Yeast Extract Broth Research Articles

Induction of mutation in Lactobacillus casei subsp. alactosus by nitrosoguanidine.

A culture of Lactobacillus casei subsp. alactosus was treated with N-methyl-N '-nitro-N-nitrosoguanidine and 107 possible mutants were isolated. There was considerable variation among mutant cultures in total acidity and production of diacetyl when grown in soymilk. Two of them coagulated reconstituted skim milk within 48 hr. About 36% of isolated mutant cultures produced larger amounts of diacetyl in soymilk than the parent strain. Only 14% of isolates produced higher amounts of acid, and the capacity to produce acid of others was partially impaired. Approximately 58% of tested mutants were found to possess unstable characters after 20 daily transfers in tryptone yeast extract broth. Cultural studies on five selected mutants indicated that two of them ferment lactose and sorbose in contrast to other tested mutants and the parent. However, their morphologies were similar to the parent.

Enrichment of linear alkylbenzenesulphonate (LAS) degrading bacteria in continuous culture

AbsractEnrichment experiments were carried out in continuous‐flow units using a mineral medium with commercial linear alkylbenzenesulphonate (LAS) as the limiting carbon‐ and energy‐source. The mixed bacterial culture originating from the waste water of a detergent plant consisted of five strains belonging to the genus Pseudomonas and two strains each of the genera Achromobacter and Acinetobacter. The cultivation conditions corresponding to dilution rates of 0.025‐0.1 h‐1 and LAS concentrations of 20–50 mg/1 were examined. During the experiments the composition of mixed cultures and the kinetics of LAS biodegradation were followed. Continuous‐flow enrichment experiments resulted in the selection of six bacterial cultures with different compositions of individual species and capability to utilize LAS. From the original seven strains at lower dilution rates (0.025 and 0.05 h‐1) six were selected, excluding Pseudomonas sp. 3, while at the highest dilution rate (0.1/h‐1) five strains were selected after eliminating Pseudomonas sp. 5 and Achromobacter sp. 1. All enriched mixed cultures were more efficient in primary than in ultimate LAS degradation. Two of the culture strains were able to achieve primary LAS degradation (Pseudomonas sp. 1 in mineral medium with LAS as the sole carbon‐ and energy‐source and Acinetobacter sp. 3 in medium supplemented by yeast extract and nutrient broth).None of the strains could degrade LAS completely, which indicates that many types of interactions based on combined metabolic attack as well as those based on provision of specific nutrients, may exist between culture members during the complete LAS bio‐oxidation.

Cocultivation of Legionella pneumophila and free-living amoebae.

Studies of the interaction of Legionella pneumophila with free-living amoebae showed that Naegleria lovaniensis and Acanthamoeba royreba could use L. pneumophila as a sole food source. However, growth of the amoebae on nonnutrient agar plates seeded with L. pneumophila was slower than growth on nonnutrient agar plates seeded with Escherichia coli. On inoculation of L. pneumophila into axenic cultures of N. lovaniensis and A. royreba, 99.9% of the L. pneumophila was destroyed within 24 h. After several weeks, however, some amoeba cultures became chronically infected and supported the growth of L. pneumophila. Amoebae exposed to L. pneumophila and containing adhered L. pneumophila, L. pneumophila antigens, or both, showed no increased pathogenic potential on intranasal inoculation of weanling mice. Similarly, L. pneumophila propagated in chronically infected amoeba cultures showed no increase in virulence on intraperitoneal inoculation of guinea pigs relative to L. pneumophila grown in yeast extract broth.

Virulence conversion of Legionella pneumophila serogroup 1 by passage in guinea pigs and embryonated eggs.

When virulent Legionella pneumophila is passaged on supplemented Mueller-Hinton agar, it remains virulent for guinea pigs and embryonated hen eggs for two passages. However, by the fifth passage the cultures become avirulent for guinea pigs. Flagella were not produced by L. pneumophila on the first passage on supplemented Mueller-Hinton agar. In contrast, 12 passages on charcoal-yeast extract agar did not result in the reduction of virulence or the loss of flagella of L. pneumophila. Growth in supplemented yeast extract broth or on Norit-A-filtered supplemented yeast extract agar also did not result in a reduction of the virulence of L. pneumophila. However, L. pneumophila did not produce flagella when grown on these two media. Thus, it appears that the production of flagella is not required for the virulence of L. pneumophila when administered by the intraperitoneal route of infection. A virulent flagellated form of L. pneumophila was recovered by passing an avirulent form six times in guinea pigs. When avirulent L. pneumophila was passaged 12 times in embryonated eggs, a nonflagellated form of the bacterium was recovered which had an increased virulence for guinea pigs and embryonated eggs. However, virulent forms were not recovered by passage of avirulent forms on commonly used laboratory media. These results support the suggestion that a suitable host is required for the selection of the virulent form of L. pneumophila from avirulent cultures.

Enterobacteria Differentiated by Gas-Liquid Chromatography of Metabolites

The short chain acids of 47 strains within the family Enterobacteriaceae grown on prereduced, anaerobically sterilized peptone yeast-extract glucose broth (PRAS PY G) under standardized conditions were determined by gas-liquid chromatography (GLC). The bacteria could be subdivided into six groups on the basis of their metabolites. Acetate, lactate, succinate and ethanol were detected from all strains and constituted a set of basic fermentation products, P, mirabilis, in addition, produced propanol, and was distinguished from P. morganii which formed propionate, pyruvate, methanol and oxalacetate, P. rettgeri only produced methanol along with the basic metabolites. Butanol and 2, 3-butanediol were characteristic of bacteria with the butanediol modification of mixed-acid fermentation, but could only be detected after anaerobic growth. Additionally, Hafnia produced propanol, which differentiated these strains from Enterobacter, Klebsiella, Serratia and Erwinia. The results indicate that GLC of metabolites may render supplementary information utilizable in identification of enteric bacteria.

The ompB locus and the regulation of the major outer membrane porin proteins of Escherichia coli K12

Insertion and spontaneous mutants defective in the production of the major outer membrane protein, OmpF § § Abbreviations used: Several nomenclature systems have been used for the major outer membrane proteins of Escherichia coli. To avoid confusion, we give the protein the same name as the structural gene. For example, the ompF gene specifies the OmpF protein. Certain ompB alleles, isolated in different laboratories, have been given different names. The ompB allele that confers an OmpF − OmpC + phenotype has also been called tpo (Wandersman et al., 1980) and perA (Wanner et al., 1979). Genetic nomenclature is from Bachmann & Low (1980). The symbol Φ designates fusions (Muller-Hill & Kania, 1974; Silhavy et al., 1976), which may be of two types: those that result in production of a hybrid protein caused by fusion of two structural genes (protein fusions) and those that do not result in the production of a hybrid protein (operon fusions) (see Fig. 1), but only place a gene under the regulatory control of a different promoter. Protein fusions and operon fusions are designated as Φ( ompF- lacZ)hyb and Φ(ompF-lacZ +), respectively. Fusion components are written in their order of transcription. The superscript c, as in ompC c, indicates a mutation causing constitutive expression of the gene. Bacteriophage λ nomenclature is from Hershey (1971). Primes, as in ompF′, indicate that the DNA for the genetic region referred to is deleted on the side the prime is written. The symbol::, as in ompF::Tn5, indicates the site of a genetic insertion. Tn5 is a translocon conferring kanamycin resistance (Berg, 1977). The symbol Δ indicates a deletion. Phenotype abbreviations are Lac, lactose; Kan, kanamycin; s, sensitive; r, resistant; Ts, temperature-sensitive. Other abbreviations are: NB, nutrient broth; TY broth, Tryptone yeast extract broth; TSB, tryptic soy broth; XG, 5-bromo-4-chloro-3-indolyl-β- d-galactoside. , were selected by tolerance to colicin L. Eight of the insertion mutations were insertions of the genome of bacteriophage Mu at ompF. One insertion was the kanamycin resistance translocon Tn5, also in ompF. The Mu c(Ts) insertions at ompF were used to construct both ompF-lac operon and protein fusions. These fusions and ompC-lac fusions isolated previously (Hall & Silhavy, 1979) enabled us to study the regulatory properties of the ompF and ompC genes and to further clarify the role of the ompB locus. The results indicate that the ompB locus performs the dual role of regulating both the absolute and relative transcriptional levels of expression of the ompF and ompC genes.

Flagella on Legionnaires' disease bacteria: ultastructural observations.

Scanning and transmission electron microscopic studies were done on five strains of flagellated Legionella pneumophila cultured for 1 to 3 days on charcoal yeast extract agar or yeast extract broth. Ultrastructurally, each strain consisted of pleomorphic, flagellated bacilli, many of which showed the typical pinching type of division as seen with other gram-negative bacteria. Most of the flagellated organisms, regardless of the strain, had a single, relatively straight or undulant polar flagellum, about 25 nm in diameter. In some instances, retraction of the bacterial cell membranes made the flagellar basal bodies (bulbs) visible when observed by transmission electron microscopy. The bulbous nature of these bodies appears to be different from the basal structures described for Escherichia coli and other gram-negative bacteria. Long "streamers," previously observed with the light microscope, appear to be fimbriae (or pili) that have a diameter approximatley half that of the flagella and are somewhat coiled and bent at irregular intervals.

Liquid medium for growth of Legionella pneumophila.

The medium described is a simple yeast extract broth capable of growing large number of Legionella neumophila, the causative organism of Legionnaires disease. Filtration was chosen as a means of sterilization, since medium that was autoclaved did not support growth without the presence of Norite A. The filtered medium gave rapid cell growth and maintained the initial antigen production. The observed generation time was 99 min with a maximum cell population of 2 X 10(2) COLONY-FORMING UNITS PER ML IN APPROXIMATELY 40 H.

Environmental Factors Influencing Growth and Sporulation of Cercospora Kikuchii

Cercospora kikuchii grew in 10 different liquid or solid media. Most growth was obtained in V-8-juice broth, yeast-extract and potato-dextrose broth, and the least growth occurred in nutrient broth or sterile distilled water. The fungus sporulated abundantly on V-8-juice agar and carrotleaf-decoction agar, moderately on nutrient agar, yeast-extract agar and no conidia formed on water agar, cornmeal agar, malt-extract agar, malt agar or potato dextrose after 14 da at 28 C. At 13 C, the fungus sporulated in continuous light and in alternating light-dark periods (12 h/12 h) on V-8 juice agar, but in continuous darkness a 60% reduction in sporulation was noted. Higher temperatures (22 or 28 C) seemed to counteract this phenomenon, as no significant differences in conidial counts were obtained at 28 C in continuous darkness, alternating light and dark periods, or continuous light.

Acetic acid and hydrogen metabolism during coculture of an acetic acid producing bacterium with methanogenic bacteria.

Two microorganisms originally existing as a mixed culture obtained from an anaerobic digester fluid were separated for pure and coculture studies. One of these was motile, Gram-negative, and non-sporeforming, and it required yeast extract for growth and acetic acid production. This isolate produced H2 and did not need H2 and (or) CO2 for growth and acetate formation. The other isolate was a methanogen whick resembled Methanobacterium arbophilicum in morphology and substrate specificity. Coculture growth of the two isolates in yeast extract broth (80% N2--20% CO2 gas phase) indicated that the non-methanogen produced up to four to five times more H2 than when grown separately. Although the growth of the non-methanogen was not enhanced by the removal of H2 by the methanogen, the hydrogen produced was essential for the growth of methanogen. Similar results were obtained when the non-methanogen was cocultured with Methanospirillum hungatti GP1. Cultivation of the non-methanogen in the presence of M. hungatti GP1 (under abundance of 80% H2--20% CO2) indicated that the acetate produced was consumed by M. hungatii, without inhibiting the growth of the other culture.

Collaborative Study of a Method for the Detection of Clostridium botulinum and Its Toxins in Foods

The mouse toxicity and protection technique for the detection and identification of Clostridium botulinum and its toxins in foods was collaboratively studied by 11 laboratories. Each laboratory received 4 samples of cream of mushroom soup; 2 contained spores and toxin of C. botulinum type A, 1 contained spores and toxin of C. botulinum type E, and 1 contained spores of C. sporogenes. The media used were cooked meat medium (beef heart or chopped liver broth) and trypticase peptone glucose yeast extract broth with trypsin. The results indicate that this method has a high degree of repeatability and reproducibility. All 11 laboratories correctly identified the toxins and the nontoxic sample in the food and detected and identified the viable spores in the samples by means of the subsequent cultures. This method has been adopted as official first action.

Experiments with haliscomenobacter hydrossis in continuous culture without and with zoogloea ramigera

A study has been made of the sheath-forming bacterium Haliscomenobacter hydrossis, by growing the organism in continuous culture. Glucose together with components from trypticase soy broth/yeast extract were used as the energy source. At low dilution rates glucose was a more important energy source than components from trypticase soy broth/yeast extract. H. hydrossis was unable to produce large amounts of reserve material even in the presence of an excess of glucose. The minimum doubling time was approximately 13 h in the complex medium. The uptake sequence of amino acids derived from the trypticase soy broth yeast extract was found to be as follows: glutamic acid, glycine, methionine, tryptophan, lysine and arginine were easily consumed by the micro-organism, whereas the other amino acids were taken up mainly at low dilution rates. Competition experiments were carried out with H. hydrossis and Z. ramigera at two different dilution rates (0.016 and 0.005 h -t). At both dilution rates Z. ramigera was dominantly present in the cultures. Finally the glucose respiration and glucose uptake of a washed cell suspension of H. hydrossis was studied. The oxygen uptake rate increased only very slightly upon addition of glucose. The difference between endogenous respiration and oxygen uptake rate in the presence of glucose was increased when the washed cells had been aerated a period of time.

Oxidation of arsenite to arsenate by Alcaligenes faecalis.

Alcaligenes faecalis, resistant to the toxic effects of 0.01 M sodium arsenite, was isolated from raw sewage and shown to be capable of oxidizing arsenite to arsenate. When the organisms were grown in chemically defined medium, this conversion was due to the appearance at stationary phase of an intracellular, oxygen-sensitive, inducible enzyme and/or component of the electron transport system; when the organisms were grown in a nutrient broth-yeast extract medium, the enzyme appeared in the late exponential phase of growth. The presence of 0.02 M arsenite in the culture medium affected neither growth rate nor final cell yield.

A Simplified Procedure for Embedding Dilute Suspensions of Biologic Material.

Electron microscopic examination of ultra-thin sections of dilute suspensions of biologic material is difficult, time consuming and unrewarding. Generally, a cell mass from larger cultures compacted in some way or tissue pieces are handled successfully. We have developed a rapid method for embedding and sectioning more dilute particulate materials, such as membrane components harvested from sucrose gradients or dilute cultures.Bacterial cells, Bacillus cereus and Bacillus subtilis were cultured using a standard growth medium (Tryptone glucose yeast extract broth, 100 ml) for 24 hours at ambient temperature. In this experiment the culture was divided in half at the conclusion of incubation. One half of the mixture (50 ml) was centrifuged to produce a pellet which was immediately frozen in liquid nitrogen. The other half was diluted and utilized during the second part of this experiment.

Isolation and characterization of membranes from a hydrocarbon-oxidizing Acinetobacter sp.

Membranes were isolated and purified from nutrient broth-yeast extract- and hexadecane-grown cells of Acinetobacter sp. strain HO1-N. Two membrane fractions were isolated from nutrient broth-yeast extract-grown cells, the cytoplasmic membrane and the outer membrane. In addition to these two membrane fractions, a unique membrane fraction was isolated from hexadecane-grown cells (band 1) and characterized as a lipid-rich, low-density membrane containing high concentrations of hexadecane. The outer membrane preparations of Acinetobacter, obtained from nutrient broth-yeast extract- and hexadecane-grown cells, exhibited a low ratio of lipid phosphorus to protein and contained phospholipase activity and 2-keto-3-deoxyoctulosonic acid. Phosphatidic acid cytidyltransferase, adenosine triphosphatase, and reduced nicotinamide adenine dinucleotide oxidase were recovered almost exclusively in the cytoplasmic membrane fractions. The cytoplasmic membrane fractions contained 20 to 25 polypeptide species on sodium dodecyl sulfate-polyacrylamide gels, and the outer membrane fractions contained 15 to 20 polypeptide species. A major polypeptide species with an apparent molecular weight of approximately 42,000 to 44,000 was found for all outer membrane fractions. The buoyant densities of the cytoplasmic membrane fractions and the outer membrane fractions were closely similar, necessitating their separation by differential centrifugation. Band 1 of hexadecane-grown cells had a ratio of lipid phosphorus to protein that was almost twice that of cytoplasmic membrane and a correspondingly low buoyant density (1.086 g/cm3). Enzyme activities associated with band 1 were identical to those associated with the cytoplasmic membrane. The electrophoretic banding pattern of band 1 was essentially identical to the banding pattern of the cytoplasmic membrane. The phospholipid and neutral lipid compositions of the isolated membrane fractions were determined as qualitatively similar, with significant quantitative differences. The ultrastructure characteristics of the respective membrane fractions were examined by the negative-stain technique.

Effect of substrate on metabolite production of Alternaria alternata

Alternariol and alternariol monomethyl ether are commonly associated with weathered grain sorghum. Production of these metabolites and altenuene by isolates of Alternaria alternata was evaluated on various sterile grain substrates. At 35% moisture content and 25 C, metabolite yields were highest on rice, intermediate on sorghums, and lowest on wheat and yellow corn. Fourteen-to 21-day cultures on milled rice were best in terms of ease of metabolite recovery, even though yields were higher on 28-day cultures of rough and brown rice. Metabolite production was reduced when rice was supplemented with yeast extract or yeast extract plus Czapek-Dox broth.

A Strain of Pseudomonas aeruginosa Resistant to a Quaternary Ammonium Compound

Cells of Pseudomonas aeruginosa able to grow in Tryptone glucose yeast extract (TGY) broth containing 750 μg/ml of quaternary ammonium compound (alkyldimethylethylbenzyl ammonium chloride-(QAC) were studied in comparison to sensitive cells unable to grow in such medium to identify factors important in determining the precise degree of resistance. The germicidal activity of QAC solutions against both sensitive and resistant cells in TGY broth was shown to be greatly affected by the concentration of Tryptone and yeast extract, but not by the amount of sugar. The pH of the broth also influenced germicidal activity; both strains were more susceptible under slightly acid conditions. A comparison of the pH range of growth of sensitive and resistant cells demonstrated the abiility of the former to grow in TGY broth at pH 4.5 while the latter could not achieve growth at pH 5.0. A study of the effects of 50 ppm QAC, buffered to various pH values, indicated that susceptibility of resistant cells was nearly the same as that of sensitive cells below pH 3.0, at pH 6.0, and above pH 9.0. The greatest difference between the two cell types occurred from pH 3.5 to pH 4.5 with a second peak of resistance being observed from pH 7.0 to pH 8.5.

Influences of Temperature on Growth of Streptococcus cremoris and Streptococcus lactis

Optimal growth temperatures in yeast extract broth for nine strains of Streptococcus cremoris, selected as representative of 45 strains, varied from 28 through 31C and averaged 29.3C. Maximal growth temperatures were 34.4 through 38.9C in broth and in litmus milk, with an average of 36.1C. The one strain of Streptococcus lactis had an optimal growth temperature of 31.5C and a maximum of 42C. Maximal growth temperatures for the nine strains of S. cremoris were 3.9 to 10.4C above the optimal temperatures, and strains with high optimal temperatures did not have necessarily correspondingly high minimal or maximal temperatures. Only three strains with low maximal growth temperatures apparently were damaged by treatment for 90min at 38.9C.

A Strain of Pseudomonas aeruginosa Resistant to a Quaternary Ammonium Compound

Pseudomonas aeruginosa cells were selected for their ability to grow in the presence of 770 ppm n-alkyl (50% C12, 30% C14, 17% C16, 3% C18) dimethyl dichlorobenzyl ammonium chloride (QAC). These cells retained resistance to the germicide throughout tri-weekly transfers for 7 months in tryptone glucose yeast extract broth containing no QAC. Comparisons of resistant and sensitive cells were made in an attempt to define the mechanism of resistance and to provide some information as to the mode of action of QAC. Broth cultures of the resistant strain displayed a distinct fruity odor. Gas chromatographic analysis showed that the QAC-resistant cells, unlike the sensitive, produced large quantities of ethyl acetate and ethyl valerate. Two bands of esterase activity were demonstrated in sensitive cell extracts by gel electrophoresis, while only one band was detected in the resistant cell extracts when alpha napthyl acetate was used as the substrate. Biochemical tests disclosed numerous differences between the two cell types, many of which appeared to be interrelated. The most significant differences were losses in the ability of resistant cells to synthesize extracellular lipase and protease enzymes. Many other biochemical tests on resistant cells were negative or became positive only after prolonged incubation. Resistant cells were also more resistant to osmotic disruption. Permeability studies indicated a reduced rate of glucose uptake by resistant cells. Furthermore, growth curve studies indicated a slower rate of growth by resistant cells and a 15-min longer generation time.

Effects of temperature and nutrients on proteinase production by Pseudomonas fluorescens and Ps. aeruginosa in broth and milk.

Temperature and the composition of the medium influenced the production of proteinase by Pseudomonas fluorescens and Pseudomonas aeruginosa isolated from raw milk. Many isolates of Ps. fluorescens digested litmus milk at 10° but not at 5° or 2°. With Ps. fluorescens proteinase production per unit of growth in a Peptone–Yeast Extract broth declined progressively as the incubation temperature was reduced from 20° to 5°. At 30° there was heavy growth in the same medium but only slight proteinase production whereas enzyme production by Ps. aeruginosa was maximal at this temperature. Proteinase production by both species in semi‐defined media was essentially a function of the organic nitrogen content of the medium; there was evidence of catabolite repression by glucose and, to a lesser extent, lactate. In milks seeded with these pseudomonads, the extent of proteolysis was either increased markedly or slightly decreased when glucose was included.