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Abstract
For the pig respiratory tract pathogens, Actinobacillus pleuropneumoniae and Pasteurella multocida, Minimum Inhibitory Concentration (MIC) of marbofloxacin was determined in recommended broths and pig serum at three inoculum strengths. MICs in both growth matrices increased progressively from low, through medium to high starting inoculum counts, 104, 106 and 108CFU/mL, respectively. P. multocida MIC ratios for high:low inocula were 14:4:1 for broth and 28.2:1 for serum. Corresponding MIC ratios for A. pleuropneumoniae were lower, 4.1:1 (broth) and 9.2:1 (serum). MIC high:low ratios were therefore both growth matrix and bacterial species dependent. The effect of alterations to the chemical composition of broths and serum on MIC were also investigated. Neither adjusting broth or serum pH in six increments over the range 7.0 to 8.0 nor increasing calcium and magnesium concentrations of broth in seven incremental steps significantly affected MICs for either organism. In time-kill studies, the killing action of marbofloxacin had the characteristics of concentration dependency against both organisms in both growth matrices. It is concluded that MIC and time-kill data for marbofloxacin, generated in serum, might be preferable to broth data, for predicting dosages of marbofloxacin for clinical use.
Highlights
The most widely used parameter of antimicrobial drug potency is Minimum Inhibitory Concentration (MIC)
European Union Committee on Antimicrobial Sensitivity testing (EUCAST) and Clinical Laboratory Standards Institute (CLSI) standardised MIC data are generated in broths, such as Cation Adjusted Mueller Hinton Broth (CAMHB), each formulated to facilitate growth in vitro, the composition differing from biological fluids in general and from the biophase in particular, in respect of chemical, immunological and cellular constituents, so that they might not be representative of bacterial growth conditions in vivo;
MICs were determined by microdilution for six isolates each of A. pleuropneumoniae and P. multocida, in accordance with CLSI guidelines, except for (1) using five sets of overlapping two-fold serial dilutions to reduce inaccuracy of individual isolate estimates from up to 100% to not greater than 20%; (2) comparing serum with broth; (3) using Columbia broth supplemented with 2μg/mL nicotinamide adenine dinucleotide (NAD) as the liquid broth; and (4) use of starting inocula of three strengths
Summary
The most widely used parameter of antimicrobial drug potency is Minimum Inhibitory Concentration (MIC). Guidelines and standards have been set by the European Union Committee on Antimicrobial Sensitivity testing (EUCAST) and the Clinical Laboratory Standards Institute (CLSI) Their methods require use of a specific broth for each organism (formulated to facilitate /optimise bacterial growth) and are conducted using two-fold dilutions. From a clinical use perspective, the principal objective in generating MIC data for each drug against each pathogenic organism harvested from each animal species is its application to determine dosage regimens that optimise bacterial kill and minimise opportunities for the emergence of resistance For this purpose the following considerations are paramount: EUCAST and CLSI standardised MIC data are generated in broths, such as Cation Adjusted Mueller Hinton Broth (CAMHB), each formulated to facilitate growth in vitro, the composition differing (for some constituents markedly) from biological fluids in general and from the biophase in particular, in respect of chemical, immunological and cellular constituents, so that they might not be representative of bacterial growth conditions in vivo; EUCAST and CLSI standardised MICs are determined using two-fold dilutions and are subject to good precision but potentially high inaccuracy - for an isolate MIC of say 4 μg/mL, the lower MIC is 2 μg/mL and the “true” MIC might be 2.01
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