Safety and Growth Optimization of Lactic Acid Bacteria Isolated From Feedlot Cattle for Probiotic Formula Design

Abstract

In order to eliminate the widespread use of antibiotics in livestock production, the research for alternatives has increased lately. This study examined the safety of 40 lactic acid bacteria (LAB) isolated from bovine feedlot environment and previously selected as potential probiotics. A high sensitivity prevalence to ampicillin (AMP, 100%), gentamicin (GEN, 96.3%), kanamycin (KAN, 96.3%), clindamycin (CLI, 85.2%), chloramphenicol (CHL, 92.6%) and streptomycin (STR, 88.9%) while moderate and high resistance against erythromycin (ERY, 48%) and tetracycline (TET, 79%) respectively, were determined. Feedlot enterococci and pediococci displayed high resistance to CLI, ERY, GEN and TET (73, 100, 54.5, and 73%, respectively). Among fifteen resistance genes investigated, seven were identified in lactobacilli; their presence not always was correlated with phenotypic resistance. STR resistance genes, <i>aad</i>A and <i>ant(6)</i> were observed in 7.4 and 3.7% of isolates, respectively; genes responsible for aminoglycosides resistance, such as <i>bla</i> (7.4%), and <i>aph(3")-III</i> (3.7%) were also recognized. In addition, resistance <i>cat</i> and <i>tetS</i> genes (3.7 and 7.4%, respectively) were harbored by feedlot lactobacilli strains. The presence of <i>erm</i>B gene in 22.3% of isolates, including two of the six strains phenotypically resistant to ERY, exhibited the highest prevalence among the assessed antibiotics. None of the feedlot lactobacilli harbored virulence factors genes, while positive PCR amplification for <i>ace</i>, <i>agg</i>, <i>fsrA</i>, and <i>atpA</i> genes was found for enterococci. With the objective of producing large cell biomass for probiotic delivery, growth media without peptone but containing glucose and skim milk powder (Mgl and Mlac) were selected as optimal. <i>Lactobacillus acidophilus</i> CRL2074, <i>L. amylovorus</i> CRL2115, <i>L. muc</i>o<i>sae</i> CRL2069, and <i>L. rhamnosus</i> CRL2084 were strains selected as free of antibiotic resistance and virulence determinants, able to reach high cell numbers in non-expensive culture media and being compatible among them.