Target site antimicrobial activity of colistin might be misestimated if tested in conventional growth media

Abstract

Cation-dependent inhibition of antimicrobial activity was reported for polymyxin antibiotics. Ca(2+) and Mg(2+) concentrations recommended by the Clinical and Laboratory Standards Institute (CLSI) for the supplementation of Müller-Hinton broth (MHB) are markedly lower than interstitial space fluid (ISF) concentrations in vivo. Hence, it was speculated that the antimicrobial activity of colistin might be overestimated if tested using conventional cation-adjusted MHB. The antimicrobial activity of colistin against n = 100 clinical isolates of Pseudomonas aeruginosa, Acinetobacter baumannii, Klebsiella pneumoniae and Escherichia coli (n = 25 each) was evaluated by broth microdilution and, for selected isolates, by time-kill curves, in MHB without cations (MHB(ONLY)), MHB supplemented with 25 mg/L Ca(2+) and 12.5 mg/L Mg(2+) according to CLSI recommendations (MHB(CLSI)), and in MHB adjusted to 50 mg/L Ca(2+) and 20 mg/L Mg(2+) simulating ISF concentrations (MHB(ISF)). The minimum inhibitory concentration (MIC) values of colistin against the vast majority of isolates of both P. aeruginosa and A. baumannii increased significantly with higher cation concentrations. The susceptibility of K. pneumoniae isolates to colistin did not show significant changes between cation-supplemented media, while the MICs of E. coli decreased with ascending cation concentrations. These findings were confirmed in time-kill studies, where colistin killing against P. aeruginosa 1514 and A. baumannii 1485 declined with increasing cation concentrations. Contrarily, the killing of selected concentrations of colistin against K. pneumoniae 15 and E. coli 16 was enhanced in the presence of increasing cation concentrations. The present data suggest that the clinical antimicrobial activity of colistin might be misestimated in vitro if tested in conventional growth media.

Colistin Heteroresistance and Involvement of the PmrAB Regulatory System in Acinetobacter baumannii

“…They might miss the HR phenotype because of the low bacterial load used in these methods, a load that is inappropriate to detect the (low-frequency) resistant subpopulation. In addition, colistin activity might be overestimated if tested in cation-adjusted Mueller-Hinton (MH) broths (as recommended) compared to interstitial space fluid-like media possibly causing a shift from susceptible to resistant in A. baumannii (37). On the other hand, adsorption of polymyxins to plastics might overestimate colistin MICs, which can be prevented by the use of low-protein-binding polypropylene (38).…”

Polymyxins: Antibacterial Activity, Susceptibility Testing, and Resistance Mechanisms Encoded by Plasmids or Chromosomes

“…Note that the calcium concentration recommended by the CLSI for determining colistin susceptibility in vitro is 2-fold higher than the concentration found in human interstitial space fluid in vivo (92). A recent study revealed that the MIC of colistin might be misestimated if tested with conventional cation-adjusted growth media (overestimation for P. aeruginosa and A. baumannii and underestimation for E. coli) (92). The use of cation-adjusted or non-cation-adjusted medium therefore remains questionable, and a consensus is still needed.…”

“…In fact, it is suspected that the colistin antimicrobial activity might be overestimated if tested using conventional cation-adjusted MH as recommended by the CLSI. Note that the calcium concentration recommended by the CLSI for determining colistin susceptibility in vitro is 2-fold higher than the concentration found in human interstitial space fluid in vivo (92). A recent study revealed that the MIC of colistin might be misestimated if tested with conventional cation-adjusted growth media (overestimation for P. aeruginosa and A. baumannii and underestimation for E. coli) (92).…”

Colistin heteroresistance and the involvement of the PmrAB regulatory system in Acinetobacter baumannii

“…They might miss the heteroresistant phenotype because of the low bacterial load used in these methods, a load that is inappropriate to detect the (low) frequency resistant subpopulation. In addition, colistin activity might be overestimated if tested in cation-adjusted Mueller-Hinton broths (as recommended) as compared to interstitial space fluid-like media possibly causing a shift from susceptible to resistant in A. baumannii (37). At the opposite, adsorption of polymyxins to plastics might overestimate colistin MICs which can be prevent by the use of low-protein-binding polypropylene (38).…”