Utilization of organic nitrogen compounds by yeasts of the genusSaccharomyces

Yeasts belonging to 27 species ofSaccharomyces were tested for their ability to used-amino acids,l-lysine and various amines and amides as nitrogen source. Most yeasts capable of growing onl-lysine or amines could utilized-amino acids.Saccharomyces (sensu strictu) have a very limited ability to grow on the organic nitrogen compounds tested. However, there is no obvious relationship between the utilization of these compounds and the proposed divisions of the genusSaccharomyces.Issued as N.R.C. No. 9845.     

Aerobic dissimilation of d-ribulose by yeasts

Aerobic dissimilation ofd-ribulose by various genera and species of yeasts was examined. Most strains tested utilizedd-ribulose fairly well, andd-arabitol was accumulated in the fermented broth but ribitol was not found. Torulopsis famata ATCC 20214,T.mannitofaciens CBS 5981 andT.versatilis CBS 1752 producedd-arabitol with a yield of 26 to 67% of the sugar used. Extracellular and intracellular formation of pentitol fromd-ribulose were compared with those fromd-xylulose.We wish to express our sincerest thanks to Prof. K. Arima and Prof. Y. Ikeda of the University of Tokyo for their kind guidance. We also wish to thank Dr. M. Mogi and Dr. N. Iguchi of this Institute for their encouragement and Mr. K. Kouchi for his technical assistance.     

A study on the inhibition of rat myocardium glutathione peroxidase and glutathione reductase by moniliformin

The yeasts of patients with oral cancer has been studied before and during Xr-therapy. Gram and PAS smears revealed an increase of yeast-like structures, during treatment, from 56% to 66% of the cases. Before radiotherapy oral yeasts were isolated from 56% of the patients with cancer represented by Candida albicans (30%); C. tropicalis (12%); C. glabrata and C. krusei (4%), besides six other different species (2%). During radiotherapy yeasts were isolated in 72% of the cases, as follow: C. albicans (36%); C. tropicalis (16%); Rhodotorula rubra (6%); C. kefyr; C. krusei and Pichia farinosa (4%), besides other nine species (2%). C. albicans serotype A represented 93% of the isolated samples, before treatment and 88,8% during Xr-therapy.     

Efficient synthesis of enantiomeric ethyl lactate by <Emphasis Type="Italic">Candida antarctica</Emphasis> lipase B (CALB)-displaying yeasts

The whole-cell biocatalyst displaying Candida antarctica lipase B (CALB) on the yeast cell surface with α-agglutinin as the anchor protein was easy to handle and possessed high stability. The lyophilized CALB-displaying yeasts showed their original hydrolytic activity and were applied to an ester synthesis using ethanol and l-lactic acid as substrates. In water-saturated heptane, CALB-displaying yeasts catalyzed ethyl lactate synthesis. The synthesis efficiency increased depending on temperature and reached approximately 74% at 50°C. The amount of l-ethyl lactate increased gradually. l-Ethyl lactate synthesis stopped at 200 h and restarted after adding of l-lactic acid at 253 h. It indicated that CALB-displaying yeasts retained their synthetic activity under such reaction conditions. In addition, CALB-displaying yeasts were able to recognize l-lactic acid and d-lactic acid as substrates. l-Ethyl lactate was prepared from l-lactic acid and d-ethyl lactate was prepared from d-lactic acid using the same CALB-displaying whole-cell biocatalyst. These findings suggest that CALB-displaying yeasts can supply the enantiomeric lactic esters for preparation of useful and improved biopolymers of lactic acid.     

Some physiological observations on the uptake of d-glucose and 2-deoxy-d-glucose by starving and exponentially-growing yeasts

Some methods for measuring the uptake of sugars by yeasts were investigated critically. A study was made of the effects of starvation of Pichia pinus, Candida utilis, Saccharomyces cerevisiae and Rhodosporidium toruloides on their uptake of d-glucose and 2-deoxy-d-glucose. Marked changes in the rates of uptake of these sugars occurred during 10 h of starvation, including (a) an immediate increase of up to 75% above that for growing cells and (b) a continuous decline to as little as 4%. Each yeast behaved differently. The rates did not remain constant during the periods of starvation often used for studies on the transport of sugars into yeasts. For Pichia pinus, there were striking differences, associated with starvation, between the transport of 2-deoxy-d-glucose and d-glucose, despite evidence that the two sugars enter this yeast by means of the same carrier. Some physiological explanations for these findings are discussed.     

The effect of respiratory inhibitors on the fermentative ability of Pichia stipitis,Pachysolen tannophilus and Saccharomyces cerevisiae under various conditions of aerobiosis

Summary The growth and ethanol production by the d-xylose-fermenting yeasts Pichia stipitis and Pachysolen tannophilus under various conditions of aerobiosis responded similarly to the addition of the respiratory inhibitors potassium cyanide (KCN), antimycin A (AA), sodium azide and rotenone. However, the d-glucose-fermenting yeast Saccharomyces cerevisiae differed markedly from these yeasts in response to the inhibitors. In general the growth of the d-xylose-fermenting yeasts was inhibited by the respiratory inhibitors while ethanol production was either stimulated (especially when oxygen was available) or unaffected or inhibited by rotenone or AA or KCN and sodium azide, respectively. However, by exception KCN and AA stimulated ethanol production under aerobic conditions by Pichia stipitis and Pachysolen tannophilus respectively. Stimulatory or inhibitory effects by respiratory inhibitors were less marked in S. cerevisiae. These data suggest that unimpaired mitochondrial function is necessary for growth on d-xylose and optimal d-xylose fermentation. A requirement for membrane generated energy during d-xylose utilisation is indicated by 2,4-dinitrophenol inhibition of growth and fermentation.     

Multiple mechanisms may contribute to the adherence ofCandida yeasts to living cells

The adherence ofCandida yeasts to monolayers of human intestinal epithelium was studied in order to determine the specific and nonspecific mechanisms that might contribute to yeast adherence. Multiple factors were shown to significantly affect the adherence of yeasts to intestinal cells. It was demonstrated that hydrophobic yeasts adhered two times greater than normal yeasts, and positively charged yeasts adhered ten times greater than normal yeasts to monolayers of intestinal epithelium. The binding of yeasts to the intestinal cells was saturable and was most effectively blocked by mucin, which caused an 83% reduction in adherence, whereas the addition ofd-glucose caused a 41% reduction in adherence. Aggregation or coadherence of yeasts occurred as the yeast inocula were increased.Candida appears to possess the ability to adhere to living tissue by several mechanisms, such as adhesin-receptor interactions, nonspecific hydrophobic and ionic bonding, and aggregation or coadherence. This is the first demonstration of multiple forces that may act simultaneously in the process of adherence of yeasts to living cells.     

A taxonomic study of soil yeasts

Summary Eighty-four samples of Minnesota soils were collected in the spring of the year. All samples yielded yeasts, and a total of 180 cultures were isolated of which 117 were studied taxonomically. Twenty-five cultures were black yeasts. Approximately one-third of the isolates were spore forming yeasts and members of the oxidative and film forming generaHansenula, Pichia, andDebaryomyces, of which a new species (D. mrakii) has been described. Not a single culture ofSaccharomyces was isolated. Perhaps later in the season when fruit was abundant members of this genus would have been present.The greatest number of imperfects belonged to the genusCandida (32 cultures), althoughPullularia (25 cultures),Rhodotorula (20 cultures), andTorulopsis (16 cultures) were well represented.The cultures ofPullularia darkened very slowly if at all, in the asbence of excess sugar. Under these circumstances they were very similar toT. pullulans.A single culture ofTrichisporon cerebriformis was isolated. The culture isolated formed both blastospores and arthrospores.I would like to express my appreciation to Dr.C. E. Skinner of Washington State College, for his help and guidance throughout this entire problem. I would also like to thank Dr.E. M. Mark, of the University of California, Berkeley, for the privilege of working under his direction, and in his laboratory during the study.     

The effect of aminopterin (4-amino folic acid) on growth and cell division of fermentative versus oxidative yeasts

In a related brewing study detailed characteristics of fermentations displaying effective yeastaminopterin interaction were presented.Fermentative yeast types (certain Saccharomyces species and Selenotila intestinalis) proved effective aminopterin reactors whereas oxidative yeasts (certain Candida, Cryptococcus, Pichia, Rhodotorula, Saccharomyces, and Trigonopsis species) proved ineffective reactors. In general effective reactors were polyploids characterized by the lack of film or pellicle formation and ineffective reactors the opposite. In stationary fermentations the Fleischmann 139 strain of S. cerevisiae proved a fair reactor. When aerated it proved an ineffective reactor and aminopterin or products there-of stimulated growth. Conversely aeration enhanced aminopterin activity of effective reactor yeasts.The positive effect of biotin on aminopterin activity and the negative effect of yeast extract, L-asparagine, adenine and thymine is shown and compared and contrasted with earlier reported studies.These findings supported by outside data suggest that oxidative yeasts (and bacteria) can readily elicit enzymes capable of inactivating aminopterin whereas fermentative types are lacking in this capability. Finally that past yeast-aminopterin studies were conducted with oxidative yeast types.Advantages of effective aminopterin reactor yeasts to be published elsewhere include improved ultrastructure using KMnO₄–OsO₄ fixation, a yeast bioassay procedure for detecting aminopterin in plasma and urine, and cell synchronization.Non-Standard Abbreviation apt aminopterin     

Species Distribution and Antifungal Susceptibility Profile of Oral Candida Isolates from HIV-infected Patients in the Antiretroviral Therapy Era

In this study, we investigated the yeasts colonization of genus Candida, including C. dubliniensis, isolated of HIV-infected patients oral cavities and we accessed in vitro susceptibility pattern of the Candida isolates to four antifungal agents. Out of 99 patients investigated, 62 (62.6%) were colonized with yeasts. C. albicans was the prevailing species (50%). C. dubliniensis isolates were not recovered in our study. We verified that 8.1% of the yeasts isolated were resistant to fluconazole, 8.1% to itraconazole and 3.2% to voriconazole. The isolates demonstrated very low voriconazole MICs, in which 79% (49/62) presented values of 0.015 μg/ml. All Candida isolates were susceptible to amphotericin B. The results reported here showed that although C. albicans continues to be present in one-half of oral Candida carriage of HIV-infected patients, Candida non-albicans species are increasing among these patients. Besides, the findings of resistant isolates endorse the role of antifungal susceptibility testing whenever antifungal treatment with azoles is planned.     

Comparison of volatile sulphur compound production by cheese-ripening yeasts from methionine and methionine–cysteine mixtures

Production of volatile sulphur compounds (VSC) was assessed in culture media supplemented with l-methionine or l-methionine/l-cysteine mixtures, using five cheese-ripening yeasts: Debaryomyces hansenii DH47(8), Kluyveromyces lactis KL640, Geotrichum candidum GC77, Yarrowia lipolytica YL200 and Saccharomyces cerevisiae SC45(3). All five yeasts produced VSC with l-methionine or l-methionine/l-cysteine, but different VSC profiles were found. GC77 and YL200 produced dimethyldisulphide and trace levels of dimethyltrisulphide while DH47(8), KL640 and SC45(3) produced mainly methionol and low levels of methional. S-methylthioacetate was produced by all the yeasts but at different concentrations. DH47(8), KL640 and SC45(3) also produced other minor VSC including 3-methylthiopropyl acetate, ethyl-3-methylthiopropanoate, a thiophenone, and an oxathiane. However, VSC production diminished in a strain-dependent behaviour when l-cysteine was supplemented, even at a low concentration (0.2 g l⁻¹). This effect was due mainly to a significant decrease in l-methionine consumption in all the yeasts except YL200. Hydrogen sulphide produced by l-cysteine catabolism did not seem to contribute to VSC generation at the acid pH of yeast cultures. The significance of such results in the cheese-ripening context is discussed.     

Mannan-hydrolyzing enzymes of yeasts and yeast-like organisms

The ability to degrade mannan in the yeastSaccharomyces cerevisiae, i.e. the ability to produce an enzyme of the α-mannosidase type was tested in 57 representatives of various genera and species of yeasts and yeast-like organisms. Their growth was simultaneously monitored on soluble mannan and on 4-nitrophenyl-α-D-mannopyranoside. The majority of strains produced α-mannosidase (EC 3.2.1.24).     

L-Arabinose metabolism in Candida arabinofermentans PYCC 5603T and Pichia guilliermondii PYCC 3012: influence of sugar and oxygen on product formation

l-Arabinose utilization by the yeasts Candida arabinofermentans PYCC 5603^T and Pichia guilliermondii PYCC 3012 was investigated in aerobic batch cultures and compared, under similar conditions, to d-glucose and d-xylose metabolism. At high aeration levels, only biomass was formed from all the three sugars. When oxygen became limited, ethanol was produced from d-glucose, demonstrating a fermentative pathway in these yeasts. However, pentoses were essentially respired and, under oxygen limitation, the respective polyols accumulated—arabitol from l-arabinose and xylitol from d-xylose. Different l-arabinose concentrations and oxygen conditions were tested to better understand l-arabinose metabolism. P. guilliermondii PYCC 3012 excreted considerably more arabitol from l-arabinose (and also xylitol from d-xylose) than C. arabinofermentans PYCC 5603^T. In contrast to the latter, P. guilliermondii PYCC 3012 did not produce any traces of ethanol in complex l-arabinose (80 g/l) medium under oxygen-limited conditions. Neither sustained growth nor active metabolism was observed under anaerobiosis. This study demonstrates, for the first time, the oxygen dependence of metabolite and product formation in l-arabinose-assimilating yeasts.     

The mycoflora of domesticated and wild bees (Apoidea)

Fungi including yeasts are common in the honey stomachs and provisions of diverse bees. They may be parasites, commensals or mutualistic. Yeasts, singly or in association with bacteria, are pioneer colonizers during a microbial succession in larval cells of many subterranean bees. They are followed by fungi such asAspergillus, Penicillium, Emericellopsis, Sartorya, Pseudoarachniotus, Gymnoascus, Carpenteles andFusarium. Aspergillus flavus andSaccharomyces spp. are pathogenic to many species of bees, and fungi are the main cause of declining alkali bee populations. There are 124 species of fungi, including 36 new records, associated with Apoidea; 49 species are associated with alkali bees.Cooperative investigations of the Plant Sciences Research (L. R. Batra) and Entomology (G. E. Bohart) Divisions, Agricultural Research Service, U.S. Department of Agriculture, and the Utah Agricultural Experiment Station, Logan, Utah.     

Torulopsis or Cryptococcus?

Summary Use of the generic name Cryptococcus Kützing should be avoided in yeast taxonomy, as it is both a nomen dubium and a nomen confusum. Dubium because it is at the least doubtful that the first Cryptococcus species described by Kützing (Cr. mollis) was a yeast (Kützing himself classified Cryptococcus among the Algae). Confusum because it has appeared from an examination of authentic herbarium material of Cr. mollis, that Kützing dit not deal with one, bur with a mixture of at least five organisms.The generic name Torulopsis Berlese, on the contrary, is a valid name for a group of asporogenous yeasts, since Berlese has purposely included, as appears from the diagnosis given, in his new genus Torulopsis the yeasts at that time brought to the genus Torula sensu Pasteur-Hansen.The objection which at first sight could be raised against the acceptance of the genus Torulopsis, namely that the first species described by Berlese: Torulopsis rosea, belongs to the present genus Rhodotorula Harrison is untenable, since this species certainly was not a yeast with a carotinoid pigment. All evidence available is in favour of the suggestion that Berlese's species was identical with Torulopsis pulcherrima (Lindner) Sacc.Herewith the validity of the generic name Torulopsis for the asporogenous, nonmycelium forming, colourless yeasts may be considered to be proved.A slightly amended diagnosis of the genus Torulopsis Berlese is given.     

Caratteristiche morfologiche biochimiche e biologiche di Cryptococcus ater Castellani 1960

Riassunto Nel 1960Castellani descrisse un lievito capsulato produttore di un pigmento nero, isolato da un caso di ulcere multiple della gamba. In seguitoCastellani ha isolato altri due ceppi simili al primo. I tre ceppi, secondoCastellani, appartengono ad una nuova specie di criptococco:Cryptococcus ater. Questo fungo è stato oggetto di una nota pubblicata dagli AA. nel 1963.In questa comunicazione sono riportati i dati morfologici, biochimici e biologici relativi ai tre ceppi studiati dagli AA. nel loro laboratorio. Dopo aver confermata la diagnosi di criptococco gli AA. concludono trattarsi di una nuova specie denominabileCryptococcus ater Castellani 1960.

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Summary In 1960Castellani described a capsulated yeast which produces a black pigment, isolated from multiple ulcers of the legs.Castellani subsequently isolated two more yeasts similar to the first one and he pointed out that all of them belong to a new species ofCryptococcus: Cryptococcus ater.This fungus was already object of a note published in 1963. In this communication the morphological, chemical, and biological data regarding this yeast obtained in our laboratory are reported. We confirm that it is a new species ofCryptococcus: Cryptococcus ater Castellani 1960.

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Read at the First Congress of International Society for Tropical Dermatology, Naples, June 8–13, 1964.     

Phenotypic and genotypic diversity of wine yeasts used for acidic musts

The aim of this study was to examine the physiological and genetic stability of the industrial wine yeasts Saccharomyces cerevisiae and Saccharomyces bayanus var. uvarum under acidic stress during fermentation. The yeasts were sub-cultured in aerobic or fermentative conditions in media with or without l-malic acid. Changes in the biochemical profiles, karyotypes, and mitochondrial DNA profiles were assessed after minimum 50 generations. All yeast segregates showed a tendency to increase the range of compounds used as sole carbon sources. The wild strains and their segregates were aneuploidal or diploidal. One of the four strains of S. cerevisiae did not reveal any changes in the electrophoretic profiles of chromosomal and mitochondrial DNA, irrespective of culture conditions. The extent of genomic changes in the other yeasts was strain-dependent. In the karyotypes of the segregates, the loss of up to 2 and the appearance up to 3 bands was noted. The changes in their mtDNA patterns were much broader, reaching 5 missing and 10 additional bands. The only exception was S. bayanus var. uvarum Y.00779, characterized by significantly greater genome plasticity only under fermentative stress. Changes in karyotypes and mtDNA profiles prove that fermentative stress is the main driving force of the adaptive evolution of the yeasts. l-malic acid does not influence the extent of genomic changes and the resistance of wine yeasts exhibiting increased demalication activity to acidic stress is rather related to their ability to decompose this acid. The phenotypic changes in segregates, which were found even in yeasts that did not reveal deviations in their DNA profiles, show that phenotypic characterization may be misleading in wine yeast identification. Because of yeast gross genomic diversity, karyotyping even though it does not seem to be a good discriminative tool, can be useful in determining the stability of wine yeasts. Restriction analysis of mitochondrial DNA appears to be a more sensitive method allowing for an early detection of genotypic changes in yeasts. Thus, if both of these methods are applied, it is possible to conduct the quick routine assessment of wine yeast stability in pure culture collections depositing industrial strains.     

Mating-type differentiation in ascosporogenous yeasts on the basis of mating-type-specific substances responsible for sexual cell-cell recognition

Summary Sexual agglutination occurred only between cells of opposite mating types of the same species in all the Sacharomyces, Hansenula, Saccharomycodes, and Pichia yeasts tested. We succeeded in solubilizing the sex-specific glycoprotein, cell wall agglutination substance responsible for sexual agglutinability by briefly autoclaving these yeasts. The agglutination substances of all the above yeasts were univalent and sensitive to the enzyme pronase. The formation of complementary complexes was observed only between agglutination substances of opposite mating types of the same species. In general, the agglutination substance of one mating type was more resistant to heat treatment at 100°C in 3% acetic acid and more sensitive to 5% 2-mercaptoethanol treatment than the agglutination substance of the other mating type in these yeasts. On the basis of these results together with the pheromone response and production, we expect that almost all ascosporogenous yeasts can be classified into the two mating types corresponding to a and mating types in Saccharomyces cerevisiae, respectively.     

Cryptococcus nodaensis sp nov, a yeast isolated from soil in Japan that produces a salt-tolerant and thermostable glutaminase

An anamorphic basidiomycetous yeast, which produced a salt-tolerant and thermostable glutaminase, was isolated from soil in Japan and classified in the genus Cryptococcus. Its substrate specificity suggests that this enzyme is an L-glutaminase asparaginase (EC 3.5.1.38). The strain, G60, resembles Cryptococcus laurentii in the taxonomic criteria traditionally employed for yeasts, however it can be distinguished as a separate species based on DNA–DNA reassociation experiments and sequence analysis of the large sub-unit rDNA. Phenotypically, the isolate can be differentiated from C. laurentii by the inability to utilize arbutin as a sole source of carbon. Based on sequence analysis, the strain is related to a group of hymenomycetous yeasts including Bulleromyces albus, Bullera unica, C. laurentii and C. skinneri. The strain, which is formally described as Cryptococcus nodaensis, is industrially important for the formation of the umami taste during production of proteolytic seasonings. Received 28 July 1998/ Accepted in revised form 04 February 1999     

Production of a potentially novel anti-microbial substance by Bacillus polymyxa

A bacterial strain, SCE2, identified as Bacillus polymyxa, produced an anti-microbial substance active against yeasts, fungi and different genera of Gram-positive and-negative bacteria, in liquid medium and in plate assays. This substance appeared to be an antibiotic different from the polymyxin group, mainly because of its action against the majority of Gram-positive bacteria tested and its lack of activity against Pseudomonas aeruginosa, a species usually killed by polymyxins. Preliminary characterization showed resistance to heat (65°C, 2 h), to proteases, trypsin, lysozyme, deoxyribonuclease I, ribonuclease A, phospholipase C, ethanol, acetone, chloroform, ether and to strong alkali treatment (2 M NaOH). The molecular weight was less than 3500. The B. polymyxa strain harboured a plasmid that did not correlate with antibiotic production; after curing experiments, a derivative strain, SCE2(46), was isolated that lacked the plasmid pES1, but showed the same inhibitory spectrum as the wild-type strain.