Selection of resistant bacteria at very low antibiotic concentrations

The widespread use of antibiotics is selecting for a variety of resistance mechanisms that seriously challenge our ability to treat bacterial infections. Resistant bacteria can be selected at the high concentrations of antibiotics used therapeutically, but what role the much lower antibiotic concentrations present in many environments plays in selection remains largely unclear. Here we show using highly sensitive competition experiments that selection of resistant bacteria occurs at extremely low antibiotic concentrations. Thus, for three clinically important antibiotics, drug concentrations up to several hundred-fold below the minimal inhibitory concentration of susceptible bacteria could enrich for resistant bacteria, even when present at a very low initial fraction. We also show that de novo mutants can be selected at sub-MIC concentrations of antibiotics, and we provide a mathematical model predicting how rapidly such mutants would take over in a susceptible population. These results add another dimension to the evolution of resistance and suggest that the low antibiotic concentrations found in many natural environments are important for enrichment and maintenance of resistance in bacterial populations.     

Selective inhibition of the potato scab pathogen by antagonistic bacteria and substrate influence on antibiotic production

Summary In a long-term field experiment a soybean cover crop and green manure incorporation prevented the build-up of common scab of potato whereas barley, employed in the same manner, increased disease incidence. When soil from these plots was assayed for organisms antagonistic toS. scabies, a bacterium indentified asBacillus subtilis, was found to be predominant. Laboratory tests showed thatS. scabies was more sensitive to the antibiotic produced by this bacterium than most non-pathogenicStreptomyces spp which were tested. The antibiotic was found to be similar to bacitracin and activity was expressed as units bacitracin. Water extracts of greenhouse-grown soybean and barley were compared, at different concentrations, as a substrate for growth and antibiotic production by this bacterium. It was found that on the soybean extract, antibiotic activity, as measured by the standard filter-paper-disc technique, was 2.5 to 3 times greater per unit of bacterial growth than when barley extract was used as the substrate. Similar results were obtained when extracts of partially decomposed tissue were used. It is suggested that when evaluating antagonistic organisms as a possible factor in the behavior of plant pathogens in soil, the relative sensitivity of the pathogen to the antibiotic activities of the antagonists as well as the substrates available to the antagonists should be considered.     

The locus of inhibition of NADH oxidation by benzothiadiazoles in beef heart submitochondrial particles

6-Chloro-1,2,3-benzothiadiazole (6-Cl-BTD) is an effective inhibitor of NADH oxidase (site I) but not of succinate oxidase in beef heart submitochondrial particles. For NADH oxidase activity maximal inhibition (80-85%) was achieved at 0.75mM 6-Cl-BTD. A similar level of inhibition was also observed (half maximal inhibitory concentration 0.5mM) towards NADH-duroquinone reductase; NADH-juglone reductase was slightly inhibited (23%) at 0.5mM 6-Cl-BTD while NADH-ferricyanide reductase was unaffected. The data suggest that 6-Cl-BTD interacts with an electron transport site on the oxygen side of NADH dehydrogenase and inhibitory studies with 6-Cl-BTD and rotenone indicate that it might correspond with one of the two sites affected by rotenone. The substituted 1,2,3-benzothiadiazoles (BTDs) are perhaps best known for their activity as inhibitors of cytochrome P-450-mediated mixed-function oxidation (MFO). In vitro, the BTDs are potent inhibitors of MFO activities in microsomes from mammalian liver and insect tissues and they have been demonstrated to inhibit aminopyrine metabolism in perfused rat liver. In vivo, they reportedly prolong hexobarbital sleeping time in mice, inhibit the irreversible binding of labeled trichloro-ethylene to microsomal protein and effectively enhance the toxicity (synergize) of pyrethrin, organophosphorus-containing and carbamate insecticides to insects.     

Combating antibiotic resistance in bacteria

Combinations of certain antibiotics select against resistant strains of bacteria. This finding may provide a strategy of combating antibiotic resistant bacteria.     

The inhibition of mucopeptide synthesis by benzylpenicillin in relation to irreversible fixation of the antibiotic by staphylococci

1. Benzylpenicillin is irreversibly fixed to staphyloccoci by a reaction that obeys second-order kinetics, whereas the progress of inhibition of mucopeptide synthesis obeys first-order kinetics after a short lag during which the antibiotic has no effect. 2. When the micro-organisms are saturated with benzylpenicillin they can still make mucopeptide in solutions containing chloramphenicol at a normal rate after a lag period. 3. About 90% of the benzylpenicillin stays fixed to the cells after mucopeptide synthesis has reached its maximum and constant rate. 4. During the phase when mucopeptide synthesis by cells saturated with benzylpenicillin is accelerating, a small number of additional sites that fix benzylpenicillin is revealed. The number of these sites reaches a maximum and constant value at about the same time as mucopeptide biosynthesis reaches a maximum and constant rate. 5. Staphylococci saturated with benzylpenicillin are exceedingly sensitive to fresh additions of the antibiotic. 6. The degree of inhibition of mucopeptide synthesis caused by these small amounts of antibiotic agrees with the degree of substitution by benzylpenicillin of the newly revealed or `sensitive' sites. 7. Since these sensitive sites are revealed during incubation of the bacteria with chloramphenicol it is unlikely that they are due to newly formed protein. 8. On the basis of these results, a hypothesis for the inhibition by penicillin of the cross-linking reaction in the terminal stages of mucopeptide synthesis is suggested.