关键词: 石碌含笑, 枯梢病, Diaporthe ueckerae, 生物学特性

Abstract: [Objective] In 2014, dieback disease was observed on the 2-3-year-old grafted twigs of Michelia shiluensis in Xuwen County, Guangdong Province. To understand the pathogen causing the dieback, the isolate was collected and identified, the pathogenicity was tested according to the Koch's postulates, and the biological characteristics were determined. This study aims to provide basic theoretical knowledge for diagnosis and control of this disease.[Method] To get the pathogen, the infested tissues were cut from symptomatic twigs of which the barks were removed, placed on potato dextrose agar (PDA), and incubated at 25-30℃ in dark. The isolates were inoculated on the healthy hosts to test the pathogenicity according to the Koch's postulates. The sequences of internal transcribed spacer (ITS) rDNA, translation elongation factor (TEF) gene, β-tubulin gene and histone (HIS) gene were amplified, sequenced and blasted in GenBank. Based on the multi-locus (ITS, HIS, TEF, TUB) phylogenetic analyses with Neighbor joining (NJ) and Bayesian inference (BI) method, the phylogenetic relationship of the pathogen with Diaporthe spp. was analyzed. The pathogen was identified according to the morphological and molecular characteristics. The effects of culture medium, temperature, pH, carbon and nitrogen source as well as light treatment on mycelial growth and conidiomata formation were determined.[Result] The isolate from infected tissues caused twigs dieback on healthy hosts, and the morphological characters was the same as Diaporthe sp. The Blast result showed that the ITS sequences of the pathogen were above 99% homology with several species of Diaporthe, and the sequences of HIS, TEF and TUB gene were 100% homology with D. ueckerae. The pathogen was clustered in the same branch with D. ueckerae in a well-supported clade with Bootstrap support values above 99% and Bayesian posterior probability values 1.00. Under the condition of culture medium, temperature, pH, carbon and nitrogen source and light treatment, the mycelial growth and conidiomata formation had significant difference. The pH values ranged from 4 to 8 had insignificant effects on mycelial growth and conidiomata formation. Different light treatments had insignificant effect on the mycelial growth, but significant effect on conidiomata formation, thus it was not conducive to form the conidiomata under two light treatments, i.e. alternation of 12 hillumination and 12 h darkness, and alternation of 12 hillumination, 12 h darkness and 10 minultraviolet radiation.[Conclusion] The pathogen causing dieback on M. shiluensis has been identified as D. ueckerae Udayanga et Castlebury 2015, with strong adaptation to various environmental factors. The fungus could not form conidiomata below 15℃ or above 35℃.

Key words: Michelia shiluensis, dieback, Diaporthe ueckerae, biological characteristics