Role of Acid pH and Deficient Efflux of Pyrazinoic Acid in Unique Susceptibility of Mycobacterium tuberculosis to Pyrazinamide

ABSTRACT Pyrazinamide (PZA) is an important antituberculosis drug. Unlike most antibacterial agents, PZA, despite its remarkable in vivo activity, has no activity against Mycobacterium tuberculosis in vitro except at an acidic pH. M. tuberculosis is uniquely susceptible to PZA, but other mycobacteria as well as nonmycobacteria are intrinsically resistant. The role of acidic pH in PZA action and the basis for the unique PZA susceptibility of M. tuberculosisare unknown. We found that in M. tuberculosis, acidic pH enhanced the intracellular accumulation of pyrazinoic acid (POA), the active derivative of PZA, after conversion of PZA by pyrazinamidase. In contrast, at neutral or alkaline pH, POA was mainly found outside M. tuberculosis cells. PZA-resistantM. tuberculosis complex organisms did not convert PZA into POA. Unlike M. tuberculosis, intrinsically PZA-resistant M. smegmatis converted PZA into POA, but it did not accumulate POA even at an acidic pH, due to a very active POA efflux mechanism. We propose that a deficient POA efflux mechanism underlies the unique susceptibility of M. tuberculosisto PZA and that the natural PZA resistance of M. smegmatis is due to a highly active efflux pump. These findings may have implications with regard to the design of new antimycobacterial drugs.

[1]  V. Mizrahi,et al.  Purification, Gene Cloning, Targeted Knockout, Overexpression, and Biochemical Characterization of the Major Pyrazinamidase fromMycobacterium smegmatis , 1998, Journal of bacteriology.

[2]  A. Scorpio,et al.  The pncA gene from naturally pyrazinamide-resistant Mycobacterium avium encodes pyrazinamidase and confers pyrazinamide susceptibility to resistant M. tuberculosis complex organisms. , 1997, Microbiology.

[3]  H. Nikaido,et al.  Active efflux of bile salts by Escherichia coli , 1997, Journal of bacteriology.

[4]  R. Gilman,et al.  Characterization of pncA mutations in pyrazinamide-resistant Mycobacterium tuberculosis , 1997, Antimicrobial agents and chemotherapy.

[5]  Ying Zhang,et al.  Mutations in pncA, a gene encoding pyrazinamidase/nicotinamidase, cause resistance to the antituberculous drug pyrazinamide in tubercle bacillus , 1996, Nature Medicine.

[6]  T. Weisbrod,et al.  Efflux pump of the proton antiporter family confers low-level fluoroquinolone resistance in Mycobacterium smegmatis. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[7]  D. Livermore,et al.  Role of efflux pump(s) in intrinsic resistance of Pseudomonas aeruginosa: resistance to tetracycline, chloramphenicol, and norfloxacin , 1994, Antimicrobial Agents and Chemotherapy.

[8]  H. Nikaido,et al.  Penetration of lipophilic agents with multiple protonation sites into bacterial cells: tetracyclines and fluoroquinolones as examples , 1993, Antimicrobial Agents and Chemotherapy.

[9]  S. Cole,et al.  The catalase—peroxidase gene and isoniazid resistance of Mycobacterium tuberculosis , 1992, Nature.

[10]  L. Heifets,et al.  Pyrazinamide sterilizing activity in vitro against semidormant Mycobacterium tuberculosis bacterial populations. , 1992, The American review of respiratory disease.

[11]  V. Bidnenko,et al.  Efflux-mediated multidrug resistance in Bacillus subtilis: similarities and dissimilarities with the mammalian system. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[12]  M. Salfinger,et al.  Determination of pyrazinamide MICs for Mycobacterium tuberculosis at different pHs by the radiometric method , 1988, Antimicrobial Agents and Chemotherapy.

[13]  S. Trivedi,et al.  Pyrazinamidase activity of Mycobacterium tuberculosis--a test of sensitivity to pyrazinamide. , 1987, Tubercle.

[14]  B. Plikaytis,et al.  Identification and drug susceptibility test results for Mycobacterium spp. , 1985 .

[15]  D. Mitchison.  The action of antituberculosis drugs in short-course chemotherapy. , 1985, Tubercle.

[16]  A. Azzi,et al.  The effect of N,N'-dicyclohexylcarbodiimide on enzymes of bioenergetic relevance. , 1984, Biochimica et biophysica acta.

[17]  A. Tsang,et al.  Use of pyrazinamidase activity on Mycobacterium tuberculosis as a rapid method for determination of pyrazinamide susceptibility , 1981, Antimicrobial Agents and Chemotherapy.

[18]  H. David,et al.  Basis for lack of drug susceptibility of atypical mycobacteria. , 1981, Reviews of infectious diseases.

[19]  H. Kaback,et al.  The electrochemical gradient of protons and its relationship to active transport in Escherichia coli membrane vesicles. , 1976, Proceedings of the National Academy of Sciences of the United States of America.

[20]  I. Tárnok,et al.  Biochemical background of some enzymatic tests used for the differentiation of mycobacteria. , 1976, Tubercle.

[21]  B. Olivera,et al.  Pyridine nucleotide metabolism in Escherichia coli. 3. Biosynthesis from alternative precursors in vivo. , 1973, The Journal of biological chemistry.

[22]  J. Youatt,et al.  Radioactive content of Mycobacterium tuberculosis after exposure to 14C-isoniazid. , 1969, The American review of respiratory disease.

[23]  W. Mcdermott,et al.  Pyrazinamide susceptibility and amidase activity of tubercle bacilli. , 1967, The American review of respiratory disease.

[24]  W. Mcdermott,et al.  THE FATE OF MYCOBACTERIUM TUBERCULOSIS IN MOUSE TISSUES AS DETERMINED BY THE MICROBIAL ENUMERATION TECHNIQUE , 1956, The Journal of experimental medicine.

[25]  W. Mcdermott,et al.  Activation of pyrazinamide and nicotinamide in acidic environments in vitro. , 1954, American review of tuberculosis.

[26]  Tarshis Ms,et al.  Lack of significant in vitro sensitivity of Mycobacterium tuberculosis to pyrazinamide on three different solid media. , 1953 .

[27]  L. Malone,et al.  The effect of nicotinic acid amide on experimental tuberculosis of white mice. , 1948, The Journal of laboratory and clinical medicine.

[28]  Susan Budavari,et al.  The Merck index , 1998 .

[29]  F. Ryan,et al.  The Forgotten Plague: How the Battle Against Tuberculosis Was Won - And Lost , 1992 .

[30]  H. Rottenberg.  The measurement of membrane potential and deltapH in cells, organelles, and vesicles. , 1979, Methods in enzymology.

[31]  L. Wayne,et al.  Simple pyrazinamidase and urease tests for routine identification of mycobacteria. , 1974, The American review of respiratory disease.

[32]  M. Tarshis,et al.  Lack of significant in vitro sensitivity of Mycobacterium tuberculosis to pyrazinamide on three different solid media. , 1953, American review of tuberculosis.