A total of 44 lactic acid bacteria (LAB) strains originally isolated from cattle feces and different food sources were screened for their potential probiotic features. The antimicrobial activity of all isolates was tested by well-diffusion assay and competitive exclusion on broth against Salmonella Montevideo, Escherichia coli O157:H7 and Listeria monocytogenes strain N1-002. Thirty-eight LAB strains showed antagonistic effect against at least one of the pathogens tested in this study. Improved inhibitory effect was observed against L. monocytogenes with zones of inhibition up to 24 mm when LAB overnight cultures were used, and up to 21 mm when cell-free filtrates were used. For E. coli O157:H7 and Salmonella maximum inhibitions of 12 and 11.5 mm were observed, respectively. On broth, 43 strains reduced L. monocytogenes up to 9.06 log 10 CFU/ml, 41 reduced E. coli O157:H7 up to 0.84 log 10 CFU/ml, and 32 reduced Salmonella up to 0.94 log 10 CFU/ml 24 h after co-inoculation. Twenty-eight LAB isolates that exhibited the highest inhibitory effect among pathogens were further analyzed to determine their antimicrobial resistance profile, adhesion potential, and cytotoxicity to Caco-2 cells. All LAB strains tested were susceptible to ampicillin, linezolid, and penicillin. Twenty-six were able to adhere to Caco-2 cells, five were classified as highly adhesive with > 40 bacterial cells/Caco-2 cells. Low cytotoxicity percentages were observed for the candidate LAB strains with values ranging from -5 to 8%. Genotypic identification by whole genome sequencing confirmed all as members of the LAB group; Enterococcus was the genus most frequently isolated with 21 isolates, followed by Pediococcus with 4, and Lactobacillus with 3. In this study, a systematic approach was used for the improved identification of novel LAB strains able to exert antagonistic effect against important foodborne pathogens. Our findings suggest that the selected panel of LAB probiotic strains can be used as biocontrol cultures to inhibit and/or reduce the growth of L. monocytogenes, Salmonella , and E. coli O157:H7 in different matrices, and environments.

Nontyphoid Salmonella strains are some of the leading causes of foodborne illnesses worldwide; however, there is very limited information on the presence and characteristics of Salmonella in the food production chain in developing countries. In this study, pulsed-field gel electrophoresis (PFGE) was used for molecular subtyping and for monitoring the ecology and transmission of Salmonella isolates in a slaughter facility in Mexico in an attempt to determine specific steps that need to be improved to reduce Salmonella contamination in beef carcasses. A total of 94 isolates from a Salmonella stock culture collection originally obtained from a single vertically integrated feedlot and beef abattoir in Mexico were analyzed. A total of 26 unique PFGE patterns were identified, 38.5% of them corresponding to a single serotype. High concordance (88.4%) was found between serotype and PFGE banding subtype. Salmonella Kentucky and Salmonella Give were the most clonal subtypes in this study, and Salmonella Muenster was the most diverse, with 11 banding patterns identified. A total of 73.7, 70.6, and 85.7% of the PFGE subtypes identified from preevisceration, precooler, and chilled carcasses, respectively, were identified only at those specific points and not at any previous or subsequent steps of the slaughter process, suggesting that each step is in itself a source of Salmonella contamination. Salmonella Mbandaka was more likely to be recovered from feces than from any of the steps of the slaughter process. The genetic diversity and distribution of PFGE subtypes in the processing facility highlight the need to implement antimicrobial interventions and improve sanitation procedures at various points to avoid further Salmonella dissemination into the meat supply.

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Focal Ulcerative Dermatitis (FUDS) is an emerging dermatological disease that affects cage-free laying flocks, it is characterized by the development of a lesion on the dorsum of the birds; FUDS is sporadic in nature and can result in a drop in egg production and up to 50% of cumulative mortality. A total of two cage-free flocks (flock 1: no history of FUDS; flock 2: birds affected with FUDS) from a commercial laying hen operation in the mid-west U.S. were sampled in this study. The microbial composition of skin, cloacal, cecal, and ileal samples from each bird was characterized through next generation sequencing (NGS). Results identified Staphylococcus aureus and Staphylococcus agnetis as the potential causative agents of FUDS, being the most predominant in FUDS positive birds. These results were confirmed by plating, with both staphylococci as the only pathogens isolated from lesions of FUDS positive birds. A total of 68 confirmed Staphylococcus isolates from skin and environmental samples were further analyzed by whole genome sequencing (WGS) for the presence of antimicrobial resistance (AMR) genes and virulence factors that could have contributed to the development of FUDS. Forty-four-point one-two percent of the isolates had between one and four acquired AMR genes encoding for macrolides, lincosamides, spectrogramines, and beta-lactams resistance. Six classes of virulence factors associated with adherence, enzyme, immune evasion, secretion system, toxin, and iron uptake were identified. The antimicrobial effect of 4 proprietary Bacillus Direct Fed Microbial (DFM) combinations was evaluated against the Staphylococcus aureus and Staphylococcus agnetis isolates, by agar well-diffusion (AWD) assay and competitive exclusion (CE) on broth culture. Through this antimicrobial screening, a particular two-strain combination of Bacillus pumilus was identified as the most effective inhibitor of both staphylococci. A customized Bacillus pumilus product is being used at different farms with history of FUDS resulting in the successful inhibition of both Staphylococcus aureus and Staphylococcus agnetis, decreasing FUDS mortalities, and improving harvestable eggs.

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