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Short communication
Notes on powdery mildews (Erysiphales) in Japan:II. Erysiphe sect. Microsphaera
Jamjan Meeboon a,b, Susumu Takamatsu a,*
a Department of Bioresources, Graduate School, Mie University, 1577 Kurima-Machiya, Tsu 514-8507, Japanb Department of Agriculture, Ministry of Agriculture and Cooperatives, 50 Phaholyothin Rd., Ladyao, Chatuchak,
Bangkok, Thailand
a r t i c l e i n f o
Article history:
Received 12 May 2014
Received in revised form
26 June 2014
Accepted 26 June 2014
Available online 14 August 2014
Keywords:
Erysiphaceae
Molecular phylogeny
Morphology
New host
Pseudoidium
* Corresponding author. Tel.: þ81 59 231 949E-mail address: [email protected]
http://dx.doi.org/10.1016/j.myc.2014.06.0071340-3540/© 2014 The Mycological Society of
a b s t r a c t
Several novelties of powdery mildews in Japan are reported, including the first Japanese
collection of the teleomorph of Erysiphe platani on Platanus � acerifolia, the first description
of ascomata of E. viburni on Viburnum sieboldii var. obovatifolium, and the first morphological
descriptions and molecular data for E. wallrothii on Leucotho€e grayana and Pseudoidium
hortensiae on Hydrangea serrata var. megacarpa based on Japanese specimens.
© 2014 The Mycological Society of Japan. Published by Elsevier B.V. All rights reserved.
This is the second part of a series dealing with new records
and data on powdery mildews in Japan. In this report, four
powdery mildew species belonging to Erysiphe sect. Micro-
sphaera and its Pseudoidium anamorphs are described and
illustrated, and supplementary molecular data were obtained.
The nucleotide sequences of the 50-end of the 28S rRNA
gene (including domains D1 and D2) and the internal tran-
scribed spacer (ITS) regions were determined according to the
procedure of Meeboon and Takamatsu (2014). Representative
new sequences determined in this study were deposited in
DNA Data Base of Japan (DDBJ) under the accession numbers
AB926018eAB926022. Sequences generated from the 28S rRNA
gene and ITS region were aligned with other sequences of the
7; fax: þ81 59 231 9540.p (S. Takamatsu).
Japan. Published by Else
Erysiphaceae that were retrieved from DNA databases (DDBJ,
EMBL, NCBI) using MUSCLE (Edgar 2004) implemented in
MEGA 5 (Tamura et al. 2011). Alignments were then manually
refined using the MEGA 5 program and were deposited in
TreeBASE (http://www.treebase.org/) under the accession
number S15710. Phylogenetic trees were obtained from the
data using maximum parsimony (MP) method and maximum
likelihood (ML) method as described in Meeboon and
Takamatsu (2014).
The 28S rRNA gene for three specimens of Erysiphe and
Pseudoidium were determined in this study. The three se-
quences were aligned with 28 Erysiphe sequences retrieved
from the DNA databases. Erysiphe glycines F.L. Tai (AB022397)
vier B.V. All rights reserved.
Erysiphe trifoliorum ex Trifolium AB103078
Erysiphe multappendicis ex Berberis AB103076
Erysiphe cruciferarum ex Cardaria AB102944
Erysiphe lycopsidis ex Anchusa AB103072
Erysiphe heraclei ex Chaerophyllum AB103067
Erysiphe heraclei ex Daucus AB022391
Erysiphe bremeri ex Alhagi AB103077
Erysiphe friesii ex Rhamnus AB022382
Erysiphe wallrothii ex Vaccinium AB015930
Erysiphe wallrothii ex Leucothoe AB926020
Erysiphe alphitoides ex Quercus AB257431
Erysiphe monascogera ex Styrax AB331645
Erysiphe nomurae ex Symplocos AB331648
Erysiphe quercicola ex Quercus AB292694
Erysiphe hypophylla ex Quercus AB292716
Erysiphe syringae ex Syringa AB015920
Erysiphe platani ex Platanus AB926022
Erysiphe abbreviata ex Quercus AB271785
Erysiphe epigena ex Quercus AB292722
Erysiphe hypogena ex Quercus AB292727
Erysiphe corylopsidis ex Corylopsis AB478988
Erysiphe pulchra var. pulchra ex Cornus AB015935
Erysiphe paeoniae ex Paeonia AB257438
Erysiphe ligustri ex Ligustrum AB015917
Erysiphe aquilegiae ex Cimicifuga AB022405
Pseudoidium hortensiae ex Hydrangea AB926018
Pseudoidium sp. ex Medicago AB102942
Erysiphe staphyleae ex Staphylea AB015922
Erysiphe vanbruntiana ex Sambucus AB015925
Erysiphe pulchra var. japonica ex Cornus AB022389
Erysiphe glycines ex Desmodium AB022397
5 changes
70/75
75/84-/95
-/76
76/87
92/89
87/80
92/99
28S rRNA gene31 sequences773 characters191 stepsCI = 0.5550RI = 0.6458RC = 0.3584
Fig. 1 e Phylogenetic analysis of the divergent domains D1 and D2 sequences of the 28S rRNA gene for 31 sequences from
the genus Erysiphe. This tree is a phylogram of one of the 1107 most parsimonious trees with 191 steps, which was found
using a heuristic search. Horizontal branch lengths are proportional to the number of substitutions that were inferred to
have occurred along a particular branch of the tree. BS (>70%) values by the maximum parsimony (MP) and maximum
likelihood (ML) methods were shown on the respective branch. Sequences determined in this study are shown in boldface.
The GenBank number for each sequence is presented after species name on the tree.
myc o s c i e n c e 5 6 ( 2 0 1 5 ) 2 3 0e2 3 6 231
was used as outgroup according to Mori et al. (2000). The
alignment data matrix consisted of 31 sequences and 773
characters, of which 91 (11.8%) characters were variable and
55 (7.1%) characters were informative for parsimony
analysis. A total of 1107 equally parsimonious trees with
191 steps were constructed using MP analysis. One of the
1107 trees with the highest log likelihood value is shown in
Fig. 1.
Erysiphe platani ex Platanus AB926022
Erysiphe platani ex Platanus KF499270
Erysiphe platani ex Platanus JX997805
Erysiphe syringae ex Syringa AB015920
Erysiphe syringae ex Ligustrum AB295461
Erysiphe magnifica ex Magnolia AF011312
Erysiphe magnifica ex Magnolia GU195046
Erysiphe liriodendri ex Liliodendron AF011302
Erysiphe abbreviata ex Quercus AB271785
Erysiphe castaneigena ex Castanea AF298545
Erysiphe quercicola ex Quercus AB292691
Erysiphe quercicola ex Quercus AB292694
Erysiphe alphitoides ex Quercus AB237783
Erysiphe pseudolonicerae ex Cocculus AB015915
Erysiphe wallrothii ex Vaccinium AB015930
Erysiphe wallrothii ex Leucothoe AB926021
Erysiphe epigena ex Quercus AB292720
Erysiphe hypogena ex Quercus AB292724
Erysiphe monascogera ex Stylax AB331646
Erysiphe hypophylla ex Quercus AB292715
Pseudoidium neolycopersici ex Lycopersicon AB032484
Pseudoidium neolycopersici ex Lycopersicon AB034722
Pseudoidium hortensiae ex Hydrangea AB926019
Erysiphe sedi ex Sedum JX173290
Pseudoidium hortensiae ex Hydrangea JQ669944
Erysiphe aquilegiae ex Cimicifuga AB000944
Erysiphe aquilegiae ex Clematis AB015929
Erysiphe macleayae ex Macleaya AB016048
Erysiphe aquilegiae ex Clematis AF154322
Erysiphe circaeae ex Circaea AB104517
Erysiphe paeoniae ex Paeonia AB257437
Erysiphe paeoniae ex Paeonia AB257436
Erysiphe glycines ex Desmodium AB015927
5 changes
100/100
93/90
100/100
100/100
100/99
100/100
-/78
ITS33 sequences579 characters217 stepsCI = 0.7604RI = 0.8632RC = 0.6563
Fig. 2 e Phylogenetic analysis of the ITS region for 33 sequences from the genus Erysiphe. This tree is a phylogram of one of
the 1414 most parsimonious trees with 217 steps, which was found using a heuristic search. Horizontal branch lengths are
proportional to the number of substitutions that were inferred to have occurred along a particular branch of the tree. BS
(>70%) values by the maximum parsimony (MP) and maximum likelihood (ML) methods were shown on the respective
branch. Sequences determined in this study are shown in boldface. The GenBank number for each sequence is presented
after species name on the tree.
my c o s c i e n c e 5 6 ( 2 0 1 5 ) 2 3 0e2 3 6232
For the ITS analysis, the three sequences determined in
this study were aligned with 30 Erysiphe sequences retrieved
from the DNAdatabases. Erysiphe glycines (AB015927)was used
as outgroup according to Takamatsu et al. (1999). The align-
ment data matrix consisted of 33 sequences and 579
characters, of which 139 (24.0%) characters were variable and
77 (13.3%) characters were informative for parsimony anal-
ysis. A total of 1414 equally parsimonious trees with 217 steps
were constructed using MP analysis. One of the 1414 trees
with the highest log likelihood value is shown in Fig. 2. The
Fig. 3 e Teleomorph of Erysiphe platani on Platanus £ acerifolia (MUMH5657). A: Chasmothecium. B: Asci. C, D: Appendages.
Bars: A 100 mm; B 60 mm; C, D 50 mm.
myc o s c i e n c e 5 6 ( 2 0 1 5 ) 2 3 0e2 3 6 233
phylogenetic trees generated using ML analysis were almost
identical to the MP trees in both 28S rRNA gene and ITS ana-
lyses, and thus only the bootstrap values were shown on the
MP trees.
Erysiphe platani (Howe) U. Braun & S. Takam., Schlechtendalia
4: 12, 2000. Fig. 3.
Description: Mycelium on leaves, amphigenous, dense, white,
forming patches, confluent, evanescent to persistent. Chas-
mothecia scattered to gregarious, (90e)97e119(e126) mm
diam. Peridium composed of irregularly polygonal cells,
10e20 mm diam. Appendages 9e16 per chasmothecium,
equatorial, straight to curved, 1e2 times as long as the chas-
mothecial diam., 7e10 mm wide, 0e1-septate, hyaline, brown
at the base, walls smooth to rough, thin in the upper half,
thick towards the base. Apices 4e6 times densely and regu-
larly dichotomously branched, rarely trichotomously, occa-
sionally forked in the lower half, tips recurved. Asci 3e6 per
chasmothecium, broadly ellipsoid-obovoid, saccate, (47e)
52e60(e64) � (38e)41e43(e46) mm, short-stalked, 4e6-spored.
Ascospores ellipsoid-ovoid, hyaline, (16e)18e21(e23) � (9.5e)
11e12(e14) mm.
Collection examined: on Platanus � acerifolia (Aiton) Willd.
(Platanaceae), Japan, Mie Pref., Mie University, 1 Dec 2012, S.
Takamatsu, MUMH5657, GenBank accession number:
AB926022 (ITS and 28S rRNA gene).
Remarks: Erysiphe plataniwas first described byHowe (1874)
as Microsphaera platani based on a specimen found on Pl. occi-
dentalis L. from U.S.A. It is likely that the fungus was then
introduced to South America, South Africa, Europe, and Asia
(Braun and Cook 2012). In Japan, powdery mildew on Platanus
spp. was first found in Tokyo in 1997 (Tanda 1999; Horie et al.
2002), but was found only in anamorphic stage. The ITS
sequence obtained from E. platani on Pl. � acerifolia was iden-
tical to that of E. platani (JX997805) collected in Korea and
differed only one base from KF499270 collected in Brazil.
These three sequences formed a distinct clade with high
bootstrap supports (100% in both MP and ML analyses; Fig. 2).
Erysiphe viburni Duby, Bot. Gall. 2: 872, 1830. Fig. 4.
Description: Mycelium amphigenous, evanescent. Chas-
mothecia (79e)84e101(e108) mm diam., scattered, globose to
subglobose, dark brown. Peridium composed of dark brown,
irregularly polygonal cells, 15e25 mm diam. Appendages 4e9
per chasmothecium, 1e1.5 times as long as the chasmothecial
diam., ± equatorial, flaccid or loosely curved, 0(e1)-septate,
completely hyaline or pigmented light brown at the base, thin-
walled, smooth. Apices 3e5 times irregularly to dichoto-
mously branched, often loosely branched, primary branches
somewhat elongated, tips straight. Asci 2e4 per chasmothe-
cium, (32e)38e63(e70) � (35e)47e54(e60) mm, 3e5-spored,
broadly obovoid to subglobose, short-stalked or subsessile.
Ascospores (18e)19.5e24(e25.5) � 10e15(e16.5) mm, ellipsoid-
ovoid, hyaline. Anamorph not observed.
Collection examined: on Viburnum sieboldii Miq. var. obo-
vatifolium (Yanagita) Sugimoto (Adoxaceae), Japan, Niigata
Pref., Mt. Yahiko, 18 Oct 1996, S. Takamatsu, MUMH263.
Remarks: Erysiphe viburni is a powdery mildew species
commonly found on Viburnum spp. (Adoxaceae) in the
Northern Hemisphere. Occurrence of powdery mildew on V.
sieboldii var. obovatifolium was reported by Hirata and Wada
(1973) and Sato (1980) as Microsphaera sp. but without a
morphological description. Thus, the morphology of chas-
mothecia of this species occurring on this host was examined
in detail. In comparison to the description of Braun and Cook
(2012), the specimen occurring on V. sieboldii var. obovatifolium
has smaller chasmothecia and ascospores, and less number of
asci. DNA sequencing of this collection failed.
Erysiphe wallrothii (U. Braun & Tanda) U. Braun & S. Takam.,
Schlechtendalia 4: 15, 2000. Fig. 5.
Description: Mycelium amphigenous, in patches or effuse,
white, evanescent to persistent. Hyphae 3e6 mm wide. Chas-
mothecia (114e)118e120(e141) mm diam., globose, dark
brown, scattered. Peridium 9e16 mm thick, composed of dark
brown, irregularly polygonal cells, 17e30 mm diam. Append-
ages 5e8 per chasmothecium, (56e)97e148(e226) � (5.5e)
Fig. 4 e Teleomorph of Erysiphe viburni on Viburnum sieboldii var. obovatifolium (MUMH263). A, B: Chasmothecium. CeF:
Ascus. GeJ: Appendages. Bars: A, B 100 mm; CeF 25 mm; GeJ 30 mm.
my c o s c i e n c e 5 6 ( 2 0 1 5 ) 2 3 0e2 3 6234
7e10 mm, 1e2 times as long as the chasmothecial diam.,
equatorial, stiff to flexuous, aseptate or with a single septum
at the base, hyaline, walls thick at the base, thin towards the
apex, smooth to somewhat rough. Apices 4e6 times regularly
and tightly dichotomously branched, rarely forked in the
lower half, tips distinctly recurved. Asci 4e6 per chasmothe-
cium, (35e)46.5e63(e67) � (37e)40e47(e50.5) mm, 5e7-spored,
broadly ellipsoid-obovoid to saccate, sessile or short-stalked.
Ascospores 17e24(e26) � 8e13 mm, ellipsoid to ovoid, hyaline.
Collections examined: on Leucotho€e grayana Maxim. (Erica-
ceae), Japan, Mie Pref., Mt. Miike, Happu Pass, 28 Oct 2007, S.
Takamatsu, MUMH4752, GenBank accession number:
AB926021 (ITS), AB926020 (28S rRNA gene); Mie Pref., Mt.
Kama-ga-take, 27 Oct 2002, S. Takamatsu, MUMH2152.
Remarks: Powderymildew occurring on L. grayanawas first
reported by Takamatsu (1982) as Microsphaera sp. without a
morphological description. Thus, morphological characteris-
tics of E. wallrothii collected from this host were examined in
detail, supplemented with rRNA gene sequence data. Molec-
ular analyses showed that the powdery mildew found on L.
grayana is 100% identical to E. wallrothii occurring Vaccinium
hirtum (AB015930) in both 28S rRNA gene and ITS sequences
(Figs. 1, 2).
Pseudoidium hortensiae (Jørst.) U. Braun & R.T.A. Cook, Taxo-
nomic Manual of the Erysiphales (Powdery Mildews): 606,
2011. Fig. 6.
Description:Myceliumamphigenous,white, indense or thin
patches, irregular, evanescentorpersistent.Hyphalappressoria
lobed, solitary or in opposite pairs. Conidiophores arising cen-
trally from the upper surface of mother cells, erect, (48.5e)
56e75.5(e91) mm long. Foot cells cylindrical, straight, (21e)
Fig. 5 e Teleomorph of Erysiphe wallrothii on Leucotho€e grayana (MUMH4752). A: Chasmothecium. BeE: Ascus. FeJ:
Appendages. Bars: A 120 mm; BeE 20 mm; FeJ 30 mm.
Fig. 6 e Pseudoidium hortensiae on Hydrangea serrata var. megacarpa (MUMH281). A: Conidia. B: Conidiophores. C:
Appressoria. Bar: 20 mm.
myc o s c i e n c e 5 6 ( 2 0 1 5 ) 2 3 0e2 3 6 235
my c o s c i e n c e 5 6 ( 2 0 1 5 ) 2 3 0e2 3 6236
23e25(e33.5) � 7e10 mm, followed by 0e3 shorter cells, with a
basal septum at the branching point of the mycelium, 1e3 mm,
forming conidia singly. Conidia ellipsoid-ovoid, (25.5e)
29.5e36.5(e39.5) � 12e16(e18) mm. Germ tube not observed.
Collections examined: on Hydrangea serrata (Thunb. ex
Murray) Ser. var. megacarpa (Ohwi) H. Ohba (Hydrangeaceae),
Japan, Niigata Pref., Mt. Yahiko, 8 Oct 1996, S. Takamatsu,
MUMH221; Niigata Pref., Mt. Yahiko, 18 Oct 1996, S. Taka-
matsu, MUMH281, GenBank accession number: AB926019
(ITS), AB926018 (28S rRNA gene).
Remarks: Hirata and Wada (1973) reported occurrence of
Microsphaera sp. on H. serrata var. megacarpa without a
morphological description. Molecular analyses showed that
the ITS sequence of the powdery mildew occurring on H. ser-
rata var.megacarpa is identical to that ofOidium hortensiae (¼Ps.
hortensiae) collected from H. macrophylla (JQ669944) (Fig. 2).
Acknowledgments
This work was financially supported in part by a Grant-in-Aid
for Scientific Research (No. 23580061) from the Japan Society
for the Promotion of Science, a grant from the Institute for
Fermentation, Osaka, Japan to ST, and MONBUKAGAKU SHO:
MEXT (Ministry of Education, Culture, Science, and Technol-
ogy) Scholarship of the Japanese Government awarded to JM.
r e f e r e n c e s
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Howe EC, 1874. New fungi. Bulletin of the Torrey Botanical Club 5:3e4.
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Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S,2011. MEGA5: molecular evolutionary genetics analysis usingmaximum likelihood, evolutionary distance, andmaximum parsimony methods. Molecular Biology andEvolution 28: 2731e2739. http://dx.doi.org/10.1093/molbev/msr121.
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