In vitro efficacy of several antimicrobial combinations against Brucella canis and Brucella melitensis strains isolated from dogs

Abstract

In vitro efficacy of tetracyclines, aminoglycosides, fluoroquinolones, rifampin and macrolides and their combinations against four Brucella canis and two B. melitensis strains isolated from dogs was evaluated by means of inhibition tests: minimal inhibitory concentration (MIC) and fractional inhibitory concentrations (FIC) and by bactericidal tests: minimal bactericidal concentrations (MBC) and kill-time experiments. In inhibition studies, tetracyclines showed the lowest MICs of all compounds tested (0.06–0.50 mg/l). However, doxycycline did not inhibit growth of B. canis D-519 strain at concentrations lesser than 1 mg/l. Most strains were resistant to macrolides. Chequerboard MIC titrations showed synergy between enrofloxacin and aminoglycosides against B. canis RM 6 66 reference strain and between doxycycline and streptomycin against B. canis D-519 strain. Combinations of tetracyclines and aminoglycosides were at the limit of synergy for B. canis RM 6 66 strain (FIC = 0.56). Combination of doxycycline and rifampin seemed to be antagonistic against B. canis M- strain (FIC = 3). Other combinations studied were indifferent or additive against all other strains. In bactericidal studies, aminoglycosides, fluoroquinolones and rifampin showed bactericidal activity at 1–4 × MIC but tetracyclines or macrolides did not kill brucellae below 8 × MIC. Results of kill-time experiments indicated that streptomycin was the antimicrobial agent tested which more rapid killed brucellae. In most cases, doxycycline did not kill brucellae in 96 h of incubation. When using rifampin alone, some inocula of B. canis strains RM 6 66 , D-519 and M- showed regrowth after 24h of incubation. Retesting of these regrowing brucellae showed an increased MIC indicating development of phenotypic resistance. Testing of antimicrobial combinations in kill-time experiments indicated synergy between enrofloxacin and streptomycin and antagonism between rifampin and doxycycline both for B. canis RM 6 66 strain.