甘肃省甜瓜黑斑病菌（Alternaria tenuissima）的分离鉴定

doi:10.16420/j.issn.0513-353x.2020-0929

摘要/Abstract

摘要：

对甘肃省白银市甜瓜黑斑病菌进行分离、鉴定和生物学特性测定。结果表明,分离物98BYT2-1菌落呈青褐色圆形,分生孢子单生或串生,倒棍棒状或卵圆形,具纵、横隔膜,大小为（21.94 ~ 52.10）μm ×（7.01 ~ 13.93）μm,ITS、CAL和gpd基因序列分析表明其与细极链格孢（Alternaria tenuissima）同源性达到100%,且在系统发育树上聚在一起,将其鉴定为细极链格孢（A. tenuissima）。生物学特性测定表明,最适菌丝生长、产孢和孢子萌发的碳源分别为淀粉、乳糖和葡萄糖;最适菌丝生长的氮源为酵母膏和牛肉膏、培养基为PDA,最适产孢和孢子萌发的氮源均为蛋白胨、培养基为PCA;最适菌丝生长和孢子萌发的pH均为7、温度为30 ℃,最适产孢的pH为6、温度为15 ℃;黑暗条件不利于菌丝生长但利于产孢和孢子萌发。该研究结果为甘肃省甜瓜黑斑病的诊断和综合防治提供了理论依据。

关键词: 甜瓜黑斑病, 细极链格孢, 鉴定, 生物学特性, 测定

Abstract:

This experiment was conducted to isolate,identify and determine the biological characteristics of black spot pathogen on melon in Baiyin City,Gansu Province. The results showed that the colony of isolate 98BYT2-1 was green-brown,round,conidia solitary or clustered,inverted rod-shaped or oval,with a vertical and horizontal septum. The size of conidia was（21.94-52.10）μm ×（7.01-13.93）μm,combining ITS,CAL and gpd gene sequence analysis showed that the strain 98BYT2-1 had 100% homology with Alternaria tenuissima and they were clustered together on the phylogenetic tree. Together,the isolate was identified as A. tenuissima. Starch,lactose and glucose were the most appropriate carbon sources for mycelia growth,spore formation and spore germination,respectively. Yeast extract and beef extract were the most suitable nitrogen sources for mycelia growth,but peptone was the most suitable source of nitrogen for sporulation and spore germination. In addition,PDA facilitated the mycelia growth of pathogen,and PCA was suitable for sporulation and spore germination. The optimum pH for mycelia growth and spore germination was 7 and the value for sporulation was 6. The optimum temperature for the growth and spore germination of mycelia was 30 ℃,and the sporulation was 15 ℃. The dark held up the growth of mycelium but benefited to sporulation and spore germination. The results obtained will provide a basis for the diagnosis and comprehensive control of melon black spot in Gansu Province.

Key words: muskmelon alternaria black spot, Alternaria tenuissima, identification, biological characteristics, determination

中图分类号:

S652

施兆荣, 张广荣, 孙述俊, 金梦军, 崔凌霄, 刘彦超, 魏立娟, 梅大海, 杨成德. 甘肃省甜瓜黑斑病菌（Alternaria tenuissima）的分离鉴定[J]. 园艺学报, 2022, 49(2): 427-436.

SHI Zhaorong, ZHANG Guangrong, SUN Shujun, JIN Mengjun, CUI Lingxiao, LIU Yanchao, WEI Lijuan, MEI Dahai, YANG Chengde. Isolation and Identification of Melon Black Spot in Gansu Province[J]. Acta Horticulturae Sinica, 2022, 49(2): 427-436.

图/表 11

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基因 Gene	引物序列（5′-3′） Primer sequence
ITS	ITS1：TCCGTAGGTGAACAYCCTGCG G	ITS4：TCCTCCGCTTATTGATATGC
CAL	CALDR1：CTTCTGCATCATCAYCTGGACG	CALDF1：AGCAAGTCTCCGAGTTCAAGG
gpd	gpd-1：CAACGGCTTCGGTCGCATTG	gpd-2：GCCAAGCAGTTGGTTGTGC

基因 Gene	引物序列（5′-3′） Primer sequence
ITS	ITS1：TCCGTAGGTGAACAYCCTGCG G	ITS4：TCCTCCGCTTATTGATATGC
CAL	CALDR1：CTTCTGCATCATCAYCTGGACG	CALDF1：AGCAAGTCTCCGAGTTCAAGG
gpd	gpd-1：CAACGGCTTCGGTCGCATTG	gpd-2：GCCAAGCAGTTGGTTGTGC

碳源 Carbon source	菌落直径/cm Colony diameter	产孢量/（× 10⁶ CFU · mL^-1） Sporulation	孢子萌发率/ % Spore germination	碳源 Carbon source	菌落直径/ cm Colony diameter	产孢量/（× 10⁶ CFU · mL^-1） Sporulation	孢子萌发率/ % Spore germination
木糖 Xylose	2.47 ± 0.02 e	13.17 ± 2.46 e	52.00 ± 3.06 de	葡萄糖 Glucose	2.20 ± 0.05 f	21.67 ± 0.44 d	90.33 ± 1.45 a
甘露醇 Mannite	2.53 ± 0.04 e	23.50 ± 0.76 d	69.33 ± 2.03 c	乳糖 Lactrose	4.13 ± 0.22 c	61.50 ± 0.50 a	82.33 ± 2.03 b
肌醇 Inosite	2.83 ± 0.07 d	24.67 ± 1.01 d	53.00 ± 4.93 de	淀粉 Starch	5.47 ± 0.03 a	56.50 ± 1.52 a	49.00 ± 1.52 c
麦芽糖 Maltose	4.43 ± 0.01 b	50.00 ± 0.76 b	58.67 ± 1.33 d	蔗糖 Sucrose	3.03 ± 0.01 d	43.33 ± 3.94 c	47.00 ± 0.58 e

碳源 Carbon source	菌落直径/cm Colony diameter	产孢量/（× 10⁶ CFU · mL^-1） Sporulation	孢子萌发率/ % Spore germination	碳源 Carbon source	菌落直径/ cm Colony diameter	产孢量/（× 10⁶ CFU · mL^-1） Sporulation	孢子萌发率/ % Spore germination
木糖 Xylose	2.47 ± 0.02 e	13.17 ± 2.46 e	52.00 ± 3.06 de	葡萄糖 Glucose	2.20 ± 0.05 f	21.67 ± 0.44 d	90.33 ± 1.45 a
甘露醇 Mannite	2.53 ± 0.04 e	23.50 ± 0.76 d	69.33 ± 2.03 c	乳糖 Lactrose	4.13 ± 0.22 c	61.50 ± 0.50 a	82.33 ± 2.03 b
肌醇 Inosite	2.83 ± 0.07 d	24.67 ± 1.01 d	53.00 ± 4.93 de	淀粉 Starch	5.47 ± 0.03 a	56.50 ± 1.52 a	49.00 ± 1.52 c
麦芽糖 Maltose	4.43 ± 0.01 b	50.00 ± 0.76 b	58.67 ± 1.33 d	蔗糖 Sucrose	3.03 ± 0.01 d	43.33 ± 3.94 c	47.00 ± 0.58 e

氮源 Nitrogen source	菌落直径/ cm Colony diameter	产孢量/（× 10⁶CFU · mL^-1） Sporulation	孢子萌发率/ % Spore germination
氯化铵 Ammonium Chloride	1.28 ± 0.06 f	6.67 ± 0.33 e	32.67 ± 1.76 d
蛋白胨 Peptone	5.77 ± 0.05 b	47.50 ± 2.36 a	92.33 ± 0.89 a
尿素 Carbamide	3.23 ± 0.13 d	37.83 ± 2.05 bc	56.33 ± 4.67 b
硫酸铵 Ammonium Sulfate	1.83 ± 0.12 e	0.13 ± 0.02 f	32.67 ± 1.76 d
硝酸钾 Potassium Nitrate	3.68 ± 0.11 c	38.17 ± 1.59 bc	52.67 ± 5.46 bc
酵母膏 Yeast extrac	6.08 ± 0.02 a	26.00 ± 0.76 d	90.33 ± 2.33 a
牛肉膏 Beef extract	6.17 ± 0.17 a	33.50 ± 0.00 c	51.33 ± 2.19 bc
硝酸钠 Sodium Nitrate	3.03 ± 0.01 d	43.33 ± 3.94 ab	47.00 ± 0.58 c