I determined MBC results for a MDR M. tuberculosis strain and I have some strange results I can't make sense of. The MBC definition I used was a greater than 99 % reduction of viable cells buy plating out onto 7H10 OADC agar and counting colonies. I investigated a cationic peptide at 16, 32, 64, 128 and 256 mg/L. I determined the following reduction; 97.12 % (16 mg/L), 98.36 % (32 mg/L), 99.00 % (64 mg/L), 97.74 % (128 mg/L), 97.46 % (256 mg/L).
I have a H37 control and it had a MBC of 256 mg/L with 99 % reduction.
Statistical analysis revealed that the MBC of H37 is not different from the MBC for MDR at 64 mg/L. However, all concentrations tested for the MDR are not statistically different from each other.
Using the MBC definition, the MBC is 64 mg/L. But this doesn't make sense since concentrations above this are less than 99 % reduction of cells. But are not statistically different?
How do I determine what the MBC is here?
I did the experiments in duplicate on the day and triplicate over three different days. The standard deviation is low enough as well. My coefficient of variation is below 10 %. In addition, I included growth controls, sterile controls and INH and RIF controls. Everything checks out fine.
Thanks in advance.
I think your experience is correct. you should attention to your cationic peptide, It may be they have adverse effect on each other , in high concentration. I think 64 mg/L is the MBC of your isolate.
Shane, how many times have you repeated the test? If you did the test in triplets and still got the same results than it could be that you are getting false positive results and it might be that it's your dissolving solvent which is reacting against the bacteria rather than your compound.
Have you measured the concentration of bacteria in each well prior to the treatment? Have you done any troubleshooting?
Did you test at concentrations lower than that tested for the cationic peptide?Also did you check the solubility behaviour of the sample in the solvent you used
HI guys, the solvent I used was dH2O. So I don't expect a problem there and my peptide easily dissolves in dH2O as it is amphiphatic. I did the experiments in triplicate and I measured the CFU/ml for the strain at the beginning of each experiment. I determined there was about 1 x 10^5 CFU/ml.
I decided to use statistics to determine my MBC. For example, since there was no statistical difference above 64 mg/L, it must be the MBC.
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