Identification and Biological Characteristics of a Colletotrichum fructicola Causing Chestnut Internal Rot Disease

doi:10.13304/j.nykjdb.2023.0871

Abstract

Abstract:

Chestnut internal rot is the main disease of chestnut during postharvest storage， which causes rot and mildew of chestnut kernel and brings huge economic losses to production. In order to clarify the pathogenic bacteria and its biological characteristics， the strain ZHZF21 was obtained by tissue isolation method. The strain was identified by morphological characteristics and ITS， TUB2 and CAL multi-gene sequence analysis. The pathogenicity of strain ZHZF21 was determined by mycelial inoculation method， and its biological characteristics were studied. The results showed that the colony of strain ZHZF21 was dark green ring concentric circles， with sexual and asexual reproduction stages. This strain was clustered with Colletotrichum fructicola numbered NDSTY31 in GenBank based on multi-gene sequences. Combined with colony morphological characteristics and molecular tree， strain ZHZF21 was preliminarily identified as Colletotrichum fructicola. The brown lesions after inoculation of strain ZHZF21 were consistent with the natural symptoms. The strain ZHZF21 could grow at 5~35 ℃， the optimum temperature was 25 ℃， the lethal temperature was 45~50 ℃， and the optimum pH was 6. Under the condition of full darkness， strain ZHZF21 grew fastest， the utilization rates of arabinose and yeast were the highest， the utilization rate of lactose was the lowest， and urea could hardly be used.

Key words: chestnut, internal rot, pathogen identification, colletotrichum fructicola, biological characteristics

摘要：

板栗内腐病是板栗采后贮藏期主要病害，引起栗仁腐烂霉变，给生产带来巨大经济损失。为明确板栗内腐病的致病菌及其生物学特性，采用组织分离法获得菌株ZHZF21，通过形态学特征及ITS、TUB2、CAL多基因序列分析对该菌株进行鉴定，以菌丝块有伤接种法测定菌株ZHZF21的致病性，并对其生物学特性进行研究。结果表明，菌株ZHZF21的菌落呈墨绿色环状同心圆圈，具备有性和无性繁殖阶段。该菌株的序列与GenBank中编号为NDSTY31的果生炭疽菌（Colletotrichum fructicola）聚为一支。结合菌落形态特征及分子树将ZHZF21初步鉴定为果生炭疽菌。将菌株ZHZF21接种栗果后产生褐色病斑，与自然发病症状一致。该菌株在5~35 ℃均可生长，最适生长温度为25 ℃，致死温度为45~50 ℃，最适pH为6；在全黑暗条件下生长最快，阿拉伯树脂糖和酵母的利用率最高，乳糖的利用率最低，几乎不能利用尿素。

关键词: 板栗, 内腐病, 病原菌鉴定, 果生炭疽菌, 生物学特性

CLC Number:

S664.2

Shifeng MU, Xiaolei WEN, Lina FENG, Dexuan ZHAO, Suhong GAO, Peng GAO, Huixia QI. Identification and Biological Characteristics of a Colletotrichum fructicola Causing Chestnut Internal Rot Disease[J]. Journal of Agricultural Science and Technology, 2024, 26(9): 122-128.

母时风, 温晓蕾, 冯丽娜, 赵德轩, 高素红, 高朋, 齐慧霞. 一种引起板栗内腐病的果生炭疽菌鉴定及其生物学特性[J]. 中国农业科技导报, 2024, 26(9): 122-128.

Figures/Tables 7

Table 1 Primers sequence used

基因Gene	引物名称Primer name	引物序列Primer sequence（5’-3’）
ITS	ITS1	TCCGTAGGTGAACCTGCGG
ITS	ITS4	TCCTCCGCTTATTGATATGC
TUB2	βt2a	GGTAACCAAATCGGTGCTGCTTTC
TUB2	βt2b	ACCCTCAGTGTAGTGACCCTTGGC
CAL	CL1	GAATTCAAGGAGGCCTTCTC
CAL	CL2	CTTCTGCATCATGAGCTGGAC

Fig. 1 Culture morphology of strain ZHZF21A： Colony on PDA； B： Mycelium； C： Ascus disc； D： Conidia； E： Conidia pile； F： Ascus shell； G： Ascus； H： Ascospores

Fig. 2 NJ phylogenetic tree based on ITS， TUB2 and CAL gene sequences

Fig. 3 Symptoms of chestnut kernelsA： Naturally diseased kernels； B： Kernels inoculated strain ZHZF21； C： CK

Fig. 4 Colony diameter of strain ZHZF21 under different temperaturesNote：Different lowercase letters indicate significant differences between different treatments at P<0.05 level.

Fig. 5 Colony diameter of strain ZHZF21 under different carbon and nitrogen sourcesNote：Different lowercase letters indicate significant differences between different treatments at P<0.05 level.

Fig. 6 Growth of strain ZHZF21 under different temperatures

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