Staphylococcus aureus is the bacterium most frequently isolated from milk of bovines with mastitis. Four allelic groups, which interfere with the regulatory activities among the different groups, have been identified in the accessory gene regulator (agr) system. The aim of this study was to ascertain the prevalence of the different agr groups in capsulated and noncapsulated S. aureus bacteria isolated from mastitic bovines in Argentina and whether a given agr group was associated with MAC-T cell invasion and in vivo persistence. Eighty-eight percent of the bovine S. aureus strains were classified in agr group I. The remainder belonged in agr groups II, III, and IV (2, 8, and 2%, respectively). By restriction fragment length polymorphism analysis after PCR amplification of the agr locus variable region, six agr restriction types were identified. All agr group I strains presented a unique allele (A/1), whereas strains from groups II, III, and IV exhibited more diversity. Bovine S. aureus strains defined as being in agr group I (capsulated or noncapsulated) showed significantly increased abilities to be internalized within MAC-T cells, compared with isolates from agr groups II, III, and IV. agr group II or IV S. aureus strains were cleared more efficiently than agr group I strains from the murine mammary gland. The results suggest that agr group I S. aureus strains are more efficiently internalized within epithelial cells and can persist in higher numbers in mammary gland tissue than S. aureus strains classified in agr group II, III, or IV.

Real-time biofilm analysis provides a promising tool to evaluate antibiotic therapy in clinical biofilm-mediated infections.

Staphylococcus aureus capsular polysaccharides (CP) have been shown to enhance staphylococcal virulence in numerous animal models of infection. Although serotype 5 CP (CP5) and CP8 predominate among S. aureus isolates from humans, most staphylococcal isolates from bovines with mastitis in Argentina are capsule negative. This study was designed to evaluate the effects of CP5 and CP8 expression on the pathogenesis of experimental murine mastitis. Lactating mice were challenged by the intramammary route with one of three isogenic S. aureus strains producing CP5, CP8, or no capsule. Significantly greater numbers of acapsular mutant cells were recovered from the infected glands 12 days after bacterial challenge compared with the encapsulated strains. Histopathological analyses revealed greater polymorphonuclear and mononuclear leukocyte infiltration and congestion in the mammary glands of mice infected with the encapsulated strains compared with the acapsular mutant, and the serotype 5 strain elicited more inflammation than the serotype 8 strain. In vitro experiments revealed that the acapsular S. aureus strain was internalized by MAC-T bovine epithelial cells in significantly greater numbers than the CP5-or CP8-producing strain. Taken together, the results suggest that S. aureus lacking a capsule was able to persist in the murine mammary gland, whereas encapsulated strains elicited more inflammation and were eliminated faster. Loss of CP5 or CP8 expression may enhance the persistence of staphylococci in the mammary glands of chronically infected hosts.

Selection pressures exerted on Staphylococcus aureus by host factors may lead to the emergence of mutants better adapted to the evolving conditions at the infection site. This study was aimed at identifying the changes that occur in S. aureus exposed to the host defense mechanisms during chronic osteomyelitis and evaluating whether these changes affect the virulence of the organism. Genome assessment of two S. aureus isolates collected 13 months apart (HU-85a and HU-85c) from a host with chronic osteomyelitis was made by whole genome sequencing. Agr functionality was assessed by qRT-PCR. Isolates were tested in a rat model of osteomyelitis and the bacterial load (CFU/tibia) and the morphometric osteomyelitic index (OI) were determined. The ability of the isolates to trigger the release of proinflammatory cytokines was determined on macrophages in culture. Persistence of S. aureus within the host resulted in an agrC frameshift mutation that likely led to the observed phenotype. The capacity to cause bone tissue damage and trigger proinflammatory cytokines by macrophages of the agr-deficient, unencapsulated derivative (HU-85c) was decreased when compared with those of the isogenic CP8-capsulated parental strain (HU-85a). By comparison, no significant differences were found in the bacterial load or the OI from rats challenged with isogenic Reynolds strains [CP5, CP8, and non-typeable (NT)], indicating that lack of CP expression alone was not likely responsible for the reduced capacity to cause tissue damage in HU-85c compared with HU-85a. The production of biofilm was significantly increased in the isogenic derivative HU-85c. Lack of agr-dependent factors makes S. aureus less virulent during chronic osteomyelitis and alteration of the agr functionality seems to permit better adaptation of S. aureus to the chronically infected host.