Identification of Pathogen Causing Bacterial Fruit Blotch of Melon in Hainan

doi:10.13304/j.nykjdb.2023.0124

Abstract

Abstract:

To identify the pathogen which seriously caused fruit blotch disease in melo， the melon fruit samples suspected bacterial fruit blotch disease were collected from Dongfang city， Hainan province. Pathogen was isolated and purified from samples， and the pathogenicity of bacterium strains was confirmed according to Koch’s postulate. The identification of the pathogenic strains was carried out according to the colony morphology， physiological and biochemical analysis and 16S rDNA sequences analysis. The results showed that the isolated bacterium with similar cultural characteristics was obtained from all diseased samples， which the separation rate of bacteria was 100%. All of tested strains showed pathogenic to melon fruits. Strains were identified as Gram-negative bacterium， which colony morphology， physiology and biochemistry shared similarly with that of Acidovorax citrull. BLAST analysis showed that the 16S rDNA sequences had 100% homology with that of A. citrulli， and the phylogenetic analysis showed the pathogen was A. citrulli. Above results determined the pathogen of melon fruit blotch，which provided a basis for effective prevention and control of this disease.

Key words: melon, bacterial fruit blotch, pathogen identification, Acidovorax citrulli, Dongfang city in Hainan province

摘要：

细菌性果斑病严重影响甜瓜生产，为确定其病原菌种类，从海南省东方市种植基地采集大量疑似细菌性果斑病的甜瓜样本，进行病原菌分离纯化，并按柯赫法则对获得的菌株进行致病性测定。通过菌落形态观察、生理生化测定及16S rDNA序列比对分析，对分离病原菌进行鉴定。结果表明，大田采集的所有甜瓜病果中均分离到培养性状一致的细菌，细菌分离率为100%；所有分离菌株均对甜瓜果实具有致病性，该病原菌革兰氏反应呈阴性，其菌落形态及生理生化测定结果与已报道的西瓜嗜酸菌（Acidovorax citrulli）一致；BLAST比对与A. citrulli菌株的同源性可达100%，系统发育分析也进一步证实该病菌为A. citrulli。研究结果确定了甜瓜果斑病的致病菌，为该病的有效防治提供了依据。

关键词: 甜瓜, 细菌性果斑病, 病原菌鉴定, 西瓜嗜酸菌, 海南省东方市

CLC Number:

S652

Pengsheng LI, Qingtai HUANG, Yongmei FAN, Meng WANG, Ye YANG. Identification of Pathogen Causing Bacterial Fruit Blotch of Melon in Hainan[J]. Journal of Agricultural Science and Technology, 2024, 26(3): 117-123.

李鹏声, 黄清泰, 范咏梅, 王萌, 杨叶. 海南省东方市甜瓜细菌性果斑病病原鉴定[J]. 中国农业科技导报, 2024, 26(3): 117-123.

Figures/Tables 5

Fig. 1 Symptoms of bacterial fruit blotch disease on melon

Fig. 2 Symptoms of infected melon fruit after inoculation with the isolated strainsNote： 1~6 indicate strains HNTG22， HNTG32， HNTG01， HNF2.3， HNF6.2， HNC03；CK1 and CK2 indicate controls.

Fig. 3 Colony morphology and Gram-negative of pathogen causing sweet melon bacterial fruit blotchA：Colony morphology；B：Gram-negative

Table 1 Physiological and biochemical characteristics of pathogens

测定项目Test item	菌株Strain
测定项目Test item	HNTG01	HNF2.3
KBA培养基产荧光Fluorescent pigment on KBA	-	-
耐盐性Salt tolerance	+	+
过氧化氢酶Catalase	+	+
淀粉水解Hydrolysis of starch	-	-
氧化酶Oxidase	+	+
甲基红Methyl red	-	-
乙酰甲基甲醇Voges-proskauer	-	-
葡萄糖氧化发酵OF test	+	+
明胶液化Gelatin liquefaction	+	+
硝酸盐还原Nitrate reduction	+	+
吲哚产生Indole production	-	-
甘露醇Mannitol	+	+
山梨醇Sorbitol	+	+
肌醇Insoitol	+	+
产生果聚糖Levan formation	-	-
精氨酸双水解Arginine reaction	-	-
吐温80分解Tween 80 hydrolysis	+	+

Fig. 4 Phylogenetic tree generated by maximum likelihood method on the basis of partial 16S rDNA sequence

References 21

1	鲁思梦,王贤磊,宁雪飞,等.甜瓜MR-1细菌性果斑病抗病基因QTL定位分析[J].分子植物育种,2019,17(9):2924-2931.
	LU S M, WANG X L, NING X F, et al.. Location analysis of bacterial fruit blotch resistance gene QTL in melon MR-1 [J]. Mol. Plant Breed., 2019, 17(9):2924-2931.
2	胡国智,冯炯鑫,张炎,等.施肥对甜瓜生长、养分吸收及产量品质的影响[J].新疆农业科学,2014,51(8):1445-1450.
	HU G Z, FENG J X, ZHANG Y, et al.. Effect of fertilization on growth,nutrient uptake, yield and quality of melons [J]. Xinjiang Agric. Sci., 2014, 51(8):1445-1450.
3	刘梦华,卢霞,李腾,等.甜瓜细菌性果斑病病菌鉴定及不同材料抗病性测定[J].安徽农业科学,2021,49(6):145-148.
	LIU M H, LU X, LI T, et al.. Identification of bacterial fruit blotch pathogen and detection of different resistant plant materials in melon [J]. J. Anhui Agric. Sci., 2021, 49(6):145-148.
4	张兴平, RHODES B B.一种毁灭性的西瓜新病害:细菌性果实腐斑病[J].中国瓜菜,1990,3(2):44-45.
5	赵廷昌,孙福在,王兵万,等.哈密瓜细菌性果斑病病原菌鉴定[J].植物病理学报,2001,31(4):357-364.
	ZHAO T C, SUN F Z, WANG B W, et al.. Pathogen identification of hami melon bacterial fruit blotch [J]. Acta Phytopathol. Sin., 2001, 31(4):357-364.
6	王雪,高洁,张静,等.63 种杀菌剂对西瓜、甜瓜细菌性果斑病菌的室内毒力测定[J].吉林农业大学学报,2012,34(6):612-617, 623.
	WANG X, GAO J, ZHANG J, et al.. Virulence evaluation of 63 bactericides to Acidovorax avenae subsp. citrulli on water melon and muskmelon [J]. J. Jilin Agric. Univ., 2012, 34(6):612-617, 623.
7	谢慧婷,李战彪,秦碧霞,等.广西甜瓜细菌性果斑病病原鉴定及16S rDNA序列分析[J].南方农业学报,2016,47(10):1698-1703.
	XIE H T, LI Z B, QIN B X, et al.. Identification of pathogen causing melon bacterial fruit blotch in Guangxi and analysis of its 16S rDNA sequence [J]. J. South. Agric., 2016, 47(10):1698-1703.
8	刘丹丹,刘畅.甜瓜细菌性果斑病菌鉴定及杀菌剂毒力测定[J].北方园艺,2017(18):6-10.
	LIU D D, LIU C. Identification of bacterial fruit blotch and toxicity test of fungicides [J]. Northern Hortic., 2017(18):6-10.
9	于海博,毕馨月,夏博,等.辽宁西瓜和甜瓜细菌性果斑病的病原鉴定[J].沈阳农业大学学报,2019,50(3):345-350.
	YU H B, BI X Y, XIA B, et al.. Identification of the pathogen causing bacterial fruit blotch of watermelon and melon in Liaoning [J]. J. Shenyang Agric. Univ., 2019, 50(3):345-350.
10	张亮,王惠林,万秀琴,等.不同甜瓜材料苗期细菌性果斑病抗性鉴定[J].北方园艺,2016(9):117-120.
	ZHANG L, WANG H L, WAN X Q, et al.. Identification of resistance to Acidovorax citrulli of different melons in the seedling stage [J]. Northern Hortic., 2016(9):117-120.
11	李俊阁,王惠林,张亮,等.种子接种果斑病菌对不同甜瓜幼苗生长的影响[J].北方园艺,2015(12):103-106.
	LI J G, WANG H L, ZHANG L, et al.. Effect of seed inoculated Actinidia citrulli on growth of muskmelon seedlings [J]. Northern Hortic., 2015(12):103-106.
12	倪玉杰.甜瓜细菌性果斑病[J].河北农业,2015,247(10):50.
13	张荣意,谭志琼,文衍堂,等.西瓜细菌性果斑病症状描述和病原菌鉴定[J].热带作物数学报,1998,19(1):70-76.
	ZHANG R Y, TAN Z Q, WEN Y T, et al.. Description and identification of the organism of bacterial fruit blotch of watermelon [J]. China J. Tropic. Crops, 1998, 19(1):70-76.
14	吉训聪,肖彤斌,黄伟明,等.海南甜瓜细菌性果斑病病原菌鉴定[J].广东农业科学,2012,39(11):89-91.
	JI X C, XIAO T B, HUANG W M, et al.. Pathogen identification of bacterial fruit blotch of melon in Hainan [J]. Guangdong Agric. Sci., 2012, 39(11):89-91.
15	季苇芹,叶云峰,张爱萍,等.我国瓜类细菌性果斑病研究新进展[J].中国瓜菜,2022,35(9):1-8.
16	曹淼欣.海南东方一年三造哈密瓜陆续上市[EB/OL].(2022-11-18) [2023-01-19]. .
17	东秀珠,蔡妙英.常见细菌系统鉴定手册[M].北京:科学出版社,2001:1-419.
18	赵斌,何绍江.微生物学实验[M].北京:科学出版社, 2002:1-285.
19	蔡馥宇,关巍,乔培,等.瓜类细菌性果斑病研究新进展[J].中国瓜菜,2017,30(11):1-5.
	CAI F Y, GUAN W, QIAO P, et al.. Research advances of bacterial fruit blotch of cucurbits [J]. China Cucurbits Veget., 2017, 30(11):1-5.
20	WILLEMS A, GOOR M, THIELEMANS S, et al.. Transfer of several phytopathogenic Pseudomonas species to Acidovorax as Acidovorax avenae subsp. avenae subsp. nov., comb. nov., Acidovorax avenae subsp. citrulli, Acidovorax avenae subsp. cattleyae, and Acidovorax konjaci [J]. Int. J. Syst. Bacteriol., 1992.42(1):107-119.
21	SCHAAD N W, POSTNIKOVA E, SECHLER A, et al.. Reclassification of subspecies of Acidovorax avenae as A. avenae (Manns 1905) emend., A. cattleyae (Pavarino, 1911) comb. nov., A. citrulli Schaadet al., 1978) comb. nov., and proposal of A. oryzae sp. nov [J]. Syst. Appl. Microbiol., 2008, 31(6-8):434-446.

[1]	Yuping SHI, Yixian LIU, Guowei LI, Yi TANG, Liming DAI, Lanlan LI, Zhiying CAI. Isolation and Identification of Pathogen of Rubber Tree Phytophthora Leaf Fall Disease and Screening of Control Fungicides [J]. Journal of Agricultural Science and Technology, 2023, 25(11): 114-122.
[2]	Lin ZHANG, Yantao YANG, Lili SONG, Shiping MAO. Current Situation and High Quality Development Countermeasures of Watermelon and Muskmelon Industry in Beijing [J]. Journal of Agricultural Science and Technology, 2023, 25(11): 20-27.
[3]	Keze YANG, Fang WU, Yujie WEI, Liangfang WANG, Hao CHANG, Zhitao WU, Xianzhong YANG. Pathogen Identification and Laboratory Drug Screening of Peony Root Rot in Cold and Cool Region of Hexi [J]. Journal of Agricultural Science and Technology, 2023, 25(10): 144-151.
[4]	Jiahuan HUO, Xiaolei WEN, Shuangmin LI, Lina FENG, Shuhui LAN, Lixin DONG, Sirou GUO, Jianing LI, Jianhua WANG, Huixia QI. Identification and Biological Characteristics of the Pathogen Causing Root Rot of Atractylodes chinensis [J]. Journal of Agricultural Science and Technology, 2022, 24(5): 137-144.
[5]	Yuxin DONG, Zhi ZHENG, Wenkang WANG, Zhiheng WANG, Xiaofeng CHEN. Effect Trials of Nano-silver and Nano-iron Oxide Against Melon Powdery Mildew [J]. Journal of Agricultural Science and Technology, 2022, 24(11): 137-147.
[6]	ZHANG Nana§, LI Shuangmin§, WEN Xiaolei, FENG Lina, WANG Junfeng, YANG Wenjie, HUO Jiahuan, LAN Shuhui, SUN Weiming, QI Huixia. Identification and Biological Characteristics of the Pathogen Causing Pink Disease of Chestnut [J]. Journal of Agricultural Science and Technology, 2021, 23(7): 145-152.
[7]	WEN Xiaolei1,2, QI Huixia1*, SUN Weiming1, LIU Yijian1, FENG Lina1, MENG Tongyao2, HAN Zhiling1, CAO Jia1, WANG Junfeng1. Identification and Biological Characteristics of the Pathogen (Fusarium equiseti) Causing Shoot Blight of Atractylodes chinensis [J]. Journal of Agricultural Science and Technology, 2020, 22(5): 115-121.
[8]	QI Xiaochen1§, ZHAO Ku1§, YE Zupeng1, WANG Ting2, CHEN Bolang1*. Effects of Sophora alopecuroides Green Manure on Growth and Nutrient Characteristics of Melon [J]. Journal of Agricultural Science and Technology, 2018, 20(6): 104-112.
[9]	LI Peng-fei, HUO Xiu-ai, CHENG Yong-qiang, DAI Liang, YANG Bing-yan, DUAN Hui-ju. Assessment of Genetic Diversity in Watermelon Based on SRAP Analysis [J]. , 2013, 15(2): 89-96.
[10]	LU Ning, WANG Qi, SONG Juan, ZHANG Yi-ming, ZHOU Hong-you. Preliminary Analysis on Genetic Improvement of Bio-control Strains X4 of Sweet Melon's Postharvest Diseases and Related Traits [J]. , 2011, 13(2): 25-30.
[11]	Gao Dong1, ZHOU Hong-pei2. The Discussion of Planting Density of Sweet Potato Intercropped with Watermelon in Greenhouse [J]. , 2009, 11(S2): 63-65.