Isolation and identification of pathogen causing anthracnose on Aspidistra elatior

doi:10.3969/j.issn.1004-1524.2021.08.13

Acta Agriculturae Zhejiangensis ›› 2021, Vol. 33 ›› Issue (8): 1461-1467.DOI: 10.3969/j.issn.1004-1524.2021.08.13

• Plant Protection • Previous Articles Next Articles

Isolation and identification of pathogen causing anthracnose on Aspidistra elatior

HUANG Yuting(), SUN Jie, ZHANG Huijuan, ZHU Yan, JIANG Ming^*()

College of Life Sciences, Taizhou University, Taizhou 318000, China

Received:2020-10-18 Online:2021-08-25 Published:2021-08-27
Contact: JIANG Ming

Abstract

Abstract:

To identify the pathogenic fungus causing anthracnose on Aspidistra elatior, tissue isolation method was applied on the samples collected and a fungus strain YYL was obtained. Acupuncture method was used to confirm its pathogenicity. Based on the morphological identification results, ITS, ACT, GAPDH, and CAL fragments were cloned for multigene analysis. The results showed that YYL had high pathogenicity, and the characteristics of colony, hyphae, and conidium were well matched to Colletotrichum lilii. The gene fragments from YYL shared the highest similarity with C. lilii, and the ITS and ACT sequences of both YYL and C. lilii were identical. By combining the morphological features and molecular biological identification results, C. lilii was identified as the pathogen causing anthracnose on Aspidistra elatior.

Key words: Aspidistra elatior, anthracnose, pathogen, Colletotrichum lilii

CLC Number:

S432.1

HUANG Yuting, SUN Jie, ZHANG Huijuan, ZHU Yan, JIANG Ming. Isolation and identification of pathogen causing anthracnose on Aspidistra elatior[J]. Acta Agriculturae Zhejiangensis, 2021, 33(8): 1461-1467.

Figures/Tables 7

Table 1 PCR Primers and their optimum annealing temperatures

基因名称 Gene name	引物名称 Primer name	引物序列(5’→3’) Primer sequence (5’→3’)	退火温度 Annealing temperature/℃
ITS	ITS1	TCCGTAGGTGAACCTGCGG	56.3
	ITS4	TCCTCCGCTTATTGATATGC
ACT	512F	ATGTGCAAGGCCGGTTTCGC	62.1
	783R	TACGAGTCCTTCTGGCCCAT
GAPDH	GDF1	GCCGTCAACGACCCCTTCATTGA	56.3
	GDR1	GGGTGGAGTCGTACTTGAGCATGT
CAL	CL1	GARTWCAAGGAGGCCTTCTC	56.3
	CL2	TTTTTGCATCATGAGTTGGAC

Fig.1 Anthracnose symptoms on leaves of Aspidistra elatior A, Infected plants; B, Leaf lesion; C, Penetration by fungal hyphae through a stomatal pore, 2 500×.

Fig.2 Morphological characteristics of pathogenic fungus YYL A, Colonial morphology; B, Hypha morphology; C, Conidia morphology; D, Morphology of hypha and spores under a scanning electron microscope.

Fig.3 Pathogenicity detection A, Control; B, Leaf inoculated with pathogenic fungus.

Fig.4 Agarose gel electrophoresis of PCR products about different gene fragments M, DL 2000 DNA marker; 1-2, ITS; 3-4, ACT; 5-6, GAPDH; 7-8, CAL。

Fig.5 Multiple alignment of ACT sequence and its homologous sequences

Fig.6 Phylogenetic tree based on gene fragments and their homologous sequences A, ITS; B, ACT; C, GAPDH; D, CAL. MH292787, Colletotrichum liriopes; MH292784, C. tofieldiae; KC843542, C. bletillum; MN752248, C. incanum; GU227908, C. lilii; MN433299, C. spaethianum; GQ856785, C. dematium; KC843538, C. guizhouensis; JQ005837, C. cereale.

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Isolation and identification of pathogen causing anthracnose on Aspidistra elatior

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