Ferment Galactose Research Articles

Isolation of an extremely thermophilic chemoorganotrophic anaerobe similar to Dictyoglomus thermophilum from new zealand hot springs

A strain similar to Dictyoglomus thermophilum, isolated from a New Zealand hot spring, is described. This strictly anaerobic, Gram-negative, non-motile and nonsporulating bacterium usually exists as long thin filaments of 5 to 25 μm by 0.35 to 0.45 μm. Rotund bodies are commonly observed. Thin sections of the cells revealed a two-layered cell wall. The optimum temperature and pH for growth was 70°C and 7.0 and 7.5 respectively. No growth was observed at 40°C and 85°C or at pH 4.5 to pH 9.0. The organism fermented glucose, maltose, mannose, xylose, lactose, cellobiose, galactose and sucrose and produced acetate as the major end-product with significant amounts of lactate, H2 and CO2 and only traces of ethanol. The doubling time on glucose was 10 h. The DNA base composition was 29.5% guanine plus cytosine as determined by the thermal denaturation method. Growth was inhibited by penicillin, tetracycline and chloramphenicol indicating that the organism was a eubacterium. These features are in common with the newly described species Dictyoglomus thermophilum to which the New Zealand isolate belongs.

Studies on the metabolic activation of diethanolnitrosamine in animal-mediated and in vitro assays using Escherichia coli K-12 343/113 as an indicator.

The mutagenic activity of diethanolnitrosamine (NDELA), a carcinogenic compound which leads to inconsistent results in standard in vitro procedures was tested in vitro and in animal-mediated assays with the indicator strain Escherichia coli (E. coli) K-12 343/113. This strain allows the simultaneous detection of forward and back mutations arising in several genes of the E. coli chromosome. In animal-mediated assays in which mice were used as hosts for i.v. injected E. coli indicator cells, s.c. application of NDELA induced a dose dependent increase of galactose fermenting mutants in cells recovered from the livers of animals exposed for 3 h to the mutagen. Comparison with results obtained with diethylnitrosamine (DENA) in the same test system revealed that the two compounds apparently cause different types of mutagenic lesions. Induction of arg+ mutations by DENA and several other aliphatic nitrosamines is mainly due to base pair substitutions, whereas NDELA is rather mutagenic in the galRs system. This latter system is, in addition, sensitive to frameshifts and deletions. These differences in mutagenic specificity suggest that NDELA and DENA, although structurally closely related, are activated via different molecular mechanisms. In fact, evidence is accumulating that alcohol dehydrogenase (ADH) could be involved in the activation of NDELA. On the other hand, the effective mutagenesis of NDELA obtained in vitro with E. coli upon addition of rat liver microsomal fraction would not be expected if ADH is involved in the activation since the S-9 Mix used in the present experiments was devoid of cofactors (NAD, NADP), necessary to accomplish oxidation by ADH. Therefore, further in vivo studies were performed, in which pyrazole, a potent blocker of ADH, was administered prior (1 and 24 h) to the injection of the mutagen. The observation that a dose dependent increase of mutants in the liver (and to a lower extent in the spleens) of treated animals takes place under conditions in which ADH activity is blocked, whereas several microsomal enzymes are stimulated, indicated that besides oxidation of NDELA by ADH other metabolic activation pathways are involved. Apparently enzymes contained in the liver homogenate, possibly NADPH dependent enzymes of the microsomal ethanol oxidizing system, play an important role in the formation of mutagenic metabolites of NDELA.

A modified scheme for biotyping Gardnerella vaginalis.

A scheme is proposed for biotyping Gardnerella vaginalis, based on detection of hippurate hydrolysis, beta-galactosidase (ONPG) and lipase, and fermentation of arabinose, galactose and xylose. Seventeen biotypes were found among 197 strains from asymptomatic women and patients with bacterial vaginosis (non-specific vaginitis). The distribution of biotypes was similar in both populations but some biotypes were found more frequently in patients. The proposed scheme is compared with those previously described.

Nonenzymatic Browning of Mozzarella Cheese

Mozzarella cheese was made with combinations of Streptococcus thermophilus nongalactose (Gal−) and galactose fermenting (Gal+) strains and Lactobacillus helveticus (Gal+) and L. bulgaricus (Gal−). Galactose was found in all Mozzarella cheese regardless of the culture used. The highest concentration was in cheese made with Streptococcus thermophilus Gal−L. bulgaricus Gal− combination with Streptococcus thermophilus Gal+Lactobacillus helveticus Gal+ having the least. Little if any lactose was found in any of the cheeses. The temperature and time during stretching of the curd inhibited fermentation of the residual galactose.Galactose accumulated in Mozzarella when either strain of Streptococcus thermophilus was used. The fermentation of accumulated galactose was the result of metabolism by Lactobacillus helveticus. There was a positive correlation between galactose content and brown color intensity when Mozzarella cheese was heated. A predictive test was effective in evaluating the browning tendency of cheese.

Factors Affecting Nonenzymatic Browning of Process Cheese

Variations in manufacturing procedures and the composition of stirred-curd Cheddar cheese were investigated for effects on nonenzymatic browning in process cheese. Using a nongalactose-fermenting Streptococcus thermophilus culture as an adjunct to mesophiic lactic starter culture resulted in increased galactose concentration in cheese and intensified browning of process cheese. The correlation between galactose content in stirred-curd cheese and brown color intensity in process cheese was very high (r=.929). Higher salt-in-moisture ratios of stirred curd cheese were related to more intense brown color in process cheese. The high salt-in-moisture ratio probably reduced the catabolic activity of the lactic starter and resulted in more residual lactose and galactose in the cheese. Increasing available amino groups in stirred curd cheese by adding casein hydrolysate proved inconclusive. Faster cooling of process cheese from 80 to 37.7°C reduced intensity of brown color. Process cheese cooled very rapidly did not develop any brown color, regardless of sugar and salt contents. Browning may be controlled by regulating salt and by using strains of Streptococcus thermophilus that ferment galactose.

The use of mini-Gal plasmids for rapid incompatibility grouping of conjugative R plasmids

The galactose operon of Escherichia coli K-12 has been used as a phenotypic marker for miniplasmids derived in vitro from R plasmids representing six incompatibility groups. This has enabled the development of a rapid incompatibility typing scheme in which the miniplasmids are used as incompatibility exemplars, their presence in strains being monitored on galactose fermentation indicator media.

Inability of Petite Mutants of Industrial Yeasts to Utilize Various Sugars, and a Comparison with the Ability of the Parent Strains to Ferment the Same Sugars Microaerophilically

A number of industrial strains of Saccharomyces cerevisiae were converted to the petite form and tested for the ability to utilize galactose, maltose, sucrose, α-methyl glucoside and raffinose. The parent strains all metabolized these sugars aerobically. Twelve of the petite forms did not utilize galactose, six failed to utilize maltose, 17 did not utilize x-methyl glucoside, and 18 did not utilize raffinose. The petites of two distiller’s yeast strains did not utilize sucrose. The respiratory-competent parent strains nearly all fermented galactose, maltose, sucrose and raffinose, though 19 strains did not ferment α-methyl glucoside microaerophilically. Three strains did not ferment galactose, two fermented it only after several days adaptation, one did not ferment raffinose, and two did not ferment sucrose under microaerophilic conditions. Six respiratory-competent strains which did not utilize galactose when in the petite form fermented higher (10%) concentrations of glucose and maltose under microaerophilic conditions, but only three of these fermented galactose. The implications of these findings for the use of such strains in industry are discussed briefly.

Galactose Fermentation and Classification of Thermophilic Lactobacilli

The ability to ferment galactose is a major characteristic which can be used to differentiate Lactobacillus helveticus (galactose positive) from Lactobacillus lactis and Lactobacillus bulgaricus (galactose negative). In milk cultures, galactose-positive strains produced d- and l-lactic acid with little galactose accumulation, whereas galactose-negative strains produced d-lactic acid, and galactose accumulated to high levels.

Use of mitochondrial mutants in the isolation of hybrids involving industrial yeast strains

A hybrid of Saccharomyces diastaticus x Hansenula capsulate was obtained by conversion of the S. diastaticus parent to a petite, protoplasting and fusion of the strains and isolation of the hybrids on selective media, containing glycerol or galactose. All of the single-cell clones obtained resembled S. diastaticus in morphology, and fermented glucose, galactose, maltose, sucrose and starch. The hybrid sporulated, producing spores resembling those of S. diastaticus, but in asci which differed somewhat morphologically. The stability of the hybrid depended on the composition of the maintenance media, as hybrid clones maintained on glycerol medium lost their ability to sporulate after a few transfers and the cellular morphology altered to a form more closely resembling H. capsulate, and those maintained on glucose-containing medium lost the ability to utilize glycerol. Cultures maintained on yeast extract-starch medium retained the ability to sporulate and utilize glycerol. However, dissection of asci from the hybrid yielded clones, some of which had lost the ability to metabolize glycerol but which still sporulated. These clones had an unaltered cytochxome spectrum, and in other ways appeared to be respiratory competent. Washed cells of hybrids grown on galactose or starch as sole carbon source metabolized starch, as shown by respirometric measurements, but cells maintained on glycerol as sole carbon source had lost this ability. There was some evidence that the addition of vitamins stimulated oxygen uptake on starch by the hybrids. The formation of sporulating hybrids in the cross may permit the genetic analysis of other Saccharomyces x Hansenula crosses.

Saccharomyces uvarum inulyticus var. nov., a new high-concentration ethanol tolerant yeast from rice wine

A new alcohol tolerant yeast which had the ability to ferment glucose, galactose, maltose, trehalose, melizitose, α-methyl-d-glucoside, melibiose and raffinose (complete) was isolated from fermented rice wine. This new isolate which was morphologically, culturally and physiologically similar to type species Saccharomyces uvarum Beijerinck was considered as a variety of the species, hence, designated as Saccharomyces uvarum inulyticus var. nov., because of its extraordinary ability to ferment inulin into ethanol and hyper-tolerance to high concentration of ethanol at a wide range of temperature. The new variant produced 21.6%, 20%, 18.6%, and 9.8% v/v ethanol at 15, 20, 30, and 40° C, respectively, when fermentation was carried out by the stepwise addition of sucrose to the synthetic medium. The addition of fungal mycelia into the synthetic medium increased the fermentation rate and ethanol concentration up to 22.4%, 21.8%, 20.1%, and 11.6% v/v at 15, 20, 30, and 40° C, respectively. The new variant was found to grow well at 40° C and did not require any vitamin or growth factor, such as biotin, calcium pantothenate, inositol, niacin, p-aminobenzoic acid, pyridoxine hydrochloride, riboflavin and thiamine hydrochloride for normal growth.

Novel Kefir yeast having remarkable sugar utilization. Part I. Identification and fermentation character of Kefir yeast.

A novel yeast KY-5 was isolated from Danish Kefir grains grown in skimmed milk and identified as Torulopsis holmii by the morphological, physiological and chemical method with analyses of its electrophoresis, coenzyme Q system and DNA base composition. Glucose, galactose, mannose and sucrose were fermented by the yeast, but lactose was not fermented. An inductive period was not required for galactose fermentation, even though the yeast was cultivated in a glucose medium. On the other hand, an inductive period was necessary for fructose fermentation. The yeast preferentially metabolized galactose in a glucose and galactose mixed medium. According to Warburg respirometric analyses of the intact cell, protoplast cell and cell free extract of the yeast, the remarkable sugar utilization might be dependent on constituent enzymes involved in the sugar intake and metabolism. The enzymes seemed to be quite different from those of usual yeasts.

The use of mitochondrial mutants in hybridization of industrial yeasts

Protoplasts of petites of strains 625-C(I) of Saccharomyces diastaticus and NCYC 1085 of Saccharomyces cerevisiae, originally obtained from the National Collection of Yeast Cultures, England, were fused with protoplasts of Candida pseudotropicalis, Saccharomyces rosei, Yaccharbmycesmontanus, Pichiamembranefaciens, Hansenula anomala, Hansenula capsulata, and Schizosac-charomyces pombe. The respiratory-competent products of the fusions were selected on the basis of using at least one of the carbon sources utilized by the petite parent and not by the other. The products of the fusion of C. pseudotropicalis x 1085(p(-)) consisted of two cell types; an oval cell which utilized both lactose and maltose and fermented lactose vigorously, and a cylindrical form which fermented maltose slowly. The S. rosei x 1085(p(-)) hybrids had acquired the ability to metabolize and ferment galactose, and to ferment maltose, from the petite parent. The P. membranaefaciens x 625(p(-)) hybrids acquired the ability to metabolize galactose, sucrose and maltose, but fermented only glucose, weakly, like the P. membranaefaciens parent strain. The H. capsulate x 625(p(-)) hybrids, unlike the hybrids with P. membranaefaciens or S. rosei, resembled the petite parent morphologically and also had the fermentative abilities of this strain (galactose, maltose, sucrose and starch), and the ability to ferment starch was considerably enhanced. The S. montanus x 625(p(-)) hybrids acquired the ability to utilize starch. Schizosaccharomyces pombe x 625(p(-)) hybrids resembled S. pombe morphologically, but had the ability to metabolize galactose and starch. Some of the asci produced by these hybrids contained abnormal numbers of spores. H. anomala x 624 x(p(-)) hybrids fermented starch, though weakly.

Clustering of phenotypic variables and construction of an identification scheme for human isolates of Haemophilus

Summary Isolates of Haemophilus from clinical laboratories were subjected to 35 biochemical tests with the data posted to a random matrix. Following rearrangement of the information, 13 clusters organisms were identified. H. influenza was found to be a species of considerable heterogenity that could be divided into a number of the clusters. Important tests for the identification of Haemophilus were determined to be indole production and fermentation of fructose, xylose, maltose, galactose, mannose and sucrose.

IMP1/imp1: a gene involved in the nucleo-mitochondrial control of galactose fermentation in Saccharomyces cerevisiae.

In some strains of Saccharomyces cerevisiae, the induction of enzymes of the Leloir pathway, galactose fermentation and growth on galactose depend on mitochondrial function; mitochondrial dependence is elicited through the recessive allele imp1 of the nuclear gene IMP1. The genetic element IMP1 is not allelic to any of the known GAL genes; IMP1 strains can grow on and ferment galactose in respiratory-deficient (RD) condition or in the presence of the mitochondrial inhibitors ethidium bromide and erythromycin; whereas, imp1 strains can grow on and ferment galactose only in respiratory-sufficient (RS) condition. The imp1 elicited mitochondrial dependence apparently involves regulation of the synthesis of the galactose catabolizing enzymes and synthesis of the galactose specific permease. IMP1 is not the only genetic determinant that elicits an interaction of the mitochondrion and the expression of the Gal system; the GAL3 gene, whose role in galactose utilization is demonstrated by the long-term adaptation phenotype of gal3 rS mutants, gives rise to a noninducible phenotype in RD condition or in the presence of mitochondrial inhibitors.

Candida guilliermondii, an Actidione‐resistant Yeast from Wine*

Candida guilliermondii var. guilliermondii, a yeast which showed high resistance to actidione, was isolated from a red wine sample. Growth in 5000 parts/106of actidione was equal to that of the control. Sorbic acid and sodium benzoate were most effective in checking the growth at 250 parts/106but growth occurred at 750 parts/106of potassium metabisulphite. Fermentation of glucose, galactose, sucrose and raffinose was not inhibited in the presence of 1000 parts/106actidione. This is the first report on the occurrence of actidione‐resistant yeasts in the genus Candida and on the isolation of C. guilliermondii var. guilliermondii from wine.

Galactose fermentation by Streptococcus lactis and Streptococcus cremoris: pathways, products, and regulation.

All of the lactic streptococci examined except Streptococcus lactis ML8 fermented galactose to lactate, formate, acetate, and ethanol. The levels of pyruvate-formate lyase and lactate dehydrogenase were elevated and reduced, respectively, in galactose-grown cells compared with glucose- or lactose-grown cells. Reduced intracellular levels of both the lactate dehydrogenase activator (fructose, 1,6-diphosphate) and pyruvate-formate lyase inhibitors (triose phosphates) appeared to be the main factors involved in the diversion of lactate to the other products. S. lactis ML8 produced only lactate from galactose, apparently due to the maintenance of high intracellular levels of fructose 1,6-diphosphate and triose phosphates. The growth rates of all 10 Streptococcus cremoris strains examined decreased markedly with galactose concentrations below about 30 mM. This effect appeared to be correlated with uptake predominantly by the low-affinity galactose phosphotransferase system and initial metabolism via the D-tagatose 6-phosphate pathway. In contrast, with four of the five S. lactis strains examined, galactose uptake and initial metabolism involved more extensive use of the high-affinity galactose permease and Leloir pathway. With these strains the relative flux of galactose through the alternate pathways would depend on the exogenous galactose concentration.

Genetic recombination between Proteus mirabilis and Providencia alcalifaciens.

Chromosome transfer occurred in plate matings between Proteus mirabilis strain PM5006 and Providencia alcalifaciens strain P29 in either direction with the use of plasmid D or R772 as sex factor. Auxotrophic chromosomal markers of recipients were converted to prototrophy and the galactose fermentation marker of donor PM5006 could also be selected. Recombination frequencies for a group of selected markers in PM5006(D) x P29 matings varied between 3 x 10(-5) (trp+) and 1.2 x 10(-7) (lys+) per donor. In the reverse cross, plasmid D mobilized markers on the P29 chromosome randomly with a recombination frequency of about 1.7 x 10(-7) per donor for all selected P29 markers. R772 produced random mobilization of markers on both chromosomes yielding recombinants at a frequency of about 1.8 x 10(-7) per donor. Unselected markers separated by no more than about 10 min from selected markers on the PM5006 chromosome were cotransferred from P29 by both plasmids. Despite the low degree of DNA homology existing between the two species, all hybrids behaved as stable haploids. Progeny from P29(D or R772) x PM5006 auxotroph matings displayed similar sets of naturally occurring P29 unselected markers irrespective of the selected prototroph allele. In reverse crosses, a similar range of PM5006 naturally occurring unselected markers registered in P29 recipients, although differences existed in the sets of markers mediated by the two plasmids. Weak linkage was detected between PM5006 gal+ allele(s) and some P29 auxotroph markers. Adsorption of donor-specific phages 5006M or PL25 to hybrids could not be demonstrated and many recombinants failed to express some or all of the plasmid markers.

Salient features of Haemophilus vaginalis

A total of 78 strains of Haemophilus vaginalis were examined for 104 features. All strains fermented dextrin, maltose, and starch. Additionally, more than 90% of the strains fermented galactose, glucose, and ribose. Arbutin, cellobiose, melibiose, rhamnose, and salicin were not fermented by any of these strains. None of the strains acidified any of 14 alcohols or alkalinized any of 25 organic salts and amides. More than 90% of the strains hemolyzed human blood agar and hydrolyzed hippurate. No strain hemolyzed sheep blood agar. A recommendation is included for those minimal features that best differentiate H. vaginalis from other oxidase- and catalase-negative, gram-negative organisms.

A Novel Pleiotropic Mutation in Escherichia coli K12 which Affects Transduction, Transformation and Rates of Mutation

A mutant strain of Escherichia coli K12, R2721, has been shown to differ from its parent strain, S491, in four associated phenotypic characters as a result of a single mutation. This strain did not give recombinants with DNA transduced by bacteriophage PI or bacteriophage Mu, nor transformats after exposure to R factor DNA: lysates of bacteriophage PI grown on this strain did not appear to contain any transducing particles when tested on normal recipients. Moreover, the reversion rates, both spontaneous and ultraviolet-induced, for two auxotrophic markers were reduced. The frequency of revertants was at least two orders of magnitude lower in cultures of R2721 than in cultures of S491I. Many of the rare revertants for one or other of the auxotrophic markers were found to have regained normal reversion frequencies for the other marker and for the capacity to be transduced. In all other respects, recombination in R2721 appeared normal, the frequency of chromosomal mobilization by and F' factor was unaffected and normal yields of recombinants were obtained from matings with Hfr strains. The only circumstance in which transduction of R2721 was observed was when the capacity to ferment galactose was selected and PI had been grown on a strain carrying lambdadgal when, presumably, integration was effected by the phage-coded gene products. The mutation has been located on the E. coli chromosone map between tonA and pro and has been given the symbol tdi (transduction inhibition). Double mutants, (tdi recA) and (tdi recB), have been isolated and show no unexpected properties.

An evaluation of D-glucosamine as a gratuitous catabolite repressor of Saccharomyces carlsbergensis

Glucose represses mitochondrial biogenesis and the fermentation of maltose, galactose and sucrose in yeast. We have analyzed the effect of D-glucosamine on these functions in order to determine if it can produce a similar repression. It was found that glucosamine represses the respiration rate (QO2) but more rapidly than glucose and to a final level slightly higher than in glucose-treated cells. Derepression of the respiration rate following either glucose or glucosamine repression was similar. A two hour lag was followed by a linear increase in QO2 to the derepressed level. Both glucose and glucosamine repressed the level of cytochrome oxidase to the same level. Glucosamine was also found to repress maltose and galactose fermentation but not sucrose fermentation. The derepression of maltase synthesis was inhibited by glucosamine. The constitutive synthesis of maltase was repressed by the addition of glucosamine. Glucosamine was judged to produce a repressed state similar to glucose repression in many respects.