Anti-staphylococcal activity and mode of action of clofazimine.

OBJECTIVES: Infections caused by Staphylococcus aureus might be treated with agents whose primary indications are for other infections. Clofazimine, an established anti-mycobacterial drug, could be such a candidate. However, the anti-staphylococcal properties of clofazimine have not been fully described and its mode of action, possibly involving inhibition of both RNA polymerase and a membrane-located target, has not been explored in detail. We have now conducted experiments to address these issues. METHODS: Using established procedures, we examined the activity of clofazimine against a range of clinical isolates of S. aureus and determined whether it was bactericidal, exhibited a post-antibiotic effect (PAE), or interacted synergically with other agents. The potential for emergence of clofazimine-resistant mutants was also examined. Mode of action studies involved macromolecular synthesis assays, cross-screening against rifampicin-resistant mutants, susceptibility of RNA polymerase to clofazimine in vitro and several methods to detect drug-induced membrane damage. RESULTS: Clofazimine demonstrated good anti-staphylococcal activity encompassing MSSA, MRSA and GISA. It was bactericidal and resistant mutants could not be isolated. Clofazimine did not exhibit a PAE and failed to act synergically with other drugs. No evidence for specific inhibition of RNA polymerase was obtained. Clofazimine caused non-specific inhibition of DNA, RNA and protein synthesis, consistent with membrane-damaging activity that was detected in three independent assays for membrane disrupting agents. CONCLUSIONS: Clofazimine is a potent anti-staphylococcal agent. It appears to be a membrane-disrupting agent and does not inhibit RNA polymerase.

Biosynthetic pathways in Mycobacterium leprae.

The scope of this paper is biosynthetic pathways which are found generally in living organisms. Many are present in M. leprae but some are lacking, perhaps reflecting the parasitic mode of existence of this microbe. The biosynthesis of nucleotides (the immediate precursors of nucleic acids) and fatty acids (intermediates in the biosynthesis of most if not all mycobacterial lipids) has been studied in detail in M. leprae. For the former, M. leprae appears incapable of synthesising its own purines de novo and thus depends upon the host for a source of the purine ring. It can synthesise pyrimidines, though enzymes for this activity are inhibited in the M. leprae organisms obtained from host tissue. For the latter, M. leprae is capable of fatty acid biosynthesis de novo, probably at a repressed level. However, a key enzyme, phosphotransacetylase, appears to be deficient.

Control of ribosome synthesis in Escherichia coli: analysis of an energy source shift-down.

The rate of ribosome synthesis and accumulation in Escherichia coli during the transition after an energy source shift-down was analyzed. The shift was imposed on cultures of stringent and relaxed strains growing in glucose minimal medium by the addition of the glucose analogue alpha-methylglucoside. In the stringent strain, ribosome synthesis was almost instantaneously reduced after the shift, whereas the relaxed strain exhibited a more gradual response. The rate of messenger ribonucleic acid (mRNA) synthesis was affected similarly, though to a smaller extent. A comparison of the rates of synthesis and accumulation of ribosomal RNA (rRNA) and ribosomal proteins showed that far more ribosomal components were synthesized after the shift than were accumulated, indicating that a substantial part of the rRNA made after the shift was unstable. A new method was used to measure relative rates of rRNA synthesis and to estimate the transcription time for the rRNA operon under different conditions. In steady states of growth with growth rates ranging from 0.75 to 2.3 doublings/h, as well as during the transition after a shift-down, the transcription time of the rRNA operon was constant. The rate of synthesis of rRNA correlated during this transition - in contrast to the rate of accumulation (M. T. Hansen et al., J. Bacteriol. 122: 585-591, 1975) - with the ppGpp pool in the same way as has been observed during partial amino acid starvation.