Two Transport Systems for Tetracycline in Sensitive
            <i>Escherichia coli</i>
            : Critical Role for an Initial Rapid Uptake System Insensitive to Energy Inhibitors

The Nine Mile, S Q217, and Priscilla isolates, representative of the three major genetic groups of Coxiella burnetii, are known to differ in their susceptibilities to antibiotics. Mechanisms potentially responsible for these differences were investigated. Accumulation of antibiotics by infected L929 cells and purified isolates was measured. In addition, C. burnetii plasmid-transformed Escherichia coli HB101 cells were used to study the possibility that different C.

Section: Discussionsupporting

confidence: 52%

Mechanisms that may account for differential antibiotic susceptibilities among Coxiella burnetii isolates

Yeaman

Baca

1991

“…An energy-dependent accumulation of the tetracyclines occurred in susceptible E. coli ML3Q8-225 grown in medium A and assayed in phosphate buffer; subsequent blocking of energy production by the uncoupler 2,4-dinitrophenol (DNP) caused the drug to flow out of the cells (e.g., see minocycline, Fig. 1A) (21)(22)(23). However, when the same strain was grown in L broth before being assayed in buffer, minocycline and tetracycline accumulation was much smaller, and the addition of DNP either decreased the uptake very little or, particularly in the case of minocycline, actually increased uptake (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…We therefore propose that most of the tetracyclines which accumulate in these cells exist in at least three cellular compartments: the membrane, the ribosome, and the efflux system. Furthermore, since binding is reversible (3,21,22), it is likely that membrane-bound drug is in equilibrium with that on the ribosomes. Most of the active net uptake seen in cells grown in minimal media may also represent increased bound drug and not free drug.…”

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Susceptible Escherichia coli cells can actively excrete tetracyclines

McMurry

Aronson

Levy

1983

Escherichia coli shows severalfold less susceptibility to tetracyclines when grown in enriched medium than in minimal medium. Transport studies with cells harvested from these media showed different handling of the drugs. Whereas an energy-dependent uptake oftetracycline and minocycline was observed in susceptible K-12 and wild-type E. coli strains grown in a medium, an active efflux of minocycline and, to a lesser extent, tetracycine was seen in cells grown in L broth and other enriched media. This efflux was replaced by an active uptake system after treatment of cells grown in L broth with EDTA. When assayed at lower temperature (270C), even cells grown in minimal medium showed an efflux of minocycline. Everted membrane vesicles pgepared from susceptible cells grown in minimal medium or L broth showed. an energy-dependent accumulation of nunocycline and tetracycline when supplied with certain divalent cations. These results suggest that an active efflux of tetracyclines occurs in susceptible E. coli but is not detected in cells grown in minimal medium because greater permeability of the outer membrane allows a more rapid active uptake. This efflux system is distinct from that specified by tetracycline resistance determinants. Since the active efflux of minocycline in cells grown in L broth disappeared at external antibiotic concentrations of >100 ,uM, it may be saturable and so mediated by a membrane carrier.More is known about how a bacterium takes up needed moeules or ions from its environment than how it excretes-them. Among the latter processes are energy-dependent efflux mechanisms for sodium (30), calcium (6, 28) and protons (15,27

“…It is of interest that the uptake of tetracycline by Lforms is only slightly sensitive to DNP. As pointed out by McMurry and Levy (20), E. coli displays two tetracycline uptake kinetics, the first one DNP insensitive and the second inhibited by DNP. The kinetics of tetracycline uptake by the parent Listeria strain appears to be biphasic with a rapid binding insensitive to DNP up to 10 min of incubation.…”

Section: Discussionmentioning

confidence: 99%

Tetracycline-resistant L-forms isolated from an antibiotic-susceptible strain of Listeria monocytogenes

Schmitt-Slomska¹,

Marmouset²,

Louis³

et al. 1982

A tetracycline-susceptible strain of Listeria monocytogenes type 4b was converted to stable L-forms by penicillin. L-form variants resistant to tetracycline were then selected from a predominantly tetracycline-susceptible L-form population on plates containing penicillin and increasing concentrations of tetracycline. The origin of tetracycline-resistant L-forms from the parent Listeria strain was confirmed biochemically, by immunofluorescence, and by polyacrylamide gel electrophoresis. Scanning and transmission electron microscopy confirmed the typical L-form structure and the complete lack of cell wall in both L-form strains. The level of [3H]tetracycline uptake was lower in tetracycline-resistant than in susceptible cells.Although the molecular basis of tetracycline resistance has been actively explored, relatively little is known about the mechanism of tetracycline accumulation in gram-positive bacteria (3). In gram-negative bacteria, tetracycline resistance appears to result from changes in the cell envelope or from intracellular protection of ribosomes or both (16,28). However, the mechanism of this phenomenon is not obvious (3). The acquisition of tetracycline resistance in Escherichia coli is accompanied by an alteration of tetracycline uptake (7). Penicillin-induced spheroplasts of E. coli accumulate about 30% less tetracycline than do intact bacteria (8). To our knowledge, there have been no reports of tetracycline resistance in wall-less forms of grampositive bacteria. Most penicillin-induced Lforms have been reported to be at least as susceptible to tetracycline as the parent bacteria (9, 11-13, 23, 27). Recently, have found the loss of plasmid-mediated resistance to several antibiotics, including tetracycline, in stable L-forms of group B streptococci carrying R plasmids. This loss was associated with the apparent physical elimination of plasmid DNA.The purpose of this study was to investigate the unusual finding of resistance to tetracycline in L-form variants isolated as satellites from a tetracycline-susceptible L-form population of an antibiotic-susceptible strain of Listeria monocytogenes. MATERIALS AND METHODSStrains and media. L. monocytogenes 5, type 4b, was isolated from the spinal fluid of a patient at the Massachusetts General Hospital. The parent Listeria strains were grown in Trypticase soy broth (TSB; BBL Microbiology Systems, Cockeysville, Md.). L-form culture media contained TSB supplemented with 3% NaCl (wt/vol) as an osmotic stabilizer, 10% (vol/vol) Todd-Hewitt broth (Difco Laboratories, Detroit Mich.), and 10% inactivated horse serum (TSB-L); for solid medium (TSA-L), 1% agar (Difco) was added. These media were occasionally supplemented with 1,000 U of penicillin per ml.L-form induction and reversion procedures. L-forms were obtained on TSA-L medium by the penicillin gradient method (S. Madoff and L. Dienes, Bacteriol. Proc., 1967, p. 70-71; 29). Subcultures of L-colonies were performed by weekly transfers on TSA-L penicillin medium. The growth of L-forms in liquid medium was ob...