生防菌Pseudomonas koreensis ZF518的分离鉴定及对黄瓜棒孢叶斑病生防效果研究

doi:10.16420/j.issn.0513-353x.2024-0985

园艺学报 ›› 2025, Vol. 52 ›› Issue (12): 3346-3362.doi: 10.16420/j.issn.0513-353x.2024-0985

生防菌Pseudomonas koreensis ZF518的分离鉴定及对黄瓜棒孢叶斑病生防效果研究

丁常宗¹^,^*, 李磊¹^,^*, 姜振群¹, 申雨荷¹, 高素芹¹, 德吉央宗², 石延霞¹, 柴阿丽¹, 范腾飞¹, 孙先花¹, 李宝聚¹^,^**(), 谢学文¹^,^**()

¹ 中国农业科学院蔬菜花卉研究所，蔬菜生物育种全国重点实验室，北京 100081
² 西藏自治区农牧科学院蔬菜花卉研究所，拉萨 850030

收稿日期:2025-09-29 修回日期:2025-11-04 出版日期:2025-12-25 发布日期:2025-12-20
通讯作者:
**（E-mail：libaojuivf@163.com；
xiexuewen@caas.cn）
作者简介:
* 共同第一作者
基金资助:
拉萨市重点科技计划项目(LSKJ202428); 中国农业科学院科技创新工程项目(CAAS-ASTIP-IVFCAAS); 农业农村部园艺作物生物学与种质创制重点实验室项目(IVF2023)

Isolation and Identification of Pseudomonas koreensis ZF518 and Its Biocontrol Effect on Cucumber Leaf Spot

DING Changzong¹, LI Lei¹, JIANG Zhenqun¹, SHEN Yuhe¹, GAO Suqin¹, DEJI Yangzong², SHI Yanxia¹, CHAI Ali¹, FAN Tengfei¹, SUN Xianhua¹, LI Baoju¹^,^**(), XIE Xuewen¹^,^**()

¹ State Key Laboratory of Vegetable Biobreeding，Institute of Vegetables and Flowers，Chinese Academy of Agricultural Sciences，Beijing 100081，China
² Institute of Olericulture，Xizang Academy of Agriculture and Animal Husbandry Sciences，Lhasa 850030，China

Received:2025-09-29 Revised:2025-11-04 Published:2025-12-25 Online:2025-12-20

摘要/Abstract

摘要：

为获得对黄瓜棒孢叶斑病具有良好防效的生防假单胞菌，通过低温富集及特定培养基筛选从中国各地56份黄瓜根际土样中得到557株能在365 nm紫外光照下具荧光的菌株。利用平板对峙法筛选出对多主棒孢菌有拮抗作用的菌株，通过活体盆栽试验得到1株对黄瓜棒孢叶斑病防效较好且稳定的菌株ZF518。其平板抑菌率为65.82%，盆栽防治效果为57.08%。通过形态学、生理生化特性及多基因系统发育树的构建，将ZF518鉴定为韩国假单胞杆菌Pseudomonas koreensis。该菌株对离体情况下的多种病原真菌具有直接抑制效果，并可产生铁载体，为黄瓜棒孢叶斑病的生物防治提供生防资源。

关键词: 黄瓜, 棒孢叶斑病, 假单胞杆菌, 生防菌鉴定, 生物防治, 生理生化, 系统进化

Abstract:

To obtain biocontrol Pseudomonas strains with high efficacy against cucumber Corynespora leaf spot，a total of 557 fluorescent strains were isolated from 56 cucumber rhizosphere soil samples collected across China. This was achieved through low-temperature enrichment and selective medium screening，with fluorescence detected under 365 nm UV light. Using the plate confrontation assay，strains antagonistic against Corynespora cassiicola were selected. Through in vivo pot experiments，one strain，designated ZF518，was identified as providing stable and effective control of cucumber Corynespora leaf spot，with an inhibition rate of 65.82% in vitro and a control efficacy of 57.08% in planta. Based on a polyphasic taxonomic approach including morphology，physiological-biochemical characteristics，and multi-gene phylogenetic analysis，strain ZF518 was identified as Pseudomonas koreensis. This strain exhibited direct inhibitory effects against various phytopathogenic fungi in vitro and was capable of producing siderophores. These findings indicate that P. koreensis ZF518 is a promising biocontrol resource for managing cucumber Corynespora leaf spot.

Key words: cucumber, Corynespora cassiicola, Pseudomonas, biocontrol agent identification, Biological control, physiological and biochemical characteristics, phylogenetic evolution

丁常宗, 李磊, 姜振群, 申雨荷, 高素芹, 德吉央宗, 石延霞, 柴阿丽, 范腾飞, 孙先花, 李宝聚, 谢学文. 生防菌Pseudomonas koreensis ZF518的分离鉴定及对黄瓜棒孢叶斑病生防效果研究[J]. 园艺学报, 2025, 52(12): 3346-3362.

DING Changzong, LI Lei, JIANG Zhenqun, SHEN Yuhe, GAO Suqin, DEJI Yangzong, SHI Yanxia, CHAI Ali, FAN Tengfei, SUN Xianhua, LI Baoju, XIE Xuewen. Isolation and Identification of Pseudomonas koreensis ZF518 and Its Biocontrol Effect on Cucumber Leaf Spot[J]. Acta Horticulturae Sinica, 2025, 52(12): 3346-3362.

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基因 Gene	引物名称 Primer name	引物序列（5′-3′） Primer sequence	扩增长度/bp Amplicon length	来源 Source
16S rDNA	27F	GGCGACATGGTCAACGG	1 465	Dutta et al.，2022
	1492R	CGGCTGGCGGCGTATAT	1 465	Dutta et al.，2022
rpoD	rpoD-F	CACGGTTGAGCACATCCTCT	1 105	Dutta et al.，2022
	rpoD-R	GGAGAGTACTTCGCGAGTCG	1 105	Dutta et al.，2022
rpoB	rpoB-F	GTCCGACTTCACTGTGAGCA	1 126	Dutta et al.，2022
	rpoB-R	GTGAAAAAGCCAGCGACGTT	1 126	Dutta et al.，2022
gyrB	gyrB-F	GACAAGCTGGTGTCTTCCGA	1 320	Dutta et al.，2022
	gyrB-R	CATCTCGCCCAGACCTTTGT	1 320	Dutta et al.，2022

基因 Gene	引物名称 Primer name	引物序列（5′-3′） Primer sequence	扩增长度/bp Amplicon length	来源 Source
16S rDNA	27F	GGCGACATGGTCAACGG	1 465	Dutta et al.，2022
	1492R	CGGCTGGCGGCGTATAT	1 465	Dutta et al.，2022
rpoD	rpoD-F	CACGGTTGAGCACATCCTCT	1 105	Dutta et al.，2022
	rpoD-R	GGAGAGTACTTCGCGAGTCG	1 105	Dutta et al.，2022
rpoB	rpoB-F	GTCCGACTTCACTGTGAGCA	1 126	Dutta et al.，2022
	rpoB-R	GTGAAAAAGCCAGCGACGTT	1 126	Dutta et al.，2022
gyrB	gyrB-F	GACAAGCTGGTGTCTTCCGA	1 320	Dutta et al.，2022
	gyrB-R	CATCTCGCCCAGACCTTTGT	1 320	Dutta et al.，2022

基因 Gene	引物序列（5′-3′） Primer sequence	扩增长度/bp Amplicon length	来源 Source
吩嗪Phenazine（phzCD）	TGCCAAGCCTCGCTCCAAC	1 400	Zhang et al.，2006
吩嗪Phenazine（phzCD）	CCGCGTTGTTCCTCGTTCAT	1 400	Zhang et al.，2006
硝吡咯菌素Pyrrolnitrin（prnC）	CCACAAGCCCGGCCAGGAGC	720	Mavrodi et al.，2001
硝吡咯菌素Pyrrolnitrin（prnC）	GAGAAGAGCGGGTCGATGAAGCC	720	Mavrodi et al.，2001
2-羟基吩嗪 2-hydroxylated（phzO）	AAGTCCAGATGCGAAAGAACG	1 400	Delaney et al.，2001
2-羟基吩嗪 2-hydroxylated（phzO）	GTGAAAAAGCCAGCGACGTT	1 400	Delaney et al.，2001
2,4-二乙酰基间苯三酚 2,4-DAPG（phlD）	GAGGACGTCGAAGACCACCA	745	Mavrodi et al.，2001
2,4-二乙酰基间苯三酚 2,4-DAPG（phlD）	AAGTGGCATGGCTCGAACAAAG	745	Mavrodi et al.，2001
藤黄绿脓菌素 Pyoluteorin（pltB）	CGGAGCATGGACCCCCAGC	900	Mavrodi et al.，2001
藤黄绿脓菌素 Pyoluteorin（pltB）	GTGCCCGATATTGGTCTTGACC	900	Mavrodi et al.，2001

基因 Gene	引物序列（5′-3′） Primer sequence	扩增长度/bp Amplicon length	来源 Source
吩嗪Phenazine（phzCD）	TGCCAAGCCTCGCTCCAAC	1 400	Zhang et al.，2006
吩嗪Phenazine（phzCD）	CCGCGTTGTTCCTCGTTCAT	1 400	Zhang et al.，2006
硝吡咯菌素Pyrrolnitrin（prnC）	CCACAAGCCCGGCCAGGAGC	720	Mavrodi et al.，2001
硝吡咯菌素Pyrrolnitrin（prnC）	GAGAAGAGCGGGTCGATGAAGCC	720	Mavrodi et al.，2001
2-羟基吩嗪 2-hydroxylated（phzO）	AAGTCCAGATGCGAAAGAACG	1 400	Delaney et al.，2001
2-羟基吩嗪 2-hydroxylated（phzO）	GTGAAAAAGCCAGCGACGTT	1 400	Delaney et al.，2001
2,4-二乙酰基间苯三酚 2,4-DAPG（phlD）	GAGGACGTCGAAGACCACCA	745	Mavrodi et al.，2001
2,4-二乙酰基间苯三酚 2,4-DAPG（phlD）	AAGTGGCATGGCTCGAACAAAG	745	Mavrodi et al.，2001
藤黄绿脓菌素 Pyoluteorin（pltB）	CGGAGCATGGACCCCCAGC	900	Mavrodi et al.，2001
藤黄绿脓菌素 Pyoluteorin（pltB）	GTGCCCGATATTGGTCTTGACC	900	Mavrodi et al.，2001

处理 Treatment	抑菌直径/cm Antibacterial diameter	抑菌率/% Inhibition rate	病情指数 Disease index	防治效果/% Control efficiency
ZF518	5.20 ± 0.10 a	65.82 ± 1.27 a	27.92 ± 0.79 f	57.08 ± 1.61 b
ZF549	4.23 ± 0.15 c	53.59 ± 1.93 c	48.92 ± 0.70 e	41.01 ± 0.84 c
ZF557	4.40 ± 0.10 bc	55.70 ± 1.27 bc	58.84 ± 2.35 cd	29.04 ± 2.84 d
ZF561	4.50 ± 0.10 bc	56.96 ± 1.27 bc	62.12 ± 3.71 bc	25.08 ± 4.47 d
ZF577	4.70 ± 0.10 b	59.49 ± 1.27 b	63.15 ± 3.70 bc	23.84 ± 4.46 d
ZF582	4.60 ± 0.10 b	58.23 ± 1.27 b	69.83 ± 2.96 b	15.79 ± 3.57 e
ZF589	4.27 ± 0.20 c	54.01 ± 4.79 c	59.92 ± 3.10 cd	27.73 ± 3.74 d
ZF593	5.07 ± 0.06 a	64.14 ± 0.73 a	8.92 ± 2.11 e	41.01 ± 2.54 c
ZF612	4.50 ± 0.10 bc	56.96 ± 1.27 bc	52.19 ± 2.06 de	37.05 ± 2.49 e
75%百菌清WP 75% Chlorothalonil WP	—	—	27.92 ± 0.79 f	66.32 ± 0.95 a
病原菌对照 Pathogen control	—	—	82.92 ± 7.34 a	—

生防菌Pseudomonas koreensis ZF518的分离鉴定及对黄瓜棒孢叶斑病生防效果研究

Isolation and Identification of Pseudomonas koreensis ZF518 and Its Biocontrol Effect on Cucumber Leaf Spot

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