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Microbiology. 2019 Feb 11. doi: 10.1099/mic.0.000777. [Epub ahead of print]

Very rapid flow cytometric assessment of antimicrobial susceptibility during the apparent lag phase of microbial (re)growth.

Jindal S1,2, Thampy H3, Day PJR4,1, Kell DB1,3,2,5.

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2​Manchester Institute of Biotechnology, The University of Manchester, 131 Princess St, Manchester M1 7DN, UK.
1​School of Chemistry, The University of Manchester, 131 Princess St, Manchester M1 7DN, UK.
3​Firsway Health Centre, 121 Firs Way, Sale, Manchester M33 4BR, UK.
4​Faculty of Faculty of Biology, Medicine and Health, The University of Manchester, Manchester M13 9PT, UK.
†​Present address: Department of Biochemistry, Institute of Integrative Biology, Faculty of Health and Life Sciences, University of Liverpool, Crown St, Liverpool L69 7ZB, UK.


Rapid changes in the number and flow cytometric behaviour of cells of E. coli taken from a stationary phase and inoculated into rich medium.Cells of E. coli were grown in LB medium, taken from a stationary phase of 2-4 h, and re-inoculated into fresh media at a concentration (105 ml-1 or lower) characteristic of bacteriuria. Flow cytometry was used to assess how quickly we could detect changes in cell size, number, membrane energization (using a carbocyanine dye) and DNA distribution. It transpired that while the lag phase observable macroscopically via bulk OD measurements could be as long as 4 h, the true lag phase could be less than 15-20 min, and was accompanied by many observable biochemical changes. Antibiotics to which the cells were sensitive affected these changes within 20 min of re-inoculation, providing the possibility of a very rapid antibiotic susceptibility test on a timescale compatible with a visit to a GP clinic. The strategy was applied successfully to genuine potential urinary tract infection (UTI) samples taken from a doctor's surgery. The methods developed could prove of considerable value in ensuring the correct prescription and thereby lowering the spread of antimicrobial resistance.


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