First Report of Diaporthe eres Causing Twig Dieback of White Bark Pine in China

Abstract

White bark pine (Pinus bungeana Zucc.) is popularly cultivated as a garden ornamental tree in China because of the high ornamental value. During July 2017, twig dieback symptoms were found on white bark pine trees in the campus of Henan Agricultural University, Zhenzhou, Henan Province, China. The rate of diseased plants was 25% (up to 40% in 2018) in this campus. Early symptoms appeared as round or oval, red-brown lesions on the twigs, which enlarged progressively, leading to dieback. Two representative Diaporthe strains (zm170130 and zm180920) were isolated and purified from two specimens collected in 2017 and 2018 on potato dextrose agar (PDA). Pycnidia scattered in PDA, dark brown to black, globose to subglobose. Conidiogenous cells phialidic, cylindrical, tapering toward the apex, with periclinal thickening. Two kinds of conidia were produced: α-conidia, unicellular, hyaline elliptical or fusiform, two-guttulate, 5.2 to 9.0 × 2 to 3 µm (n = 50); β-conidia hyaline, aseptate, without guttules, filiform, curved, with obtuse ends, 15.5 to 31 × 0.8 to 1.5 µm (n = 50). These morphological characteristics of the isolates agreed with those of Diaporthe eres (Dissanayake et al. 2015; Liu and Tang 1995; Udayanga et al. 2014). The identity of the two fungal isolates was confirmed by DNA sequencing of the ITS rDNA region (GenBank accession nos. MK265702 and MK265701), TUB gene (GenBank accession nos. MK265704 and MK265703), and histone H3 gene (GenBank accession nos. MK265700 and MK265699), which were over 99% homologous to those consensus sequences of the ex-epitype strain D. eres (KJ210529, KJ420799, KJ210550) (Guarnaccia et al. 2018). The pathogenicity of the isolate zm180920 was tested on 2-year-old healthy twigs of white bark pine in July 2018. Five 2-year-old healthy twigs (30 cm long) cut from 5-year-old trees were used for each treatment. Fifteen mycelial plugs harvested from the periphery of a 7-day-old colony were placed on the surface of the wounded twigs, after the twigs had been surface sterilized with 75% ethanol for 2 min; an equal number of noncolonized plugs of PDA served as the control treatment. All test twigs were kept in a clear plastic box with wet sterilized cotton to maintain high relative humidity at 26 to 30°C in the greenhouse. Symptoms similar to those originally observed on the naturally infected twigs were observed 10 days after inoculation, and the same fungus was reisolated from each of the symptomatic twigs; control twigs remained asymptomatic, and no fungus was isolated. The isolates from the symptomatic twigs were identified as D. eres on the basis of the morphology and the same gene sequences. Koch’s postulates were repeated three times with the same results. The fungus has been reported on peach, grapevine, jujube, Red Robin, and so on in China (Dissanayake et al. 2015; Liu and Tang 1995; Song et al. 2019; Yang et al. 2018; Zhang et al. 2018). To our knowledge, this is the first report of D. eres causing twig dieback of white bark pine in China. Disease control strategies in the region are being further studied.