Activity of Pyrazinamide against Mycobacterium tuberculosis at Neutral pH in PZA-S1 Minimal Medium.

Wanliang Shi

doi:10.3390/antibiotics10080909

Abstract

Susceptibility testing of tuberculosis (TB) drugs on Mycobacterium tuberculosis is essential for the rapid detection of strains resistant to the drugs, providing the patient with effective treatment, and preventing the spread of drug-resistant TB strains. Pyrazinamide (PZA) is one of the first-line agents used for the treatment of TB. However, current phenotypic PZA susceptibility testing is unreliable due to its performance in acidic pH conditions. The aims of this study were to develop minimal media to determine the activity of PZA at a neutral pH at 37 °C to avoid problems caused by an acidic pH, which is currently used in PZA susceptibility tests, and to identify PZA-resistant M. tuberculosis in media with reproducibility and accuracy. Different minimal media were used to determine the activity of PZA using the broth microdilution method with M. tuberculosis H37Ra as the reference strain. The PZA-S1 minimal medium was proposed as the most suitable medium. PZA inhibited the growth of M. tuberculosis in PZA-S1 at a neutral pH of 6.8, which is the optimal pH for M. tuberculosis growth. Moreover, PZA showed activity at a neutral pH on a PZA-S1 agar plate when using the disk diffusion method. PZA-resistant M. tuberculosis could be identified at a neutral pH in PZA-S1 minimal medium. This study establishes valuable information regarding the testing of PZA’s susceptibility in relation to M. tuberculosis at a neutral pH of 6.8 with reliability and accuracy in clinical settings.

Highlights

Tuberculosis (TB) is the leading infectious disease worldwide and the pandemic could make it worse [1]
In Group I, the minimal inhibitory concentration (MIC) of PZA against M. tuberculosis was greater than 1000 mg/L and one of five kinds of amino acids, either L-Aspartic acid (L-Asp) (Figure 1D), L-Glutamic acid (L-Glu) (Figure 1E), L-Asparagine (L-Asn) (Figure 1N), L-Glutamine (L-Gln) (Figure 1Q), or L-Cysteine (L-Cys) (Figure 1C), was included as the nitrogen source in the minimal media
In Group III, the MIC of PZA was not detectable since M. tuberculosis failed to grow in the minimal media, which included either L-Methionine (L-Met) (Figure 1M), L-Phenylalanine (L-Phe) (Figure 1F), L-Tryptophan (L-Trp) (Figure 1O) or L-Tyrosine (L-Tyr) (Figure 1J) as the nitrogen source

Summary

Introduction

Tuberculosis (TB) is the leading infectious disease worldwide and the pandemic could make it worse [1]. PZA is playing a role in new regimens designed to further shorten the treatment duration and more effectively treat multidrugresistant tuberculosis (MDR-TB) [3,4]. PZA is a prodrug that is converted to its active form, pyrazinoic acid (POA), by pyrazinamidase/nicotinamidase encoded by the pncA gene in Mycobacterium tuberculosis [6,7]. Several target proteins of PZA were suggested to inhibit fatty acid synthase (Fas I) [8], ribosomal protein S1 (RpsA) [9,10,11], aspartate decarboxylase (PanD) [12,13,14], caseinolytic protease (ClpC1) [15] and guanosine pentaphosphate synthetase (GpsI) [16,17]. There are controversies regarding the target proteins of PZA, Fas I [18], RpsA [19] and PanD [20]

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