Chapter 26
Determining Minimum Inhibitory Concentrations in Liquid
Cultures or on Solid Medium
Qinglan Wang and Helena I. M. Boshoff
Abstract
Antimicrobial susceptibility testing is the mainstay of tuberculosis drug development programs. In this
chapter, we describe methods for determination of the minimum inhibitory concentration of compounds
against Mycobacterium tuberculosis growing in liquid media as a function of carbon source, detergent, and
environmental stress imposed by acidic pH as well as reactive nitrogen intermediates. Methods for determining the effect of bovine serum albumin in the growth medium on antimicrobial susceptibility are also
described. Finally, we provide a method for antimicrobial susceptibility testing on agar medium.
Key words Minimum inhibitory concentration (MIC), Agar MIC, Carbon source, Detergent, Liquid
MIC, Nitrosative stress, Acidified medium
1 Introduction
The goal of current drug discovery efforts for tuberculosis is to
accelerate the decline in global incidence of this disease by developing regimens that are superior to frontline and second line drug
combinations in terms of safety and duration of treatment while
remaining affordable and less prone to the development of drug
resistance in the face of non-adherence. Whole cell screening
against Mycobacterium tuberculosis (M. tuberculosis) has been key
to the discovery of all drug candidates in the tuberculosis drug
discovery pipeline over the last few decades. Several drug discovery
efforts have attempted to recapitulate aspects of in vivo pathogenesis of the organism that were deemed critical for the replication or
survival of the pathogen in humans including growth in macrophages [1–3], survival at acidic pH [4], hypoxia [5], biofilm formation [6], nutrient starvation [7], growth on fatty acids [8–10],
glucose, cholesterol [1], multi-stress conditions [11], as well as
growth under nutrient-rich conditions [12, 13]. The importance
of recapitulating in vivo relevant growth conditions was emphasized by the discovery of several hits that lacked in vivo efficacy due
Tanya Parish and Anuradha Kumar (eds.), Mycobacteria Protocols, Methods in Molecular Biology, vol. 2314,
https://doi.org/10.1007/978-1-0716-1460-0_26, © Springer Science+Business Media, LLC, part of Springer Nature 2021
595
Determining Minimum Inhibitory Concentrations in Liquid
Cultures or on Solid Medium
Qinglan Wang and Helena I. M. Boshoff
Abstract
Antimicrobial susceptibility testing is the mainstay of tuberculosis drug development programs. In this
chapter, we describe methods for determination of the minimum inhibitory concentration of compounds
against Mycobacterium tuberculosis growing in liquid media as a function of carbon source, detergent, and
environmental stress imposed by acidic pH as well as reactive nitrogen intermediates. Methods for determining the effect of bovine serum albumin in the growth medium on antimicrobial susceptibility are also
described. Finally, we provide a method for antimicrobial susceptibility testing on agar medium.
Key words Minimum inhibitory concentration (MIC), Agar MIC, Carbon source, Detergent, Liquid
MIC, Nitrosative stress, Acidified medium
1 Introduction
The goal of current drug discovery efforts for tuberculosis is to
accelerate the decline in global incidence of this disease by developing regimens that are superior to frontline and second line drug
combinations in terms of safety and duration of treatment while
remaining affordable and less prone to the development of drug
resistance in the face of non-adherence. Whole cell screening
against Mycobacterium tuberculosis (M. tuberculosis) has been key
to the discovery of all drug candidates in the tuberculosis drug
discovery pipeline over the last few decades. Several drug discovery
efforts have attempted to recapitulate aspects of in vivo pathogenesis of the organism that were deemed critical for the replication or
survival of the pathogen in humans including growth in macrophages [1–3], survival at acidic pH [4], hypoxia [5], biofilm formation [6], nutrient starvation [7], growth on fatty acids [8–10],
glucose, cholesterol [1], multi-stress conditions [11], as well as
growth under nutrient-rich conditions [12, 13]. The importance
of recapitulating in vivo relevant growth conditions was emphasized by the discovery of several hits that lacked in vivo efficacy due
Tanya Parish and Anuradha Kumar (eds.), Mycobacteria Protocols, Methods in Molecular Biology, vol. 2314,
https://doi.org/10.1007/978-1-0716-1460-0_26, © Springer Science+Business Media, LLC, part of Springer Nature 2021
595