Applying reporter strains to probe mode of action in Gram negative bacteria
Author(s): ,
Bonnie Evans
Affiliations:
Public Health England
,
Mark Sutton
Affiliations:
Public Health England
,
Lucy Bock
Affiliations:
Public Health England
Charlotte Hind
Affiliations:
Public Health England
PHE ePoster Library. Evans B. Mar 20, 2018; 205930; 12606
Bonnie Evans
Bonnie Evans
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Abstract
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Abstract Drug-resistant infections pose an ever increasing threat to human health, and novel antibiotics are required to meet this challenge. A major bottleneck in the development of new antimicrobial compounds is limited knowledge about their mode of action. This results in lack of understanding on how to improve and progress these compounds.We have purchased a library of fluorescent reporter strains, which have GFP fused to each of around 2,000 Escherichia coli promoters. The library will be screened by measuring fluorescence induction, and therefore promoter activity, of each reporter strain when exposed to sub-inhibitory concentrations of antibiotics. This will allow us to determine genes activated in response to antibiotics with different modes of action, such as cell wall synthesis, DNA replication, transcription and translation. The GFP-promoter(s) that best indicate each mode of action will be taken forward to establish a reporter strain assay. We validate the assay by using a panel of antibiotics with established mechanisms of action, to create a ‘fingerprint' of fluorescence induction for each mode of action across the chosen reporter strains. The reporter strain assay is then used to determine the mode of action of novel compounds in a high-throughput screen. This also has the potential to uncover antibiotics with novel modes of action, which are urgently needed in the antibiotic development pipeline. The assay will be a toolkit for mode of action studies of novel antimicrobials in Gram negative bacteria, which is essential for further investigations of relevant compounds. Funding Placement student 2017/18
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