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Strange minimum inhibitory concentrations for M. tuberculosis, possible growth r

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#1 Shanevanbreda



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Posted 17 October 2013 - 02:44 AM

Hi guys,

I have a problem interpreting my MIC (minimum inhibitory concentration) results determined by the the alamar blue assay. Briefly, I am investigating the MIC of a cationic peptide against M. tuberculosis (H37Ra ATCC 25177). I have investigated the MIC of my compound in three media (7H9 with OADC (oleic acid, albumin, dextrose, catalase) enrichment, 7H9 without OADC enrichment and Sauton media [cation adjusted]), for the reason that I believe OADC enrichment causes an interaction with my cationic peptide, falsely elevating it's MIC.

I perform the assay as published previously by Franzblau et al. with a starting inoculum of 5 x 10^5 CFU/ml. For 7H9 with OADC and 7H9 without OADC, alamar blue is added on day 5 with color change from blue to pink indicating sufficient cellular growth in control wells and my MIC results are read. For Sauton media, alamar blue is added on day 9 and results are read. I recorded the following trend in MICs: 7H9 with OADC = > 100 ug/ml, 7H9 without OADC = 25 - 50 ug/ml and Sauton media = 0.39 - 0.77 ug/ml.

It must be noted that all my controls check out for my experiment i.e., no contamination etc

What could explain this trend of decreasing MIC results? Is the decrease in MICs observed due to different growth rates and longer lag phases or possibly due to less drug interactions with the media?

I have not investigated how the MICs of isoniazid and rifampicin change with the different media. I guess this is the most logical starting point since these MICs are known.

If there is a change in MIC results (including isoniazid and rifampicin) does this mean there is a problem with growth rates? If I determine that 7H9 and OADC are in fact causing an drug interaction, but Sauton is not, how would it be possible to increase my growth rate to get comparable MIC results for isoniazid and rifampicin so that I can determine the correct MIC for my cationic peptide?

Any suggestions and help will be greatly appreciated.

Thank you

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