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Phylogenetic studies on Vialaeaceae reveals a novel species, Vialaea

mangiferae

Article  in  Sydowia -Horn- · December 2014

DOI: 10.12905/0380.sydowia66(2)2014-0203

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 DOI ????????????????

Phylogenetic studies on Vialaeaceae reveals a novel species,

Vialaea mangiferae

Indunil C. Senanayake 1,2,3,4, Sajeewa S.N. Maharachchikumbura 2,4,

Peter E. Mortimer 1,3, Jianchu Xu 1,3, Jayarama D. Bhat 1,2,5,6,
Kevin D. Hyde 1,2,3,4,*.

1
  Key Laboratory for Plant Biodiversity and Biogeography of East Asia (KLPB), Kunming
Institute of Botany, Chinese Academy of Science, Kunming 650201, Yunnan, PR China
2
  Institute of Excellence in Fungal Research, Mae Fah Luang University,
Chiang Rai 57100, Thailand
3
  World Agroforestry Centre, East and Central Asia, 132 Lanhei Road, Kunming 650201,
PR China
4
  School of Science, Mae Fah Luang University, Chiang Rai. 57100, Thailand
5
  No. 128/1-J, Azad Housing Society, Curca, P.O. Goa Velha-403108, India
6
  Formerly at Department of Botany, Goa University, Goa 403 206, India

Senanayake I. C., Maharachchikumbura S. S. N., Mortimer P. E., Xu J., Bhat J. D. &

Hyde K. D. (2014) Vialaeaceae; introducing a novel species Vialaea mangiferae – Sydowia
66 (1): pp–pp.
Vialaea comprises endophytic as well as phytopathogenic species, the later forming
cankers on two economically important and ornamental host plant species. A strain of Vi-
alaea was isolated from dead Mangifera indica twigs in Chiang Rai Province, in northern
Thailand. Phylogenetic analysis of ITS and LSU sequence data shows that this is a novel
species, which groups with Vialaea species forming a distinct clade, and supports Vialae-
aceae as a distinct family in Xylariales. The new species, introduced as Vialaea mangiferae
is described, illustrated and compared with other species in the genus. Vialaea mangiferae
differs from other Vialaea species in having 3–5 clustered perithecia in pseudo-stromatic
tissues and a peridium with globose cells. The asexual state of Vialaea, which formed in
culture, is reported for the first time.
Keywords: isthmoid ascospore, ITS, LSU, pathogen, Xylariales.

Vialaeaceae is an ascomycete family in the order Xylariales of the class

Sordariomycetes. It was invalidly introduced by Schrantz (1960) as “Vialaea-
cees” and formally established as Vialaeaceae by Cannon (1995) with Vialaea
insculpta (Fr.) Sacc. as the generic type. Members of this family are endo-
phytes or mild pathogens causing cankers on young twigs and older branch-
es of host trees. It is not yet clear whether the fungal members in Vialaeaceae
are plant pathogens or simply colonize previously infected or necrotic
branches (Redlin 1989). Members of Vialaeaceae form stromatic or non-stro-

* e-mail: kdhyde3@gmail.com
204 Senanayake et al.: Vialaea mangiferae, sp. nov.

matic papillate perithecia, scattered on the host surface (Cannon 1995). Asci
are cylindrical with a complex, J+ apical apparatus and ascospores are elon-
gate, strongly isthmoid, hyaline and one to three septate (Cannon 1995). An
asexual morph has not been reported for the genus (Cannon 1995).
Vialaea was introduced by Saccardo (1896) and placed in Amphispha-
eriaceae (Xylariales) based on the presence of an amyloid annulus in the
ascus apex (Chadefaud 1957, Shoemaker et al. 2013). However, Cannon
(1995) excluded Vialaea from Amphisphaeriaceae based on a difference in
the iodine reaction in the ascus apex. Vialaeaceae was assigned to Sordari-
ales based on the presence of peridial pores on the ascomatal wall (Cannon
1995). Peridial pores have for instance been reported in family Lasiospha-
eriaceae of Sordariales. However, asci in the Sordariales never have a blue
apical ring when stained with Melzer’s reagent (Cannon 1995). Chadefaud
(1960) noted that ascospores of Vialaea are similar to some members in Dia-
porthales, especially Pleuroceras (Shoemaker et al. 2013). Müller & Von Arx
(1962) placed Vialaea along with Pleuroceras in the family Diaporthaceae.
However later Pleuroceras was placed in the family Gnomoniaceae. Cannon
(1995) proposed a new family, Vialaeaceae, for Vialaea species, based on stro-
mata, perithecia and ascal characteristics, as well as nutritional strategy as
a weak pathogen, which he placed in the order Diaporthales. The above
placements were based on morphology. Shoemaker et al. (2013) analyzed the
LSU gene and showed that Vialaeaceae is a distinct family in the order Xy-
lariales. McTaggart et al. (2013) also used LSU data analysis of a Vialaea
minutella Petr. isolate from mango in Australia to show that the family Vial-
aeaceae belongs in Xylariales, rather than the Diaporthales. These observa-
tions also supported the status of Vialaeaceae as a distinct family.
A collection of Vialaea was made from mango (Mangifera indica) in
northern Thailand and proved to be new to science. The objectives of this
study are to 1) introduce a new species in the genus, 2) revisit the phylogeny
of the family Vialaeaceae based on molecular data and 3) illustrate the asex-
ual state of Vialaea.

Materials and methods

Specimens and cultures
Dead twigs of Mangifera indica (Anacardiaceae) were collected from a
garden at Baan Khuakhae, near Bandu, Muang District, Chiang Rai Prov-
ince, Thailand. The samples were brought to the laboratory in paper bags
and examined under a Motic SMZ 168 stereo-microscope to observe disease
symptoms and fungal ascomata. Sections of perithecia were taken by hand
under a stereo-microscope and mounted in water. Specimens were photo-
graphed using a Nikon Eclipse 80i compound microscope with a Canon 450D
digital camera. Ascomata on the host surface were photographed using a
Carl Zeiss Discovery V8 stereo-microscope with an AxioCam ERc 5s camera.
 Sydowia 66 (2014) 205

Canker symptoms on plant twigs were sterilized using 70 % ethanol

(Doilom et al. 2013). Contents of the perithecia (opened by slicing off the top
with a sterilized blade) were transferred to a few drops of sterilized distilled
water mounted on a glass slide. The contents was crushed using a sterilized
forceps to break down the ascus wall and release the ascospores (Chomnunti
et al. 2014). The spore suspension was inoculated on malt extract agar (MEA)
plates and incubated at 25 °C for spore germination. Germinated ascospores
were aseptically transferred to fresh MEA plates. Colony characteristics
were recorded from such plates. Illustration of plates and measurements fol-
low those of Liu et al. (2012).
The holotype is deposited at MFLU and the isotype at KUN. Living cul-
tures are deposited at the Mae Fah Luang Culture Collection (MFLUCC),
Mae Fah Luang University, Chiang Rai, Thailand and at the International
Collection of Microorganisms from Plants (ICMP), New Zealand (Wijaya-
wardene et al. 2013).

Establishing the asexual state

A few small, circular, agar blocks from the colony margin were cut and
placed on fresh water agar (Becton, Dickinson & Co., France) plates. Steri-
lized pine needles were laid on the plates as substrate for fungal growth
(Doilom et al. 2013). Conidiomata produced on both water agar and pine
needles were observed after six weeks of incubation at 25 °C. Morphological
characteristics were studied following the same procedure mentioned above
(Doilom et al. 2013).

DNA extraction and PCR amplification

Genomic DNA was extracted from fresh fungal mycelia (500 mg),
scraped from the colony margin on MEA plates incubated at 25 °C for 7 days
(Maharachchikumbura et al. 2013) using the Biospin Fungus Genomic DNA
Extraction Kit (BioFlux®) (BSC14S1) following the manufacturer’s proto-
col. The ITS4 and ITS5 primers were used to amplify the region of internal
transcribed spacers (ITS) (White et al. 1990). LR5 and LROR were used to
amplify a region spanning the large subunit rDNA (Vilgalys & Hester 1990).
PCR was performed with the 25 μl reaction system consisting of 9.5 μl of
double-distilled water, 12.5 μl of 2× Master Mix (0.1 U Taq DNA polymerase/
μl, 500 μM dNTP each, 20 mM Tris-HCl [pH 8.3], 100 nM KCl, 3 mM MgCl2),
1 μl of each primer (10 μM) and 1 μl of DNA template. The amplification was
performed with an initial denaturing step of 95 °C for 5 min, followed by 35
amplification cycles of 94 °C for 45 s and 58 °C for 45 s and 72 °C for 90 s and
a final extension step of 72 °C for 10 min (Phillips et al. 2008). The PCR prod-
ucts were verified by staining with ethidium bromide on 1 % agarose electro-
phoresis gel. Purification and sequencing of PCR products were carried out
at the SinoGenoMax Co., Beijing, China.
206 Senanayake et al.: Vialaea mangiferae, sp. nov.

Phylogenetic analysis
Sequence homologies for the assembled consensus sequences were ana-
lyzed using the BLAST search engine of the National Center for Biotechnol-
ogy Information (NCBI) for the rough identification of the new isolate used
in the analysis (Udayanga et al. 2012). Sequences of the available ex-type
cultures were obtained from GenBank (Tab 1). Sequences were optimized
manually to allow maximum alignment and maximum sequence similarity
as detailed in Maharachchikumbura et al. (2011, 2012). A maximum parsi-
mony analysis (MP) was performed using PAUP v. 4.0b10 (Swofford 2002).
Ambiguously aligned regions were excluded and gaps were treated as a fifth
character state. All characters were unordered and given equal weight. Trees
were inferred using the heuristic search option with TBR branch swapping
and 1000 random sequence additions. MaxTrees were set up to 1000, branch-
es of zero length were collapsed and all multiple parsimonious trees were
saved. Tree length (TL), consistency index (CI), retention index (RI), rescaled
consistency index (RC), homoplasy index (HI), and log likelihood (-ln L)
(HKY model) were calculated for trees generated under different optimality
criteria. The robustness of the most parsimonious trees was evaluated by
1000 bootstrap replications resulting from maximum parsimony analysis,
each with ten replicates of random step-wise addition of taxa (Felsenstein
1985). The Kishino–Hasegawa tests (Kishino & Hasegawa 1989) were per-
formed to determine whether the trees inferred under different optimality
criteria were significantly different. Trees were rooted to Hypocrea america-
na and visualized with TreeView (Page 1996). Sequences derived in this study
were deposited in GenBank.

Results
A phylogenetic tree was constructed using ITS and LSU sequences of
Vialaea, and related families with Hypocrea americana as outgroup (Tab. 1).
The alignment comprised 55 taxa and 1461 characters (including gaps). Par-
simony analysis indicates that 814 characters were constant; 89 variable
characters are parsimony-uninformative; and 558 characters are parsimony
informative. The parsimony analysis of the data matrix resulted in 185
equally parsimonious trees and the best tree (TL= 2988, CI= 0.389, RI= 0.683,
HI= 0.611, RC= 0.266) is shown in Fig. 1. Five strains of Vialaea clustered in
a well-supported clade (100 %) representing the family Vialaeaceae, and was

Fig. 1. Maximum parsimonious (MP) tree of Vialaea and related genera generated from the
combined ITS and LSU dataset. Parsimony analysis bootstrap support values (above 50)
are given at the nodes. Hypocrea americana was used as outgroup. New sequence generated
in this study is in red bold. Type strains are in black bold.
 Sydowia 66 (2014) 207

a sister group to Amphisphaeriaceae and Xylariaceae in Xylariales. The Vi-

alaea mangiferae isolate clustered with other species of Vialaea, but was
separated from other species of the genus with relatively high bootstrap sup-
port (94 %).
208 Senanayake et al.: Vialaea mangiferae, sp. nov.

Tab. 1. Sequences representing the closest taxa to Vialaea available in GenBank. Ex-type
strains are in bold.

Culture GenBank accession

Family Taxon accession number
number ITS LSU

Amphisphaeriaceae Bartalinia robillardoides BRIP 14180 AF405301 AF382366

Amphisphaeriaceae Discosia pseudoartocreas CPC 21117 KF777161 KF777214
Amphisphaeriaceae Discostroma fuscellum NBRC 32625 AB594794 AB593726
Amphisphaeriaceae D. fuscellum NBRC 32680 AB594806 AB593739
Amphisphaeriaceae D. stoneae NBRC 32690 AB594797 AB593729
Amphisphaeriaceae Pestalotiopsis disseminata CPC 10950 DQ195782 DQ195794
Amphisphaeriaceae P. theae DMC 698a EU833970 EU833969
Amphisphaeriaceae Seimatosporium obtusum CPC 12935 JN871206 JN871215
Amphisphaeriaceae S. walkeri CPC 17644 JN871207 JN871216
Amphisphaeriaceae Truncatella angustata TAPL1 KC241879 KC241880
Apiosporaceae Apiospora montagnei C27 FJ903318 –
Apiosporaceae A. montagnei H3 83 JN688916 –
Apiosporaceae Arthrinium jatrophae MMI00052 JQ246355 –
Boliniaceae Apiorhynchostoma curreyi UAMH 11088 JX460984 JX460989
Boliniaceae Endoxyla mallochii UAMH 11087 JX460986 JX460991
Calosphaeriaceae Calosphaeria pulchella CCTU 316 JX876610 JX876611
Calosphaeriaceae Jattaea mookgoponga STE U 6401 EU367450 EU367459
Calosphaeriaceae J. mookgoponga STE U 6184 EU367449 EU367458
Clypeosphaeriaceae Clypeosphaeria uniseptata 6349 (HKUCC) – DQ810219
Clypeosphaeriaceae C. uniseptata – – AY083830
Cryphonectriaceae Cryphonectria parasitica ATCC 38755 AY141856 EU199123
Cryphonectriaceae C. radicalis CMW 10477 AF368328 AY194102
Diaporthaceae Diaporthe phaseolorum KP 2012a JQ697843 JQ697869
PL42
Diaporthaceae D. pseudolongicolla PS69 JQ697847 JQ697873
Diatrypaceae Cryptosphaeria ATCC MYA FJ172278 FJ430586
pullmanensis 4414
Diatrypaceae Diatrype stigma ATCC MYA FJ430596 FJ430588
4418
Diatrypaceae Diatrypella verrucaeformis ATCC MYA FJ172276 FJ430584
4409
Diatrypaceae Eutypella sp. ATCC MYA FJ172282 FJ430589
4421
Gnomoniaceae Gnomonia fragariae CBS 146.64 EF212844 EF212856
Graphostromataceae Graphostroma sp. SGLMf27 EU715682 –
Graphostromataceae G. platystoma CBS 270.87 JX658535 DQ836906
Hypocreaceae Hypocrea americana AFTOL ID 52 – AY544649
 Sydowia 66 (2014) 209

Culture GenBank accession

Family Taxon accession number
number ITS LSU
Hyponectriaceae Monographella nivalis ATCC MYA FJ614641 –
3968
Hyponectriaceae M. lycopodina LL JF440979 –
Melanconidaceae Melanconis stilbostoma E00153 AY577811 AY577810
Microascaceae Kernia pachypleura 6807 S5 KC009507 KC009271
Microascaceae Microascus cirrosus ATCC MYA JQ906771 JQ906766
4885
Schizoparmeaceae Schizoparme acericola MFLUCC 13 KC951998 KC951999
0299
Schizoparmeaceae S. straminea STE U 3932 AY339348 AY339296
Sordariaceae Apiosordaria BW 2013 11 KC702789 KC702790
pentagonospora. 1352
Sordariaceae Sordaria fimicola CBS 508.50 AY681188 AY681160
Sordariaceae S. macrospora ATCC MYA KC171332 KC171305
4828
Togniniaceae Togninia MR 3064 HQ878591 HQ878595
fraxinopennsylvanica
Valsaceae Valsa ambiens ATCC 52280 AY347345 AF277146
Valsaceae Waydora typica PDD 103894 KF727412 KF727413
Vialaeaceae Vialaea insculpta DAOM 240860 JX123570 –
Vialaeaceae V. insculpta DAOM 240257 JX139726 –
Vialaeaceae V. mangiferae MFLUCC 12 KF724974 KF724975
0808
Vialaeaceae V. minutella BRIP 56960 KC181927 KC181925
Vialaeaceae V. minutella BRIP 56959 KC181926 KC181924
Xylariaceae Halorosellinia oceanica SGLAf82 EU715635 –
Xylariaceae Rosellinia limonispora CBS 382.86 KF719199 KF719211
Xylariaceae R. limonispora CBS 283.64 KF719198 KF719210
Xylariaceae Xylaria acuta 5220 JQ862676 JQ862637
Xylariaceae X. hypoxylon ATCC 42768 AY327477 AY327480

Taxonomy

Vialaea Sacc., Bull. Soc. mycol. Fr. 12: 66 (1896)

Probable synonym (from www.indexfungorum.org 2014):
Boydia A. L. Sm., Trans. Br. Mycol. Soc. 6(2): 151 (1919) [1918]
Facesoffungi number: 000011
Parasitic or endophytic on twigs of trees. Sexual state: Pseudostromata
(pseudoclypeus) absent or if present, immersed in host tissues, with brown to
210 Senanayake et al.: Vialaea mangiferae, sp. nov.

black tissues around immersed neck, containing one to nine ascomata. Asco-
mata perithecial, immersed solitary or aggregated in circular groups with
ostioles converging in their centre, globose or subglobose, somewhat flat-
tened, black to brown, Ostioles periphysate. Necks elongate dark brown to
black. Peridium composed of several layers of slightly flattened, strongly
melanized, thick-walled cells of textura globulosa or textura angularis with
peridial pores between cells. Paraphyses few in number, sometimes poorly
developed, deliquescent at maturity, much shorter than the asci, thin-walled,
hyaline, septate or not. Asci 8-spored, unitunicate, cylindrical, sometimes ta-
pering towards the apex or base, short stalked or sessile, thin-walled except
apex, apex obtuse or truncate, apical ring subconical or subapical, J+. As-
cospores biseriate, triseriate to fasciculate; sometimes weakly helically
coiled, strongly isthmoid, ends fusiform to rhombic, apical portion some-
times slightly larger than the basal part, 1–3-septate, hyaline, smooth-walled.
Asexual state: coelomycetous; pycnidial conidiomata, superficial, solitary,
globose. Conidiophores erect, branched, septate, hyaline. Conidiogenous cells
proliferating percurrently to produce small, hyaline, ellipsoidal conidia. Co-
nidia oblong to fusiform, one-celled, hyaline and smooth-walled.

Vialaea mangiferae I. C. Senanayake & K. D. Hyde, sp. nov. – Figs. 2–24

MycoBank: MB 808311
Facesoffungi number: FoF00005
E t y m o l o g y. – The specific epithet mangiferae is based on the host
genus from which the fungus was isolated.
H o l o t y p e . – MFLU 13–0342.
Parasitic on dead twigs of Mangifera indica. S e x u a l s t a t e . – Pseu-
dostromata visible as black dots on the host surface with covering of asco-
mata, not darkened, with 3–5 perithecia aggregated in a circle, with a single
central opening. Perithecia 155–265 µm high, 50–160 µm wide (x̄ = 206 ×
92 µm, n = 30), immersed, aggregated in circular groups, globose, dark brown,
ostioles converging and fusing into the centre of the pseudostroma, with a
barely raised papilla. Papilla 125–130 μm long, 25–35 μm wide (x̄ = 128 ×
30 µm, n = 20), periphysate. Periphyses 8–12 μm long (x̄ = 10 µm, n = 30), nu-
merous, hyaline. Peridium 15–25 μm wide (x̄ = 20 µm, n = 20), composed of 3–7
layers of slightly flattened, melanized, dark greenish-brown to blackish,
thick-walled cells of textura globulosa. Paraphyses 40–75 µm long, 5–10 µm
wide (x̄ = 55 × 7 µm, n = 10), hyaline, septate, blunt at the ends, shorter than
the asci. Asci 165–205 × 10–15 µm (x̄ = 187 × 12 µm, n = 10), 8-spored, unitu-
nicate, cylindrical, sometimes tapering towards the apex, usually short-
stalked, thin-walled, with a conical apical ring, 5–7.5 µm high × 2.5–4.0 µm
wide (x̄ = 6.1 × 3.2 µm, n = 10); with a small, oval, basal region with 2–3 µm
high, 1–2 µm wide (x̄ = 2.6 × 1.5 µm, n=10) is J+. Ascospores 75–90 µm long (x̄ =
83 µm, n = 15), overlapping biseriate to fasciculate, 1-septate, hyaline, with
two fusiform end cells, 45–50 × 4–5.5 µm (x̄ = 48.6 × 4.8 µm, n = 15), tapering
gradually to acute ends and towards the central portion, joined by a narrow,
 Sydowia 66 (2014) 211

Figs. 2–16. Vialaea mangiferae (holotype) 2. Dead twigs of Mangifera indica. 3. Appearance
of pseudostromata on the natural substrate. 4. Horizontal section of perithecia with central
neck. 5. Vertical section of perithecia with central neck. 6. Periphyses. 7. Peridium. 8. Pa-
raphyses. 9. Apical ring, blueing at the base in Melzer’s reagent. 10–12. Asci in water. 13–16.
Ascospores. Note: The central ostiolar canal and septum of ascospore arrowed in 4 and 14.
Scale bars: 4 200 µm, 5–7 50 µm, 8, 9 10 µm, 10–16 20 µm.
212 Senanayake et al.: Vialaea mangiferae, sp. nov.

long isthmus which is 2–3 µm wide, smooth-walled. A s e x u a l s t a t e . –

Conidiomata 300 × 500 µm diam., pycnidia, superficial, solitary, scattered,
globose, with slimy, shining spore mass and basal mycelium forming thick
black strands. Conidiophores 5–15× 1.5–2 µm (x̄ = 10.2 × 1.7 µm, n = 10), erect,
branched, septate, hyaline. Conidiogenous cells phialidic, discrete or in small
whorls, lageniform to cylindrical, hyaline. Conidia 2.5–3.5 × 1.5–2.0 µm, (x̄ =
3.1 × 1.7 µm, n = 10), oblong to ellipsoidal, one-celled, hyaline, smooth, with
± truncate abscission scar.
C o l o n y c h a r a c t e r i s t i c s . – Colonies on WA greenish black,
with sparse aerial mycelium, with irregular, dendritic or coralloid margin,
slow growing, attaining 2 cm diam within 14 days at 25 °C, producing co-
nidiomata after 45 days.
Material examined. – THAILAND, Chaing Rai Province, Muang District, near Bandu,
Baan Khuakhae, at 31M. 17, (19° 59’ 52.05” N; 99° 49’ 25.15” E), on twigs of Mangifera in-
dica, 15 Nov. 2012, leg. K. D. Hyde, CHUNI001 (MFLU13–0342, holotype); (HKAS 81790,
isotype), ex-type living culture = MFLUCC 12–0808 = ICMP.

Key to Vialaea species

1. Ascomata solitary, non-stromatic, globose.................................V. insculpta
1. Ascomata aggregated, pseudostromatic, globose to subglobose .............. 2
2. Peridium with angular cells, ascospores 96–112 × 4–4.5 µm........................
....................................................................................................... V. minutella
2. Peridium with globose cells, ascospores 75–90 × 4–6 µm.............................
.................................................................................................... V. mangiferae

Discussion
Vialaeaceae is a monotypic family representing the genus Vialaea, com-
prising the generic type, V. insculpta, and V. minutella. Vialaea bambusae
Hara (1913) and V. ingae Rehm (1901) were excluded from this family (Can-
non 1995). The protologue of V. bambusae is in Japanese, was translated by
Cannon (1995) and there are no records concerning the holotype specimen.
Thus Cannon (1995) considered it as a doubtful species and excluded it from
Vialaea. Vialaea ingae was treated as a synonym of Diatractium ingae (Rehm)
Syd. & P. Syd. (Cannon 1989).
Vialaea mangiferae is a new species collected from twigs of Mangifera
indica in Chaing Rai, Thailand. Besides V. minutella this is the second species
occurring on Mangifera indica. The twigs had died on the mango trees due to
insect borer damage and the taxon had probably developed as secondary
invaders on the dead twigs. A synopsis of the characters of V. insculpta, V.
mangiferae and V. minutella is provided in Tab. 2. Vialaea mangiferae shares
many similarities with other species in the genus. However V. insculpta dif-
fers from V. mangiferae in having non-stromatic, less globose, larger perithe-
cia (280–400 µm) with long, non-convergent ostiolar necks, a thick peridium
(20–30 µm) and 1 or 2-septate ascospores. Vialaea minutella contains subglo-
 Sydowia 66 (2014) 213

Figs. 17–24. Coelomycetous asexual state of Vialaea mangiferae (holotype). 17. Culture
from above. 18. Culture from below. 19–21. Conidiomata forming on WA. 22. Conidia form-
ing on phialides attached to conidiophores. 23. Conidia attached to lageniform intercalary
cells. 24. Conidia. Scale bars: 19, 21 500 µm, 20 1000 µm, 22–24 =10 µm.

bose, small perithecia (130–180 µm) within stromatic tissues, a thinner pe-
ridium comprising cells arranged in a textura angularis and 96–112 µm long
and 4–4.5 µm wide ascospores. Vialaea mangiferae has slightly larger, glo-
bose perithecia (150–260 µm), which form within stromatic tissues, a thick
peridium comprising cells that form globose cells and 75–90 µm long and
4.2–5.6 µm wide ascospores. However the V. mangiferae differences from V.
minutella are subtle.
There are no records concerning the asexual state of species of the fam-
ily Vialaeaceae. Cannon (1995), however, suggested that asexual state was
coelomycetous. In this study, conidiomata were produced within six weeks
after incubation on WA at 25 °C.
Phylogenetic analysis of the ITS and LSU sequence data provides evi-
dence that Vialaea mangiferae is a new species with a high bootstrap support
(94 %) and can be included in Vialaeaceae, where species form a distinct
clade (100 %) (Fig.1). In addition, the family placement of Vialaceaceae in
214 Senanayake et al.: Vialaea mangiferae, sp. nov.

Tab. 2. Synopsis of morphological features of Vialaea species.

Species Pseudostro- Perithecia Peridium Ascal ring Ascospores

mata

V. insculpta Indistinct 280–400 µm 20–30 µm thick, 3–4 µm 78–94 ×

diam., scattered composed of several diam., 5.5–6, 1–2
singly or layers of slightly 3.5–9.5 µm septate
aggregated in flattened, strongly high
small groups, melanized, thick
roughly globose walled forming
textura globulosa
V. minutella Black to 130–180 µm 14–18 µm thick, 3–4 3–4.5 µm 96–112 ×
brown, diam., aggre- layers of rather diam., 4–4.5,
immersed in gated, subglo- flattened, thick 4–6 µm 1-septate
bark, bose walled, dark brown high
containing cells forming
2–9 ascomata textura angularis
V. mangif- Dark brown, 150–260 µm 15–24 μm diam., 3–7 2.5–4.0 µm 75–91 ×
erae immersed, diam., aggre- layers of slightly diam., 4.2–5.6,
containing gated, globose flattened, mel- 5.2–7.5 µm 1-septate
3–5 ascomata anized, thick- high
walled globose cells

the Xylariales, rather than the Diaporthales is supported by LSU and ITS
data in this study, which concurs with Shoemaker et al. (2013) and McTag-
gart et al. (2013). The three species of Vialaea formed a monophyletic clade
within the Xylariales.

Acknowledgements
We would like to thank the CGIAR Research Program 1.2 – Humidtrop-
ics: Integrated systems for the humid tropics, for partially funding this work.
Liu Ende, Assistant Curator, Herbarium, Kunming Institute of Botany, Chi-
nese Academy of Sciences (KUN), Kunming, China and Saranyaphat Boon-
mee, Curator, MFLU Herbarium, The Institute of Excellence in Fungal Re-
search, Mae Fah Luang University, Chiang Rai, Thailand were thanked for
keeping herbariums. Chinthani Senanayake thanks S. C. Karunarathana, N.
N. Wejayawardhana, D. N. Wanasingha and Yee Yee Than at Mae Fah Luang
University, Chiang Rai, Thailand for their valuable suggestions and help. K.
D Hyde thanks The Chinese Academy of Sciences, project number
2013T2S0030, for the award of Visiting Professorship for Senior Internation-
al Scientists at Kunming Institute of Botany.

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(Manuscript accepted 14 May 2014; Corresponding Editor: I. Krisai-Greilhuber)

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