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Watching DNA replication inhibitors in action: Exploiting time-lapse microfluidic microscopy as a tool for target-drug interaction studies in Mycobacterium .
Trojanowski, Damian Kołodziej, Marta Hołówka, Joanna Zakrzewska-Czerwińska, Jolanta
Trojanowski, Damian
Kołodziej, Marta
Hołówka, Joanna
Zakrzewska-Czerwińska, Jolanta
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2019-08-05
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Spreading resistance to antibiotics and the emergence of multidrug-resistant strains have become frequent in many bacterial species, including mycobacteria - a causative agents of severe diseases and have profound impacts on global health. Here, we used a system of microfluidics, fluorescence microscopy and target-tagged fluorescent reporter strains of Mycobacterium smegmatis to perform real-time monitoring of replisome and chromosome dynamics following the addition of replication-altering drugs (novobiocin, nalidixic acid and griselimycin) at the single-cell level. We found that novobiocin stalled replication forks and caused relaxation of the nucleoid, nalidixic acid triggered rapid replisome collapse and compaction of the nucleoid, while griselimycin caused replisome instability with subsequent over-initiation of chromosome replication and over-relaxation of the nucleoid. In addition to study target-drug interactions, our system also enabled to observe how the tested antibiotics affected the physiology of mycobacterial cells (i.e., growth, chromosome segregation, etc.).
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Antimicrob Agents Chemother. 2019 Aug 5. pii: AAC.00739-19. doi: 10.1128/AAC.00739-19.
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1098-6596
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Attribution-NonCommercial-ShareAlike 4.0 International