Biomedical subjects

I R Hamilton

Publications and source records attributed to I R Hamilton.

At least 37 records · Page 2Linked to original sources

PubMed1994

Biochemical change exhibited by oral streptococci resulting from laboratory subculturing.

The intent of this study was to assess the effects of continued laboratory subculturing on selected biochemical properties of oral streptococci freshly isolated from dental plaque. Six fresh isolates (3 Streptococcus mutans and 3 non-mutans) and 2 laboratory strains were subcultured daily for a total of 225 transfers, and cells were harvested every 75 transfers from duplicate batch cultures grown with glucose at a constant pH. Eleven biochemical properties were assayed with cells, membranes and cell-free extracts and the results subjected to statistical analysis for differences between the duplicate cultures and the various subcultures. In addition, the activity of 19 hydrolytic enzymes was assayed with the semiquantitative apiZYM system (Analytab products). The activity of zero-time samples varied by as much as 241-fold for a single property with particularly low activity for EIIglc of the phosphoenolpyruvate phosphotransferase system and cell-associated extracellular polysaccharide synthesis. The 3 S. mutans fresh isolates had higher activity in 8 of the 11 assays compared with the 3 non-mutans strains, with extracellular polysaccharide synthesis the most significant trait. Statistical analysis of the 2816 assays of the 11 traits for the 8 test strains at the 4 selected time intervals revealed considerable change in the activity of the test parameters. The most notable changes in the S. mutans strains over the 225 subcultures were significant increases in glycolytic activity and decreases in hydrophobicity and extracellular polysaccharide synthesis activity. Of the measured properties, lactate dehydrogenase and cell-associated extracellular polysaccharide synthesis activity were the most stable and H+/ATPase activity was the most variable.(ABSTRACT TRUNCATED AT 250 WORDS)

Adaptation, Biological↗

PubMed1994

Sequence and expression of the genes for HPr (ptsH) and enzyme I (ptsI) of the phosphoenolpyruvate-dependent phosphotransferase transport system from Streptococcus mutans.

We report the sequencing of a 2,242-bp region of the Streptococcus mutants NG5 genome containing the genes for ptsH and ptsI, which encode HPr and enzyme I (EI), respectively, of the phosphoenolpyruvate-dependent phosphotransferase transport system. The sequence was obtained from two cloned overlapping genomic fragments; one expresses HPr and a truncated EI, while the other expresses a full-length EI in Escherichia coli, as determined by Western immunoblotting. The ptsI gene appeared to be expressed from a region located in the ptsH gene. The S. mutans NG5 pts operon does not appear to be linked to other phosphotransferase transport system proteins as has been found in other bacteria. A positive fermentation pattern on MacConkey-glucose plates by an E. coli ptsI mutant harboring the S. mutans NG5 ptsI gene on a plasmid indicated that the S. mutans NG5 EI can complement a defect in the E. coli gene. This was confirmed by protein phosphorylation experiments with 32P-labeled phosphoenolpyruvate indicating phosphotransfer from the S. mutans NG5 EI to the E. coli HPr. Two forms of the cloned EI, both truncated to varying degrees in the C-terminal region, were inefficiently phosphorylated and unable to complement fully the ptsI defect in the E. coli mutant. The deduced amino acid sequence of HPr shows a high degree of homology, particularly around the active site, to the same protein from other gram-positive bacteria, notably, S. salivarius, and to a lesser extent with those of gram-negative bacteria. The deduced amino acid sequence of S. mutans NG5 EI also shares several regions of homology with other sequenced EIs, notably, with the region around the active site, a region that contains the only conserved cystidyl residue among the various proteins and which may be involved in substrate binding.

Amino Acid Sequence↗

PubMed1991

Adaptation by Streptococcus mutans to acid tolerance.

Our previous continuous culture studies with strains of Streptococcus mutans have indicated that the organism has the capacity of adapt to growth in acidic environments. This study was undertaken to examine this question in more detail. S. mutans Ingbritt and the phosphotransferase system (PTS)-defective mutant, S. mutans DR0001/6, were grown in continuous culture at pH 7.5 and 5.5 or 5.1, and the pH optimum for glucose uptake and glycolysis and the capacity of the cells to generate pH gradients were determined over the pH range 4.5 to 8.0 with steady state, washed cells. In addition, the proton permeability of the cells was measured over the pH range by an acid pulse technique. The results indicate that the pH optimum for glucose uptake by S. mutans Ingbritt grown at pH 7.5 was 7.5 and this optimum shifted to 7.0 and 6.0 for cells grown at pH 5.5 and 5.1, whereas with the S. mutans DR0001/6, the optimum shifted from 7.5 to 7.0 for the pH 5.5 cells. A similar shift in the pH optimum for glycolysis was observed for the 2 strains, and this was particularly pronounced for cells incubated with glucose in the presence of gramicidin to dissipate proton gradients. The capacity of the cells to generate pH gradients was related to their metabolic activity, and although larger gradients were not formed by the pH 5.5 cells, these cells were nevertheless capable of maintaining gradients at a lower pH; S. mutans DR0001/6 generated 2-fold larger pH gradients at pH 5.5 than S. mutans Ingbritt.(ABSTRACT TRUNCATED AT 250 WORDS)

Adaptation, Physiological↗

PubMed1991

Sorbitol inhibition of glucose metabolism by Streptococcus sanguis 160.

Clinical studies in Sweden have shown that the proportion of sorbitol-utilizing strains of Streptococcus sanguis increases in dental plaque from individuals using sorbitol-containing products for prolonged periods. We have undertaken to study the metabolism of glucose and sorbitol by S. sanguis 160, isolated from a subject consuming sorbitol-containing chewing-gum 4 times a day for 4 years. Growth on glucose was inhibited by the presence of sorbitol in the growth medium and sorbitol was utilized in the presence of glucose, albeit, at a slower rate than glucose. In addition, pulses of glucose added to cultures growing on sorbitol resulted in the expulsion of sorbitol from the cell. In order to examine further the relationship of sorbitol and glucose, uptake assays were carried out with S. sanguis 160 grown in continuous culture (pH 7.0, dilution rate = 0.1 h-1) with glucose, sorbitol or nitrogen (sorbitol excess) limitations. The uptake of [14C]-glucose by sorbitol-limited cells, but not by glucose-limited cells, was inhibited by sorbitol, as was glycolysis. Kinetic experiments with glucose-limited cells showed 2 transport systems for glucose with Ks values of 5.2 and 40 microM, and glucose phosphorylation activity by decryptified cells indicated transport by the P-enolpyruvate (PEP) phosphotransferase system (PTS) with lesser activity for an ATP-dependent transport process. Transition from glucose-limited growth to sorbitol-limited growth revealed repression of total [14C]-glucose uptake by intact cells and activity for Enzyme II for glucose (Ellglc) of the PTS measured in membrane preparations in the presence of an excess of the soluble PTS proteins in crude cell-free supernatant fractions.(ABSTRACT TRUNCATED AT 250 WORDS)

Dental Plaque↗

PubMed1991

Sorbitol transport by Streptococcus sanguis 160.

Sorbitol metabolism was examined with a sorbitol-fermenting strain (160) of Streptococcus sanguis isolated from the dental plaque of a subject using sorbitol-containing chewing-gum for 4 years. S. sanguis 160 was grown in continuous culture (pH, 7.0; dilution rate, 0.1 h-1) with glucose, sorbitol and nitrogen (sorbitol-excess) limitations. Cells grown with a glucose limitation exhibited low, but detectable, uptake of [14C]-sorbitol and transition to medium limiting in sorbitol resulted in a 5-fold increase in sorbitol uptake. Kinetic data revealed that both glucose and sorbitol-limited cells possessed 2 transport systems for sorbitol (Ks = 3.3-6.7 and 36-64 microM), but continued growth of the organism on limiting sorbitol resulted in the loss of the high-affinity system. Decryptified, sorbitol-limited cells phosphorylated sorbitol in the presence of phosphoenolpyruvate (PEP), but not with ATP, indicating sorbitol transport solely via the PEP phosphotransferase (PTS) system. PEP-dependent activity in glucose-limited and sorbitol-excess cells was 6- and 4-fold lower than that of the sorbitol-limited cells. Uptake of [14C]-sorbitol and activity for Ell for sorbitol [Ellsor] of the PTS in cells in transition from a glucose to sorbitol limitation confirmed the induction of the sorbitol-PTS and the repression of the glucose-PTS in the presence of sorbitol. Cells grown with an excess of sorbitol exhibited very low Ellsor activity. A crossover experiment with membranes and soluble fractions from glucose-, sorbitol- and nitrogen-limited cells of S. sanguis 160 demonstrated the induction of a soluble PTS component in sorbitol-limited cells essential for sorbitol transport via the PTS.(ABSTRACT TRUNCATED AT 250 WORDS)

Dental Plaque↗

PubMed1991

Effect of growth rate and pH on intracellular levels and activities of the components of the phosphoenolpyruvate: sugar phosphotransferase system in Streptococcus mutans Ingbritt.

The growth of Streptococcus mutants Ingbritt in continuous culture at low pH or high growth rates repressed the biosynthesis of the components of the phosphoenolpyruvate:sugar phosphotransferase system (PTS). The cellular concentrations of the soluble components HPr, enzyme I (EI), and EIII for mannose (IIIman) and EII activity for glucose, mannose, 2-deoxyglucose (2DG), and fructose were determined in membrane preparations from cells grown at pHs from 8.0 to 5.0 and at dilution (D) or growth rates from 0.1 to 1.0 h-1. The cellular levels of HPr and EI varied less than threefold under all of the growth conditions tested. On the other hand, EII activity in membranes from cells grown at D = 0.1 h-1 was repressed by growth at pHs below 8.0, with cells grown at pH 5.0 completely devoid of EII activity. In addition, cells grown at D = 0.5 and 1.0 h-1 exhibited little PTS activity for glucose, mannose, and 2DG and twofold-lower activity for fructose. These activities were stimulated by the addition of a membrane-free cytoplasmic fraction, and this activating activity was shown to be due to the presence of IIIman. Estimation of the cellular content of IIIman indicated that the synthesis of this factor was repressed by growth above and below pH 7.0 and was particularly sensitive to growth at high rates. These results indicate that with S. mutans Ingbritt, both pH and growth rate regulate the genes for the synthesis of EIIs involved in the phosphorylation of glucose, mannose, 2DG, and fructose and the gene for the formation of IIIman.

Bacterial Proteins↗

PubMed1991

The stability of outer-membrane protein and antigen profiles of a strain of Bacteroides intermedius grown in continuous culture at different pH and growth rates.

The stability of the outer-membrane proteins and antigens of a strain of Bacteroides intermedius (VPI 8944 group genotype II) grown in continuous culture at varying pH and growth rates (D = 0.025-0.2 h-1, pH 6.0-7.3) has been measured. The membranes showed nine major proteins (greater than 67-19.55 kilodaltons) and six major antigens (65-28 kilodaltons). Membrane proteins and antigens were stable under the conditions tested; the major proteins were detected in all membranes, and the antigen profiles tested with different antisera showed maximum similarities of 82-95%. Differences did occur in the amounts of membrane proteins synthesized; cells at high growth rates and those growing on the surfaces in the chemostat showed increased amounts of two proteins (40 and 32 kilodaltons) and possibly novel proteins of 24 and 25 kilodaltons. In addition, these membranes reflected increased synthesis or a change to increased reactivity of antigens between 20.5 and 24 kilodaltons. The results indicate stability of the expression of outer-membrane proteins and antigens in environments of differing pH and under different growth rates. However, the amount of these molecules synthesized can vary, and increases in certain proteins and antigens occur as the growth rate increases and the organisms grow on surfaces.

Antigenic Variation↗

PubMed1990

Maintenance of proton motive force by Streptococcus mutans and Streptococcus sobrinus during growth in continuous culture.

The components of the transmembrane electrochemical proton gradient, or proton motive force (PMF, delta p), were determined in cells of Streptococcus mutans Ingbritt and Streptococcus sobrinus ATCC 27352 growing in continuous culture under conditions of changing glucose concentration, growth rate and growth pH. The pH gradient (delta pH) and membrane electrical potential (delta psi) were assayed with the weak acid, salicyclic acid, and the lipophilic cation, methyltriphenylphosphonium iodide, respectively. S. mutans Ingbritt growing in continuous culture (pH 7.0, dilution rate (D) = 0.1 h-1) at 8 glucose concentrations ranging from 2.8 to 288 mM maintained a relatively constant delta p of 58.3 mV (SD +/- 5.8) in spite of a transition from glucose to nitrogen-limited growth and significant changes in cell physiology. Changes included a decreasing yield constant, increasing glucose uptake rates in the chemostat, repression of Ellglc of the PEP phosphotransferase sugar transport system and decreasing glycolytic capacity of the cells as the medium glucose concentration increased. Changes in the dilution or growth rate of S. mutans Ingbritt from 0.1 to 1.0 h-1 and S. sobrinus from 0.1 to 0.8 h-1, when growing at pH 7.0 with limited glucose and lactose, respectively, resulted in significantly lower delta p values due to the dissipation of the delta psi. When the cells of S. mutans Ingbritt were grown with excess glucose (nitrogen limitation), lower delta p values were observed at pH 5.5, but not at pH 7.0.(ABSTRACT TRUNCATED AT 250 WORDS)

Biological Transport, Active↗

PubMed1990

Biochemical effects of fluoride on oral bacteria.

Fluoride inhibition of carbohydrate metabolism by the acidogenic plaque microflora is well-established, although it has not always been appreciated that oral bacteria vary considerably in their susceptibility to fluoride. Early studies demonstrated that the F-induced reduction in acid production was due, in part, to the inhibition of the glycolytic enzyme, enolase, which converts 2-P-glycerate to P-enolpyruvate. The decreased output of PEP in the presence of F, in turn, results in the inhibition of sugar transport via the PEP phosphotransferase system (PTS). Bacterial accumulation of fluoride involves the transport of HF, a process requiring a transmembrane pH difference or pH gradient, which is generated only by metabolically active cells. The uptake of HF into the more alkaline cytoplasm results in the dissociation of HF to H+ and F- and, if allowed to continue, the accumulation of protons acidifies the cytoplasm, causing a reduction in both the proton gradient and enzyme activity. Current information indicates that in addition to enolase, F- also inhibits the membrane-bound, proton-pumping H+/ATPase, which is involved in the generation of proton gradients through the efflux of protons from the cell at the expense of ATP. Thus, fluoride has the dual action of dissipating proton gradients and preventing their generation through its action on H+/ATPase. The collapse of transmembrane proton gradient, in turn, reduces the ability of cells to transport solutes via mechanisms involving proton motive force. In spite of these known effects on the bacterial cell, there is no general agreement that the anti-microbial effects of F contribute to the anti-caries effect of fluoride.(ABSTRACT TRUNCATED AT 250 WORDS)

Bacteria↗

PubMed1989

Competition between Streptococcus mutans and Lactobacillus casei in mixed continuous culture.

In natural oral ecosystems, succession of Streptococcus mutans and Lactobacillus species occurs in bacterial communities which can be associated with the development of carious lesions. The objective of this study was to explore the interaction between 2 strains of S. mutans (SF2452) and Lactobacillus casei (RB1014) when grown together in continuous culture under conditions of varying environmental pH and in the presence of fluoride. When the strains, isolated from approximal carious lesions, were grown initially at a dilution rate of 0.1 h-1 under a glucose limitation at pH 7.0, a stable mixed community developed (Days 5-10) with S. mutans as the dominant population. When the pH control was removed (Day 10), the binary culture lowered the pH to 5.1 causing a small reduction in the numbers of S. mutans and allowing L. casei to predominate. A more dramatic effect was seen when the environmental pH was rapidly lowered to 4.8 by the addition of acid. In this case, the S. mutans population was significantly reduced and the L. casei population rose to the level previously held by S. mutans. Re-adjustment of the pH to 7.0 resulted in a re-establishment of the population ratios prior to the addition of acid. The possibility that S. mutans was capable of aciduric adaptation during a biologically-generated pH reduction was examined by mixing cultures of both organisms after each had been grown to steady state at pH 5.5 in separate chemostats.

Fluorides↗

PubMed1989

Growth and metabolic properties of Bacteroides intermedius in anaerobic continuous culture.

Two strains of Bacteroides intermedius BH20/30 and BH18/23, have been grown in anaerobic continuous culture under various conditions for periods up to 54 days. Strain BH20/30 grew over a relatively wide pH range from 5-8 with a maximum at pH 7.0 at a dilution rate (D) of 0.1 h-1 with a glucose limitation, while strain BH18/23 had an optimum between 5.8 and 7.3 and would not grow above and below this range. The maximum growth rate (mu max) for the latter strain was shown to be 0.23 h-1, or a doubling time of 3.0 at the upper limit of pH 7.3. The yield values (Y glucose) for strain BH18/23 reached 187-177 g cells (dry weight) per mole of glucose in the optimum pH range (6.0-7.0) and amino acid analysis of the spent medium indicated that these high values were the results of the combined use of glucose and amino acids; the cultures also exhibited proteolytic activity. The major acid end-products in the same pH range were formate and succinate with lesser concentrations of acetate, isovalerate and fumarate; small amounts of lactate appeared as the cells were stressed at pH values above 7.5 when the culture was 'washing out' of the chemostat. Glucose metabolism appeared to function through the glycolytic pathway in B. intermedius BH18/23 since the glycolytic inhibitors, sodium fluoride and sodium iodoacetate, completely inhibited glucose utilization as did the proton ionophore, gramicidin, and the ATPase inhibitor, N,N1-dicyclohexylcarbodiimide (DCCD). Inhibition by these latter compounds indicated that the saccharolytic Bacteroides utilize proton gradients generated by proton-extruding ATPase (H+/ATPase) to conserve energy.(ABSTRACT TRUNCATED AT 250 WORDS)

Bacteroides↗

PubMed1989

Concentration-dependent repression of the soluble and membrane components of the Streptococcus mutans phosphoenolpyruvate: sugar phosphotransferase system by glucose.

Growth of Streptococcus mutans Ingbritt in continuous culture (pH 7.0, dilution rate of 0.1 h-1) at medium glucose concentrations above 2.6 mM resulted in repression of the sugar-specific membrane components, enzyme IIGlc (EIIGlc) and EIIMan, of the phosphoenolpyruvate:sugar phosphotransferase system (PTS). In one experiment, significant repression (27-fold) was observed with 73 mM glucose when the glycolytic capacity of the cells was reduced by only 2-fold and when the culture was still glucose limited. In a more comprehensive experiment in which cells were grown in continuous culture at eight glucose concentrations from 2.6 to 304 mM, in addition to repression of specific EII activities for glucose, mannose, 2-deoxyglucose, and fructose, synthesis of the general protein, EI, was repressed at all glucose levels above 2.6 mM to a maximum of 4-fold at 304 mM glucose when the culture was growing with excess glucose (i.e., nitrogen limited). The other PTS general protein, HPr, was less sensitive to the exogenous glucose level but was nevertheless repressed fourfold under glucose-excess conditions. The Km for glucose for EIIGlc increased from 0.22 mM during growth at 3.6 mM glucose (glucose limited) to 0.48 mM at 271 mM glucose (glucose excess). The shift from heterofermentation to homofermentation during growth with increasing glucose levels suggests the involvement of glycolytic intermediates, ATP, or another high-energy phosphate metabolite in regulation of the synthesis of the PTS components in S. mutans.

Bacterial Proteins↗

PubMed1987

Effect of growth conditions on levels of components of the phosphoenolpyruvate:sugar phosphotransferase system in Streptococcus mutans and Streptococcus sobrinus grown in continuous culture.

The membrane-bound, sugar-specific enzyme II (EII) component of the phosphoenolpyruvate:sugar phosphotransferase system (PTS) in Streptococcus mutans Ingbritt is repressed by growth on glucose under various conditions in continuous culture. Compared with optimal PTS conditions (i.e., glucose limitation, dilution rate [D] of 0.1 h-1, and pH 7.0), EII activity for glucose (EIIGlc) and mannose (EIIMan) in cells grown at a D of 0.4 h-1 and pH 5.5 with the same glucose concentration was reduced 24- to 27-fold. EII activity with methyl alpha-glucoside and 2-deoxyglucose was reduced 6- and 26-fold, respectively. Growth with excess glucose (i.e., nitrogen limitation) resulted in 26- to 88-fold repression of EII activity with these substrates. The above conditions of low pH, high dilution rate, and excess glucose also repressed EII activity for fructose (EIIFru), but to a lesser extent (two- to fivefold). Conversely, growth of S. mutans DR0001 at a D of 0.2 h-1 and pH 5.5 resulted in increased EIIGlc and EIIMan activity. Unlike the EII component, the HPr concentration in S. mutans Ingbritt varied only twofold (5.5 to 11.4 nmol/mg of protein) despite growth at pH 5.5 with limiting and excess glucose. The HPr concentrations in S. mutans DR0001 and the glucose-PTS-defective mutant DR0001/6 were similar. In a companion study, the soluble components of the PTS (i.e., HPr, EI, and EIIILac) in Streptococcus sobrinus grown on limiting lactose in a chemostat were not influenced significantly by growth at various pHs (7.0 and 5.0) and growth rates (D of 0.1, 0.54, and 0.8 h-1). However, growth on lactose resulted in repression of both EIIGlc and EIIFru, confirming earlier results with batch-grown cells. Thus, the glucose-PTS in some strains of S. mutans is regulated at the level of EII synthesis by certain environmental conditions.

Glucose↗

PubMed1987

Environmental pH as a factor in the competition between strains of the oral streptococci Streptococcus mutans, S. sanguis, and "S. mitior" growing in continuous culture.

Strains of Streptococcus mutans (biotype 1), Streptococcus sanguis, and Streptococcus mitior have been grown in mixed continuous culture in a semidefined medium under glucose limitation at a growth rate of D = 0.1 h-1. The effect of varying the environmental pH on the proportions of the different populations within the community has been determined. Initially the populations were allowed to reach steady state at pH 7.0 when S. sanguis was dominant with S. mutans and "S. mitior" maintaining similar populations. The medium pH was then lowered in steps of 0.5 pH units from pH 7.0 to 4.5, and the community was grown at each step for at least 15 generations. Viable counts of each species were made at 24-h intervals. The population ratios established at pH 7.0 remained relatively stable when the environmental pH was set at pH 6.5. However, after the medium pH was lowered to 6.0 (days 18-27), the population of S. mutans began to increase and the S. mitior population began to decline. A further change was seen at pH 5.5 (days 27-34) when S. mutans became dominant, S. sanguis declined, and S. mitior was not detectable. At pH 4.5, both S. mutans and S. sanguis were reduced in numbers, but survived until the experimental run was terminated (44 days). Samples of culture fluid were taken throughout the experiment and analyzed for the presence of the acid products of glucose metabolism. The amounts of lactic acid produced by the community increased as the environmental pH was lowered.(ABSTRACT TRUNCATED AT 250 WORDS)

Culture Media↗

PubMed1985

Influence of pH and fluoride on properties of an oral strain of Lactobacillus casei grown in continuous culture.

A freshly isolated oral strain, Lactobacillus casei RB1014, was grown in continuous culture to compare the effects of pH and fluoride on growth and metabolism. The cells were grown at pH 7.0 to 3.2 in the absence of fluoride and from pH 7.0 to 5.4 with 20 mM NaF. Cell numbers varied from 3 X 10(9) to 30 X 10(9)/ml on blood agar during alterations in the growth pH from 7.0 to 4.27. Only when the culture was stressed by lowering the pH to 3.2 were cell numbers drastically reduced. Cells growing at pH 7.0 without fluoride were unable to grow when plated on fluoride agar (10.5 mM) at pH 5.5; however, when the growth pH was allowed to decrease to 4.94, cells grew on the fluoride plates in numbers equal to those growing on blood agar. This fluoride tolerance trait appeared rapidly once pH control was removed and was lost when the culture was returned to pH 7.0. The addition of 20 mM NaF to the culture medium did not adversely affect growth, provided that the pH was maintained at 6.0 or above; cells tolerant to 10.5 and 16 mM NaF appeared on pH 5.5 plates during this phase. In cells removed from the chemostat throughout the experiment and incubated at the pH of growth in a pH stat, glycolytic activity was optimum at pH 5.5 in the absence of NaF. Fluoride stimulated glycolytic activity by cells incubated at pH 7.0 and by cells growing with 20 mM NaF, provided that the pH of growth remained at or above 6.0. A more detailed examination of the adaptation to fluoride tolerance during shifts to acidic pH values revealed that cells capable of growth on acidic fluoride agar plates appeared within 2 h of the start of the fall in pH of the chemostat culture. Estimation of the intracellular pH during the period of the initial pH fall revealed that the intracellular pH was identical to the extracellular pH (i.e., no pH gradient [delta pH]), indicating that fluoride would not be transported into the cells to inhibit metabolism. However, once the pH of the medium was stabilized, delta pHs were generated, with the delta pH increasing as the pH declined. The inhibition of glycolysis by fluoride increased in proportion to the delta pH. Cells grown at pH 5.5 generated larger delta pHs than did cells grown at pH 7.0, although the values were normally small (approximately 0.9 U). The data suggest that the inherent fluoride tolerance of L. casei RB1014 was associated with relatively small delta pHs.

Fluorides↗

PubMed1985

Description of an unusual gram-negative anaerobic rod isolated from periodontal pockets.

A Gram-negative rod which grew with an unusual colonial "water-drop" form was isolated from periodontal pocket samples from 12 patients. Six strains were characterized by biochemical tests, cell wall analyses, malate dehydrogenase mobilities, protein profiles, and serology. By these criteria, the organisms formed a group of similar strains which were anaerobic, nonmotile, nonsporing, Gram-negative rods resembling Bacteroides. Comparison of the isolates to American Type Culture Collection strains of Bacteroides showed that they represented a closely related group, distinct from the described species of oral Bacteroides. Initial results on the DNA of the isolates suggested a base ratio of 54-57% G + C. Despite the DNA G + C base ratios currently accepted for the Bacteroides (28-61 mol% G + C), many species fall into a narrower range of 40-52 mol% G + C. This range would exclude the organisms described here and suggests that placing them into the genus Bacteroides may be inappropriate.

Bacteroides↗

PubMed1985

Effect of NaF and pH on the growth and glycolytic rate of recently isolated strains of oral Lactobacillus species.

Twenty-eight fresh and seven type strains of Lactobacillus species were tested for their ability to grow and metabolize glucose in the presence of fluoride. While there was variation between the test organisms, freshly isolated strains were generally more fluoride-tolerant than were the type strains, and a group of seven strains would grow well at pH 4.5 with 5.3 mm NaF. Glucose metabolism by representative strains of this latter group required average fluoride concentrations of 204, 64, and 9.3 mM to inhibit glycolysis completely at pH 7.0, 6.0, and 5.0, respectively. These values are ten times higher than those required to inhibit glucose metabolism by Streptococcus mutans.

Acids↗

PubMed1984

Comparison of polyvinyl chloride membrane electrodes sensitive to alkylphosphonium ions for the determination of the electrical difference (delta psi) of Streptococcus mutans and Lactobacillus casei.

Polyvinyl chloride membrane electrodes sensitive to tetraphenyl phosphonium (TPP+), butyltriphenyl phosphonium ( bTPP +), and methyltriphenyl phosphonium ( mTPP +) ions have been compared for the determination of the electrical potential difference (delta psi) of the oral bacteria, Streptococcus mutans DR0001 /6 and Lactobacillus casei RB1014 . All three types of electrode proved suitable for determining delta psi, although the TPP+-sensitive electrode was particularly susceptible to interference by protonmotive force (delta p) dissipators known to inhibit sugar uptake by the bacteria. The mTPP +-sensitive electrode was the least affected. Similarly, both strains had a high nonspecific binding capacity for TPP+ and bTPP + ions, and this increased for all three ions when the bacteria were heated to 80 degrees C for 1 h to abolish glucose uptake and metabolism. This heat-treatment procedure is therefore not a suitable control for determination of nonspecific binding to cells. However, 1% (v/v) toluene, 20 microM gramicidin, or 10 microM valinomycin effectively depolarized the bacteria without interfering with nonspecific binding. The ionophores were therefore used subsequently for the determination of nonspecific binding of the lipid-soluble cations. The mTPP + ion and corresponding electrode proved the most effective system, and delta psi values of -89 and -107 mV were obtained for S. mutans and L. casei, respectively, harvested from glucose-limited continuous cultures and incubated in 100 mM Hepes-KOH buffer (pH 7.0), containing 1 mM dithiothreitol and 10 mM glucose. Although the delta psi of S. mutans decreased significantly in the presence of Mes-KOH and potassium phosphate buffers at pH 7.0, it increased to -119 mV in Tris-HCl buffer (pH 7.0).(ABSTRACT TRUNCATED AT 250 WORDS)

Biological Transport, Active↗