Mycologia
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Services
Right arrow Similar articles in this journal
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Põldmaa, K.
Right arrow Search for Related Content
PubMed
Right arrow Articles by Põldmaa, K.
Agricola
Right arrow Articles by Põldmaa, K.
Mycologia, 95(5), 2003, pp. 921-933.
© 2003 by The Mycological Society of America

Three species of Hypomyces growing on basidiomata of Stereaceae


Kadri Põldmaa 1

     Institute of Zoology & Botany, Estonian Agricultural Academy, Riia 181, EE-51014, Tartu, Estonia and Estonian Biocenter, Riia 23, EE-51010, Tartu, Estonia

   ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 TAXONOMY
 DISCUSSION
 LITERATURE CITED
 

Of the eight species of Hypomyces that occur on basidiomata of Stereum species, only H. sympodiophorus grows exclusively on members of this genus. Morphologically similar fungi were found on species of Xylobolus, a genus closely related to Stereum. These are described as two new species of Hypomyces: H. thailandicus, collected on Xylobolus cf. illudens in Thailand; and H. xyloboli, on X. frustulatus and X. subpileatus in the eastern United States. These three species are unusual in Hypomyces because of their almost indistinguishable anamorphs. In parsimony analysis of LSU nuclear rDNA sequences, the three species growing only on Stereaceae do not form a monophyletic group but their constrained monophyly is not rejected either. A morphologically similar anamorphic species, Sibirina gamsii, included in the study, is transferred to the genus Cladobotryum.

Key words: Cladobotryum, fungicolous fungi, Hypocreales, rDNA, Sibirina, Stereum, systematics, Xylobolus


   INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
 RESULTS
 TAXONOMY
 DISCUSSION
 LITERATURE CITED
 
A number of fungicolous ascomycetes occur only or mainly on basidiomata of members of the genus Stereum Pers. (Stereaceae, Hymenomycetes, Basidiomycota), viz. Ascocodinaea stereicola Samuels et al (Samuels et al 1997Citation), species of Hypomyces (Fr.) Tul. outlined below, Nectriopsis oropensoides Rehm (Samuels 1976Citation), Gliocladium polyporicola (Henn.) Seifert & W. Gams, Sphaerostilbella aureonitens (Tul.) Seifert et al, Sph. berkeleyana (Plowr. & Cooke) Samuels & Candoussau, Stilbella sebacea (Ellis & Everh.) Seifert and Sti. stereicola Seifert (Seifert 1985Citation). Different species often grow together on the same basidioma, e.g., in Europe, in the cases of the anamorphs of H. sympodiophorus Rogerson & Samuels and Sph. aureonitens (Gliocladium penicillioides Corda). In North America their teleomorphs are common and frequently accompanied by Ascocodinaea stereicola and Gliocladium polyporicola.

Species of Hypomyces vary in host specificity. Aphyllophoricolous species of Hypomyces and Cladobotryum Nees (presumed asexual Hypomyces) are generally less host-specific than Hypomyces species found on discomycetes, boletes and agarics. Exceptions include H. sympodiophorus, a species recorded only on species of Stereum, and H. albidus Rehm, a taxon that produces the teleomorph almost exclusively on St. sanguinolentum (Alb. & Schwein.: Fr.) Fr. Curiously, the anamorph of the latter occurs on a variety of aphyllophorous hosts (Rogerson and Samuels 1993Citation, Põldmaa and Samuels 1999Citation). Most aphyllophoricolous species of Hypomyces and Cladobotryum with wider host ranges also have been recorded on species of Stereum.

Hypomyces aurantius on Xylobolus subpileatus (Berk. & M. A. Curtis) Boidin (Arnold 1976Citation) is the only record of Hypomyces on Xylobolus P. Karst., a genus considered to be closely related to Stereum (Chamuris 1988Citation). It recently has been shown that the two genera form a monophyletic group with several corticioid taxa nested in the russuloid clade of the homobasidiomycetes (Hibbett and Thorn 2001Citation, Wu et al 2001Citation, Larsson and Larsson in pressCitation). The term aphyllophourous is retained here also for the hosts belonging to Stereaceae, despite their classification in the Russulales (Kirk et al 2001Citation). It refers to the life form of hymenomycetes with basidiomata that mostly are nongilled, leathery or woody and in many cases perennial, as opposed to the usually soft, fleshy and annual fruiting-bodies of agarics.

The aim of this study was to clarify the systematics of species of Hypomyces found exclusively on basidiomata of Stereaceae, using phenotypic and molecular characters. Another objective was to test the monophyly of such species. A number of recently collected specimens of Hypomyces on the basidiomata of species of Stereum and Xylobolus were examined. Sibirina gamsii D. J. Gray & Morgan-Jones, an anamorphic fungus known from a single collection on Polyporus Fr., was included in the study because of its similarity to the anamorphs of Hypomyces inhabiting Stereaceae.


   MATERIALS AND METHODS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
RESULTS
 TAXONOMY
 DISCUSSION
 LITERATURE CITED
 
Morphological characterization – Colony diameters were measured from 9 cm diam plastic Petri dishes, into which a 4 mm diam plug taken from the edge of an actively growing colony was placed. Cultures were grown on malt-extract agar (1.5% malt extract, Becton Dickinson, 1.5% agar, Merck, Darmstadt, Germany) and cornmeal agar (Sigma Chemical Co., St. Louis, Missouri) at 15, 20, 25, 30 C in the dark for 1 wk, after which they were exposed to daylight to induce additional pigment formation. Microscopic measurements were made from cultures grown on MEA at 25 C and from herbarium specimens in 3% aqueous KOH solution. Thirty ascospores and conidia generally were measured from each specimen. Color references are taken from Kornerup and Wanscher (1967)Citation. The abbreviations C.T.R. and G.J.S. stand for the numbers in the culture collections of Clark T. Rogerson and Gary J. Samuels, both preserved at the Systematic Botany and Mycology Lab, U.S.D.A. (Beltsville, Maryland, USA.).

Isolates used for sequencing – Sequences in this study were obtained from 14 isolates of Hypomyces. The hosts and collecting data for the respective specimens not treated in this paper are cited by Gray and Morgan-Jones (1980)Citation, Rogerson and Samuels (1993)Citation, Põldmaa (1999)Citation and Põldmaa and Samuels (1999)Citation except for H. albidus TFC 99–163: U.S.A. Maryland, Anne Arundel County, Patuxent Wildlife Refuge, on St. hirsutum (Willd. : Fr.) Gray, 17 Oct 1999, K. Põldmaa, TAA 170339. The EMBL/GenBank accession numbers for the new sequences are: H. albidus G.J.S. 90–33—AJ459297, TFC 99–163—AJ459298; H. polyporinus Peck G.J.S. 88–40—AJ459301; H. semitranslucens G. Arnold C.T.R. 74–28—AJ459302, TFC 96–35—AJ459303; H. sibirinae Rogerson & Samuels CBS 744.88—AJ459304; H. sympodiophorus TFC 97–155—AJ459305, TFC 99–143—AJ459306, TFC 99–198—AJ459307; H. subiculosus (Berk. & Curt.) Höhn. G.J.S. 83–288—AJ459309; H. thailandicus TFC 97–142—AJ459310; H. xyloboli TFC 99–152—AJ459299, TFC 00–65—AJ459300; Sibirina gamsii CBS 598.88—AJ459308. Data for the 35 earlier published sequences from the Hypocrea-Hypomyces clade, included in this study, are available in papers dealing with the phylogeny of this group (Rehner and Samuels 1994Citation, 1995Citation, Põldmaa et al 1999Citation, Põldmaa 2000Citation).

DNA extraction, amplification and sequencing – The isolates were grown in 100 mL 1.5% potato-dextrose broth (Difco) in 200 mL flasks on a shaker at 125 rpm for 5–7 d at room temperature under ambient light. Mycelium was harvested by vacuum filtration on Whatman No. 1 filter paper and freeze-dried before DNA extraction. DNA was extracted with the PureGene genomic DNA isolation kit (Gentra Systems, Minneapolis, Minnesota, U.S.A.) according to the manufacturer's instructions.

The 5' end of the large subunit of the nuclear ribosomal DNA (LSU rDNA) was amplified in 50 µL reactions on a GeneAmp 9700 thermal cycler (Applied Biosystems, Foster City, California, U.S.A.) under these reaction conditions: 10–15 ng of genomic DNA, 200 mM each dNTP, 2.5 units Amplitaq (Applied Biosystems, Foster City, California, U.S.A.), 25 pmoles each of primers LR0R and LR7 (Vilgalys and Hester 1990Citation) and the supplied 10(PCR buffer with 15 mM MgCl2. The thermal cycler program was: 10 min at 95 C, followed by 35 cycles of 30 s at 94 C, 30 s at 55 C, 1 min at 72 C, with a final extension period of 10 min at 72 C. After amplification, PCR products were purified with QIAQuick columns (Qiagen Inc., Chatsworth, California, U.S.A.) according to the manufacturer's instructions. Amplified products were sequenced with the BigDye terminator kit (Applied Biosystems, Foster City, California, U.S.A.) on an ABI 310 automated DNA sequencer using these primers: LR0R, LR3R, LR5, LR7 (Vilgalys and Hester 1990Citation).

Phylogenetic analysis – Sequences comprising approximately 1350 bp from the 5' end of the LSU rDNA were edited using Sequencher 3.1 (Gene Codes, Ann Arbor, Michigan) and aligned manually using the program Genedoc 2.5 (Nicholas et al 1997Citation). An ambiguously aligned region of 10 bp was excluded from the analysis. The aligned sequences of the 49 taxa examined are available in the EMBL-Align database (ALIGN 000387).

Maximum-parsimony (MP) analysis was performed with PAUP* 4.0 b10 (Swofford 1999Citation), using unweighted characters, stepwise addition (addition sequence random, 1000 replications) and tree bisection-reconnection (TBR) swapping. Gaps were treated as missing data. The sequences of two species of Verticimonosporium Matsush. were used as outgroup. The sequences of the two specimens of H. thailandicus were identical and thus only that of the holotype was included in the analyses. Support for clades was evaluated by 1000 bootstrap replications with PAUP*; Bremer support was estimated with AutoDecay 4.0.2 (Eriksson 1997Citation). A heuristic search was conducted while forcing the species growing only on Stereaceae into one clade. The likelihood scores and tree lengths of the resulting MP trees were compared to the most likely unconstrained MP tree.


   RESULTS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
TAXONOMY
 DISCUSSION
 LITERATURE CITED
 
Morphological characterization – Differences in morphological characters (Table I), ascospore and conidial measurements (Fig. 1) and growth rates (Fig. 2) distinguish two new species growing on species of Xylobolus from H. sympodiophorus. Hypomyces sympodiophorus has been considered exceptional among aphyllophoricolous Hypomyces because its perithecia have a two-layered wall and often are seated directly on the host with no obvious subiculum (Figs. 3a–c, 4a). Hypomyces xyloboli, and to some extent H. thailandicus, described here, share these features (Figs. 3f, h, i, k, 5a, 6a). The size of perithecia varies in the same range in all species, with perithecia up to 360 µm high and 300 µm wide found in H. sympodiophorus, in contrast to the smaller dimensions reported earlier (Rogerson and Samuels 1993Citation). The ascospores of H. sympodiophorus are atypical in the genus, being nonapiculate and finely verrucose to smooth-walled. However, small apiculi <1 µm long, were found at both ends of the ascospores in TAA 169681 (Fig. 4c), TAA 169710 and J. Fournier 01251. In H. thailandicus and H. xyloboli, the apiculi are larger and the ascospores clearly ornamented (Figs. 5c, 6c). Ascospore length and width are different among the three species (ANOVA: F2,15 = 10.8, P = 0.0013 and F2,15 = 29.9, P < 0.0001, respectively), with H. thailandicus having larger ascospores than the other two species (Fig. 1a).


View this table:
[in this window]
[in a new window]
  		TABLE I. Characters distinguishing three of species of Hypomyces growing on Stereaceae

 


View larger version (19K):
[in this window]
[in a new window]
  		FIG. 1. Scatterplots of ascospore (a) and conidial (b) measurements of three species of Hypomyces and C. gamsii. The points represent mean values of specimens. {diamond} = H. sympodiophorus, {circ} = H. thailandicus, {square} = H. xyloboli, x = C. gamsii

 


View larger version (31K):
[in this window]
[in a new window]
  		FIG. 2. Colony diameter of three species of Hypomyces and C. gamsii grown on 1.5% MEA in the dark for 7d. Continuous lines—H. sympodiophorus, marked lines: • = H. thailandicus, {square} = H. xyloboli, x = C. gamsii. The diameter of the plug used for inoculations is 4 mm, thus plugs below this size indicate that no growth was observed

 


View larger version (146K):
[in this window]
[in a new window]
  		FIG. 3. Perithecia and anamorphs of three Hypomyces species on Stereaceae on their natural substrata. a–e. H. sympodiophorus, f, g. H. thailandicus, h–m. H. xyloboli. a–c, f, h, i. Perithecia on the hymenophore of the host (in a. together with perithecia of Ascocodinaea stereicola). d, e, g, l, m. Anamorph on the host. j. Conidia from cultures on MEA. k. Section through two perithecia and the host, mounted in lactic acid. a = BPI 745789; b, c = TAA 170083; d = TAA 170354; e = G. Arnold 00–122; f, g = TAA 169692; h, i, j = TAA 170300; k = TAA 170546; l, m = TAA 170429. Scale bars: a = 0.5 mm, b, h = 1 mm, c, e–g, i, m = 0.2 mm, d =1 cm, j = 10 µm, k = 100 µm, l = 1 mm

 


View larger version (129K):
[in this window]
[in a new window]
  		FIG. 4. Hypomyces sympodiophorus (a–c, e–i) and C. gamsii (d). a. Section through a perithecium. b. Ascus. c. Ascospores. d. Conidiophores. e, f. Conidia attached to the conidiogenous cells. g. Tip of a conidiophore. h. Conidia. i. Chlamydospores. a = BPI 841960; b = TAA 170082; c = TAA 169681; d = CBS 598.88; e = TFC 97–155; f = TFC 99–198; g = G. Arnold 00–105; h = G.J.S. 95–167 (=CBS 828.95); i = TFC 97–122. a–c from the natural substratum, others from cultures on MEA. Scale bars: a, d = 50 µm, b, c, g–i = 10 µm, e, f = 20 µm

 


View larger version (103K):
[in this window]
[in a new window]
  		FIG. 5. Hypomyces thailandicus, TAA 169692. a. Section through a perithecium. b. Asci. Ascospores. d. Upper part of a conidiophore. e. Conidiogenous cells, the arrow pointing to the refractive area under a conidiogenous locus. f. Conidia. g. Chlamydospore. a–c from the natural substratum, others on MEA. Scale bars: a, g = 50 µm; b, c, e, f = 10 µm; d = 20 µm

 


View larger version (127K):
[in this window]
[in a new window]
  		FIG. 6. Hypomyces xyloboli. a. Section through a perithecium; b. Asci. c. Ascospores. d, e. Upper parts of conidiophores. f. Conidia. g. Chlamydospores. h. Conidiophores. a, f–h = TAA 170300; b, c = 170546; d = 170416; e = 170520. a–c, e from the natural substratum, others from cultures on MEA. Scale bars: a, d, h = 50 µm; b, c, e–g = 10 µm

 
Most of the anamorph characters, e.g., the size and morphology of conidiogenous (Figs. 4g, 5d–e, 6d–e) and chlamydospore-like cells (Figs. 4i, 5g, 6g) and the shape of conidia (Figs. 4h, 5f, 6f), show no variation among the three species growing on Stereaceae, as well as S. gamsii. Only H. xyloboli is easily distinguished by its green conidia (Fig. 3j). The production of a single conidium from each conidiogenous locus that is formed at the tip of a refractive area (Figs. 4d–g, 5d–e, 6e, h) is also characteristic of all the anamorphs. The ranges of length and width of conidia of the four species overlap, although the means are different (ANOVA: F3,34 = 11.6, P < 0.0001 and F3,34 = 7.5, P = 0.0005, respectively; Fig. 1b). As with the size of ascospores, it remains unkown whether the ranges of mean values in H. thailandicus also overlap with those of the other species because only two collections from very close localities are known.

The colony growth rates show high infraspecific variation and overlap among the three species on Stereaceae (Fig. 2). In H. sympodiophorus, the strains appear to be adapted to the climatic conditions of the localities of their origin (data not shown) because isolates from the tropics usually have higher temperature optima than isolates from areas with cooler climates. Strains from Estonia, the northernmost locality of strains studied, grow more slowly than those from other localities, reaching a diam of 10–17 mm at 20 C and 7–10 mm at 25 C in 7 d on MEA. Despite of the high variability of colony growth rates in H. sympodiophorus, all strains grow much slower at 15 and 30 C than at 20 and 25 C. In H. xyloboli the strains TFC 99–152, 00–65 and 00–120 grow slightly faster at 20 C than at 25 C, but in TFC 00–89 and 00–53 the situation is reversed. Hypomyces thailandicus is distinguished by having the fastest growth at 30 C. Growth rates of the isolates of the three species on CMA (data not shown) are similar to those on MEA in their values, infraspecific variation and overlap among the species.

Infraspecific variability is found also in colony pigmentation. The formation of orange and salmon pigments unequivocally distinguishes H. xyloboli. However, in the cultures derived from its holotype (TFC 99–152) and from 00–53, the submerged mycelium remains uncolored. The isolates of H. thailandicus and that of S. gamsii do not form any pigment on MEA. Isolates of H. sympodiophorus also remain uncolored or the colony reverse often turns yellow.

Molecular analysis – The alignment consisted of 1333 characters, of which 111 were parsimony informative. MP analysis resulted in 116 MP trees of 496 steps in length (CI 0.35; -ln likelihood 4686.64–4689.73). The tree with the highest likelihood is presented in Fig. 7 (the remaining MP trees were not significantly worse, P = 0.474–0.988). The strict-consensus tree differed in that several of the unsupported branches were collapsed.



View larger version (39K):
[in this window]
[in a new window]
  		FIG. 7. The tree with the best likelihood obtained in parsimony analysis. Tree length = 496, CI = 0.35, -ln likelihood = 4686.64. Bootstrap values (shown for values >50%) and decay indices are shown above the branches. Asterisks mark the clades that are collapsed in the strict consensus. The names of species that grow on members of Stereaceae are followed by #, whereas those that are found only on Stereaceae are given in bold. The ex-type strain of Helminthophora uniseptata is indicated with a. Scale bar = five steps

 
The genus Hypomyces is not monophyletic as demonstrated in previous studies based on the analyses of the same gene region (Rehner and Samuels 1994Citation, 1995Citation, Põldmaa et al 1999Citation, Põldmaa 2000Citation). In the phylogeny presented here, the species growing on Stereaceae are not monophyletic either and are distributed among all major clades of aphyllophoricolous taxa (Fig. 7). The KOH-positive species with a wide host range, including members of Stereaceae (H. aurantius (Pers.: Fr.) Tul., H. odoratus G. Arnold, H. rosellus (Alb. & Schwein.) Tul. and H. subiculosus), form a poorly supported group with morphologically similar species (clade I). Clade II, which is sister to clade I and unresolved in the strict consensus, received good support. It includes four isolates of H. sympodiophorus, Helminthophora uniseptata Castañedam, recently was synonymized with the anamorph of H. sympodiophorus (Põldmaa and Samuels 1999Citation) and Sibirina gamsii. The three Hypomyces isolates from basidiomata of Xylobolus species form a poorly supported subclade within clade III. This clade comprises the majority of the analyzed KOH-negative aphyllophoricolous taxa, including H. albidus, H. orthosporus K. Põldmaa and H. semitranslucens, which also are found on Stereaceae. Clade III also encompasses H. tremellicola (Ell. & Ever.) Rogerson, found mostly on Crepidotus (Fr.) Staude and H. sibirinae, which is recorded from different polypores. Cladobotryum stereicola (G. Arnold) Rogerson & Samuels and Hypomyces mycophilus Rogerson & Samuels, two species reported occasionally from Stereaceae, are members of clade IV, a basal group that receives strong bootstrap support. Sphaerostilbella aureonitens and Sph. berkeleyana that grow mostly on species of Stereum, are not closely related to any of the stereicolous Hypomyces, being part of the weakly supported clade containing other members of Sphaerostilbella Sacc.

The constrained analysis that forced the monophyly of H. sympodiophorus clade (including S. gamsii) and the three isolates on Xylobolus resulted in 676 trees of 504 steps in length (-ln likelihoods 4708.14–4718.41). The Kishino-Hasegawa test revealed that 650 of the constrained trees did not differ significantly from the unconstrained MP tree with the best-likelihood score and rejected 26 of the trees (P = 0.023–0.204). The Templeton test rejected 34 of the constrained trees (P = 0.033–0.349).


   TAXONOMY
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 TAXONOMY
DISCUSSION
 LITERATURE CITED
 
Hypomyces sympodiophorus Rogerson & Samuels, Mycologia 85:241. 1993. Figs. 3a–e, 4a–c, e–i


Anamorph: Cladobotryum uniseptatum (Castañeda) K. Põldmaa, Mycologia 91:192. 1999.

New records not cited by Rogerson and Samuels (1993)Citation, Põldmaa and Samuels (1999)Citation and Põldmaa (1999)Citation:

Both teleomorph and anamorph present: AUSTRALIA. QUEENSLAND: ca 10 km S of Atherton, Wongabell State Forest, on Stereum ostrea (Bl. & Nees) Fr. s.l., 30 Aug 1999, K. Põldmaa, TAA 170236, TFC 99–141; Crater Lakes National Park, along Wrights Creek, on St. ostrea s.l., 30 Aug 1999, K. Põldmaa, TAA 170251, TFC 99–143. FRANCE. Ariège, Rimonte, Saurine, on St. hirsutum, 21 Oct 2001, J. Fournier 01251 (TAA), TFC 02–40. THAILAND. PHETCHABURI PROVINCE: Kaeng Krachan National Park, along road to Torthip Falls, on Stereum sp., 15 Jul 1997, G. J. Samuels, P. Chaverri & D. Pfister, BPI 74570, G.J.S. 97–144, TFC 97–155; NAKHORN NAYOK PROVINCE: Khao Yai National Park, on St. ostrea s. l., between the Visitors' Center and Kong Khaeon Camp, 29 Jul 1997, K. Põldmaa, G. J. Samuels & P. Chaverri, BPI 745622, TAA 169681, TFC 97–44, CBS 100360; Wang Jumpee trail to Lap Tha Kong Creek, on Stereum sp., 31 Jul 1997, K. Põldmaa, G. J. Samuels & P. Chaverri, TAA 169709; the same collecting data, on St. ostrea s. l., BPI 745685, TAA 169710, TFC 97–149; vicinity of park headquarters along a trail, on St. ostrea s. l., 10 Aug. 1997, G. J. Samuels, K. Põldmaa & P. Chaverri, BPI 745687, G.J.S. 97–44; Phaeodai, on St. ostrea s. l., 12 Aug. 1997, G. J. Samuels & P. Chaverri, BPI 745738; Darn Chang, on St. ostrea s. l., G. J. Samuels & P. Chaverri, BPI 745789; a trail from the main road to Bun Phai, north of Khao Yai Forest headquarters, on Stereum ostrea, 6 Sep 2001, G. J. Samuels, M. Reblova & R. Nasit, BPI 841960, G.J.S. 01–252; 4 km S of park headquarters, Princess Trail, on Stereum ostrea, 6 Sep 2001, G. J. Samuels, M. Reblova & R. Nasit, BPI 842049, G.J.S. 01–266; W of park headquarters at the separation point of Mo Sing To and Nong Pak Chi trails, on Stereum ostrea, 7 Sep 2001, G. J. Samuels, BPI 842053, G.J.S. 01–241. UNITED STATES. KENTUCKY: Daniel Boone National Forest, Menifee County, Tarr Ridge, on St. ostrea, 29 Jul 1999, G. J. Samuels & K. Põldmaa, TAA 170082, 170083. MARYLAND: Anne Arundel County, Patuxent Wildlife Refuge, northern region, on St. hirsutum, 17 Oct 1999, K. Põldmaa, TAA 170346, TFC 99–228; Prince George's County, golf field of University of Maryland, on Stereum sp., 19 Feb 2000, K. Põldmaa, TAA 170381. VIRGINIA: Shenandoah County, George Washington National Forest, on Stereum sp., 3 Sep 2000, K. Põldmaa & G. Arnold, TAA 170507, TAA 170512, G. Arnold 00–122.

Only anamorph present: THAILAND. NAKHORN NAYOK PROVINCE: Khao Yai National Park, trail to Tad Tha Phu falls, on Stereum sp., 5 Aug 1997, K. Põldmaa, G. J. Samuels, P. Chaverri & P. Lutthisungneon, TAA 169746, TFC 97–27; same collecting data, on St. ostrea s. l., BPI 745841, TAA 169748. UNITED STATES. KENTUCKY: Rowan County, Daniel Boone State Forest, a trail W from Stoney Cove, on St. ostrea, 28 Jul 1999, K. Põldmaa, TAA 170076, TFC 99–196; same collecting data, on St. hirsutum, 29 Jul 1999, TAA 170079, TFC 99–198. MARYLAND: Howard County, Patapsco Valley State Park, near River Road, 12 Oct 1999, K. Põldmaa, TAA 170301; Prince George's County, Patuxent Wildlife Refuge, northern region, on St. hirsutum, 17 Oct 1999, K. Põldmaa, TAA 170339, TFC 99–164; southern region, on St. hirsutum, 13 Nov 1999, K. Põldmaa, TAA 170372, TAA 170373, TFC 99–240, on St. ostrea TAA 170376. MISSOURI: Castlewood State Park, on St. ostrea, 29 Jul 1999, K. Põldmaa, TAA 170087. VERMONT: Chittenden County, near Burlington, Indian Brook Conservation Area, on Stereum sp., 29 Jul 2000, K. Põldmaa, TAA 170462. WEST VIRGINIA: Marion County, SE of Fairmont near Taiga river, Valley Falls State Park, on St. hirsutum, 30 Oct 1999, K. Põldmaa, TAA 170354; same collecting data, on St. cf. hirsutum, 170358; same collecting data, on St. ostrea, TAA 170353, TFC 99–234.

Hypomyces sympodiophorus was described based on a collection from Pennsylvania, U.S.A. (Rogerson and Samuels 1993Citation), and records from Europe and the United States were added by Põldmaa and Samuels (1999)Citation and Põldmaa (1999)Citation. The anamorph of H. sympodiophorus is common wherever old basidiomata of Stereum spp. occur. As with several other species of Hypomyces, the teleomorph is rarer in Europe than in North America and the tropics. The many collections of H. sympodiophorus from different parts of the world indicate that the species is probably cosmopolitan.

Cladobotryum gamsii (D. J. Gray & Morgan-Jones) K. Põldmaa, comb. nov. Fig. 4d


Basionym: Sibirina gamsii D. J. Gray & Morgan-Jones, Mycotaxon 10:396. 1980.

This anamorphic species, known only from its type collection on Polyporus sp. from Alabama, U.S.A., morphologically is similar to the anamorphs of the three species growing on Stereaceae. Several differences were found between our subcultures of the ex-type strain (CBS 598.88) and the protologue of C. gamsii (Põldmaa and Samuels 1999Citation). Compared to the majority of H. sympodiophorus isolates, it is characterized by less frequent intercalary conidiogenous loci, slightly larger conidia and faster growth on MEA, being thus most similar to the anamorph of H. thailandicus. The analysis of LSU rDNA data, however, reveals that C. gamsii is nested within the clade of isolates of H. sympodiophorus (Fig. 7) that cannot be resolved based on this gene alone. While there is strong likelihood that C. gamsii is the anamorph of a H. sympodiophorus-like teleomorph, more rapidly evolving gene regions need to be analyzed to resolve the question of the possible conspecificity of these two species.

The species is transferred to the anamorph genus Cladobotryum, accepting its expanded concept presented by Rogerson and Samuels (1993)Citation. The anamorph genus Sibirina G. Arnold was defined by the formation of a single conidium from each conidiogenous locus and dense verticils of conidiogenous cells (Arnold 1970Citation, Gams 1973Citation). These characters are common to all four anamorphs treated in this paper. In the phylogenetic tree, these four species and those with anamorphs initially described in the anamorph genus Sibirina (H. orthosporus, H. semitranslucens, H. sibirinae) do not form a monophyletic group. Even though most of these species are closely related, this does not advocate retaining a separate genus name for their anamorphs.

Hypomyces thailandicus K. Põldmaa & Samuels, sp. nov. Figs. 3f–h, 5


Anamorph: Cladobotryum sp.

Subiculum effusum, pallidum; hyphae hyalinae, 3–5 µm latae. Perithecia obpyriformia, 250–360 x 200–280 µm; fere superficialia, solitaria, griseo- vel brunneo-aurantiaca, KOH ope colore immutato; papilla 40–80 µm alta. Asci cylindrici, 100–130 x 6.5–8 µm, octospori, apice paulum incrassato. Ascosporae fusiformes, 16.5–20 x 5–6 µm, aeque bicellulares, hyalinae, prominenter verrucosae, apiculatae; apiculi 2–3 µm longi. Conidiophora in natura erecta, septata, hyalina. Cellulae conidiogenae in verticillis dispositae, subulatae, 15–20 µm longae, ad basim 2.5 µm latae, 1–3 locis conidiogenis e protrusionibus refringentibus ortis. Conidia cylindrica, 11–15 x 5.5–6.5 µm, bicellularia, hyalina. Chlamydosporae absentes.

Subiculum effuse, thin, scant, buff; hyphae hyaline, smooth, 3–5 µm wide, KOH-. Perithecia obpyriform, 250–360 x 200–280 µm; almost superficial on the subiculum, solitary; grayish to brownish orange (5–6 B–C 5–6), KOH-; inner cells of the perithecial wall flattened, outer cells subglobose; papilla 40–80 µm high, with outer cells swollen to 12 µm. Asci cylindrical, 100–130 x 6.5–8 µm, apex thickened, no pore observed; ascospores uniseriate with ends overlapping. Ascospores fusiform, 16.5–20 x 5–6 µm, hyaline; 1-septate, septum median or sometimes lacking; grossly warted with warts up to 1.5 µm high, apiculate with apiculi 2.5–3 µm long and blunt tips.

Characteristics of the associated anamorph. Conidiophores arising from scant mycelium, erect, septate, hyaline. Conidiogenous cells in verticils, subulate, 15–18 µm long and 2–2.5 µm wide at the base, attenuating gradually to 0.5–1 µm at the tip, with 1–3 conidiogenous loci formed on refractive protrusions. Conidia cylindrical, straight, 11–15 x 5.5–6.5 µm, hyaline, 1-septate, not constricted at the septum, cells of equal size, basal hilum centrally placed. Chlamydospores not observed.

Characteristics in culture. Colonies spreading slowly, reaching 31–44 mm diam at 25 C in 7 d; cottony, margin regular; white, reverse not colored. Odor absent. Aerial mycelium scant; hyphae hyaline, 2.5–5.5 µm wide, smooth-walled. Submerged mycelium abundant, fasciculate, cells often becoming swollen. Conidiophores arising from aerial hyphae, not differentiated from these, unbranched or giving a few side branches near the top that bear conidiogenous cells. Conidiogenous cells 3–8 in a verticil; subulate, 14–30 µm long, attenuating gradually from 2–2.5 µm at the base to 1–2 µm at the tip; with 1–2 conidiogenous loci at the tips of refractive protrusions, forming one or rarely two conidia from each. Conidia ellipsoidal to cylindrical, sometimes clavate or obovate, straight, 13.5–19 x 6.5–8 µm, hyaline, 1-septate, cells of equal size or one being larger than the other; with a protuberant central basal hilum, which is 1 µm wide; produced singly and symmetrically at the tips of conidiogenous cells or seldom two joined at the base in imbricate position. Chlamydospore-like cells rare, intercalary on submerged hyphae, 2–10 in a chain or occasionally aggregated into irregular clusters; sometimes by two, formed terminally at the end of short lateral branches; cells sublobose to ellipsoidal, 12–16 x 9–11 µm, hyaline, wall 0.5–1 µm thick.

HOLOTYPE. THAILAND. NAKHORN NAYOK PROVINCE: Khao Yai National Park, Wang Jumpee trail to Lamp Tha Kong Creek, on Xylobolus cf. illudens (Berk.) Boidin, 31 Jul 1997, K. Põldmaa, G. Samuels & P. Chaverri, TAA 169692, culture TFC 97–142, CBS 110401. PARATYPE. THAILAND. Khao Yai National Park, on Xylobolus cf. illudens, Aug 1997, K. Põldmaa, G. Samuels & P. Chaverri, TAA 169772, culture TFC 97–165.

Among the KOH-negative aphyllophoricolous species of Hypomyces, H. thailandicus can be recognized by its comparatively short and wide ascospores, the walls of which are covered with low and broad warts. In this regard, H. thailandicus resembles H. favoli Samuels et al, H. mycophilus, H. puertoricensis Samuels et al and H. subiculosus, all of which can be distinguished through conspicuous differences in their anamorphs. Moreover, H. subiculosus differs in having KOH-positive perithecia and H. favoli and H. puertoricensis possess aseptate ascospores. The size, color and wall anatomy of the perithecia in H. thailandicus is similar to that of H. sympodiophorus and H. xyloboli. However, in H. thailandicus the cells of the outer region of the perithecial wall are not thick-walled and are arranged more loosely, as in most aphyllophoricolous Hypomyces.

The anamorph of H. thailandicus is almost identical to the ex-type culture of C. gamsii. In addition to its similarity to the other two species treated here, the anamorph of H. thailandicus also is reminiscent of the anamorph of H. orthosporus but differs from it in the higher frequency of intercalary loci and the refractive tips of conidiogenous cells.

Hypomyces xyloboli K. Põldmaa, sp. nov. Figs. 3i–m, 6


Anamorph: Cladobotryum sp.

Subiculum absens. Perithecia obpyriformia, 300–400 x 250–340 µm, superficialia in substrato naturali, solitaria, brunnea, KOH ope colore immutato; papilla 40–95 µm alta. Asci cylindrici, 110–125 x 5–6 µm, octospori, apice incrassato. Ascosporae ellipsoideae vel fusiformes, 15–18 x 3.5–4.5 µm, hyalinae, aeque bicellulares, minute verrucosae, apiculatae; apiculi 1–1.5(–2) µm longi. Velutum glaucum formans. Conidiophora in natura erecta, septata, hyalina vel virescentia. Cellulae conidiogenae 3–13 in verticillis dispositae, subulatae, 13–16(–22) µm longae, ad basim 2–3 µm latae, 1–3 locis conidiogenis e protrusionibus refringentibus ortis. Conidia cylindrica, aliquando obovata vel clavata, 8–13 x 5–6 µm, bicellularia, virescentia. Chlamydosporae absentes.

Subiculum absent. Perithecia seated on the hymenophore of the host; obpyriform, 300–400 x 250–340 µm; solitary; brown (5–6 E 4), KOH-; inner cells of the perithecial wall flattened, thin-walled; cells of the outer region subglobose, thick-walled; papilla conical, 40–95 µm high, with outer cells 8–11 x 5.5–7 µm. Asci cylindrical, 110–125 x 5–6 µm, apex thickened, 1.2–1.7 µm, no pore observed; ascospores uniseriate with ends overlapping. Ascospores ellipsoidal to fusiform, 15–18 x 3.5–4.5 µm, hyaline; 1-septate, septum median; finely verrucose with warts <0.5 µm high, apiculate with apiculi 1–1.5(–2) µm long, and 1–1.5 µm wide at the widest place in the middle and blunt tips. Chlamydospores absent.

Characteristics of the associated anamorph. Mycelium that bears the conidial apparatus forming a delicate grayish green (28–29 C 4) mat that is effused over the hymenophore and sometimes the upper surface of the host's basidiomata. Conidiophores arising from the mycelium, erect, branched once or twice in the upper part; hyaline to pale green; with 6–11 septa; 300–480 µm long, 5.5–7, 4.1–4.8 and 2.7–3.4 µm wide at the base, in the middle and below the topmost verticil, respectively. Conidiogenous cells borne on short side branches (7–14 x 2.5–3 µm) or directly from the conidiophore (the topmost one or two whorls), in verticils by 3–13; subulate, 13–16(–22) µm long and 2–3 µm wide at the base, attenuating gradually to 0.5–0.7 µm at the tip, sometimes the upper part that bears the denticles swollen; with 1–3(–5) conidiogenous loci, formed on refractive protrusions that are 1–4 µm long, loci sometimes also on intercalary cells. Conidia cylindrical, occasionally slightly obovate or clavate, straight, 8–13 x 5–6 µm, green; 1-septate, sometimes constricted at the septum; cells of equal size or the upper or lower being larger; with a centrally placed basal hilum, hilum < 1 µm wide. Chlamydospores not observed.

Characteristics in culture. Colonies spreading slowly, reaching 25–40 mm diam at 25 C in 7 d; cottony, margin regular to fasciculate; white but becoming green when conidia are formed, reverse not colored or becoming light orange or salmon (5–6 A 3–5) to brownish orange (6 C 4–5). Odor absent. Aerial mycelium scant; hyphae hyaline, 2–3 µm wide, smooth-walled. Submerged mycelium abundant, fasciculate, cells becoming swollen in many places. Conidiophores arising from aerial hyphae, not differentiated from these, unbranched or branching irregular, near the top giving single or opposite side branches that bear conidiogenous cells. Conidiogenous cells 3–7 in a verticil; subulate, 14–30 µm long, attenuating gradually from 2–2.5(–3) µm at the base to 0.5–1 µm at the tip; with 1–2(–3) conidiogenous loci, formed on 1–3 µm long refractive protrusions, loci sometimes formed on intercalary cells, each locus forming one conidium. Conidia ellipsoidal to cylindrical, sometimes clavate, straight, (8–)10–13(–15) x (4.5–)5–6(–6.5) µm, green, 1-septate, not or slightly constricted at the septum, with a protuberant central basal hilum, 1–1.2 µm wide; produced singly and symmetrically at the tips of conidiogenous cells. Chlamydospore-like cells abundant, intercalary on submerged hyphae, in chains of a few or in long chains of ca 20 cells that may extend also above the agar surface, sometimes forming terminally at the ends of hyphae (or few in a lateral position); cells subglobose to ellipsoidal, 8–16 x 7.5–11 µm, hyaline, wall 0.5–1 µm thick.

HOLOTYPE. U.S.A. MARYLAND: Howard County, Patapsco Valley State Park, near River Road, on Xylobolus frustulatus (Pers. : Fr.) Boidin, 12 Oct 1999, K. Põldmaa, TAA 170300, isotype BPI 747898; culture TFC 99–152, CBS 110280. PARATYPE. U.S.A. PENNSYLVANIA: Westmoreland County, E edge of Laughlintown, on X. frustulatus, 17 Sep 2000, K. Põldmaa, TAA 170546, culture TFC 00–89, CBS 110321.

Additional specimens examined in which only the anamorph was present. U.S.A., MARYLAND: Anne Arundel County, Patuxent Wildlife Refuge, central part, SW part of River Road, on X. frustulatus, 2 Jul 2000, K. Põldmaa, TAA 170416, TFC 00–53; Prince George's County, Greenbelt Park, X. frustulatus, 9 Jul 2000, K. Põldmaa, TAA 170429, BPI 841673, TFC 00–65; Anne Arundel County, Patuxent Wildlife Refuge, central part, near Duvall Bridge, on X. subpileatus, 11 Sep 2000, K. Põldmaa, TAA 170520.

Hypomyces xyloboli is one of the few species in the genus that has green conidia. However, it is distinguished easily from the other two species in the group that share this feature. In particular, H. viridigriseus K. Põldmaa & Samuels (Põldmaa et al 1997Citation) forms 1–3-septate conidia on long sympodially proliferating conidiogenous branches while C. virescens G. Arnold (Arnold 1987Citation) produces purple pigments in culture. Although the green color is the only character distinguishing the anamorph of H. xyloboli from the other two species dealt with here, its perithecial color and ascospores are characteristic. The brown perithecia of H. xyloboli are among the darkest found in the genus, but the perithecial wall anatomy is similar to that of H. sympodiophorus and H. thailandicus.


   DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 TAXONOMY
 DISCUSSION
LITERATURE CITED
 
Hypomyces sympodiophorus and the two new species, H. thailandicus and H. xyloboli, that have been found exclusively on basidiomata of Stereaceae share several morphological features absent in the rest of aphyllophoricolous Hypomyces. These three species possess a scant subiculum that often is lacking, two-layered perithecial wall and conidiogenous cells with refractive areas below the loci. Only H. albidus and C. gamsii are known to have similar conidiogenous cells. Hypomyces on Stereaceae represent the only members of the genus that cannot be identified on the basis of the characters of the anamorphs alone. In Hypomyces the perithecial phenotype typically is conserved with the anamorph phenotype being more diagnostic of a species. Apart from the green conidia of H. xyloboli and the somewhat larger conidia of H. thailandicus, anamorphs of the three species and C. gamsii are indistinguishable. The most reliable characters for distinguishing among their teleomorphs are the ascospore morphology and the color of perithecia. The four species also differ in the fungi on which they occur, but additional collections are needed to determine whether their host ranges, indeed, are mutually exclusive.

The morphological similarity and the close relatedness of hosts of Hypomyces species found only on members of Stereaceae suggest that they would represent a monophyletic group. Unfortunately, the LSU rDNA sequences analyzed proved to be inadequate for testing this hypothesis. Although H. sympodiophorus and the two xylobolicolous species fall into different poorly supported larger clades, their monophyly is not rejected in the majority of the constrained topologies. Additional genes need to be analyzed to understand whether the morphological data are misleading due to convergence or retention of ancestral states.

When considering the 10 species of Hypomyces and two of Cladobotryum that have been reported on Stereaceae, they are distributed among all major clades of aphyllophoricolous Hypomyces, with the majority being part of the largest clade of KOH-negative species. Moreover, two species of Sphaerostilbella, nested in the Hypomyces-Hypocrea clade also grow on Stereum spp. The ancestral hypocreaceous species thus could have been generalists with species of Stereum and their close relatives among their hosts. However, it equally is possible that the ancestral Hypomyces inhabited nonstereoid hosts and that there have been several independent events of colonizing stereoid taxa, as well as of developing restricted host specificity. Obviously, additional characters, including other loci, are to be studied to disprove erroneous scenarios of the evolution of host preference in the fungicolous Hypocreaceae.


   ACKNOWLEDGMENTS
 
My sincere thanks to Dr. Gary Samuels for improving the text and making the microtome sections of perithecia, to Prof. Erast Parmasto for identifying the hosts and revising the manuscript and to Dr. Walter Gams who corrected the Latin descriptions. I am grateful to Dr. Lisa Castlebury for the opportunity to work under her supervision in the molecular lab at the Systematic Botany and Mycology Laboratory, Beltsville, and to Dr. David Farr for sharing his great expertise in digital imaging. Dr. Sarah Dodd also gave valuable hints for the work with DNA. Samuels and Jacques Fournier are thanked for providing specimens, and Dr. Wendy Untereiner is thanked for her many editorial suggestions. This study was supported in part by the United States National Science Foundation (PEET) Grant 9712308, "Monographic Studies of Hypocrealean Fungi: Hypocrea and Hypomyces" to the Pennsylvania State University Department of Plant Pathology and the Estonian Science Foundation Grant No. 4994.


   FOOTNOTES
 
1 kadri{at}zbi.ee Back

Accepted for publication March 26, 2003.


   LITERATURE CITED
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 TAXONOMY
 DISCUSSION
 LITERATURE CITED
 
Arnold GRW., 1970 Über Sibirina und Sympodiophora, zwei neue Gattungen der Moniliales. Nova Hedwigia 19:299-304

Arnold GRW., 1976 Internationale Bibliographie der Hypomycetaceae. Jena: Bibliographische Mitteilungen der Universitätsbibliothek Jena No 25. 129 p

Arnold GRW., 1987 Beitrag zur Kenntnis der Pilze Kubas. III. Feddes Repertorium 98:351-355

Chamuris GP., 1988 The nonstipitate stereoid fungi in the northeastern United States and adjacent Canada. Mycologia Memoir 14. Berlin, Stuttgart: J. Cramer. 247 p

Eriksson T., 1997 AutoDecay. Version 4.0. Distributed by the author. Department of Botany, University of Stockholm, Sweden

Gams W., 1973 Phialides with solitary conidia? Remarks on conidium ontogeny in some hyphomycetes. Persoonia 7:161-169

Gray DJ, Morgan-Jones G., 1980 Notes on Hyphomycetes. XXXIV. Some mycoparasitic species. Mycotaxon 10:375-404

Hibbett DS, Thorn RG., 2001 Basidiomycota: Homobasidiomycetes. In: McLaughlin DJ, McLaughlin EG, Lemke PA, eds. The mycota. Vol. VII. Systematics and evolution. Berlin, Germany: Springer-Verlag. p 121–168

Kirk PM, Cannon PF, David JC, Stalpers JA., 2001 Ainsworth & Bisby's dictionary of the fungi. 9th ed. Great Britain: CAB International. 655 p

Kornerup A, Wanscher JH., 1967 Methuen handbook of colour. 2nd ed. London: Methuen & Co Ltd. 243 p, 30 pl

Larsson E, Larsson K-H., 2003 Phylogenetic relationships of russuloid basidiomycetes with emphasis on aphyllophoralean taxa. Mycologia (in press)

Nicholas KB, Nicholas HBJr, Deerfield DWII., 1997 Genedoc: analysis and visualization of genetic variation. EMBNEW.NEWS 4:14

Põldmaa K., 1999 The genus Hypomyces and allied fungicolous fungi in Estonia. I. Species growing on aphyllophoralean basidiomycetes. Folia Cryptogamica Estonica 34:15-31

Põldmaa K., 2000 Generic delimitation of the fungicolous Hypocreaceae. Stud Mycol 45:83-94

Põldmaa K., Larsson E, Kõljalg U., 1999 Phylogenetic relationships in Hypomyces and allied genera, with emphasis on species growing on wood-decaying homobasidiomycetes. Can J Bot 77:1756-1768

Põldmaa K., Samuels GJ., 1999 Aphyllophoricolous species of Hypomyces with KOH-negative perithecia. Mycologia 91:277-299

Põldmaa K., Samuels GJ., Lodge DJ., 1997 Three new polyporicolous species of Hypomyces and their Cladobotryum anamorphs. Sydowia 49:80-93

Rehner SA, Samuels GJ., 1994 Taxonomy and phylogeny of Gliocladium analyzed from nuclear large subunit ribosomal DNA sequences. Mycol Res 98:625-634

Rehner SA, Samuels GJ., 1995 Molecular systematics of the Hypocreales: a teleomorph gene phylogeny and the status of their anamorphs. Canad J Bot 73:S816-S823

Rogerson CT, Samuels GJ., 1993 Polyporicolous species of Hypomyces. Mycologia 85:231-272

Samuels GJ., 1976 Revision of the Fungi Formerly Classified as Nectria Subgenus Hyphonectria. Mem New York Bot Gard 26:1-119

Samuels GJ., Candoussau F, Magni J-F., 1997 Fungicolous pyrenomycetes 2. Ascocodinaea, gen. nov., and reconsideration of Litschaueria. Mycologia 89:156-162

Seifert KA., 1985 A monograph of Stilbella and some allied hyphomycetes. Stud. Mycol 27:1-235

Swofford DL., 1999 PAUP* 4.0 phylogenetic analysis using parsimony (*and other methods), version 4.0b10. Sunderland, Massachusetts: Sinauer Associates, Inc

Vilgalys R, Hester M., 1990 Rapid genetic identification and mapping of enzymatically amplified ribosomal DNA from several Cryptococcus species. J Bacteriol 172:4238-4246[Abstract/Free Full Text]

Wu S-H, Hibbett DS, Binder M., 2001 Phylogenetic analyses of Aleurodiscus s.l. and allied genera. Mycologia 93:720-731





This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Services
Right arrow Similar articles in this journal
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Põldmaa, K.
Right arrow Search for Related Content
PubMed
Right arrow Articles by Põldmaa, K.
Agricola
Right arrow Articles by Põldmaa, K.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS