Thioridazine and chlorpromazine inhibition of ethidium bromide efflux in Mycobacterium avium and Mycobacterium smegmatis.
OBJECTIVE
Therapy of AIDS patients infected with Mycobacterium avium is problematic due to its intrinsic resistance to antibiotics. We have characterized the efflux pump activity of M. avium wild-type strain through an automated fluorometric method and correlated it with intrinsic resistance to antibiotics.


METHODS
M. avium ATCC 25291(T) and Mycobacterium smegmatis mc(2)155 were evaluated for accumulation and efflux of ethidium bromide in the presence or absence of the efflux pump inhibitors (EPIs) thioridazine, chlorpromazine, verapamil and the proton uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP). For this purpose, a new automated fluorometric method was used that separately assesses accumulation and extrusion of ethidium bromide.


RESULTS
The automated fluorometric method described in this paper allowed the detection and quantification of ethidium bromide transport across M. avium and M. smegmatis cell walls. Accumulation of ethidium bromide was found to be temperature-dependent and significantly increased by EPIs thioridazine, chlorpromazine, verapamil and CCCP in a concentration-dependent manner. Efflux of ethidium bromide under optimum conditions of temperature and glucose is inhibited by the above agents. At half their intrinsic MICs, both thioridazine and chlorpromazine, similarly to verapamil and CCCP, significantly increased the susceptibility of M. avium to erythromycin, suggesting an effect upon an efflux pump with ethidium bromide and erythromycin as substrates. A similar effect was observed for M. smegmatis with verapamil only.


CONCLUSIONS
M. avium and M. smegmatis intrinsic resistance is affected by EPIs such as thioridazine or chlorpromazine, an effect that might be important in research and development of new, more effective antimycobacterial therapies.
Rodrigues
Liliana
L
Unit of Mycobacteriology, Instituto de Higiene e Medicina Tropical, Universidade Nova de Lisboa, Rua da Junqueira 96, 1349-008 Lisboa, Portugal.
Wagner
Dirk
D
Viveiros
Miguel
M
Sampaio
Daniela
D
Couto
Isabel
I
Vavra
Martina
M
Kern
Winfried V
WV
Amaral
Leonard
L
D000900:Anti-Bacterial Agents / Q000378:metabolism / Q000494:pharmacology*; D001331:Automation; D001693:Biological Transport, Active / Q000187:drug effects; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D002746:Chlorpromazine / Q000494:pharmacology*; D004996:Ethidium / Q000378:metabolism*; D005470:Fluorometry / Q000379:methods; D026901:Membrane Transport Proteins / Q000378:metabolism; D009162:Mycobacterium avium / Q000187:drug effects*; D020102:Mycobacterium smegmatis / Q000187:drug effects*; D013881:Thioridazine / Q000494:pharmacology*; D013997:Time Factors; D014700:Verapamil / Q000494:pharmacology
D000900:Anti-Bacterial Agents; D026901:Membrane Transport Proteins; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D014700:Verapamil; D004996:Ethidium; D013881:Thioridazine; D002746:Chlorpromazine