Botryosphaeriaceae associated with the die-back of ornamental trees in the Western Balkans

Abstract

Extensive die-back and mortality of various ornamental trees and shrubs has been observed in parts of the Western Balkans region during the past decade. The disease symptoms have been typical of those caused by pathogens residing in the Botryosphaeriaceae. The aims of this study were to isolate and characterize Botryosphaeriaceae species associated with diseased ornamental trees in Serbia, Montenegro, Bosnia and Herzegovina. Isolates were initially characterized based on the DNA sequence data for the internal transcribed spacer rDNA and six major clades were identified. Representative isolates from each clade were further characterized using DNA sequence data for the translation elongation factor 1-alpha, b-tubulin-2 and large subunit rRNA gene regions, as well as the morphology of the asexual morphs. Ten species of the Botryosphaeriaceae were identified of which eight, i.e., Dothiorella sarmentorum, Neofusicoccum parvum, Botryosphaeria dothidea, Phaeobotryon cupressi, Sphaeropsis visci, Diplodia seriata, D. sapinea and D. mutila were known taxa. The remaining two species could be identified only as Dothiorella spp. Dichomera syn-asexual morphs of D. sapinea, Dothiorella sp. 2 and B. dothidea, as well as unique morphological characters for a number of the known species are described. Based on host plants and geographic distribution, the majority of Botryosphaeriaceae species found represent new records. The results of this study contribute to our knowledge of the distribution, host associations and impacts of these fungi on trees in urban environments.

Shot hole disease on Prunus laurocerasus caused by Neofusicoccum parvum in Serbia

“…Ten symptomatic leaves were collected from each of the 30 diseased plants; pycnidia were removed with a sterile needle, examined using light microscopy and plated on malt extract agar (MEA) as described in Zlatković, Keča, Wingfield, Jami, and Slippers (2016). One week later, wooly, fast growing, grey fungal colonies had developed and these were purified using hyphal tip transfers.…”

“…The internal transcribed spacer (ITS) region of rDNA operon was amplified using primers ITS-1 and ITS-4 (White et al, 1990); part of the translation elongation factor 1-alpha (TEF-1-α) gene was amplified using primers EF1-728F and EF1-986R (Carbone & Kohn, 1999); the β-tubulin-2 (BT2) gene was amplified using primers Bt2a and Bt2b (Glass & Donaldson, 1995) and | 667 part of the second largest subunit of RNA polymerase II (RPB2) gene was amplified using primers RPB2bot6F and RPB2bot7R (Sakaladis, 2004). The conditions and procedures for PCR amplification and PCR sequencing were the same as those described in Zlatković et al (2016).…”

“…Sequence alignments were conducted following the method described by Zlatković et al (2016). The phylogenetic analyses were performed using maximum parsimony (MP) and maximum-likelihood (ML) analyses.…”

“…The discovery of N. parvum on P. laurocerasus in Serbia is not surprising, as the pathogen is a plurivorous species found on ornamental trees worldwide (Phillips et al, 2013). N. parvum has been recently isolated from branches of Chamaecyparis lawsoniana and the stem of Aesculus hippocastanum in Serbia and from branches of P. laurocerasus in the nearby Montenegro (Zlatković et al, 2016).…”

Dothiorella omnivora isolated from grapevine with trunk disease symptoms in Hungary

“…Sequences were manually assembled from chromatograms using the Staden Package (Staden et al 2000). The EF1-α sequences obtained in this way were deposited in GenBank under accession numbers KY681037 and KY681038, and included the fragment spanned by primers EF446f/ EF1035r, reported by Linaldeddu et al (2016) and Zlatkovic et al (2016) for some D. omnivora strains.…”