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Antimicrobial Resistance in Direct-Fed Microbials used in Cattle
The use of antimicrobials in animal feed has come under increasing scrutiny from the public and regulatory agencies. Direct-fed microbials (DFM) or probiotics are considered valuable alternatives to antimicrobials in food animal nutrition. They are live organisms, when given orally in the feed, have proven beneficial in improving the gut microbial balance. Studies in Europe have reported antimicrobial resistance (AMR) in DFM organisms. This is of serious concern because of their potential for transferring resistance to pathogenic bacteria in the gut. The aim of the present study was to characterize phenotypic and genotypic AMR profiles of bacterial strains isolated from 9 commercially available DFM used in cattle to 20 different antimicrobials. Two methods, disc diffusion and broth micro-dilution based assay (MIC) were performed to determine phenotypic AMR. Based on our disc diffusion and MIC assays, Enterococcus faecium isolates showed resistance towards metronidazole (n=8) with an MIC of > 64 µg/mL, erythromycin (n=3) with an MIC of ≥ 8 µg/mL, rifampin (n=5) with an MIC of > 4 µg/mL, and clindamycin (n=4) with an MIC of > 4 µg/mL. A Propionibacterium freudenreichii isolate showed resistance towards kanamycin with an MIC of > 64 µg/mL. The same strain also had an MIC of 21.33 µg/mL for levofloxacin. A Lactobacillus plantarum isolate showed no inhibition zone for vancomycin, ciprofloxacin and metronidazole. An isolate of Bacillus subtilis showed resistance towards clindamycin with an MIC of > 4 µg/mL, erythromycin with an MIC of > 8µg/mL, and chloramphenicol with an MIC of > 8 µg/mL. Spotted DNA microarray was used to detect the diversity of resistance genes in the DFM. Microarray analysis revealed the following resistance genes in E. faecium strains isolated from 3 different DFMs: Strain 1 carried an aminoglycoside resistance gene [ant(4’)-Ia)], erythromycin resistance gene [ere(A2)] and three tetracycline resistance genes (tet39, tet31, tetK); Strain 2 carried a beta-lactam resistance gene (pbp5) and two tetracycline resistance genes (tet31, tet39); Strain 3 had an erythromycin resistance gene (ermB) and a tetracycline resistance gene (tetC). These studies show that AMR is prevalent among probiotic bacteria used in cattle industry in the U.S., and justifies further characterization and detection of such resistance.
Keywords: Antimicrobial Resistance, Direct-Fed Microbials, Cattle