A recycling fermentor (a chemostat with 100% biomass feedback) was used to study glucose-limited behavior of Escherichia coli B. The expectation from mass transfer analysis that growth would asymptotically approach a limit mass determined by the glucose provision rate (GPR) and the culture's maintenance requirement was not met. Instead, growth proceeded at progressively lower rates through three distinct phases. After the fermentor was seeded, but before glucose became limiting, growth followed the usual, exponential path (phase 1). About 12 h postseeding, residual glucose in the fermentor fell below 1 jig-ml`' and the growth rate (dx/dt) became constant and a linear function of GPR (phase 2). The specific growth rate, ,I, therefore fell continuously throughout the phase. Biomass yield and glucose assimilation (13%) were near the level for exponential growth, however, and independent of GPR over a broad range. At a critical specific growth rate (0.04 h-1 for this strain), phase 2 ended abruptly and phase 3 commenced. In phase 3, the growth rate was again constant, although lower than in phase 2, so that,u continued to fall, but growth rates and yields were paraboloid functions of GPR. They were never zero, however, at any positive value of GPR. By inference, the fraction of metabolic energy used for maintenance functions is constant for a given GPR, although different for phases 2 and 3, and independent of biomass. In both phases 2 and 3, orcinol, diphenylamine, and Lowry reactive materials were secreted at near-constant rates such that over 50% as much biosynthetic mass was secreted as was retained by the cells. ment of the requirements for the Ph.D.). MATERIALS AND METHODS Culture, medium, and fermentor seeding. E. coli B was maintained on Davis and Mingioli (5) minimal medium slants containing 1.25 mg of glucose. 625
SUMMARYGrowth systems appropriate for studying mass transfer in different bacterial environments are reviewed. Fed batch and recycling fermentors are suited to modelling nutrient limitation and slow growth. Use of these two growth systems reveals the existence of three growth rate regions, or domains, defined by maintenance energy demands, nucleotide regulation, metabolism, and physiological behavior. They are exemplified in Escherichia coil by domain-dependent synthesis of attachment antigens, heat-labile toxin, and inducible enzymes. Distribution of the bacterial population among cell cycle stages changes with growth rate domain because lengths of the stages differ in their dependence on growth rate. This produces subpopulafions whose ratios vary with growth rate and that are likely to differ in both molecular composition and stress resistance.
Previous studies have compared the adhesion of [3H]thymidine-labeled Streptococcus sanguis to salivacoated hydroxyapatite (SHA) and buffer-coated hydroxyapatite (HA) beads. Although the hypotonic buffer used in these assays was adjusted to simulate saliva, it does not necessarily provide the optimal parameters for the quantitative estimate of adhesion under in vitro conditions. Optimization is necessary to provide the maximum sensitivity of the assay for detecting the effects of various salivas as well as for quantitating the effect of environmental growth conditions on the adhesion of S. sanguis to SHA and HA. A major distinction between the adhesion of S. sanguis to SHA and HA was observed when the bacterial concentration was varied. At high cell concentrations, the number of cells adhering to SHA was twice the number adhering to HA. Such differences were not detected at low cell concentrations. The optimal pH for the adsorption to both SHA and HA was 6. Changes in the ionic strength or addition of monoor divalent cations found in saliva had little effect on adhesion to HA. In contrast, high concentrations of monovalent cations inhibited adhesion to SHA. Anions such as carbonate, chloride, and sulfate did not have specific effects on adhesion, whereas acetate inhibited adhesion to both SHA and HA. Fluoride inhibited adhesion to both SHA and HA, suggesting an interaction between fluoride and hydroxyapatite. These results indicated that 2 mM phosphate buffer at a pH of 6 containing 5 mM KCl and 1 mM CaCl2 was the optimal buffer for studying the in vitro adhesion of S. sanguis to SHA.
MATERIALS AND METHODS Chemicals. Sodium pyruvate, reduced nicotinamide adenine dinucleotide (NADH), FDP, and ATP were purchased from the Sigma Chemical Co., St. Louis, Mo., as were all other components of the LDH assay system. Diethylaminoethyl (DEAE)-cellulose (DE 52) was purchased from Reeve Angel, Clifton, N. J., and 0.5 M Agarose A was obtained from BioRad Laboratories, Richmond, Calif. All components of the A. viscosus growth medium were obtained from Difco Laboratories, Detroit, Mich. Growth of organisms. A. viscosus strain T-6-1600 was grown anaerobically at 37 C in a medium composed of tryptone (0.5% wt/vol), yeast extract (0.5% wt/vol), dibasic potassium phosphate (0.5% wt/vol), and Tween-80 (0.05% wt/vol). The medium was supplemented with sucrose (0.5% wt/vol) as the primary energy source. The organism was routinely grown in 6-liter batches of the complex medium described above, and cells were harvested after an incubation period of 16 h at 40 C. All cells were washed once with 0.01 M potassium phosphate buffer, pH 7.0, after harvesting them by centrifugation, and the cell pellets were frozen at-20 C until used. Laboratory cultures of A. viscosus strain T-6-1600 were kept in NIH fluid thioglycolate medium which contained fluid thioglycolate, 2.9% (wt/vol), and beef extract, 1.2% (wt/vol), supplemented with calcium carbonate, 0.2% (wt/vol). Culture transfers were made
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.