Leaf spot disease on seedlings of Quercus acutissima caused by Tubakia dryina in Korea
Australasian Plant Disease Notes
Leaf spot disease on seedlings of Quercus acutissima caused by Tubakia dryina in Korea
Dong-Hyeon Lee 0
Sang-Tae Seo 0
Seung Kyu Lee 0
Sun Keun Lee 0
0 Division of Forest Diseases & Insect Pests, National Institute of Forest Science , Seoul 02455 , South Korea
1 Sun Keun Lee
Severe attack by a leaf spot disease was found on seedlings of Quercus acutissima in Jeungpyeong, Korea in 2017. Based on the morphological characteristics of the fungus, it was identified as a member of Tubakia. Sequence comparisons based on the ITS and 28S rDNA regions confirmed the identity of the fungus as Tubakia dryina. This is the first report of Tubakia leaf spot caused by T. dryina on Q. acutissima.
Fagaceae; Melanconidaceae; Oak
-
Tubakia species have been reported as endophytes or causing
leaf spots on Castanea spp., Quercus spp., and some other
members of Fagaceae in Asia, Europe and North America
(Yun and Rossman 2011; Harrington et al. 2012; Boroń and
Grad 2017)
. Tubakia dryina is a known leaf spotting fungus
which has been reported in many countries. Although a wide
range of host species have been recorded for T. dryina, the
fungus is most frequently found on oaks
(Proffer 1990;
Kowalski 2006)
, including at least 40 species according to
Farr and Rossman (2017).
During a recent survey of plant diseases carried out to
identify the fungal pathogens infecting oaks in 2017, a damaging
leaf spot disease was consistently observed on seedlings of
Quercus acutissima (sawtooth oak) (Fagaceae), in
Jeungpyeong, Korea. Quercus acutissima is a deciduous
broad-leaved tree, native to eastern Asia with a wide
distribution and is one of the important forest components in hillsides
of South Korea.
Leaf spots were brown necrotic, circular to irregular,
49–74 mm diam, occasionally coalescing to form large
blotches (Fig. 1a–b). Fresh specimen were mounted in
water and examined under the following microscopes:
Olympus BX51 and Zeiss AX10 (equipped with a Carl
Zeiss AxioCam MRc5 camera). At least 30 measurements
were made for each structure examined. Fungal
morphology was as follows: Conidiomata (pycnothyria) scutelloid,
convex, 40–75 μm in diam, membranaceous, consisting
of brown to blackish, thick-walled cells radiating from a
central point, blackish; conidiogenous cells, 5 to 7 × 9 to
13 μm, cylindrical to clavate, narrowing toward the neck,
brown to dark brown, radiating from the center below the
developing scutellum; conidia solitary, acrogenous,
ellipsoidal, 5–7 × 15–18 μm, with a basal frill, pale to light
brown, walls smooth; microconidia absent (Fig. 1d–f).
Pure cultures were obtained by single conidia transfer
onto 2% malt extract-agar (MEA; 20 g Bacto malt extract
and 20 g Bacto Agar) plates (Fig. 1c). All isolates
obtained in this study were deposited in the culture collection
(CFPR) of the Forest Pathology Research Laboratory,
National Institute of Forest Science, Seoul, South Korea
(Accession Nos. CFPR-QA1TD and CFPR-QA2TD) as
well as in the Korean Agricultural Culture Collection
(KACC) of the National Academy of Agricultural
Science in South Korea (Accession Nos. KACC48481
for CFPR-QA1TD and KACC48482 for CFPR-QA2TD).
In addition to the morphological identifications, DNA
barcode sequences were compared with sequences
deposited in GenBank to assure the correct identity of the
fungus. DNA sequence analyses were carried out following
the techniques described by
Lee et al. (2015)
for internal
transcribed spacer (ITS) regions including the 5.8S rDNA
gene region and large-subunit (LSU) 28S rDNA. A
BLASTn search with the isolates obtained in this study
further revealed >99% similarity with those of T. dryina
isolates from Poland (KR362909 for ITS) and from Italy
(JF704187 for LSU). All sequence data obtained in this
study were deposited in the NCBI database (accession
n u m b e r s ; M F 5 7 9 7 5 5 a n d M F 5 7 9 7 5 6 f o r I T S ;
MG798658 and MG793660 for LSU).
To construct a phylogenetic tree, sequences obtained
were aligned with ten published sequences of Tubakia
species retrieved from GenBank using the online version
of MAFFT ver.7.215 (http://mafft.cbrc.jp/alignment/
server/)
(Katoh et al. 2002)
. A Maximum likelihood
(ML) analysis was performed using RAxML HPC
BlackBox ver.8.1.11
(Stamatakis 2006; Stamatakis et al.
2008)
, by using the default option with the GTR
substitution model implemented in the CIPRES cluster server
( h t t p s : / / w w w . p h y l o . o r g / ) a t t h e S a n D i e g o
Supercomputing Center. The ML analysis based on the
ITS region resulted in a marginally supported placement
of isolates obtained in this study (CFPR-QA1TD,
CFPRQA2TD) with authenticated isolates of T. dryina retrieved
from GenBank (Fig. 2).
Pathogenicity of CFPR-QA1TD and CFPR-QA2TD to
Q. acutissima, was tested through inoculations of healthy
detached leaves of Q. acutissima. Three weeks-old culture
formed on 2% MEA plates at 25 °C were flooded with
sterile distilled water and the su (...truncated)