ITS and LSU-rDNA nucleotide sequences based confirmation of Cytospora chrysosperma and Chondrostereum purpureum from symptomatic cankered tissues of Populus sp. trees in Turkey

Abstract

The fungi Cytospora chrysosperma and Chondrostereum purpureum were isolated from orange-brown inner bark with pycnidia in the bark surface and underlying wood tissues of infected poplar plants (Populus sp.) with symptoms of stem and branch canker in Doğanşehir, Malatya, in 2016, respectively. Twigs of poplar trees were inoculated during their first season of growth by removing the fourth fully expanded leaves and placing agar plugs colonized by representative isolates of C. chrysosperma and C. purpureum over the resulting wounds. Three months after inoculation, cankers in 6.4 and 3.3 cm length formed by C. chrysosperma and C. purpureum, respectively, and twigs were girdled. Pathogenicity tests in a greenhouse experiment by shallow wounds made into the bark tissue and inoculation with these isolates in a similar manner also resulted in canker formation in and around inoculated wounds 14 days after inoculation. Subsequent re-isolations of C. chrysosperma and C. purpureum confirmed that these fungi were the causal agents of the disease, and no cankers formed in wounds that received only sterile plugs. DNA was extracted from representative isolates of each fungal species. Extracted DNA templates were amplified and sequenced for rDNA internal transcribed spacer (ITS) and the large subunit (LSU) rDNA gene regions using ITS6/ITS4 and NL1/NL4 primer pairs, respectively. NCBI BLAST results showed 99% similarity with the ITS and LSU sequences of C. chrysosperma and C. purpureum in GenBank. The sequences were submitted to GenBank. Given accession numbers of C. chrysosperma and C. purpureum were MF536529 and MF536531 for ITSrDNA; MF536530 and MF536532 for LSU-rDNA, respectively. Existence of these fungi in Turkey was previously reported. However, this is a first report of molecular characterization of C. chrysosperma and C. purpureum based on ITS and LSU-rDNA nucleotide sequences of these fungi in Turkey.