Chapter 105

Bulleromyces Boekhout & A. Fonseca (1991)

Teun Boekhout

DIAGNOSIS OF THE GENUS

Anamorph: Bullera.Asexual reproduction: Yeast cells are ellipsoid, subglobose or cylindric, and monokaryotic. Budding is polar, with buds emerging enteroblasti-cally, sessile or on short denticles and with percurrent or sympodial proliferation. Ballistoconidia are rotationally symmetrical and subglobose.Colonies are cream, smooth, butyrous or mucoid.Sexual reproduction: The teleomorphic state may be heterothallic or self-sporulating. Basidiocarps are absent. Dikaryotic hyphae are regularlybranched, septate, with clamp connections and haustorial branches. Hyphal septa have dolipores with cupulate (namely, Tremellales type)parenthesomes (Fig. 105.1). Basidia are subglobose, clavate or ovoid, and after karyogamy become longitudinally, obliquely or transversely sep-tate with two to four cells. Germination of basidia occurs by yeast cells or hyphae on which yeast cells or ballistospores originate.Physiology/biochemistry: Fermentation is absent. D-Glucuronate and myo-inositol are assimilated, but nitrate is not. Starch-like compounds areproduced. Diazonium blue B and urease reactions are positive. Coenzyme Q-10 is formed. Xylose is present in whole-cell hydrolysates.Phylogenetic placement: Tremellales, Tremellomycetes, Agaricomycotina, Basidiomycota (Fig. 105.2).

TYPE SPECIES

Bulleromyces albus Boekhout & Á. Fonseca

SPECIES ACCEPTED

1. Bulleromyces albus Boekhout & Á. Fonseca (1991)

FIGURE 105.1 Bulleromyces albus CBS 5003CBS 7440. TEM image of a hyphal septum showing a dolipore with cupulate parenthesomes(Tremellales-type) occurring on both sides of the pore channel. Note the presence of electron-dense banding material in the center of thepore channel and just outside it, as well as the electron-dense fibrillar material connecting the parenthesomes and the electron-dense bandscovering the dolipore. Bar5500 nm.

1391The Yeasts, a Taxonomic Study© 2011 Elsevier B.V. All rights reserved.

SYSTEMATIC DISCUSSION OF THE SPECIES

105.1. Bulleromyces albus Boekhout &A. Fonseca (Boekhout et al. 1991a)

Anamorph:

Bullera alba (Hanna) Derx.

Synonyms:

Sporobolomyces albus Hanna (Bisby et al. 1929)Bullera alba (Hanna) Derx (1930)

Growth of yeast cells on 5% malt extract agar: After 7 days at 17�C,colonies are mucoid, flat or slightly raised, smooth, dull or shiny, cream tobrownish-cream, and have an entire margin. Yeast cells are ellipsoid,ovoid or subglobose, 5�1133�6 μm, and single. Budding is polar andbuds are sessile or occur on short denticles (Fig. 105.3). Cells may formhyphal outgrowths of up to ca. 80 μm long and 2�5 μmwide.Growth on the surface of 2% glucose in yeast nitrogen base: After1 week at 25�C, a thin film, islets, a ring and sediment are formed.Cells are ellipsoid, fusoid to cylindrical, 7�1033�4 μm, with polarbudding showing sympodial proliferation. Cells may adhere in shortchains. Inflated cells, 9�11 μm in diameter, occur as well, and mayform endospores.

Rhodosporidium diobovatum CBS 6085T / AF0704210.1

100

87

9988

84

70

76100

100

10058

76 100

9089

10084

69

99

10099

61

90

709788

50

Cryptococcus albidus CBS 142T / AF075474Holtermannia corniformis CBS 6979 / AF189843

Cryptococcus watticus CBS 9496T / AY138478

Cryptococcus festucosus CBS 10162T / AY462119Cryptococcus nyarrowii CBS 8804T / AY006480Cryptococcus mycelialis CBS 7712T / AJ311450

Bulleribasidium oberjochense PYCC 5741T / AF416646

Sirobasidium magnum CCJ1289 / AF042240Trimorphomyces papilionaceus CBS 444.92 / AF416645

Tremella giraffa CCJ1553 / AF042271

Tremella foliacea CBS 6969 / AF189868

Tremella neofoliacea CCJ1401 / AF042236

Tremella simplex FO31782 / AF042246Tremella mycophaga RB6539-4 / AF042249

Sirobasidium intermedium CBS 7805 / AF075492

Tremella exigua RB6623-15 / AF042248

Tremella indecorata HBZ 194 / AF042250Tremella nivalis CCJ1192 / AF042232

Tremella moriformis CBS 7810 / AF075493

Tremella resupinata CBS 8488 / AF042239Tremella cinnabarina CBS 8237 / AF189866

Tremella flava CBS 8471 / AF042221

Tremella fuciformis CBS 6970 / AF075476Tremella globispora CBS 6972 / AF189869

Tremella taiwanensis CCJ1151 / AF042230

Tremella brasiliensis CBS 6966 / AF189864Tremella tropica CBS 8483 / AF042251

Tremella coalescens CBS 6967 / AF189865

Tremella mesenterica CCJ1543 / AF042252

Bulleromyces albus CBS 500T / AF416643Papiliotrema bandonii PYCC 5743T / AF416642

Auriculibuller fuscus PYCC 5690T / AF444762

Tremella encephala CBS 6968 / AF189867Tremella microspora BPI702328 / AF042253Tremella aurantia CBS 6965 / AF189842

Cuniculitrema polymorpha PYCC 5647 / AY032662

Filobasidiella neoformans CBS 132T / AF075484

Fibulobasidium murrhardtense PYCC 5744T / AF416648

Fibulobasidium inconspicuum CBS 8238 / AF416649Fibulobasidium sirobasidioides RJB 12787 / AF416644

FIGURE 105.2 Phylogenetic tree based on neighbor-joining analysis of D1/D2 LSU rRNA gene sequences showing the position of Bulleromycesalbus among teleomorphic Tremellales. Figures on the branches indicate bootstrap values (.50%) after 1000 replicates.

1392 PART | VB Descriptions of Teleomorphic Basidiomycetous Genera and Species

Dalmau plate culture on morphology agar: After 7 days at 17�C,hyphae, ca. 100 μm long and 2�4 μm wide, may be present.Subglobose, somewhat thick-walled cells occur with the outer cell-wall layer peeling off. Aerobic growth is cream, pale yellowish orpale yellowish-brown, shiny, mucoid, smooth or slightly venose, flat,and the margin is entire, straight or crenulate.Formation of ballistoconidia: Ballistoconidia form on corn mealagar and malt extract agar and are ellipsoid to subglobose,4�9.533.5�7 μm (Fig. 105.4).Formation of basidiospores: Conjugation occurs after mixing strainsof different mating types on corn meal agar at 17�C or at room tem-perature. Self-sporulating strains also occur. Dikaryotic hyphae aremade up of cells measuring 40�13031.5�3 μm, with clamp connec-tions and haustorial branches. Basidia, which occur laterally and ter-minally on hyphae or hyphal branchlets, measure 5�103 4�7 μm.

Basidia are two to four-celled, due to longitudinal, transverse or obli-que septation (Fig. 105.5). Basidial cells germinate by forming yeastcells, which may represent basidiospores, on short denticles or peg-like structures or with hyphae on which yeast cells or ballistosporesare formed.

Fermentation (17�): Absent.

Growth (in Liquid Media, 17�C)

Glucose 1

Inulin 1

Sucrose 1

Raffinose 1

Melibiose vGalactose 1

Lactose 1

Trehalose 1

Maltose 1

Melezitose 1

Methyl-α-D-glucoside 1

Soluble starch vCellobiose 1

Salicin 1

L-Sorbose vL-Rhamnose 1

D-Xylose 1

L-Arabinose 1

D-Arabinose 1

D-Ribose 1

Methanol 2

Ethanol vGlycerol 1

Erythritol vRibitol vGalactitol vD-Mannitol 1

D-Glucitol 1

myo-Inositol 1

DL-Lactate 1

Succinate 1

Citrate 1

D-Gluconate 1

D-Glucosamine vN-Acetyl-D-glucosamine nHexadecane nNitrate 2

Vitamin-free 2

FIGURE 105.3 Bulleromyces albus CBS 501. SEM image of yeast cellswhen grown on yeast extract peptone glucose agar (YPGA) after5 days, 25�C. Bar55 μm. Figure courtesy of J. Stalpers, CBS.

FIGURE 105.4 Bulleromyces albus CBS 501. SEM image of ballisto-conidia on corn meal agar after 10 days, 25�C. Bar55 μm.Figure courtesy of J. Stalpers, CBS.

FIGURE 105.5 Bulleromyces albus CBS 63023CBS 7441. Dikaryotichyphae with clamp connections and phragmobasidia on corn mealagar after 20 days, 25�C. Note the peg-like outgrowths of the basidialcell which may represent epibasidia that may form yeast-like basi-diospores at their apex. Bar510 μm.

1393Chapter | 105 Bulleromyces Boekhout & A. Fonseca (1991)

Additional Growth Tests and Other Characteristics

2-Keto-D-gluconate 1

50% Glucose 2

Starch formation 1

Urease 1

Nitrite 2

Growth at 25�C 1

Growth at 30�C vGrowth at 37�C 2

CoQ: 10 (Nakase and Suzuki 1986a).Mol% G1C: 53.3�54.4, four strains, CBS 500, CBS 501, CBS 502, CBS6302 (Tm: Boekhout 1991a); 54.5 (Tm: Nakase et al. 1990c); 54.4 (Tm:Nakase and Komagata 1971g).Gene sequence accession numbers, type strain: LSU D1/D25AF416643, ITS5AF444662.Cell carbohydrates: Glucose, mannose, galactose and xylose arepresent (Gorin and Spencer 1970, von Arx and Weijman 1979).Origin of the strains studied: CBS 500 (PYCC4532, JCM 9878, MUCL30302, NRRL Y-5487, UCD-FST 415), type of Sporobolomyces albusHanna, isolated from straw of Hordeum jubatum infected by rust fungi,USA; CBS 501 (ATCC 18568, CCRC 21320, DBVPG 3786, DBPVG 6655,IFO 1192, JCM 2954, MUCL 30301, NCYC 425, NRRL Y-6655), type ofBullera alba Lodder & Kreger-van Rij, isolated from air in a dairy, B.W.Hammer, USA; CBS 502 (JCM 9877), from air in a dairy, USA; CBS 6097,isolated from frass of a beetle (Sinoxylon ruficornis), South Africa; CBS6302 (PYCC 4536, JCM 9879, MUCL 30303, NCYC 2523), from grass,USA; CBS 7440 (PYCC 4539, JCM 9880), from leaf of poplar (Populussp.), Portugal; CBS 7441 (PYCC 4538, JCM 9881, NCYC 2595), from leafof poplar (Populus sp.), Portugal; CBS 7503 (PYCC 4560, JCM 9882),from leaf of walnut (Juglans sp.), Portugal (Boekhout et al. 1991a).Complementary mating types: CBS 500 (mt A), CBS 7441 (mt A),CBS 7503 (mt A), CBS 501 (mt B), CBS 6097 (mt B), CBS 6302 (mt B)(Boekhout et al. 1991a).Type strain: CBS 5003CBS 6302 (preserved as dried material in theherbarium at CBS. Both complementary mating type strains are pre-served as living cultures at CBS, see above).Systematics: Based on LSU D1/D2 rRNA gene sequence analysis,Bulleromyces albus belongs to the Bulleromyces lineage of theTremellales branch of the Tremellomycetes, Agaricomycotina (Fell et al.2000, Hibbett et al. 2007, Scorzetti et al. 2002), with Bullera unica as itsclosest relative (100% bootstrap support). Other species that maybelong to this lineage are Tremella moriformis, T. nivalis, T. indecorataand Cryptococcus species CBS 8363 (Scorzetti et al. 2002). Using ITSsequences, B. albus and Bullera unica are also closely related (100%bootstrap support), but the three above mentioned Tremella speciesseem more distantly related (Fell et al. 2000, Scorzetti et al. 2002).Bullera variabilis, which also forms dikaryotic hyphae with clamp con-nections upon mating, is only distantly related, and clusters weaklywith the luteolus clade when using both D1/D2 and ITS sequences (Fellet al. 2000, Scorzetti et al. 2002). However, using Bayesian analysis ofthe LSU D1/D2 domains, B. albus, Bullera unica, Tremella moriformis,T. nivalis and T. indecorata formed a well supported cluster withCryptococcus laurentii as a basal lineage (Sampaio et al. 2004a).Ecology: The teleomorphic state of Bulleromyces albus has not yetbeen observed to occur in nature. In contrast, the anamorph, Bulleraalba, occurs widely on plant leaf surfaces and is considered an impor-tant phyllosphere-inhabiting yeast. This species has also been isolated

from air and insect frass, and is known too from Japan (Nakase andSuzuki 1985c, 1987c), New Zealand (Hamamoto and Nakase 1996),Canada, Portugal, S. Africa and the USA (Boekhout et al. 1991a).Biotechnology: Bulleromyces albus (cited as Bullera alba) was foundto contain beta-carotenoids, gamma-carotenoids and torulene(Fiasson 1972).Agriculture and food: Unknown.Clinical importance: Unknown.

COMMENTS ON THE GENUS

CBS 500, originally isolated by Hanna (in Bisby 1929), is the originaltype strain of Sporobolomyces albus, the basionym of Bullera alba andanamorph of B. albus. Lodder and Kreger-van Rij (1952), however,designated strain No 16, isolated by B.W. Hammer (5CBS 501) as theneotype for Bullera alba. As the original isolate of Sporobolomycesalbus made by Hanna, namely CBS 500, is available for comparison, itseems preferred to consider this latter isolate to represent the typestrain of Bullera alba.

Bandoni (1987) reported the occurrence of mating and formationof dikaryotic hyphae with clamp connections in Bullera. Boekhoutet al. (1991a) observed the formation of Tremella-type phragmobasi-dia after mating strains of Bullera alba. Bullera alba var. lactis wasfound to be conspecific with Bullera sinensis, and was reclassified asBullera sinensis var. lactis (Bai et al. 2001b, Boekhout 1991a, Nakaseet al. 1990c). This taxonomic decision was supported by the lack ofmating between isolates of B. sinensis var. lactis and Bullera alba, theanamorph of B. albus. The suggested relationship of Bullera withthe Tremellales (Boekhout et al. 1991a, Phaff 1970d) is supportedby the septal pore structure, which is a dolipore with cupulate par-enthesomes (Fig. 105.1), the basidial morphology (Fig. 105.5) andmolecular phylogeny (Fig. 105.2).

Three teleomorphic genera are presently known for the anamor-phic genus Bullera, namely Bulleromyces, Bulleribasidium andAuriculibuller (Boekhout et al. 1991a, Sampaio et al. 2002, 2004a).Bulleromyces has cruciate-septate basidia, and those of Auriculibullerand Bulleribasidium are auricularioid (Sampaio et al. 2002, 2004a).Based on a Bayesian analysis of the LSU D1/D2 domains, B. albusbelongs to a cluster comprising teleomorph taxa, such as T. moriformisT. nivalis, T. indecorata, and the anamorph Bullera unica (Sampaioet al. 2004a). Auriculibuller fuscus forms a well supported lineagewith Bullera japonica, Cryptococcus flavescens, C. aureus andCryptococcus species CBS 8372 and CBS 8358 (Sampaio et al. 2004a).Bulleribasidium oberjochense forms a supported lineage with Bulleravariabilis and Bullera miyaginana (Sampaio et al. 2004a). Interestingly,formation of dikaryotic hyphae and probasidium-like cells wasobserved among strains CBS 7347 and CBS 7367 of Bullera variabilis(Boekhout et al. 1991a), but no development of basidia was observed.The distant position between Bulleromyces albus and Bullera variabiliswas supported by SSU rRNA gene sequences (Bai et al. 2001b). Fromthe foregoing, it is clear that additional phylogenetic studies areneeded to clarify the taxonomic relationships of the speciesdiscussed.

1394 PART | VB Descriptions of Teleomorphic Basidiomycetous Genera and Species