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Myxozyma geophila and Myxozyma lipomycoides spp. nov., two new anamorphic, lipomycetaceous yeasts from Southern Africa
System. Appl. Microbial. 9, 121-124 (1987)
Myxozyma geophiJa and Myxozyma Jipomycoides spp. nov., Two New Anamorphic, Lipomycetaceous Yeasts from Southern Africa j. P. VAN DER WALT', Y. YAMADA2, T. NAKASE 3, and P. D. G. RICHARDS 4 1 1 3 4
M icrob iology Research Group, Cou ncil for Scient ific and Indu stri al Research, Pretoria, South Africa Labor ator y of Applied Mic rob iology, Department of Agricultural Chemistry, Shizuoka Universit y, Shizuok a, Japan Jap an Collection of Microorgani sms, Institute of Physical and Chemical Research, (RIKEN ), Wak o, Sairama, Japan Histological Services, Biological Evaluat ion Division, National Institut e for Food Research, Council for Scientific and Industrial Research, Pretoria, South Africa
Received June 30 , 1986
Summary Two unde scribed species of the genus Myxozyma have been recovered from sur face soils and arbo rico lous lichen , fro m South African hab itats. The two new species, Myxozyma geophila an d Myxozyma lipomycoides, are describ ed and their possible relation ship with the genera Lipomyces and Zygozyma is considered. A key for th e genus Myxozyma is given.
Key words : Myxozyma geophila - Myxozyma lipomycoides - Lipomyces - Wa/tomyces - ZygozymaYeasts - Taxonom y
Introduction When introduced, the genus Myxozyma van der Walt et a!. (1981) co mprised onl y two insect-associated species fro m North American habitats, viz. Mvxozvma melibiosi (Shifrine et Phaff) van der Walt et al. (198 1)' and Myxozyma mucilagina (Phaff er al.) van der Walt er a!. (1981). With the recent discovery of a further two unde scribed species fro m South African sources, th e genus appears to have a wider distribution th an was previou sly anticipated. The two new species ar e describ ed.
clad od es of Opuntia ficus-indica (Gro blersdal District, Trans, vaa l) CBS 7037 : isolated from soil (Preto ria District, Tran svaal ) CBS 72 19 = JCM 5220: isolat ed fro m soil (Bro nkho rstspruit District, T ran svaal) CBS 7038: = JCM 5 198 : isola ted from ar bo ricolous lichen (Nylstroorn District, Tr an svaal) CBS 604 = JCM 1785 : type of Candida parapsilosis (as refer, ence for mol % G + C determinations)
Characterization of strains Materials and Methods Strains The stra ins studied are held by the Yeast Collection of Her Centraalbureau voor Schimmel culture s (CBS), in Delft , Th e Netherl ands , and the Jap an Co llection of M icroorganisms OCM ), in Wako, Saitama, Jap a n. CBS 2102: type of Myxozyma melibiosi = Candida melibiosopbila Golubev CBS 7071 = JCM 1834: type of Myxozyma mucilagina CBS 7058: Myxozyma mucilagina, isolated from decaying
Th e morpholo gical and ph ysiological characters of the strai ns were determ ined by the conventio nal methods described by Van der Walt and Yarrow (1984). The DNA base composition of the strains was det ermined acco rding to the method described by Nakase and Suzuki (1985). The results are given in Table 1. Th e Coenzyme Q systems were determ ined according to the method described by Yamada and Kondo (1971, 1973 ). Observations by TEM were based on material gro wn on YM agar (Wickerham, 1951 ) at 25 °C and fixed in glut ar aldeh yde and osmium tetroxide and stained with uranyl acetat e and lead citrate as described by Van der Walt et al. (1974).
122
] . P. van der Walt , Y. Yam ad a, T . Nakase, and P. D. G. Richard s
Table 1. % G+C of M. melibiosi, M. mucilagina, M. geophila and M. /ipomycoides
Strains
Number of determ inations Tm (M + S.D.) % GC
M. melibiosi CBS 2102 4 M. mucilagina CBS 7071 4 M. mucilagina CBS 7058 5 M.geophila CBS 7037 7 M.geophila CBS 7219 3 M.lipomycoides CBS 7038 3 C. parapsilosis CBS 604 10 1
2
3
89.91 ± 0.24 86.68 ± 0.15 86.63 ± 0.26 88.47 ± 0.19 88.15 ± 0.10 86.07 ± 0.18 86.03 ± 0.19
50.3 l 42.4 2 42.3 46.8 46.0 40.9 40.83
The value of 61.0% reported by Storck et al. (1969) could not be confirmed . et al. (1980) report a value 43.2-44.0% for this species. Stenderup and Bak (1968) report a value of 40.8%.
Pharr
2 Fig. 2. Myxozyma lipomycoides CBS 703 8. T EM micrograph of a cell sho wing hol oblastic budding and characteristic ascomyceto us cell-wall ultrastructure. Insert bar rep resent s 0.5 urn (M aterial fixed in glut ar aldehyde and po stfixed with os mium tetroxide ). confe cto hyphae et pseudohyphae null ae. Ferrnenratio nulla. Dgalactos e L-sorboso D-riboso (lente) D-xyloso L-arabinoso Dara binoso (lente) glycerolo ribitolo xylito lo galaetita lo m-inositolo D-glu cono-l,5-lactono 2-keto-D- gluc onato 5-keto-Dgluconato D-gluconata D-galacturnato DL-Iactato (lente) suecinat o ethanolo eth ylamino hydrochlorico et imidazolo utitur nequ e D-gluco samino L-rh amnoso sucroso maltoso, a,a-treha loso meth yl-a-D-glucosido cellobi oso salicino melib ioso raffinoso melezitoso inulino amylo solub ili i-erythrito lo D-glucuronato citrata methanolo nitrate nee nit rite. Usio lactosi D-glucitoli et Dmann itoli variabile. Vitamina ext erna ad crescentiam necessaria sunr, Crescere non potest Gelatinum non liquiscit. Ureum non finditur. Materia amyloidea form atur. G+C acidi deoxy rib onucleati 46.0-46.8 mol per centum. Systerna coenzymatis Q8 adest . Typu s CBS 7219 in collectione zymotica Centraalbureau voor Schimmelcultur es Delph is Batavorurn.
3rc.
Fig. I. Myxozyma geophila CBS 72 19. TEM micro graph of a cell showing holobl astic budding and charac teristic asc omycetous cell-w all ultrastructure. Insert bar repre sents 0.5 urn (M aterial fixed in glutara ldehyde and po stfixed with osmium tetroxide ).
Descriptions Myxozyma geophila van der Walt, Yamada et Nakase sp. nov. Geophila , loving the soil; from the Gr. yi] soil and qJ(AO~ loving; referring to the species' association with sur face soils. In extracto malti post dies 3 25 °C cellulae spheroidales vel ellipsoidales , 3.0-7.5 x 2.5-7.0 urn, incap sulata, gemmatae multilate rale holoblastice singulae vel binac. Post hebdomades 4 temper atura ambeunte sedimentum et annulus adsunt. In agaro malti post dies 3 25 °C for mae et dim ensione s cellu larum eaedem sunt quae in extracto ma lti. Cultura po st hebdomades 4 temperatura ambuente viscosa raro butyro sa crem ea-bruneoli glabra nitida raro parum hebetata; margo integer. In agaro farina Z eae maydis
Growth in malt extract: After 3 days at 25 °e, the cells are glob ose to ellipsoidal, 3.0-7.5 x 2.5-7.0 11m, encap sulated , reproducing by multil ateral, holoblastic budding (Fig. 1), occurring singly or in pairs. A slight ring and sediment are formed. After 4 weeks at ambient temperature a sediment and ring are present. Growth on malt agar: After 3 days at 25 "C the cells are of the same form and dimen sion s as in malt extract. The streak culture is viscous to muco id, brownish-cream, partly hyaline, smooth, glistening with an entire margin. After 4 weeks at ambient temperature, the culture is viscous to mucoid, rarel y butyrous, creamish-brown and glistening. The margin is entire. Dalmau plate culture on corn meal agar: After 10 da ys at 25 °C neither hyphae nor pseudophyphae are formed . Fermentation: Absent. Utilization of carbon sources: D-galactose, L-sor bose, D-ribose (slow), D-xylose, L-ar abinose, D-ara binose, glycerol, ribitol, xylitol, galactitol, m-inositol, D-glucono-
Myxozyma geophila and Myxozyma lipomycoides spp. nov.
1,5-lactone, 2-keto-D-gluconate, 5-keto-D-gluconate, Dgalacturonate, DL-Iactate (slow), succinate and ethanol are utilized, but not D-glucosamine, L-rhamnose " sucrose maltose, a,a-trehalose, me-a-D-glucoside, cellobiose, salicin, melibiose, raffinose, melezitose, inulin, soluble starch, i-erythritol, D-glucuronate, citrate or methanol. The utilization of lactose, D-glucitol and D-mannitol is variable. Utilization of nitrogen sources: Ethylamine hydrochloride and imidazol are utilized, but not potassium nitrate or sodium nitrite. Growth in vitamin-free medium: Absent. Growth at different temperatures: 30°C +, 3 JOC -. Liquifaction of gelatin: Absent. Formation of amyloid material: Positive, colouring blue-green with iodine. Growth on 50% (m/m) glucose-yeast extract agar: Absent. Hydrolysis of urea: Absent, or very weak. Colour reaction with Diazonium Blue B: Absent. Mol % G+C: 46.0-46.8. Coenzyme Q system: Q8. The type of Myxozyma geophila is maintained as CBS 7219 in the Yeast Collection of Het Centraalbureau voor Schimmelcultures in Delft, The Netherlands. It is also maintained as jCM 5220 in the japan Collection of Microorganisms, Wako, Saitama, japan. The non-living holotype has been deposited as Specimen No. PREM 48580 in Herbarium for Fungi of the Research Institute for Plant Protection in Pretoria, South Africa. The morphological and physiological characters listed for M. geophila are based on the study of CBS 7219, CBS 7037 and five additional strains recovered from soil samples from the Louis Trichardt and Messina districts (Northern Transvaal). Myxozyma lipomycoides van der Walt, Yamada et Nakase sp. nov. Lipomycoides, Lipomyces-like, from Lipomyces and the Gr. hoo£ form or shape; referring to the resemblance between this species and the genus Lipomyces in respect of their common cultural charcters and Coenzyme Q9 system. In extracto malti post dies 3 25°C cellulae globosae ellipsoidales raro ovoideae, 3.0-8.0 X 3.0-7.5 urn, incapsulatae gemmatae multilaterale holoblastice singulae binae vel aggregatae. Post hebdomades 4 temperatura ambeunte annulus pellicula tenuis et sedimentum adsunt. In agaro malti post dies 3 25°C formae et dimensiones cellularum eaedum sunt quae in extracto malti. Cultura post hebdomades 4 temperatura ambeunte viscosa cremea-bruneoli glabra nitida; margo integer. In agaro farina Zeae maydis confecto post dies 10 25°C hyphae et pseudophyphae nullae. Fermentatio nulla. Dvgalactoso L-sorboso D-xyloso L-arabinoso D-arabinoso (lente) L-rhamnoso u.o-trehaloso lactoso glycerolo ribitolo xylitolo D-glucitolo (lente) Dmannitolo (lente) D-glucono-1,5-lactono 2-keto-D-gluconato Dgalacturnato succinato ethanolo ethylamino hydrochlorico et imidazolo utitur neque D-glucosamino D-riboso sucroso maltoso me-a-D-glucosido cellobioso salicino melibioso raffinoso melezitoso inulino amylo solubili i-erythritolo galactitolo m-inositolo 5-keto-D-gluconato D-gluconato D-glucuronato DL-lactato citrato methanolo nitrate nee nitrite. Vitamina externa ad crescentiam necessariae sunt. Crescere non potest in 37°C.
123
Gelatinum non liquiscit. Ureum non finditur. Materia amyloidea formatur. G+C acidi deoxyribonucleati 40.9 mol per centum. Systema coenzymatis Q9 adest. Typus CBS 7038 in collectione zymotica Centraalbureau voor Schimmelcultures Delphis Batavorum.
Growth in malt extract: After 3 days at 25°C the cells are globose to ellipsoidal, rarely ovoid, 3.0-8.0 X 3.0-7.5 urn, encapsulated, reproducing by multilateral holoblastic budding (Fig. 2), occurring singly, in pairs, or small aggregates. A slight incomplete ring and sediment are formed. Growth is scant. After 4 weeks at ambient temperature a ring, thin pellicle and sediment are present. Growth on malt agar: After 3 days at 25°C the cells are of the same form and dimensions as in malt extraxt. The streak culture is mucoid, partly hyaline, brownish-cream, smooth, shiny with an entire margin. After 4 weeks at ambient temperature, the culture is viscous, creamishbrown, smooth and glisterning with an entire margin. Dalmau plate cultures on corn meal agar: After 10 days at 25°C neither hyphae nor pseudohyphae are formed. Fermentation: Absent. Utilization of carbon sources: D-galactose, L-sorbose, D-xylose, L-arabinose, D-arabinose (slow), L-rhamnose, a,a-trehalose, lactose, glycerol, ribitol, xylitol, D-glucitol (slow), D-mannitol (slow), D-glucono-1,5-lactone, 2-ketogluconate, D-galacturonate, succinate and ethanol are utilized, but not D-glucosamine, D-ribose, sucrose, maltose, me-a-D-glucoside, cellobiose, salicin, melibiose, raffinose, melezitose, inulin, soluble starch, i-erythritol, galactitol, m-inositol, 5-keto-D-gluconate, D-gluconate, D-glucuronate, DL-lactate, citrate and methanol. Utilization of nitrogen sources: Ethylamine hydrochloride and imidazol are utilized, but not potassium nitrate or sodium nitrite. Growth in vitamin-free medium: Absent. Growth at different temperatures: 30°C +, 37°C -. Liquifaction of gelatin: Absent. Formation of amyloid material: Positive, staining bluegreen with iodine. Growth on 50% (m/m) glucose-yeast extract-agar: Absent. Hydrolysis of urea: Absent. Colour reaction with Diazonium Blue B: Absent.
Mol % G+G: 40.9. Coenzyme Q system: Q9. The type Myxozyma lipomycoides is maintained as CBS
7038 in the Yeast Collection of Het Centraalbureau voor Schimmelcultures in Delft, The Netherlands. It is also maintained as jCM 5198 in the Japan Collection of Microorganisms, Wako, Saitama, japan. The non-living holotype has been deposited as Specimen No. PREM 48579 in the Herbarium for Fungi of the Research Institute for Plant Protection in Pretoria, South Africa. The ~escription of the species is based on the type strain, which IS the only strain currently available.
Discussion The two new species, on the basis of their (i) characteristic ascomycetous cell-wall ultrastructure, (ii) multilateral,
124
J. P. van der Walt,
Y. Yamada, T. Nakase, and P.D. G. Richards
holoblastic budding, (iii) formation of extracellular polysaccharides that colour blue-green with iodine, (iv) utilization of imidazol as sole source of nitrogen and (v) strictly oxidative metabolism, possess the definitive characters of the genus Myxozyma. The combination of these characters effectively distinguishes all species of this genus from the type species of both Candida Berkhout and Cryptococcus Kiitzing emend Phaff et Spencer. Consequently, the recent inclusion of M. melibiosi in Cryptococcus (Rodrigues de Miranda, 1984), and M. mucilagina in Candida (Meyer et al., 1984), lacks the constructive basis for the rational classification of the anamorphic yeasts along natural lines, and exemplifies the limitations which denote the current classification of these yeasts (Kreger-van Rij, 1984). Candida must be restricted to anamorphs related to the Endomycetales which (i) bud multilaterally on narrow base and (ii) do not form extracellular amyloid material, and Cryptococcus, to anamorphs of heterobasidiomycetous affinity. As pointed out by Van der Walt et a1. (1981, 1987), the genus Myxozyma, on the basis of its cultural, morphological and physiological characters, appears to represent lipomycetaceous anamorphs which, depending on their Coenzyme Q systems, could be related to either Zygozyma (Q8), Lipomyces (Q9), or to the recently introduced genus Waltomyces Yamada et Nakase (1985) which is characterized by the Q10 system. On the basis of this character M. geophila, M. melibiosi and M. mucilagina show agreement with Zygozyma, and M. lipomycoides, with
Lipomyces.
All investigated strains of the genus Myxozyma utilze Dgalacturonic acid - a physiological character which seemingly serves to distinguish this anamorphic genus from the related, budding, teleomorphic genera Lipomyces, Waltomyces and Zygozyma. Like M. melibiosi, M. geophila utilizes inositol but not D-glucuronic acid - a physiological character which is also shared by certain unrelated species, e. g. Candida edax (Barnett et al., 1983). The four species which currently comprise the genus can be recognized by the following key: Key to the genus Myxozyma 2 1. Inositol utilized 3 Inositol not utilized M. melibiosi 2. Melibiose utilized M. geophila Melibiose not utilized M. mucilagina 3. D-Gluconate utilized M. lipomycoides D-Gluconate not utilized
References
Barnett, J. A., Payne, R. W., Yarrow, D.: Yeasts: Characteristics and Identification, Cambridge, Cambridge University Press 1983 Kreger-van Rij, N. J. W.: Classification of the imperfect yeasts. In: The Yeasts, a Taxonomic Study (N. J. W. Kreger-van RII, ed.), 3rd ed., pp. 35-42. Amsterdam, Elsevier Science Publishers, B.V. 1984 Meyer, S. A., Ahearn, D. G., Yarrow, D.: Genus 4. Candida Berkhout. In: The Yeasts, a Taxonomic Study (N. J. W. Kreger-uan Rij, ed.), 3rd ed., pp. 585-844. Amsterdam, Elsevier Science Publishers, B.V. 1984 Phaf], H. J., Starmer, W. T., Miranda, M., Miller, M. W.: Candida mucilagina, a new species of yeast found in decaying of Opuntia inermis and in necrotic tissue of Cereoid cacti. J. system. Bact. 30, 596-600 (1980) Nakase, T., Suzuki, M.: Taxonomic studies on Debaryomyces hansenii (Zopf) Lodder et Kreger-van Rij and related species. I. Chemotaxonomic Investigations. J. gen. app!. Microbio!' 31, 49-69 (1985) Rodrigues de Miranda, 1.: Genus 5. Cryptococcus Kiitzing emend. Phaff et Spencer. In: The Yeasts, a Taxonomic Study (N. J. W. Kreger-van Rij, ed.), 3rd ed., pp. 845-872. Amsterdam, Elsevier Science Publishers, B.V. 1984 Stenderup, A., Bak, A. 1.: Deoxyribonucleic acid base composition of some species within the genus Candida. J. gen. Microbio!. 52, 231-236 (1968) Storck, R., Alexopoulus, C. J., Phaff, H. J.: Nucleotide composition of some species of Cryptococcus, Rhodotorula and Sporobolomyces. J. Bact. 98, 1069-1072 (1969) Van der Walt, J. P., Johannsen, E., Liebenberg, N. V. D. W.: Cell-wall structure, mitosis and urease activity in Torulopsis species of high GC content. Antonie v. Leeuwenhoek 40, 417-426 (1974) Van der Walt, J. P., Yarrow, D.: Methods for the isolation, maintenance, classification of yeasts. In: The Yeasts, a Taxonomic Study (N. J. W. Kreger-van Rij, ed.), 3rd ed., pp. 45-104. Amsterdam, Elsevier Science Publishers B.V 1984 Van der Walt, J. P., Weijman, A. C. M., Von Arx, J. A.: The anamorphic yeast genus Myxozyma gen. nov. Sydowia, Ann. Myco!. Ser. II 34,191-198 (1981) Van der Walt,]. P., Von Arx, A. j., Ferreira, N. P., Richards, P. D. G.: Zygozyma gen. nov., a New Genus of the Lipomycetaceae. System. App!. Microbio!' 9, 115-120 (1987) Wickerham, 1. j.: Taxonomy of Yeasts, U.S. Dept. Agricult., Techn. Bul!. No. 1029, WashingtonlD.C. 1951 Yamada, Y., Kondo, K.: Significance of Coenzyme Q systems in yeasts and yeast-like fungi (1). Proc.lst Inst. Symp!. Yeasts, pp. 363-373, Smolenice 1971 Yamada, Y., Kondo, K.: Coenzyme Q system in the classification of the yeast genera Rbodotorula and Cryptococcus, and the yeast-like genera Sporobolomyces and Rhodosporidium. J. gen. appl, Microbio!' 19,59-77 (1973) Yamada, Y., Nakase, T.: Waltomyces, a new ascosporogenous yeast for the Qwequipped, slime-producing organisms whose asexual reproduction is by multilateral budding and whose ascospores have smooth surfaces. J. gen. app!. Microbio!' 31, 491-492 (1985)
Professor Dr. J. P. Van der Walt, Microbiology Research Group, Council for Scientificand Industrial Research, P.O. Box 395, Pretoria 0001, South Africa
Myxozyma geophiJa and Myxozyma Jipomycoides spp. nov., Two New Anamorphic, Lipomycetaceous Yeasts from Southern Africa j. P. VAN DER WALT', Y. YAMADA2, T. NAKASE 3, and P. D. G. RICHARDS 4 1 1 3 4
M icrob iology Research Group, Cou ncil for Scient ific and Indu stri al Research, Pretoria, South Africa Labor ator y of Applied Mic rob iology, Department of Agricultural Chemistry, Shizuoka Universit y, Shizuok a, Japan Jap an Collection of Microorgani sms, Institute of Physical and Chemical Research, (RIKEN ), Wak o, Sairama, Japan Histological Services, Biological Evaluat ion Division, National Institut e for Food Research, Council for Scientific and Industrial Research, Pretoria, South Africa
Received June 30 , 1986
Summary Two unde scribed species of the genus Myxozyma have been recovered from sur face soils and arbo rico lous lichen , fro m South African hab itats. The two new species, Myxozyma geophila an d Myxozyma lipomycoides, are describ ed and their possible relation ship with the genera Lipomyces and Zygozyma is considered. A key for th e genus Myxozyma is given.
Key words : Myxozyma geophila - Myxozyma lipomycoides - Lipomyces - Wa/tomyces - ZygozymaYeasts - Taxonom y
Introduction When introduced, the genus Myxozyma van der Walt et a!. (1981) co mprised onl y two insect-associated species fro m North American habitats, viz. Mvxozvma melibiosi (Shifrine et Phaff) van der Walt et al. (198 1)' and Myxozyma mucilagina (Phaff er al.) van der Walt er a!. (1981). With the recent discovery of a further two unde scribed species fro m South African sources, th e genus appears to have a wider distribution th an was previou sly anticipated. The two new species ar e describ ed.
clad od es of Opuntia ficus-indica (Gro blersdal District, Trans, vaa l) CBS 7037 : isolated from soil (Preto ria District, Tran svaal ) CBS 72 19 = JCM 5220: isolat ed fro m soil (Bro nkho rstspruit District, T ran svaal) CBS 7038: = JCM 5 198 : isola ted from ar bo ricolous lichen (Nylstroorn District, Tr an svaal) CBS 604 = JCM 1785 : type of Candida parapsilosis (as refer, ence for mol % G + C determinations)
Characterization of strains Materials and Methods Strains The stra ins studied are held by the Yeast Collection of Her Centraalbureau voor Schimmel culture s (CBS), in Delft , Th e Netherl ands , and the Jap an Co llection of M icroorganisms OCM ), in Wako, Saitama, Jap a n. CBS 2102: type of Myxozyma melibiosi = Candida melibiosopbila Golubev CBS 7071 = JCM 1834: type of Myxozyma mucilagina CBS 7058: Myxozyma mucilagina, isolated from decaying
Th e morpholo gical and ph ysiological characters of the strai ns were determ ined by the conventio nal methods described by Van der Walt and Yarrow (1984). The DNA base composition of the strains was det ermined acco rding to the method described by Nakase and Suzuki (1985). The results are given in Table 1. Th e Coenzyme Q systems were determ ined according to the method described by Yamada and Kondo (1971, 1973 ). Observations by TEM were based on material gro wn on YM agar (Wickerham, 1951 ) at 25 °C and fixed in glut ar aldeh yde and osmium tetroxide and stained with uranyl acetat e and lead citrate as described by Van der Walt et al. (1974).
122
] . P. van der Walt , Y. Yam ad a, T . Nakase, and P. D. G. Richard s
Table 1. % G+C of M. melibiosi, M. mucilagina, M. geophila and M. /ipomycoides
Strains
Number of determ inations Tm (M + S.D.) % GC
M. melibiosi CBS 2102 4 M. mucilagina CBS 7071 4 M. mucilagina CBS 7058 5 M.geophila CBS 7037 7 M.geophila CBS 7219 3 M.lipomycoides CBS 7038 3 C. parapsilosis CBS 604 10 1
2
3
89.91 ± 0.24 86.68 ± 0.15 86.63 ± 0.26 88.47 ± 0.19 88.15 ± 0.10 86.07 ± 0.18 86.03 ± 0.19
50.3 l 42.4 2 42.3 46.8 46.0 40.9 40.83
The value of 61.0% reported by Storck et al. (1969) could not be confirmed . et al. (1980) report a value 43.2-44.0% for this species. Stenderup and Bak (1968) report a value of 40.8%.
Pharr
2 Fig. 2. Myxozyma lipomycoides CBS 703 8. T EM micrograph of a cell sho wing hol oblastic budding and characteristic ascomyceto us cell-wall ultrastructure. Insert bar rep resent s 0.5 urn (M aterial fixed in glut ar aldehyde and po stfixed with os mium tetroxide ). confe cto hyphae et pseudohyphae null ae. Ferrnenratio nulla. Dgalactos e L-sorboso D-riboso (lente) D-xyloso L-arabinoso Dara binoso (lente) glycerolo ribitolo xylito lo galaetita lo m-inositolo D-glu cono-l,5-lactono 2-keto-D- gluc onato 5-keto-Dgluconato D-gluconata D-galacturnato DL-Iactato (lente) suecinat o ethanolo eth ylamino hydrochlorico et imidazolo utitur nequ e D-gluco samino L-rh amnoso sucroso maltoso, a,a-treha loso meth yl-a-D-glucosido cellobi oso salicino melib ioso raffinoso melezitoso inulino amylo solub ili i-erythrito lo D-glucuronato citrata methanolo nitrate nee nit rite. Usio lactosi D-glucitoli et Dmann itoli variabile. Vitamina ext erna ad crescentiam necessaria sunr, Crescere non potest Gelatinum non liquiscit. Ureum non finditur. Materia amyloidea form atur. G+C acidi deoxy rib onucleati 46.0-46.8 mol per centum. Systerna coenzymatis Q8 adest . Typu s CBS 7219 in collectione zymotica Centraalbureau voor Schimmelcultur es Delph is Batavorurn.
3rc.
Fig. I. Myxozyma geophila CBS 72 19. TEM micro graph of a cell showing holobl astic budding and charac teristic asc omycetous cell-w all ultrastructure. Insert bar repre sents 0.5 urn (M aterial fixed in glutara ldehyde and po stfixed with osmium tetroxide ).
Descriptions Myxozyma geophila van der Walt, Yamada et Nakase sp. nov. Geophila , loving the soil; from the Gr. yi] soil and qJ(AO~ loving; referring to the species' association with sur face soils. In extracto malti post dies 3 25 °C cellulae spheroidales vel ellipsoidales , 3.0-7.5 x 2.5-7.0 urn, incap sulata, gemmatae multilate rale holoblastice singulae vel binac. Post hebdomades 4 temper atura ambeunte sedimentum et annulus adsunt. In agaro malti post dies 3 25 °C for mae et dim ensione s cellu larum eaedem sunt quae in extracto ma lti. Cultura po st hebdomades 4 temperatura ambuente viscosa raro butyro sa crem ea-bruneoli glabra nitida raro parum hebetata; margo integer. In agaro farina Z eae maydis
Growth in malt extract: After 3 days at 25 °e, the cells are glob ose to ellipsoidal, 3.0-7.5 x 2.5-7.0 11m, encap sulated , reproducing by multil ateral, holoblastic budding (Fig. 1), occurring singly or in pairs. A slight ring and sediment are formed. After 4 weeks at ambient temperature a sediment and ring are present. Growth on malt agar: After 3 days at 25 "C the cells are of the same form and dimen sion s as in malt extract. The streak culture is viscous to muco id, brownish-cream, partly hyaline, smooth, glistening with an entire margin. After 4 weeks at ambient temperature, the culture is viscous to mucoid, rarel y butyrous, creamish-brown and glistening. The margin is entire. Dalmau plate culture on corn meal agar: After 10 da ys at 25 °C neither hyphae nor pseudophyphae are formed . Fermentation: Absent. Utilization of carbon sources: D-galactose, L-sor bose, D-ribose (slow), D-xylose, L-ar abinose, D-ara binose, glycerol, ribitol, xylitol, galactitol, m-inositol, D-glucono-
Myxozyma geophila and Myxozyma lipomycoides spp. nov.
1,5-lactone, 2-keto-D-gluconate, 5-keto-D-gluconate, Dgalacturonate, DL-Iactate (slow), succinate and ethanol are utilized, but not D-glucosamine, L-rhamnose " sucrose maltose, a,a-trehalose, me-a-D-glucoside, cellobiose, salicin, melibiose, raffinose, melezitose, inulin, soluble starch, i-erythritol, D-glucuronate, citrate or methanol. The utilization of lactose, D-glucitol and D-mannitol is variable. Utilization of nitrogen sources: Ethylamine hydrochloride and imidazol are utilized, but not potassium nitrate or sodium nitrite. Growth in vitamin-free medium: Absent. Growth at different temperatures: 30°C +, 3 JOC -. Liquifaction of gelatin: Absent. Formation of amyloid material: Positive, colouring blue-green with iodine. Growth on 50% (m/m) glucose-yeast extract agar: Absent. Hydrolysis of urea: Absent, or very weak. Colour reaction with Diazonium Blue B: Absent. Mol % G+C: 46.0-46.8. Coenzyme Q system: Q8. The type of Myxozyma geophila is maintained as CBS 7219 in the Yeast Collection of Het Centraalbureau voor Schimmelcultures in Delft, The Netherlands. It is also maintained as jCM 5220 in the japan Collection of Microorganisms, Wako, Saitama, japan. The non-living holotype has been deposited as Specimen No. PREM 48580 in Herbarium for Fungi of the Research Institute for Plant Protection in Pretoria, South Africa. The morphological and physiological characters listed for M. geophila are based on the study of CBS 7219, CBS 7037 and five additional strains recovered from soil samples from the Louis Trichardt and Messina districts (Northern Transvaal). Myxozyma lipomycoides van der Walt, Yamada et Nakase sp. nov. Lipomycoides, Lipomyces-like, from Lipomyces and the Gr. hoo£ form or shape; referring to the resemblance between this species and the genus Lipomyces in respect of their common cultural charcters and Coenzyme Q9 system. In extracto malti post dies 3 25°C cellulae globosae ellipsoidales raro ovoideae, 3.0-8.0 X 3.0-7.5 urn, incapsulatae gemmatae multilaterale holoblastice singulae binae vel aggregatae. Post hebdomades 4 temperatura ambeunte annulus pellicula tenuis et sedimentum adsunt. In agaro malti post dies 3 25°C formae et dimensiones cellularum eaedum sunt quae in extracto malti. Cultura post hebdomades 4 temperatura ambeunte viscosa cremea-bruneoli glabra nitida; margo integer. In agaro farina Zeae maydis confecto post dies 10 25°C hyphae et pseudophyphae nullae. Fermentatio nulla. Dvgalactoso L-sorboso D-xyloso L-arabinoso D-arabinoso (lente) L-rhamnoso u.o-trehaloso lactoso glycerolo ribitolo xylitolo D-glucitolo (lente) Dmannitolo (lente) D-glucono-1,5-lactono 2-keto-D-gluconato Dgalacturnato succinato ethanolo ethylamino hydrochlorico et imidazolo utitur neque D-glucosamino D-riboso sucroso maltoso me-a-D-glucosido cellobioso salicino melibioso raffinoso melezitoso inulino amylo solubili i-erythritolo galactitolo m-inositolo 5-keto-D-gluconato D-gluconato D-glucuronato DL-lactato citrato methanolo nitrate nee nitrite. Vitamina externa ad crescentiam necessariae sunt. Crescere non potest in 37°C.
123
Gelatinum non liquiscit. Ureum non finditur. Materia amyloidea formatur. G+C acidi deoxyribonucleati 40.9 mol per centum. Systema coenzymatis Q9 adest. Typus CBS 7038 in collectione zymotica Centraalbureau voor Schimmelcultures Delphis Batavorum.
Growth in malt extract: After 3 days at 25°C the cells are globose to ellipsoidal, rarely ovoid, 3.0-8.0 X 3.0-7.5 urn, encapsulated, reproducing by multilateral holoblastic budding (Fig. 2), occurring singly, in pairs, or small aggregates. A slight incomplete ring and sediment are formed. Growth is scant. After 4 weeks at ambient temperature a ring, thin pellicle and sediment are present. Growth on malt agar: After 3 days at 25°C the cells are of the same form and dimensions as in malt extraxt. The streak culture is mucoid, partly hyaline, brownish-cream, smooth, shiny with an entire margin. After 4 weeks at ambient temperature, the culture is viscous, creamishbrown, smooth and glisterning with an entire margin. Dalmau plate cultures on corn meal agar: After 10 days at 25°C neither hyphae nor pseudohyphae are formed. Fermentation: Absent. Utilization of carbon sources: D-galactose, L-sorbose, D-xylose, L-arabinose, D-arabinose (slow), L-rhamnose, a,a-trehalose, lactose, glycerol, ribitol, xylitol, D-glucitol (slow), D-mannitol (slow), D-glucono-1,5-lactone, 2-ketogluconate, D-galacturonate, succinate and ethanol are utilized, but not D-glucosamine, D-ribose, sucrose, maltose, me-a-D-glucoside, cellobiose, salicin, melibiose, raffinose, melezitose, inulin, soluble starch, i-erythritol, galactitol, m-inositol, 5-keto-D-gluconate, D-gluconate, D-glucuronate, DL-lactate, citrate and methanol. Utilization of nitrogen sources: Ethylamine hydrochloride and imidazol are utilized, but not potassium nitrate or sodium nitrite. Growth in vitamin-free medium: Absent. Growth at different temperatures: 30°C +, 37°C -. Liquifaction of gelatin: Absent. Formation of amyloid material: Positive, staining bluegreen with iodine. Growth on 50% (m/m) glucose-yeast extract-agar: Absent. Hydrolysis of urea: Absent. Colour reaction with Diazonium Blue B: Absent.
Mol % G+G: 40.9. Coenzyme Q system: Q9. The type Myxozyma lipomycoides is maintained as CBS
7038 in the Yeast Collection of Het Centraalbureau voor Schimmelcultures in Delft, The Netherlands. It is also maintained as jCM 5198 in the Japan Collection of Microorganisms, Wako, Saitama, japan. The non-living holotype has been deposited as Specimen No. PREM 48579 in the Herbarium for Fungi of the Research Institute for Plant Protection in Pretoria, South Africa. The ~escription of the species is based on the type strain, which IS the only strain currently available.
Discussion The two new species, on the basis of their (i) characteristic ascomycetous cell-wall ultrastructure, (ii) multilateral,
124
J. P. van der Walt,
Y. Yamada, T. Nakase, and P.D. G. Richards
holoblastic budding, (iii) formation of extracellular polysaccharides that colour blue-green with iodine, (iv) utilization of imidazol as sole source of nitrogen and (v) strictly oxidative metabolism, possess the definitive characters of the genus Myxozyma. The combination of these characters effectively distinguishes all species of this genus from the type species of both Candida Berkhout and Cryptococcus Kiitzing emend Phaff et Spencer. Consequently, the recent inclusion of M. melibiosi in Cryptococcus (Rodrigues de Miranda, 1984), and M. mucilagina in Candida (Meyer et al., 1984), lacks the constructive basis for the rational classification of the anamorphic yeasts along natural lines, and exemplifies the limitations which denote the current classification of these yeasts (Kreger-van Rij, 1984). Candida must be restricted to anamorphs related to the Endomycetales which (i) bud multilaterally on narrow base and (ii) do not form extracellular amyloid material, and Cryptococcus, to anamorphs of heterobasidiomycetous affinity. As pointed out by Van der Walt et a1. (1981, 1987), the genus Myxozyma, on the basis of its cultural, morphological and physiological characters, appears to represent lipomycetaceous anamorphs which, depending on their Coenzyme Q systems, could be related to either Zygozyma (Q8), Lipomyces (Q9), or to the recently introduced genus Waltomyces Yamada et Nakase (1985) which is characterized by the Q10 system. On the basis of this character M. geophila, M. melibiosi and M. mucilagina show agreement with Zygozyma, and M. lipomycoides, with
Lipomyces.
All investigated strains of the genus Myxozyma utilze Dgalacturonic acid - a physiological character which seemingly serves to distinguish this anamorphic genus from the related, budding, teleomorphic genera Lipomyces, Waltomyces and Zygozyma. Like M. melibiosi, M. geophila utilizes inositol but not D-glucuronic acid - a physiological character which is also shared by certain unrelated species, e. g. Candida edax (Barnett et al., 1983). The four species which currently comprise the genus can be recognized by the following key: Key to the genus Myxozyma 2 1. Inositol utilized 3 Inositol not utilized M. melibiosi 2. Melibiose utilized M. geophila Melibiose not utilized M. mucilagina 3. D-Gluconate utilized M. lipomycoides D-Gluconate not utilized
References
Barnett, J. A., Payne, R. W., Yarrow, D.: Yeasts: Characteristics and Identification, Cambridge, Cambridge University Press 1983 Kreger-van Rij, N. J. W.: Classification of the imperfect yeasts. In: The Yeasts, a Taxonomic Study (N. J. W. Kreger-van RII, ed.), 3rd ed., pp. 35-42. Amsterdam, Elsevier Science Publishers, B.V. 1984 Meyer, S. A., Ahearn, D. G., Yarrow, D.: Genus 4. Candida Berkhout. In: The Yeasts, a Taxonomic Study (N. J. W. Kreger-uan Rij, ed.), 3rd ed., pp. 585-844. Amsterdam, Elsevier Science Publishers, B.V. 1984 Phaf], H. J., Starmer, W. T., Miranda, M., Miller, M. W.: Candida mucilagina, a new species of yeast found in decaying of Opuntia inermis and in necrotic tissue of Cereoid cacti. J. system. Bact. 30, 596-600 (1980) Nakase, T., Suzuki, M.: Taxonomic studies on Debaryomyces hansenii (Zopf) Lodder et Kreger-van Rij and related species. I. Chemotaxonomic Investigations. J. gen. app!. Microbio!' 31, 49-69 (1985) Rodrigues de Miranda, 1.: Genus 5. Cryptococcus Kiitzing emend. Phaff et Spencer. In: The Yeasts, a Taxonomic Study (N. J. W. Kreger-van Rij, ed.), 3rd ed., pp. 845-872. Amsterdam, Elsevier Science Publishers, B.V. 1984 Stenderup, A., Bak, A. 1.: Deoxyribonucleic acid base composition of some species within the genus Candida. J. gen. Microbio!. 52, 231-236 (1968) Storck, R., Alexopoulus, C. J., Phaff, H. J.: Nucleotide composition of some species of Cryptococcus, Rhodotorula and Sporobolomyces. J. Bact. 98, 1069-1072 (1969) Van der Walt, J. P., Johannsen, E., Liebenberg, N. V. D. W.: Cell-wall structure, mitosis and urease activity in Torulopsis species of high GC content. Antonie v. Leeuwenhoek 40, 417-426 (1974) Van der Walt, J. P., Yarrow, D.: Methods for the isolation, maintenance, classification of yeasts. In: The Yeasts, a Taxonomic Study (N. J. W. Kreger-van Rij, ed.), 3rd ed., pp. 45-104. Amsterdam, Elsevier Science Publishers B.V 1984 Van der Walt, J. P., Weijman, A. C. M., Von Arx, J. A.: The anamorphic yeast genus Myxozyma gen. nov. Sydowia, Ann. Myco!. Ser. II 34,191-198 (1981) Van der Walt,]. P., Von Arx, A. j., Ferreira, N. P., Richards, P. D. G.: Zygozyma gen. nov., a New Genus of the Lipomycetaceae. System. App!. Microbio!' 9, 115-120 (1987) Wickerham, 1. j.: Taxonomy of Yeasts, U.S. Dept. Agricult., Techn. Bul!. No. 1029, WashingtonlD.C. 1951 Yamada, Y., Kondo, K.: Significance of Coenzyme Q systems in yeasts and yeast-like fungi (1). Proc.lst Inst. Symp!. Yeasts, pp. 363-373, Smolenice 1971 Yamada, Y., Kondo, K.: Coenzyme Q system in the classification of the yeast genera Rbodotorula and Cryptococcus, and the yeast-like genera Sporobolomyces and Rhodosporidium. J. gen. appl, Microbio!' 19,59-77 (1973) Yamada, Y., Nakase, T.: Waltomyces, a new ascosporogenous yeast for the Qwequipped, slime-producing organisms whose asexual reproduction is by multilateral budding and whose ascospores have smooth surfaces. J. gen. app!. Microbio!' 31, 491-492 (1985)
Professor Dr. J. P. Van der Walt, Microbiology Research Group, Council for Scientificand Industrial Research, P.O. Box 395, Pretoria 0001, South Africa