Antimicrobial Activity of Sorghum Phenolic Extract on Bovine Foodborne and Mastitis-Causing Pathogens.

Antimicrobial resistance in bacterial pathogens associated with bovine mastitis and human foodborne illnesses from contaminated food and water have an impact on animal and human health. Phenolic compounds have antimicrobial properties and some specialty sorghum grains are high in phenolic compounds, and the grain extract may have the potential as a natural antimicrobial alternative. The study's objective was to determine antimicrobial effects of sorghum phenolic extract on bacterial pathogens that cause bovine mastitis and human foodborne illnesses. Bacterial pathogens tested included Escherichia coli, Salmonella Typhimurium, Campylobacter jejuni, Campylobacter coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, Klebsiella oxytoca, Staphylococcus aureus, and Enterococcus faecalis. Antibacterial activities of sorghum phenolic extracts were determined by agar-well diffusion assay. Sorghum phenolic extract was added to the wells in concentrations of 0, 100, 200, 500, 1000, or 4000 µg/mL. The control wells did not receive phenolic extract. Plates were incubated for 18-24 h, and the diameter of each zone of inhibition was measured. The results indicated that sorghum phenolic extract had inhibitory effects on Staphylococcus aureus, Enterococcus faecalis, Campylobacter jejuni, and Campylobacter coli.

Leukotoxin production by Fusobacterium necrophorum strains in relation to severity of liver abscesses in cattle.

Fusobacterium necrophorum, a Gram-negative anaerobe, is the primary etiologic agent of liver abscesses of beef cattle. The bacterium, a member of the microbial community of the rumen, travels to the liver via portal circulation to cause abscesses. The severity of liver abscesses vary from mild with one or two small abscesses to severe with medium to large multiple abscesses. Leukotoxin, a secreted protein, is the critical virulence factor involved in the infection. Our objective was to compare leukotoxin production between strains of F. necrophorum isolated from mild and severe liver abscesses collected from slaughtered cattle. The quantification of leukotoxin was based on assays to measure cytotoxicity and protein antigen concentration. One-hundred strains, 50 from mild and 50 from severe abscesses, were utilized in the study. Cell-free supernatants were prepared from cultures grown in anaerobic broth at 9 and 24 h incubations. The leukotoxic activity was quantified by measuring cytotoxicity based on the release of lactic dehydrogenase from bovine lymphocyte cells, BL3, treated with the culture supernatant. Leukotoxin protein concentration was quantified by a sandwich ELISA assay with a leukotoxin-specific monoclonal antibody as the capture antibody. The leukotoxin activity and concentration were highly variable among the strains within each severity of liver abscesses. Although the leukotoxic activity was unaffected by incubation time, leukotoxin protein concentration was consistently higher at 24 h compared to 9 h incubation. Strains from severe liver abscesses had significantly higher leukotoxic activity and higher protein concentration compared to strains from mild liver abscesses (P < 0.0001) at both 9 and 24 h culture supernatants. Across all strains, the correlation coefficients between leukotoxic activity and leukotoxin concentration at 9 and 24 h were 0.14 (P = 0.17) and 0.47 (P < 0.0001), respectively. In conclusion, strains isolated from severe liver abscesses had significantly higher leukotoxic activities and leukotoxin protein concentrations compared to strains isolated from mild liver abscesses.

Leukotoxic activity of Fusobacterium necrophorum of cattle origin.

Fusobacterium necrophorum is a Gram negative, rod-shaped and aero tolerant anaerobe. In animals, it is an opportunistic pathogen frequently associated with necrotic infections, generally called necrobacillosis, such as calf diphtheria, foot rot and liver abscesses in cattle. Two subspecies exist: subsp. necrophorum and subsp. funduliforme. Among several virulence factors, leukotoxin (Lkt) is considered to be a major factor and a protective antigen. The objective of the study was to utilize BL3 cells and measure the release of lactic dehydrogenase to quantify Lkt activity of F. necrophorum. The assay was used to examine the effects of storage and handling conditions, growth media, polymyxin B addition on the cytotoxicity and evaluate Lkt activities of F. necrophorum strains isolated from bovine liver abscesses and foot rot. The Lkt activity peaked at 9 h of incubation. There was a significant decrease in the cytotoxicity measured in the samples after each freeze and thaw cycle. No difference was observed in the cytotoxicity for the samples handled aerobically versus anaerobically. Lkt activities of strains grown in anaerobic Brain-Heart Infusion broth were higher compared to Vegitone broth. A small reduction in the cytotoxicity activity was observed after the addition of polymyxin. The Lkt activity was consistently higher in strains of subsp. necrophorum than subsp. funduliforme of liver abscess origin. Among the strains isolated from cattle foot rot, Lkt activities of subsp. necrophorum strains appear to be much more variable. Use of BL3 cells in combination of lactic acid dehydrogenase assay appears to be a simple and valid assay to measure Lkt activity of F. necrophorum.

Total Synthesis of Reniochalistatin E.

Reniochalistatin E (1) is one of the five related cyclic peptides isolated from the marine sponge Reniochalina stalagmitis. The discovery of these compounds resulted from a screening program directed toward the identification of proline-rich bioactive compounds. Reniochalistatin E is the only member of the family to possess a tryptophan amino acid residue. Given the cytotoxicity observed for 1, efforts were directed toward developing a synthetic route to 1. The first total synthesis of 1 has been accomplished in a 15-step route in an overall 5.0% yield. The synthetic sample of reniochalistatin E was shown to have similar activity toward HeLa and RPMI-8226 cell lines compared to the natural sample, with IC50 values of 16.9 vs 17.3 µM and 4.5 vs 4.9 µM, respectively. Interestingly, both of the fully deprotected octapeptides constructed toward the synthesis of reniochalistatin E were shown to have cytotoxicity. The route provides a means to probe the structure-activity relationship of 1 and further biological investigations.