Abstract
In animal production, probiotics seek to replace the use of antibiotics, while diminishing mortality and morbidity rates to raise productivity. Probiotics constitute a natural alternative that, in contrast with antibiotics, neither produces pathogen resistance, nor leaves chemical residues in the final product. Several bacteria, including some belonging to the genus Lactobacillus have been described as probiotics with high potential. A non-invasive bioprospecting protocol aimed for the isolation and characterization of lactobacilli from chicken feces was established. Fecal samples were collected from the ground. These were diluted and cultured in LAB selective medium. Colonies were identified by three methods: Gram stain, MALDI-TOF MS and sequencing of 16S rRNA gene. An initial probiotic potential of lactobacilli isolates was determined via antagonism tests using five enteropathogen reference strains: Staphylococcus aureus, Enterococcus faecium, Candida albicans, Pseudomonas spp. and Salmonella spp. 24 isolates belonging to four Lactobacillus species were identified by MALDITOF MS. BLAST of 16S rRNA gene of eight randomly selected isolates, confirmed MALDI-TOF MS identification. Five of these eight isolates inhibited the growth of at least one of the pathogenic strains used, three isolates of Lactobacillus plantarum and two of Lactobacillus salivarius. Our protocol achieved 21 lactobacilli per 100 isolates performance, greatly surpassing the normal percentage of lactobacilli in chicken gut microbiome, that so, its implementation would facilitate the isolation and identification of new probiotic strains from feces.
Highlights
Antibiotics are widely used in the livestock industry for treatment of infections, disease prevention, and as growth promoters in subtherapeutic dosage
The 31 isolates belonged to the order Lactobacillales, distributed in three families (Enterococcaceae, Streptococcaceae, and Lactobacillaceae), four genera (Enterococcus, Pediococcus, Streptococcus, and Lactobacillus), and in eight species of bacteria (Figure 1). 24 isolates belonged to the genus Lactobacillus were distributed in four species (Figure 1), which accounts for 77.4% of the 31 isolates identified by mass spectrometry and 21% of the original 114 colonies
This suggests an effective selection by the MRS medium, and by morphological selection using Gram stain, given the fact that the majority of isolates belonged to the genus Lactobacillus, and all of them are grouped under the order Lactobacillales and the family Lactobacillaceae
Summary
Antibiotics are widely used in the livestock industry for treatment of infections, disease prevention (prophylaxis), and as growth promoters in subtherapeutic dosage Their use began during the fifties in response to the rapidly growing demand of meat worldwide [1]. Results proved that after antimicrobial use in livestock ceased, the prevalence of antibiotic-resistant pathogens was significantly reduced, with little to no changes in productivity and animal health [6],[2]. In this context, there has been a fast emergence of alternatives to antibiotics in livestock production within the last three decades [1]. One promising alternative consists in the use of probiotics as prevention tools from infectious diseases, and as growth promoters
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