Antonie van Leeuwenhoek 41 (1975) 361-365
Trichosporon terrestre J. P.
361
sp. nov.
VAN DER W A L T A N D E L Z B I E T A J O H A N N S E N
Microbiology Research Group, Council for Scientific and hMustrial Research, Pretoria, South Africa
VAN DER WALT, J. P. and JOHANNSEN, E. 1975. Trichosporon terrestre sp. nov. A n t o n i e van L e e u w e n h o e k 41: 361-365. A single strain representative o f an undescribed Trichosporon species, was recovered f r o m soil. The cell wall of the strain corresponds in structure to cell walls o f the ascomycetous yeasts. On the g r o u n d o f X-ray inactivation data the strain is assumed to be diploid. A description o f the new species is given.
A single strain of an undescribed Trichosporon species was recovered f r o m a grassland soil sample collected in the vicinity o f Pretoria. Th e following description o f the new species is based on the standard techniques e m p l o y e d in yeast t a x o n o m y . Triehosporon terrestre van der Walt et J o h a n n s e n sp. nov. A natura isolata in forma cellularum diploidearum. In medio liquido cure glucoso et peptono et extracto levendinis cellulae zymoticae ovoidiae ellipsoidales vel cylindricae, (2.5-6.5) >~, (3.0-11.0) p.m, singulae, binae aut in racemis parvis. Mycelium verum (2.0-3.5) ~.tm in diameter, pseudomycelium et arthrosporae. Sedimentum floccosum et annulus incohatus formantur. Post unum mensem in aere officinae sedimentum floccosumet annulus incohatus. Cultura in agaro cure glucoso et peptono et extracto Ievendinis post unum mensem in aere officinae mollis, non tenax, eburna, restricta, elevata, crispulata non nitida. Margo Iobatus. Oriuntur endosporae asexuales per fissionem endoplasmicas. In agaro Zea mai's post dies 7 pseudomycelium, mycelium verum et arthrosporae abunde formanmr. Ultrastructura parietis sicut in fermentis ascomycetoideis. Fermentatio nulla. Glucosum, galactosum, L-sorbosum, sucrosum, maltosum, cellobiosum (lente), trehalosum, raffinosum, D-xylosum, L-arabinosum, D-axabinosum (lente), D-ribosum (lente), L-rhamnosum, ethanolum, glycerolum, i-erythritolum, ribitolum, galactitolum (lente), D-glucitolum, D-mannitolum, 0c-methyI-D-glucosidum(lente), salicinum (lente), DL-acidum lacticum, acidum succinicum, acidum citricum et i-inositolum assimilantur at non lactosum, melibiosum, melezitosum, inulinum nec amylum solubile. Arbutinum lentissime finditur. Nitras kalii assimilatur. Ad crescentiam vitaminae externae non necessariae sunt.
362
J . P . VAN DER WALT AND E. JOHANNSEN
Crescere potest in agaro extracto fermenti qui quinquaginta partes glucosi per centum pondere continent. Crescere potest in 37 C. Materia amyloidea iodophila non formatur. Ureurrt non finditur. Isolata ex terra inculta ab territorio Pretoriensi. Typus: cultura 6697 in collectione zymotica Centraalbureau voor Schimmelcultures, Delphi Batavorum. G r o w t h in glucose - yeast extract - p e p t o n e water: After 3 d a y s at 28 C true mycelium, (2.0-3.5) ixm in diameter, a r t h r o s p o r e s a n d p s e u d o m y c e l i u m are a b u n d a n t l y formed. The b u d d i n g yeast cells are ovoid, ellipsoidal to cylindrical, m e a s u r i n g (2.5-6.5) • (3.0-11.0) ~zm, a n d occur singly, in pairs or small clusters (Fig. 1A). A floccose sediment and i n c o m p l e t e ring are formed. After 1 m o n t h at r o o m t e m p e r a t u r e a floccose sediment and i n c o m p l e t e ring are present. G r o w t h on glucose - yeast e x t r a c t - p e p t o n e agar: After 1 m o n t h at r o o m temperature the streak culture is soft, non-tenacious, c r e a m - c o l o u r e d , crispulate, restricted, raised and s o m e w h a t dull. The m a r g i n is lobate. D a l m a u plate cultures on corn meal a g a r : P s e u d o m y c e l i u m , true m y c e l i u m and a r t h r o s p o r e s are a b u n d a n t l y f o r m e d (Fig. 1B).
6)
Fig. 1. Trichosporon terrestre. A. In glucose - yeast extract - peptone water (3 days at 28 C). B. On corn meal agar (7 days at 28 C).
TRICHOSPORON TERRESTRE SP. NOV.
363
F o r m a t i o n of endospores : Asexual endospores formed by endoplasmic cleavage, were observed in old cultures on M Y agar. Fermentation : Absent. Assimilation of carbon c o m p o u n d s : Glucose + D-Ribose + (slow) Galactose + L-Rhamnose + L-Sorbose + Ethanol + Sucrose + Glycerol + Maltose + Erythritol + Cellobiose + (slow) Ribitol + Trehalose + Galactitol + (slow) Lactose -D-Glucitol + Melibiose -o-Mannitol + Raffinose + tz-Methyl-D-glucoside + (slow) Melezitose -Salicin + (slow) Inulin -DL-Lactic acid + Soluble starch (Merck's) -Succinic acid + D-Xylose + Citric acid + L-Arabinose + i-Inositol + D-Arabinose + (slow) Splitting of arbutin : Positive, but very slow. Assimilation of potassium nitrate: Positive. G r o w t h in vitamin-free medium: Positive. G r o w t h on 50 ~o (w/w) glucose-yeast extract agar: Positive. G r o w t h at 37 C: Positive. F o r m a t i o n o f amyloid material: Absent. Hydrolysis of urea: Absent. Cell wall structure by electron microscopy: G r o w t h from a 7 day old culture on corn meal agar was fixed for 6 hr at 4 C in 5 (v/v) ~o glutaraldehyde in 0.1M phosphate buffer (pH 7.3) and then postfixed for 2 hr in 1 ~ osmium tetroxide in the same buffer, also at 4 C. Ultra-thin sections o f the material imbedded in an A r a l d i t e - E p o n mixture were cut with a d i a m o n d knife and stained by consecutive treatments with saturated aqueous uranyl acetate and Reynold's lead citrate solution. Sections through the cells showed the cell wall to be of the ascomycetous type, consisting of a rather broad, light inner layer and a thin, dark outer layer (Fig. 2). Ploidy determination by X-ray inactivation: The ploidy o f the strain was determined according to the procedure described by Beam et al. (1954), using a suspension o f a 4-day-old culture on M Y agar. The X-ray source (Siemens Dermipan) was operated at 50 K V and 25 m A the dose rate being 340 R per second with the material 100 m m f r o m the tube target. The survival curve of the culture which has an LDso o f 6 . l K R , is shown in Fig. 3. As the linear portion of this curve, when extrapolated to zero, does not intercept at the origin, the strain is presumed to be diploid.
364
J . P . VAN DER WALT AND E. JOHANNSEN
Fig. 2. Tr. terrestre grown on corn meal agar. Electron micrograph of a section through a hyphal element The wall consists of a broad, light inner layer and a thin, dark outer layer (34000 •
DOSE
KILOROENTGENS io
5
~00 90
\\ \\
80 7O .J
X 6o
> riD
O3 50 FZ L~ L~ OC
30
i I0
i 2O
SECONDS
Fig. 3. Tr. terrestre. X-ray survival curve of a 4-day-old cult'ure on MY agar. The linear portion of this curve, when extrapolated to zero does not intercept at the origin. H a b i t a t : O n e strain w a s isolated f r o m grassland soil collected in the Pretoria district. T y p e : T h e strain m a i n t a i n e d as N o . 6697 in the Y e a s t C o l l e c t i o n o f the C e n t r a a l b u r e a u v o o r S c h i m m e l c u l t u r e s in D e l f t is the type for the species.
TRICHOSPORON TERRESTRE SP. NOV.
365
Etymology: terrestre L. adj., pertaining to the soil; referring to the species' occurrence in soil. DISCUSSION Although on the basis of their ability to utilize i-inositol and nitrate there is superficial agreement between Tr. terrestre and Tr. pulhdans, the two taxa are nevertheless distinguished by differences in their abilities to utilize lactose, melibiose and melezitose and to grow at 37 C (cf. do C a r m o Sousa, 1970). However, the cardinal criterion which differentiates these two species is the difference in the ultrastructure of their cell walls as observed by electron microscopy. Whereas the ceil wall of Tr. pulhdans is reported to be similar in structure to the cell walls of the yeast genera of the saprobic Hemibasidiomycetes (Kreger-van Rij and Veenhuis, 1971 ; Donk, 1973), that o f Tr. terrestre corresponds in structure to the cell walls of the ascomycetous yeasts. The data on the ploidy of CBS 6697 excludes the possibility that the type for 7"1". terrestre is representative of a haploid mating type.
The authors thank Miss M a u d y Th. Smith o f the Yeast Division o f the Centraalbureau voor Schimmelcultures for checking the new species, M r N. V. D. W. Liebenberg for his services at the electron microscope and Mrs Wilhelmina C. Hiemstra for preparing Figure 1. Received 18 February 1975.
REFERENCES BEAM,C. A., MORTIMER,R. K., WOLFE,R. O. and TOBIAS,C. A. 1954. The relation of radioresistance to budding in Saccharomyces cerevisiae. - - Arch. Biochem. Biophys. 49:110-122. do CARMOSOLISA,L. 1970. The genus Trichosporon Behrend, p. 1309-1352. In J. Lodder, (ed.), The Yeasts, 2nd Ed. - - North Holland Publ. Co., Amsterdam. DONK, M. A. 1973. The Heterobasidiomycetes: a reconnaissance IV. - - Kon. Ned. Akad. Wetensch. Proc, (Ser. C) 76: 109-125. KREGER-VAN RIJ, N. J. W. and VEENHUIS, M. 1971. A comparative study of the cell wall structure of basidiomycetous and related yeasts. - - J . Gen. Microbiol. 68: 87-95.
Trichosporon terrestre J. P.
361
sp. nov.
VAN DER W A L T A N D E L Z B I E T A J O H A N N S E N
Microbiology Research Group, Council for Scientific and hMustrial Research, Pretoria, South Africa
VAN DER WALT, J. P. and JOHANNSEN, E. 1975. Trichosporon terrestre sp. nov. A n t o n i e van L e e u w e n h o e k 41: 361-365. A single strain representative o f an undescribed Trichosporon species, was recovered f r o m soil. The cell wall of the strain corresponds in structure to cell walls o f the ascomycetous yeasts. On the g r o u n d o f X-ray inactivation data the strain is assumed to be diploid. A description o f the new species is given.
A single strain of an undescribed Trichosporon species was recovered f r o m a grassland soil sample collected in the vicinity o f Pretoria. Th e following description o f the new species is based on the standard techniques e m p l o y e d in yeast t a x o n o m y . Triehosporon terrestre van der Walt et J o h a n n s e n sp. nov. A natura isolata in forma cellularum diploidearum. In medio liquido cure glucoso et peptono et extracto levendinis cellulae zymoticae ovoidiae ellipsoidales vel cylindricae, (2.5-6.5) >~, (3.0-11.0) p.m, singulae, binae aut in racemis parvis. Mycelium verum (2.0-3.5) ~.tm in diameter, pseudomycelium et arthrosporae. Sedimentum floccosum et annulus incohatus formantur. Post unum mensem in aere officinae sedimentum floccosumet annulus incohatus. Cultura in agaro cure glucoso et peptono et extracto Ievendinis post unum mensem in aere officinae mollis, non tenax, eburna, restricta, elevata, crispulata non nitida. Margo Iobatus. Oriuntur endosporae asexuales per fissionem endoplasmicas. In agaro Zea mai's post dies 7 pseudomycelium, mycelium verum et arthrosporae abunde formanmr. Ultrastructura parietis sicut in fermentis ascomycetoideis. Fermentatio nulla. Glucosum, galactosum, L-sorbosum, sucrosum, maltosum, cellobiosum (lente), trehalosum, raffinosum, D-xylosum, L-arabinosum, D-axabinosum (lente), D-ribosum (lente), L-rhamnosum, ethanolum, glycerolum, i-erythritolum, ribitolum, galactitolum (lente), D-glucitolum, D-mannitolum, 0c-methyI-D-glucosidum(lente), salicinum (lente), DL-acidum lacticum, acidum succinicum, acidum citricum et i-inositolum assimilantur at non lactosum, melibiosum, melezitosum, inulinum nec amylum solubile. Arbutinum lentissime finditur. Nitras kalii assimilatur. Ad crescentiam vitaminae externae non necessariae sunt.
362
J . P . VAN DER WALT AND E. JOHANNSEN
Crescere potest in agaro extracto fermenti qui quinquaginta partes glucosi per centum pondere continent. Crescere potest in 37 C. Materia amyloidea iodophila non formatur. Ureurrt non finditur. Isolata ex terra inculta ab territorio Pretoriensi. Typus: cultura 6697 in collectione zymotica Centraalbureau voor Schimmelcultures, Delphi Batavorum. G r o w t h in glucose - yeast extract - p e p t o n e water: After 3 d a y s at 28 C true mycelium, (2.0-3.5) ixm in diameter, a r t h r o s p o r e s a n d p s e u d o m y c e l i u m are a b u n d a n t l y formed. The b u d d i n g yeast cells are ovoid, ellipsoidal to cylindrical, m e a s u r i n g (2.5-6.5) • (3.0-11.0) ~zm, a n d occur singly, in pairs or small clusters (Fig. 1A). A floccose sediment and i n c o m p l e t e ring are formed. After 1 m o n t h at r o o m t e m p e r a t u r e a floccose sediment and i n c o m p l e t e ring are present. G r o w t h on glucose - yeast e x t r a c t - p e p t o n e agar: After 1 m o n t h at r o o m temperature the streak culture is soft, non-tenacious, c r e a m - c o l o u r e d , crispulate, restricted, raised and s o m e w h a t dull. The m a r g i n is lobate. D a l m a u plate cultures on corn meal a g a r : P s e u d o m y c e l i u m , true m y c e l i u m and a r t h r o s p o r e s are a b u n d a n t l y f o r m e d (Fig. 1B).
6)
Fig. 1. Trichosporon terrestre. A. In glucose - yeast extract - peptone water (3 days at 28 C). B. On corn meal agar (7 days at 28 C).
TRICHOSPORON TERRESTRE SP. NOV.
363
F o r m a t i o n of endospores : Asexual endospores formed by endoplasmic cleavage, were observed in old cultures on M Y agar. Fermentation : Absent. Assimilation of carbon c o m p o u n d s : Glucose + D-Ribose + (slow) Galactose + L-Rhamnose + L-Sorbose + Ethanol + Sucrose + Glycerol + Maltose + Erythritol + Cellobiose + (slow) Ribitol + Trehalose + Galactitol + (slow) Lactose -D-Glucitol + Melibiose -o-Mannitol + Raffinose + tz-Methyl-D-glucoside + (slow) Melezitose -Salicin + (slow) Inulin -DL-Lactic acid + Soluble starch (Merck's) -Succinic acid + D-Xylose + Citric acid + L-Arabinose + i-Inositol + D-Arabinose + (slow) Splitting of arbutin : Positive, but very slow. Assimilation of potassium nitrate: Positive. G r o w t h in vitamin-free medium: Positive. G r o w t h on 50 ~o (w/w) glucose-yeast extract agar: Positive. G r o w t h at 37 C: Positive. F o r m a t i o n o f amyloid material: Absent. Hydrolysis of urea: Absent. Cell wall structure by electron microscopy: G r o w t h from a 7 day old culture on corn meal agar was fixed for 6 hr at 4 C in 5 (v/v) ~o glutaraldehyde in 0.1M phosphate buffer (pH 7.3) and then postfixed for 2 hr in 1 ~ osmium tetroxide in the same buffer, also at 4 C. Ultra-thin sections o f the material imbedded in an A r a l d i t e - E p o n mixture were cut with a d i a m o n d knife and stained by consecutive treatments with saturated aqueous uranyl acetate and Reynold's lead citrate solution. Sections through the cells showed the cell wall to be of the ascomycetous type, consisting of a rather broad, light inner layer and a thin, dark outer layer (Fig. 2). Ploidy determination by X-ray inactivation: The ploidy o f the strain was determined according to the procedure described by Beam et al. (1954), using a suspension o f a 4-day-old culture on M Y agar. The X-ray source (Siemens Dermipan) was operated at 50 K V and 25 m A the dose rate being 340 R per second with the material 100 m m f r o m the tube target. The survival curve of the culture which has an LDso o f 6 . l K R , is shown in Fig. 3. As the linear portion of this curve, when extrapolated to zero, does not intercept at the origin, the strain is presumed to be diploid.
364
J . P . VAN DER WALT AND E. JOHANNSEN
Fig. 2. Tr. terrestre grown on corn meal agar. Electron micrograph of a section through a hyphal element The wall consists of a broad, light inner layer and a thin, dark outer layer (34000 •
DOSE
KILOROENTGENS io
5
~00 90
\\ \\
80 7O .J
X 6o
> riD
O3 50 FZ L~ L~ OC
30
i I0
i 2O
SECONDS
Fig. 3. Tr. terrestre. X-ray survival curve of a 4-day-old cult'ure on MY agar. The linear portion of this curve, when extrapolated to zero does not intercept at the origin. H a b i t a t : O n e strain w a s isolated f r o m grassland soil collected in the Pretoria district. T y p e : T h e strain m a i n t a i n e d as N o . 6697 in the Y e a s t C o l l e c t i o n o f the C e n t r a a l b u r e a u v o o r S c h i m m e l c u l t u r e s in D e l f t is the type for the species.
TRICHOSPORON TERRESTRE SP. NOV.
365
Etymology: terrestre L. adj., pertaining to the soil; referring to the species' occurrence in soil. DISCUSSION Although on the basis of their ability to utilize i-inositol and nitrate there is superficial agreement between Tr. terrestre and Tr. pulhdans, the two taxa are nevertheless distinguished by differences in their abilities to utilize lactose, melibiose and melezitose and to grow at 37 C (cf. do C a r m o Sousa, 1970). However, the cardinal criterion which differentiates these two species is the difference in the ultrastructure of their cell walls as observed by electron microscopy. Whereas the ceil wall of Tr. pulhdans is reported to be similar in structure to the cell walls of the yeast genera of the saprobic Hemibasidiomycetes (Kreger-van Rij and Veenhuis, 1971 ; Donk, 1973), that o f Tr. terrestre corresponds in structure to the cell walls of the ascomycetous yeasts. The data on the ploidy of CBS 6697 excludes the possibility that the type for 7"1". terrestre is representative of a haploid mating type.
The authors thank Miss M a u d y Th. Smith o f the Yeast Division o f the Centraalbureau voor Schimmelcultures for checking the new species, M r N. V. D. W. Liebenberg for his services at the electron microscope and Mrs Wilhelmina C. Hiemstra for preparing Figure 1. Received 18 February 1975.
REFERENCES BEAM,C. A., MORTIMER,R. K., WOLFE,R. O. and TOBIAS,C. A. 1954. The relation of radioresistance to budding in Saccharomyces cerevisiae. - - Arch. Biochem. Biophys. 49:110-122. do CARMOSOLISA,L. 1970. The genus Trichosporon Behrend, p. 1309-1352. In J. Lodder, (ed.), The Yeasts, 2nd Ed. - - North Holland Publ. Co., Amsterdam. DONK, M. A. 1973. The Heterobasidiomycetes: a reconnaissance IV. - - Kon. Ned. Akad. Wetensch. Proc, (Ser. C) 76: 109-125. KREGER-VAN RIJ, N. J. W. and VEENHUIS, M. 1971. A comparative study of the cell wall structure of basidiomycetous and related yeasts. - - J . Gen. Microbiol. 68: 87-95.
