0.1 g,酚红0.01 g,KH2PO4 9.1 g, pH (6.8±0.2) 1 L medium contains 20.0 g urea, 9.5 g Na2HPO4, 0.1 g yeast extract, 0.01 g phenol red, 9.1 g KH2PO4, and pH (6.8±0.2) |
LB培养基 LB medium | 1 L培养基含胰蛋白胨10 g、酵母提取物5 g、氯化钠10 g 1 L medium contains 10 g tryptone, 5 g yeast extract, and 10 g sodium chloride |
Table 1 Test the required medium formulation
培养基 Culture medium | 组分 Component |
---|---|
筛选培养基 Screening medium | 1 L培养基含(NH2)SO₄ 2 g、CaCl₂ 0.1 g、NaH2PO4 0.5 g、K2HPO4 0.5 g、MgSO4·7H2O 0.2 g、碳源(水杨酸、没食子酸、邻苯二甲酸、对羟基苯甲酸、丁香酸、香草酸、肉桂酸、魏酸)500 mg,1 000 mL蒸馏水,pH 7.2,固体培养基加入20 g琼脂粉 1 L medium contains 2 g (NH2)SO₄, 0.1 g CaCl₂, 0.5 g NaH2PO4, 0.5 g K2HPO4, 0.2 g MgSO4·7H2O, 500 mg carbon source (salicylic acid, gallic acid, phthalic acid, p-hydroxybenzoic acid, butyric acid, vanillic acid, cinnamic acid and ferulic acid), 1 000 mL water,and pH 7.2, the solid medium contains 20 g agar powder |
Hugh-Leifson 培养基 Hugh-Leifson medium | 1 L培养基含葡萄糖10 g,蛋白胨2 g,磷酸氢二钾0.3 g,氯化钠5 g,琼脂3~5 g, 质量浓度为1% 溴麝香草酚蓝3 mL 1 L medium contains 10 g glucose, 2 g peptone, 0.3 g dipotassium hydrogen phosphate, 5 g sodium chloride, 3~5 g agar and 3 mL 1% bromomuscohol blue by mass |
明胶培养基 Gelatin medium | 1 L培养基含蛋白胨5 g,牛肉膏3 g,明胶120 g,pH 6.8~7.0 1 L medium contains 5 g peptone, 3 g beef paste, 120 g gelatin, pH 6.8~7.0 |
硝酸盐培养基 Nitrate medium | 1 L培养基含KNO3(不含NO2-)0.2 g,蛋白胨5 g,pH 7.4 1 L medium contains 0.2 g KNO3 (NO2- free), 5 g peptone, pH 7.4 |
尿素培养基 Urea medium | 1 L培养基含尿素20.0 g,Na2HPO4 9.5 g,酵母浸膏0.1 g,酚红0.01 g,KH2PO4 9.1 g, pH (6.8±0.2) 1 L medium contains 20.0 g urea, 9.5 g Na2HPO4, 0.1 g yeast extract, 0.01 g phenol red, 9.1 g KH2PO4, and pH (6.8±0.2) |
LB培养基 LB medium | 1 L培养基含胰蛋白胨10 g、酵母提取物5 g、氯化钠10 g 1 L medium contains 10 g tryptone, 5 g yeast extract, and 10 g sodium chloride |
编号Number | 比对结果Alignment result | 相似度Similarity/% |
---|---|---|
14XC | 铜绿假单胞菌Pseudomonas aeruginosa | 99.52 |
19AW | Pseudomonas furukawaii | 99.67 |
BH1AW | Pandoraea morbifera | 99.87 |
BH2XC | 木糖氧化无色杆菌Achromobacter xylosoxidans | 99.55 |
D2 | Pseudomonas qingdaonensis | 99.86 |
DXH1 | 德莱无色杆菌Achromobacter deleyi | 99.22 |
L1 | Methylorubrum populi | 99.85 |
MSZ | 涅斯特连科氏菌Nesterenkonia | 99.72 |
S1 | 伯克霍尔德菌Burkholderia FNTGs | 100.00 |
RG1 | 副伯克霍尔德氏菌Paraburkholderia phytofirmans | 99.86 |
Table 2 Sequence alignment based on 16S rRNA gene
编号Number | 比对结果Alignment result | 相似度Similarity/% |
---|---|---|
14XC | 铜绿假单胞菌Pseudomonas aeruginosa | 99.52 |
19AW | Pseudomonas furukawaii | 99.67 |
BH1AW | Pandoraea morbifera | 99.87 |
BH2XC | 木糖氧化无色杆菌Achromobacter xylosoxidans | 99.55 |
D2 | Pseudomonas qingdaonensis | 99.86 |
DXH1 | 德莱无色杆菌Achromobacter deleyi | 99.22 |
L1 | Methylorubrum populi | 99.85 |
MSZ | 涅斯特连科氏菌Nesterenkonia | 99.72 |
S1 | 伯克霍尔德菌Burkholderia FNTGs | 100.00 |
RG1 | 副伯克霍尔德氏菌Paraburkholderia phytofirmans | 99.86 |
项目Item | 结果Result | 项目Item | 结果Result |
---|---|---|---|
革兰氏染色Gram stainning method | - | 硝酸盐还原试验Nitratereduction test | - |
吲哚试验Indole test | - | 明胶液化试验Gelatinliquefaction test | - |
甲基红试验Methylred test | + | 尿素酶试验Urease test | + |
V⁃P试验 V⁃P test | + | 氧化-发酵试验Oxidation⁃fermentation test | + |
Table 3 Physiological and biochemical test results of strain S1
项目Item | 结果Result | 项目Item | 结果Result |
---|---|---|---|
革兰氏染色Gram stainning method | - | 硝酸盐还原试验Nitratereduction test | - |
吲哚试验Indole test | - | 明胶液化试验Gelatinliquefaction test | - |
甲基红试验Methylred test | + | 尿素酶试验Urease test | + |
V⁃P试验 V⁃P test | + | 氧化-发酵试验Oxidation⁃fermentation test | + |
酚酸种类 Type of phenolic acid | 标准曲线 Standard curve | 检测波长 Wave length/nm | 菌株 Strain | 降解率 Degradation rate/% |
---|---|---|---|---|
水杨酸Salicylic acid | y=0.005 74x-0.011 91,R2=0.995 0 | 300 | S1 | 65.32 |
香草酸Vanillic acid | y=0.006 57x+0.041 87,R2=0.996 2 | 260 | 14XC | 55.00 |
丁香酸Syringic acid | y=0.004 47x+0.035,R2=0.990 8 | 280 | DXH1 | 48.00 |
对羟基苯甲酸 Para-hydroxybenzoic acid | y=0.107 90x-0.019 97,R2=0.999 7 | 225 | D2 | 43.00 |
邻苯二甲酸Phthalic acid | y=0.004 19x+0.001 86,R2=0.998 9 | 216 | L1 | 41.37 |
肉桂酸Cinnamic acid | y=0.016 66x-0.039 72,R2=0.990 8 | 278 | RG1 | 34.00 |
Table 4 Degradation effects of six strains of ginseng autotoxic substances degrading bacteria
酚酸种类 Type of phenolic acid | 标准曲线 Standard curve | 检测波长 Wave length/nm | 菌株 Strain | 降解率 Degradation rate/% |
---|---|---|---|---|
水杨酸Salicylic acid | y=0.005 74x-0.011 91,R2=0.995 0 | 300 | S1 | 65.32 |
香草酸Vanillic acid | y=0.006 57x+0.041 87,R2=0.996 2 | 260 | 14XC | 55.00 |
丁香酸Syringic acid | y=0.004 47x+0.035,R2=0.990 8 | 280 | DXH1 | 48.00 |
对羟基苯甲酸 Para-hydroxybenzoic acid | y=0.107 90x-0.019 97,R2=0.999 7 | 225 | D2 | 43.00 |
邻苯二甲酸Phthalic acid | y=0.004 19x+0.001 86,R2=0.998 9 | 216 | L1 | 41.37 |
肉桂酸Cinnamic acid | y=0.016 66x-0.039 72,R2=0.990 8 | 278 | RG1 | 34.00 |
Fig. 3 Degradation rate of strain S1on salicylic acid under different cultured conditionsNote:Different lowercase letter indicate significant differences at P<0.05 level.
处理 Treatment | 胚根长 Radicle length /mm | 胚轴长/mm Hypocotyl length /mm | RI胚根长 RI of radicle length | RI胚轴长 RI of hypocotyl length |
---|---|---|---|---|
CK | 53.42±5.70 a | 27.95±8.76 a | — | — |
SA0.025 | 46.87±12.04 ab | 13.57±4.16 bc | -0.128±0.18 b | -0.514±0.03 b |
SA2.5 | 46.61±3.58 ab | 12.30±1.01 bc | -0.124±0.06 b | -0.539±0.10 b |
SA250 | 35.77±4.55 b | 7.10±1.60 c | -0.330±0.05 b | -0.731±0.10 c |
SA250+S1 | 60.13±8.71 a | 18.31±2.89 b | 0.154±0.13 a | -0.145±0.02 a |
Table 5 Radicle and hypocotyl lengths of ginseng seed under different treatments
处理 Treatment | 胚根长 Radicle length /mm | 胚轴长/mm Hypocotyl length /mm | RI胚根长 RI of radicle length | RI胚轴长 RI of hypocotyl length |
---|---|---|---|---|
CK | 53.42±5.70 a | 27.95±8.76 a | — | — |
SA0.025 | 46.87±12.04 ab | 13.57±4.16 bc | -0.128±0.18 b | -0.514±0.03 b |
SA2.5 | 46.61±3.58 ab | 12.30±1.01 bc | -0.124±0.06 b | -0.539±0.10 b |
SA250 | 35.77±4.55 b | 7.10±1.60 c | -0.330±0.05 b | -0.731±0.10 c |
SA250+S1 | 60.13±8.71 a | 18.31±2.89 b | 0.154±0.13 a | -0.145±0.02 a |
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