Development and Optimization of Chromosomally-Integrated Fluorescent Mycobacterium tuberculosis Reporter Constructs

TY - JOUR

T1 - Development and Optimization of Chromosomally-Integrated Fluorescent Mycobacterium tuberculosis Reporter Constructs

AU - Kolbe, Katharina

AU - Bell, Alice C.

AU - Prosser, Gareth A.

AU - Assmann, Maike

AU - Yang, Hee Jeong

AU - Forbes, He Eun

AU - Gallucci, Sophia

AU - Mayer-Barber, Katrin D.

AU - Boshoff, Helena I.

AU - Barry, Clifton E.

N1 - Funding Information: This work was funded by the Intramural Research Program of the National Institute of Allergy and Infectious Disease, National Institutes of Health. The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication. Publisher Copyright: © Copyright © 2020 Kolbe, Bell, Prosser, Assmann, Yang, Forbes, Gallucci, Mayer-Barber, Boshoff and Barry.

PY - 2020/12/9

Y1 - 2020/12/9

N2 - Mycobacterium tuberculosis resides in the lungs in various lesion types with unique microenvironmental conditions. This diversity is in line with heterogeneous disease progression and divergent drug efficiency. Fluorescent reporter strains can be used to decipher the micromilieu and to guide future treatment regimens. Current reporters using replicating plasmids, however, are not suitable for long-term mouse infections or studies in non-human primates. Using a combination of recombinant DNA and protein optimization techniques, we have developed reporter strains based on integrative plasmids, which exhibit stimulus-response characteristics and fluorescence intensities comparable to those based on replicating plasmids. We successfully applied the concepts by constructing a multi-color reporter strain able to detect simultaneous changes in environmental pH, Mg2+ concentrations, and protein expression levels.

AB - Mycobacterium tuberculosis resides in the lungs in various lesion types with unique microenvironmental conditions. This diversity is in line with heterogeneous disease progression and divergent drug efficiency. Fluorescent reporter strains can be used to decipher the micromilieu and to guide future treatment regimens. Current reporters using replicating plasmids, however, are not suitable for long-term mouse infections or studies in non-human primates. Using a combination of recombinant DNA and protein optimization techniques, we have developed reporter strains based on integrative plasmids, which exhibit stimulus-response characteristics and fluorescence intensities comparable to those based on replicating plasmids. We successfully applied the concepts by constructing a multi-color reporter strain able to detect simultaneous changes in environmental pH, Mg2+ concentrations, and protein expression levels.

KW - fluorescent protein

KW - magnesium

KW - Mycobacterium tuberculosis

KW - pH

KW - reporter strain

KW - riboswitch

UR - https://www.scopus.com/pages/publications/85098191513

U2 - 10.3389/fmicb.2020.591866

DO - 10.3389/fmicb.2020.591866

M3 - Article

C2 - 33362741

AN - SCOPUS:85098191513

SN - 1664-302X

VL - 11

JO - Frontiers in Microbiology

JF - Frontiers in Microbiology

M1 - 591866

ER -