Simple filter paper procedure for estimation of glucose uptake via group translocation by whole-cell suspensions of bacteria

Germaine, G R; Tellefson, L M

doi:10.1128/aem.41.3.837-839.1981

Cited by 9 publications

(12 citation statements)

References 9 publications

(6 reference statements)

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“…Estimation of glucose uptake was performed on whole cells suspended in 50 mM potassium phosphate buffer (unless noted otherwise), pH 7.0. Estimation of glucose phosphorylation by toluene-acetone-treated S. mutans has also been described (9). [U-'4C]glucose (1.5 mCi/mmol) was used at a final concentration of 200 ytM.…”

Section: Methodsmentioning

confidence: 99%

Effect of human saliva on the fluoride sensitivity of glucose uptake by Streptococcus mutans

Germaine¹,

Tellefson²

1981

Infect Immun

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The fluoride (F) sensitivity of glucose uptake by whole cell suspensions of Streptococcus mutans in the presence and absence of human whole salivary supernatant was studied. It was observed that dithiothreitol (DTT) and other thiols markedly reduced the F sensitivity of cells when saliva (50%, vol/vol) was present during glucose uptake. In the absence of saliva, cells were sensitive to 2 to 2.5 mM F regardless of the presence of thiols. Supplementation of cells in phosphate or tris(hydroxymethyl)aminomethane-hydrochloride buffers with physiological concentrations of calcium or phosphate had no effect on the F sensitivity of the organism. Experiments with permeabilized cells suggested that thiols themselves had no direct effect on the F sensitivity of enolase (a principal F target). Cells pretreated with DTT subsequently exhibited decreased F sensitivity when examined in the presence of saliva but not in the absence of saliva. Cells pretreated with whole salivary supernatant were found to be subsequently less sensitive to F in the absence of saliva during glucose uptake. Furthermore, in cases where cells were pretreated with saliva, subsequent additions of DTT were unnecessary to obtain maximal reduction in the F sensitivity of glucose uptake. It was concluded that the saliva-dependent reduction in F sensitivity of glucose uptake was not due to sequesteration of available F by salivary constituents. The data suggest that a salivary component(s) interacts directly with the microorganism in some manner which results in reduced F sensitivity of the process under study. Possible mechanisms of saliva action are discussed. 'Dialyzed for 18 h at 4°C against 200 volumes of distilled water. INFECT. IMMUN.

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Section: Methodsmentioning

confidence: 99%

Effect of human saliva on the fluoride sensitivity of glucose uptake by Streptococcus mutans

Germaine¹,

Tellefson²

1981

Infect Immun

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“…In all cases reported here, salivary supernatant was used at the desired final concentrations in 50 mM potassium phosphate buffer (pH 7.0) (PB) or in PB supplemented with 1 mM MgCl2 (PBMg). [U-14C]glucose uptake was estimated by a filter paper method (4,5). Glucose was present in the uptake mixtures at 200 ,uM (1.5 mCi/mmol).…”

Section: Methodsmentioning

confidence: 99%

Glucose Uptake by Streptococcus mutans, Streptococcus mitis , and Actinomyces viscosus in the Presence of Human Saliva

Greg

Tellefson

1982

Infect Immun

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Glucose uptake was examined by using whole-cell suspensions of Streptococcus mutans (strains BHT, Ingbritt, and GS-5), Streptococcus mitis (strains 9811 and 72X41), and Actinomyces viscosus (strains T6 and WVU626) incubated for up to 90 min in 0 to 82% (vol/vol) human whole salivary supernatant. Glucose uptake by the S. mutans strains was completely inhibited at all saliva concentrations. Dithiothreitol (DTT), present during saliva incubation, prevented saliva inhibition. Glucose uptake was also restored when saliva-inhibited cells were subsequently exposed to DTT. The inclusion of catalase in the saliva incubation mixtures resulted in protection equal to that obtained with DTT. The S. mitis strains were also inhibited by saliva but to a far lesser extent that S. mutans. DTT and catalase also protected S. mitis from saliva inhibition. Both A. viscosus strains were completely refractory to saliva inhibition of glucose uptake. Based on (i) the sensitivity of the catalase-negative streptococci and the resistance of catalase-positive actinomyces to saliva inhibition and (ii) the equal and complete protection to saliva inhibition afforded by DTT and catalase, we conclude that the lactoperoxidase-SCN-H202 system in saliva was the only antibacterial system expressed under our experimental conditions. The relative resistance of S. mitis 9811 (compared with S. mutans BHT) to saliva inhibition was shown not to result from poor H202 production in either glucose-supplemented buffer or saliva solutions. S. mitis produced inhibitory quantities of H202 that equaled or exceeded S. mutans H202 accumulation. It is suggested that S. mitis might possess a greater ability to repair lactoperoxidase-mediated damage than does S. mutans. Every organism studied exhibited a saliva concentration-dependent, cell growth-independent stimulation of glucose uptake after 60 to 90 min of incubation. The A. viscosus and S. mitis strains showed saliva stimulation (or stabilization) of glucose uptake with unsupplemented saliva. In the case of S. mutans, saliva stimulation was only observed when DTT was present. The possible role of salivary lactoperoxidase as a modulator of the intraoral site specificities exhibited by S. mutans is discussed.

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“…was inoculated and grown in citrate enrichment medium at room temperature 24 h prior to 0n c r the experiment. The cells were centrifuged (6,000 x g for 10…”

Section: Tions S 80mentioning

confidence: 99%

“…The duration of counting ' \ for each sample was programmed such that the 2 sigma error E 60 was less than 6% for each determination. Estimation of the E 14C in the cells was performed as described by Germaine and Tellefson (6). Samples (0.5 ml) were placed on 3 MM filter Q 40 -< W * * paper pieces (3.7 by 4.5 cm; Whatman) and immediately placed in a beaker of methanol.…”

Section: _<8mentioning

confidence: 99%

Effects of Glucose Concentrations on Cadmium, Copper, Mercury, and Zinc Toxicity to a Klebsiella sp

Brynhildsen

Lundgren

Allard

et al. 1988

Appl Environ Microbiol

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The influence of glucose concentration on Cd, Cu, Hg, and Zn toxicity to a Klebsiella sp. was studied by following the degradation of 14C-labeled glucose at pH 6.0. Uptake of '4C into the cells was also determined. The carbon concentrations ranged from 0.01 to 40 mg liter-', which are equivalent to soluble C concentrations in natural environments. The toxicity of Cu, Cd, and Zn to a KlebsieUa sp. was affected considerably by the C concentration. Copper at 10-s M was toxic when the carbon concentration was 10 or 40 mg liter-', while at 0.01 to 1.0 mg liter-' no toxicity was observed. Cadmium and zinc were toxic at 10-2 M in media containing 0.01 to 1.0 mg of C liter-'. At C concentrations greater than 1.0 mg liter-', the inhibition of glucose degradation and carbon assimilation was observed at 10-3 M Cd and Zn. The toxicity of mercury seemed to be independent of the C concentration. Results of this study showed that the nutritional state of an organism may have a profound effect on its sensitivity to metals. Metals taken up by an energy-driven transport system may be less toxic under conditions of C starvation. The C concentration should be taken into account when evaluating results from toxicity studies, especially as most microorganisms in nature live under energy-limited conditions.

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Simple filter paper procedure for estimation of glucose uptake via group translocation by whole-cell suspensions of bacteria

Cited by 9 publications

References 9 publications

Effect of human saliva on the fluoride sensitivity of glucose uptake by Streptococcus mutans

Effect of human saliva on the fluoride sensitivity of glucose uptake by Streptococcus mutans

Glucose Uptake by Streptococcus mutans, Streptococcus mitis , and Actinomyces viscosus in the Presence of Human Saliva

Effects of Glucose Concentrations on Cadmium, Copper, Mercury, and Zinc Toxicity to a Klebsiella sp

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