Characterization and fungicides sensitivity of Colletotrichum species causing Hydrangea macrophylla anthracnose in Beijing, China

Abstract

Hydrangea macrophylla (Thunb.) Ser. is one of the widely cultivated plants in home gardens and scenic areas of China. Anthracnose disease is commonly observed during the normal growth of H. macrophylla, significantly impacting its ornamental and economic values. From 2021 to 2023, an investigation on H. macrophylla anthracnose was carried out in nine parks of Beijing, China, and a total of 114 Colletotrichum isolates were obtained from the diseased leaves with typical anthracnose symptoms. Based on morphological characteristics and phylogenetic analysis of six genomic loci including rDNA-ITS, ACT, TUB2, CAL, CHS-1, and GAPDH, these isolates were identified as belonging to six Colletotrichum species. Among which, C. gloeosporioides was the most abundant (65 isolates, 57.0%), followed by C. fructicola (33 isolates, 28.9%), C. aenigma (8 isolates, 7.0%), C. truncatum (4 isolates, 3.5%), C. subacidae (2 isolates, 1.8%) and C. sojae (2 isolates, 1.8%). Pathogenicity test conducted on detached leaves of H. macrophylla revealed a distinct variation in virulence among isolates from different Colletotrichum species, and wounding was either essential or conducive to successful infection. Specifically, C. gloeosporioides exhibited greater aggressiveness, resulting in larger lesions, while C. subacidae induced lesions most quickly. Fungicide sensitivity assays demonstrated that prochloraz exerted a remarkable inhibitory effect on the mycelial growth of representative isolates belonging to the three predominant Colletotrichum species. In contrast to difenoconazole and tebuconazole, the mean EC₅₀ values for prochloraz against C. gloeosporioides, C. fructicola, and C. aenigma were 0.062, 0.033, and 0.023 μg/ml, respectively. This is the first report of C. aenigma, C. truncatum, C. subacidae and C. sojae causing H. macrophylla anthracnose worldwide including China. These findings have elucidated the Colletotrichum species associated with H. macrophylla anthracnose as well as their fungicides sensitivities in Beijing, China. This provides a scientific foundation for the accurate diagnosis and local management of H. macrophylla anthracnose.

Keywords: Colletotrichum species; Hydrangea macrophylla; anthracnose; fungicide sensitivity; multi-loci phylogeny.

Figure 1

Typical symptoms of anthracnose on the leaves of Hydrangea macrophylla in Beijing, China. (A, B). Symptoms of small spots on the leaves; (C, D). Coalesce of brown necrotic lesions on the leaves; (E, F). Anthracnose symptoms on the whole plants.

Figure 2

Phylogenetic tree based on the concatenated sequences of rDNA-ITS, ACT, TUB2, CAL, CHS-1, and GAPDH genomic regions of 35 representative Colletotrichum isolates obtained from Hydrangea macrophylla in Beijing, China. The species Monilochaetes infuscans CBS 869.96 was selected as an outgroup. Maximum likelihood boot strap support values (ML≥50) and Bayesian posterior probability values (PP≥0.90) were shown at the nodes, respectively. Colored blocks indicated clades containing isolates from different Colletotrichum species in this study.

Figure 3

Culture characteristics and microscopic features of six Colletotrichum species obtained from Hydrangea macrophylla in Beijing, China. Each species was represented in four pictures (A-D). (A, B). Upper and reverse view of colony on potato dextrose agar at 28°C; (C). Conidia, scale bars = 20 µm; (D). Appressoria, scale bars = 20 µm. Plates 1-6 referred to C. gloeosporides JZB1040-3-1, C. fructicola JZB1241-2-7, C. aenigma JZB1040-11-2, C. truncatum JZB1564-1-2, C.subacidae JZB1490-1-4, and C. sojae JZB1124-3-1, respectively.

Figure 4

Symptoms of Hydrangea macrophylla leaves induced by inoculation of representative isolates from six Colletotrichum species under wounded conditions. The inoculation was conducted by dropping 5 mm mycelia disks of representative isolates on the detached leaves of H. macrophylla. Each leaf was wounded by pin-pricking with a sterilized needle. Control leaves were treated with sterilized agar blocks with the same size. The lesions on leaves were photographed 7 days post inoculation.