Abstract
For clinical isolates of bovine Mannheimia haemolytica and Pasteurella multocida, this study reports minimum inhibitory concentration (MIC) differences for tetracycline, oxytetracycline and doxycycline between cation-adjusted Mueller-Hinton broth (CAMHB), foetal bovine serum (FBS) and Roswell Park Memorial Institute (RPMI) medium. MICs were determined according to CLSI standards and additionally using five overlapping sets of twofold dilutions. Matrix effect: (a) free drug MICs and minimum bactericidal concentrations (MBC) for all drugs were significantly higher in FBS than in CAMHB for both pathogens (p<0.001); (b) MICs and MBCs were higher for CAMHB and FBS compared to RPMI for P.multocida only. Net growth rate for P.multocida in CAMHB was significantly slower than in FBS and higher than in RPMI, correlating to MIC and MBC ranking. Drug effect: doxycycline MICs and MBCs were significantly lower (p<0.001) in both CAMHB and FBS than tetracycline and oxytetracycline for both pathogens. Only for M.haemolytica were oxytetracycline MIC and MBC significantly lower than tetracycline, precluding the use of tetracycline to predict oxytetracycline susceptibility in this species. Determining potencies of tetracyclines in a physiological medium, such as FBS, is proposed, when the objective is correlation with pharmacokinetic data for dosage determination.
Highlights
The bovine pathogens Mannheimia haemolytica and Pasteurella multocida have been linked to cases of bovine calf pneumonia (Davies et al, 2004; Griffin et al, 2010; Welsh et al, 2004)
P. multocida) minimum inhibitory concentration (MIC) in foetal bovine serum (FBS) were significantly higher with ratios (FBS:cation-adjusted Mueller-Hinton broth (CAMHB)) of 6.7:1, 7.0:1 and 1.3:1 for tetracycline, oxytetracycline and doxycycline, respectively
MICs in Roswell Park Memorial Institute (RPMI) were significantly lower than those determined in both FBS
Summary
The bovine pathogens Mannheimia haemolytica and Pasteurella multocida have been linked to cases of bovine calf pneumonia (Davies et al, 2004; Griffin et al, 2010; Welsh et al, 2004). The high prevalence of these infections has necessitated the widespread use in veterinary medicine of tetracyclines, especially oxytetracycline and doxycycline Susceptibility to these AMDs is most commonly measured using the minimum inhibitory concentration (MIC). The physiological relevance of in vitro methods using artificial media, such as cation-adjusted Mueller-Hinton broth (CAMHB), has been questioned for some drug classes, including tetracyclines and macrolides. For these classes, MICs for most pathogens are markedly dependent on the growth medium (Brentnall et al, 2012; Buyck et al, 2012; Dorey et al, 2016; Lees et al, 2015, 2016; Toutain et al, 2017). For the same drug and similar testing conditions (inoculum size and incubation time), the differences in MIC (on a free-concentration basis) between the afore-mentioned media could be related solely to rates of bacterial growth and death in each medium (Mouton and Vinks, 2005)
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