Staphylococcus aureus remains one of the most common causative agents of bovine mastitis because of its pathogenicity, contagiousness, capability to persist in the mammary gland, colonization of skin and poor cure rates when causing intramammary infections with the currently available therapies. On the other hand, non-aureus staphylococci (NAS) originating from bovine milk and teat apices have beneficial traits, potentially including the production of metabolites to prevent or target S. aureus infections. Therefore, the main focus of this thesis was to explore some elements of the regulation of S. aureus growth and virulence by bovine NAS species, including S. chromogenes, S. epidermidis and S. simulans, originating from both milk and teat apices.
A few recent studies have suggested that interactions between S. aureus and other colonizing staphylococci via the accessory gene regulator (agr) quorum-sensing (QS) system might affect the ability of S. aureus to produce virulence factors such as biofilm and cause infection. In co-culture with S. aureus the 3 NAS species in general downregulated the expression of rnaIII, the effector molecule of the QS system, yet this effect was more pronounced in S. chromogenes and S. simulans isolates than in S. epidermidis isolates. In vitro growth inhibition of S. aureus by NAS resulted in a small underestimation of the downregulating effect of NAS on rnaIII expression of S. aureus. Additionally, the culture supernatant of the NAS isolates and supernatant treated with proteinase K expressed greater regulatory activity over S. aureus virulence genes rnaIII, hla, and spa than washed NAS cells suspended in sterile water.
The capacity of an agr-positive S. aureus strain to form biofilm was increased more in the presence of S. chromogenes than in the presence of S. simulans and S. epidermidis isolates and in the presence of NAS isolates that did not harbor biofilm related genes. Conversely, biofilm dispersion of this S. aureus strain was suppressed by NAS as a group, an effect that was more pronounced by isolates from teat apices. In general, the effects were not associated with the capacity of NAS to repress the agr system.
A quantitative method was developed to determine the ability of NAS to inhibit the growth of S. aureus, Escherichia coli and Streptococcus uberis and the results were compared with those from a semiquantitative method. Overall, in vitro growth inhibition of the major mastitis pathogens, including E. coli, was confirmed for all NAS, an effect that varied highly among NAS isolates, a finding that was not evident from the semiquantitative method. As well, S. simulans, NAS originating from teat apices and NAS with a strong or moderate capacity to repress the agr system required lower concentrations to inhibit S. aureus growth (including methicillin-resistant S. aureus isolates). The inhibitory effects of teat apex isolates was more pronounced in S. epidermidis than in S. simulans and S. chromogenes.
Our findings indicate that bovine-associated NAS are able to regulate the growth and virulence of S. aureus in vitro. Isolation and identification of NAS metabolites that affect these microbial interactions might help to develop alternative strategies for treatment and control of S. aureus mastitis in the future.
Dr. Bruno Toledo Silva studied Veterinary Medicine at the State University of North Paraná in Brazil and later he obtained his postgraduate Master in Veterinary Clinical Sciences at the University of São Paulo. He recently received his PhD from the Faculty of Veterinary Medicine at Ghent University. Bruno is currently working as a postdoctoral researcher at the “Mastitis and Milk Quality Research Unit” of the same university.