Plant-derived Compounds, Trans-cinnamaldehyde and Eugenol, Reduce Adhesion and Invasion of Staphylococcus aureus in Bovine Mammary Epithelial Cells in vitro
Staphylococcus aureus is one of the major bacterial pathogens involved in ruminant mastitis globally. The disease has major impacts on animal health and milk quality leading to severe economic losses to the dairy industry. S. aureus adhesion to and invasion of mammary epithelial cells (MEC) is critical for the establishment of the disease. We hypothesized that two plant-derived antimicrobials, trans-cinnamaldehyde (TC) and eugenol (EG), would reduce S. aureus adhesion to and invasion of primary bovine MEC in vitro. Mammary epithelial cells were isolated from mammary glands of lactating cows post-slaughter and were characterized using multi-cytokeratin immunostaining. Confluent MEC monolayers were inoculated with four S. aureus strains (Strain ST 35, Thorn 17, 15, and M9175) at mid log phase separately (multiplicity of infection – 100:1) either in the presence or absence (control) of sub-inhibitory concentrations (SICs: greatest concentration that did not inhibit bacterial growth) of TC (0.006%) and EG (0.03%), and incubated for 2 h for bacterial adhesion. Infected cells were treated with gentamicin (100 µg/mL) for 1 h at 37°C for enumerating invaded bacteria. All experiments included duplicate samples and were repeated three times. Data were analyzed using PROC ANOVA and significance determined at P < 0.05. For strain Thorn 17, pre-exposure of S. aureus (5 h) and MEC (12 h) to EG reduced S. aureus adhesion to and invasion of MEC by ~1.6 ± 0.01 log10 CFU/mL (control = 6.67 ± 0.01 log10 CFU/mL; P < 0.0001) and 2.8 ± 0.11 log10 CFU/mL (control = 3.9 ± 0.02 log10 CFU/mL; P < 0.0001), respectively. Pre-exposure of S. aureus and MEC to TC reduced S. aureus adhesion to and invasion of MEC by ~2.2 ± 0.02 log10 CFU/mL (control = 6.7 ± 0.02 log10 CFU/mL; P < 0.0001) and 2.85 ± 0.08 log10 CFU/mL (control = 3.94 ± 0.01 log10 CFU/mL P<0.0001), respectively. Similar results were observed with strains ST 35, M9175 and Thorn 15. In conclusion, SICs of TC and EG reduced S. aureus adhesion to and invasion of MEC. In vivo studies using a mammalian model to validate these results are warranted.
Mammary epithelial cells, Plant compound, S.aureus