First Report of Valsa nivea Causing Valsa Canker on Korla Pear (Pyrus sinkiangensis) in the World

Abstract

Korla pear (Pyrus sinkiangensis Yü) is an important commercial fruit tree that originated in China (Zhou et al. 2020). In April 2020, a survey was conducted in Aksu region, Xinjiang (40°55'37"N, 80°28'42"E), China. Some Korla pear trees (>15 years old) exhibited symptoms of branch dieback and branch cankers. Cankers observed on the trunk and branches of the tree were sunken, dark ulcerative lesions sometimes exhibiting signs of stromata erumpent through the bark and exuding yellow to reddish-orange spore tendrils. Of the 180 plants surveyed, 80% were symptomatic. Thirty samples of symptomatic tissues of infected branches were taken to the laboratory. Bark and cortical wood samples containing necrotic and healthy tissue were excised with flame-sterilized scalpels, surface disinfected with 75% ethanol and 1% NaClO, placed on PDA plates, and incubated at 25°C. A total of 30 fungal isolates were obtained. Among them, 28 isolates were identified as Valsa mali var. pyri (Lu. 1992) based on morphological and molecular identification, and two isolates (ALE6T-GP21 and ALE7T-GP23) were identified as Valsa nivea (Hoffm.) Fr. Valsa nivea isolates had a fine villi form mycelium that was initially white, turned grayish-green over time and grew close to the medium surface. Cultures also contained black ostiolate pycnidia in a stroma that consisted of multiple irregular locules. Conidiophores were hyaline, occasionally branched at the bases and (15.50-)16.48-17.94(-18.50)×(1.00-)1.13-1.37(-1.50) μm (n=20). Conidiogenous cells were phialidic and subcylindrical that taper towards the apex. Conidia were hyaline, banana-like and (5.47-)6.13-6.97(-7.64)×(1.02-)1.06-1.20(-1.23) μm (n=10). The molecular characteristics are consistent with the previous description of V. nivea (Adams et al. 2006). The internal transcribed spacer (ITS), transcription elongation factor (tef-1α) and β-tubulin (Tub2) gene were sequenced using ITS1/ITS4, EF1-728F/EF1-986R and Bt2a/Bt2b primers, respectively (Zhang et al. 2014). BLAST (Basic Local Alignment Search Tool) searches against the NCBI database revealed that the ITS sequence had 99.83% homology (ON843984.1 and ON843987.1), tef-1α gene had 99.22% homology (MH015266.1 and MH015267.1), and the Tub2 sequence had 99.57% and 100% homologies (KT934364.1 and KT934364.1) with V. nivea sequences. The amplified sequences of ITS region (OK442665 and OK442666), tef-1α (OK510871 and OK510872) and Tub2 (OK510869 and OK510870) were deposited in the GenBank. A phylogenetic analysis was performed using MEGA7 that shows 100% bootstrap support that ALE6T-GP21 and ALE7T-GP23 were V. nivea. A pathogenicity trial was conducted with isolate ALE6T-GP21 inoculated onto 1-year-old shoots of 15-year-old Korla pear trees in Alar city, Xinjiang, China. Five shoots were inoculated by making 5-mm deep wounds using a sterile scalpel then inoculating with a 50 μL conidia suspension (1×106 mL−1). Additionally, five shoots served as the negative control and were inoculated in the same way using 50 μL ddH2O. The trees were kept under ambient conditions. Inoculated branches developed symptoms 18 days post inoculation, whereas the control branches showed no symptoms. V. nivea was re-isolated from the symptomatic areas and the isolate confirmed as ALE6T-GP21 by sequence analysis. Currently, the proven hosts of V. nivea are Populus, Elaeagnus, Juglans, Malus and Salix (Adams et al. 2006; Wang et al. 2020). To our knowledge, this is the first report of pathogenic V. nivea occurring on P. sinkiangensis in the world. It will provide a basis for research into the occurrence, distribution of V. nivea on Korla Pear.