Although opportunistic pathogens, coagulase-negative staphylococci (CoNS), including Staphylococcus epidermidis and Staphylococcus haemolyticus, have long been regarded as avirulent organisms. The role of toxins in the development of infections caused by CoNS is still controversial. The objective of this study was to characterize the presence of enterotoxin and cytotoxin genes in S. epidermidis and S. haemolyticus isolates obtained from blood cultures. Cytotoxin genes were detected by PCR using novel species-specific primers. Among the 85 S. epidermidis and 84 S. haemolyticus isolates, 95.3% and 79.8%, respectively, carried at least one enterotoxin gene. The most frequent enterotoxin genes were sea (53.3%), seg (64.5%) and sei (67.5%). The seg gene was positively associated with S. epidermidis (p = 0.02), and this species was more toxigenic than S. haemolyticus. The hla/yidD gene was detected in 92.9% of S. epidermidis and the hla gene in 91.7% of S. haemolyticus isolates; hlb was detected in 92.9% of the S. epidermidis isolates and hld in 95.3%. Nosocomial Staphylococcus epidermidis and S. haemolyticus isolates exhibited a high toxigenic potential, mainly containing the non-classical enterotoxin genes seg and sei. The previously unreported detection of hla/yidD and hlb in S. epidermidis and S. haemolyticus using species-specific primers showed that these hemolysin genes differ between CoNS species and that they are highly frequent in blood culture isolates.

This study aimed to correlate the presence of ica genes, biofilm formation and antimicrobial resistance in 107 strains of Staphylococcus epidermidis isolated from blood cultures. The isolates were analysed to determine their methicillin resistance, staphylococcal cassette chromosome mec (SCCmec) type, ica genes and biofilm formation and the vancomycin minimum inhibitory concentration (MIC) was measured for isolates and subpopulations growing on vancomycin screen agar. The mecA gene was detected in 81.3% of the S. epidermidis isolated and 48.2% carried SCCmec type III. The complete icaADBC operon was observed in 38.3% of the isolates; of these, 58.5% produced a biofilm. Furthermore, 47.7% of the isolates grew on vancomycin screen agar, with an increase in the MIC in 75.9% of the isolates. Determination of the MIC of subpopulations revealed that 64.7% had an MIC ≥ 4 μg mL-1, including 15.7% with an MIC of 8 μg mL-1 and 2% with an MIC of 16 μg mL-1. The presence of the icaADBC operon, biofilm production and reduced susceptibility to vancomycin were associated with methicillin resistance. This study reveals a high level of methicillin resistance, biofilm formation and reduced susceptibility to vancomycin in subpopulations of S. epidermidis. These findings may explain the selection of multidrug-resistant isolates in hospital settings and the consequent failure of antimicrobial treatment.

The objective of the present study was to determine the antimicrobial resistance profile of planktonic and biofilm cells of Staphylococcus aureus and coagulase-negative staphylococci (CoNS). Two hundred Staphylococcus spp. strains were studied, including 50 S. aureus and 150 CoNS strains (50 S. epidermidis, 20 S. haemolyticus, 20 S. warneri, 20 S. hominis, 20 S. lugdunensis, and 20 S. saprophyticus). Biofilm formation was investigated by adherence to polystyrene plates. Positive strains were submitted to the broth microdilution method to determine the minimum inhibitory concentration (MIC) for planktonic and biofilm cells and the minimal bactericidal concentration for biofilm cells (MBCB). Forty-nine Staphylococcus spp. strains (14 S. aureus, 13 S. epidermidis, 13 S. saprophyticus, 3 S. haemolyticus, 1 S. hominis, 3 S. warneri, and 2 S. lugdunensis) were biofilm producers. These isolates were evaluated regarding their resistance profile. Determination of planktonic cell MIC identified three (21.4%) S. aureus strains that were resistant to oxacillin and six (42.8%) that were resistant to erythromycin. Among the CoNS, 31 (88.6%) strains were resistant to oxacillin, 14 (40%) to erythromycin, 18 (51.4%) to gentamicin, and 8 (22.8%) to sulfamethoxazole/trimethoprim. None of the planktonic isolates were resistant to vancomycin or linezolid. MICs were 2-, 4-, 8-, and up to 16-fold higher for biofilm cells than for planktonic cells. This observation was more common for vancomycin and erythromycin. The MBCB ranged from 8 to >256 µg/mL for oxacillin, 128 to >128 µg/mL for vancomycin, 256 to >256 µg/mL for erythromycin and gentamicin, >64 µg/mL for linezolid, and 32/608 to >32/608 µg/mL for sulfamethoxazole/trimethoprim. The results showed considerably higher MICs for S. aureus and CoNS biofilm cells compared to planktonic cells. Analysis of MBCM confirmed that even high concentrations of vancomycin were unable to eliminate the biofilms of S. aureus and CoNS species. Linezolid was the most effective drug in inhibiting staphylococci in the biofilm, without an increase in the MIC, when compared to planktonic cells. None of the isolates were resistant to this drug.

Epidemiological studies have identified Staphylococcus aureus as the most common agent involved in food poisoning. However, current research highlights the importance of toxigenic coagulase-negative staphylococci (CoNS) isolated from food. The aim of this study was to characterize Staphylococcus spp. isolated from cows with bovine subclinical mastitis regarding the presence of genes responsible for the production of staphylococcal enterotoxins and of the tst-1 gene encoding toxic shock syndrome toxin 1, and to determine the clonal profile of the isolates carrying any of the genes studied. A total of 181 strains isolated in different Brazilian states, including the South, Southeast, and Northeast regions, were analyzed. The sea gene was the most frequent, which was detected in 18.2% of the isolates, followed by seb in 7.7%, sec in 14.9%, sed in 0.5%, see in 8.2%, seg in 1.6%, seh in 25.4%, sei in 6.6%, and ser in 1.6%. The sej, ses, set, and tst-1 genes were not detected in any of the isolates. The typing of the isolates by pulsed-field gel electrophoresis revealed important S. aureus and S. epidermidis clusters in different areas and the presence of enterotoxin genes in lineages isolated from animals that belong to herds located geographically close to each other.

Bacterial biofilms play an important role in urinary tract infections (UTIs), being responsible for persistent infections that lead to recurrences and relapses. Staphylococcus saprophyticus is one of the main etiological agents of UTIs, however, little is known about biofilm production in this species and especially about its response to the antimicrobial agents used to treat UTIs when a biofilm is present. For this reason, the aim of this work was to evaluate the response of S. saprophyticus biofilms to five antimicrobial agents. Staphylococcus saprophyticus was evaluated for antimicrobial susceptibility in its planktonic form by means of minimum inhibitory concentration (MIC) and in biofilms by means of minimum inhibitory concentration in biofilm (MICB) against the following antimicrobial agents by the microdilution technique: vancomycin, oxacillin, trimethoprim/sulfamethoxazole, ciprofloxacin, and norfloxacin. Of the 169 S. saprophyticus studied, 119 produced a biofilm as demonstrated by the polystyrene plate adherence method. Biofilm cells of S. saprophyticus exhibited a considerable increase in MICB when compared to the planktonic forms, with an increase of more than 32 times in the MICB of some drugs. Some isolates switched from the category of susceptible in the planktonic condition to resistant in the biofilm state. Statistical analysis of the results showed a significant increase in MICB (p < 0.0001) for all five drugs tested in the biofilm state compared to the planktonic form. Regarding determination of the minimum bactericidal concentration in biofilm (MBCB), there were isolates for which the minimum bactericidal concentration of all drugs was equal to or higher than the highest concentration tested.

Infections with coagulase-negative staphylococci are often related to biofilm formation. This study aimed to detect biofilm formation and biofilm-associated genes in blood culture isolates of Staphylococcus epidermidis and S. haemolyticus. Half (50.6%) of the 85 S. epidermidis isolates carried the icaAD genes and 15.3% the bhp gene, while these numbers were 42.9% and 0 for S. haemolyticus, respectively. According to the plate test, 30 S. epidermidis isolates were biofilm producers and 40% of them were strongly adherent, while only one (6%) of the 17 S. haemolyticus biofilm-producing isolates exhibited a strongly adherent biofilm. The concomitant presence of icaA and icaD was significantly associated with the plate and tube test results (P ≤ 0.0004). The higher frequency of icaA in S. epidermidis and of icaD in S. haemolyticus is correlated with the higher biofilm-producing capacity of the former since, in contrast to IcaD, IcaA activity is sufficient to produce small amounts of polysaccharide. Although this study emphasizes the importance of icaAD and bhp for biofilm formation in S. epidermidis, other mechanisms seem to be involved in S. haemolyticus.

The use of antimicrobial agents has led to the emergence of resistant bacterial strains over a relatively short period. Furthermore, Staphylococcus spp. can produce β-lactamase, which explains the survival of these strains in a focus of infection despite the use of a β-lactam antibiotic. The aim of this study was to evaluate the resistance of Staphylococcus spp. isolated from bovine subclinical mastitis to oxacillin and vancomycin (by minimum inhibitory concentration) and to detect vancomycin heteroresistance by a screening method. We also evaluated β-lactamase production and resistance due to hyperproduction of this enzyme and investigated the mecA and mecC genes and performed staphylococcal cassette chromosome mec typing. For this purpose, 181 Staphylococcus spp. isolated from mastitis subclinical bovine were analyzed. Using the phenotypic method, 33 (18.2%) of Staphylococcus spp. were resistant to oxacillin. In contrast, all isolates were susceptible to vancomycin, and heteroresistance was detected by the screening method in 13 isolates. Production of β-lactamase was observed in 174 (96%) of the Staphylococcus spp. isolates. The mecA gene was detected in 8 isolates, all of them belonging to the species Staphylococcus epidermidis, and staphylococcal cassette chromosome mec typing revealed the presence of type I and type IV isolates.

The aim of this study was to evaluate the antimicrobial susceptibility profile of 85 Staphylococcus epidermidis and 84 Staphylococcus haemolyticus strains isolated from blood cultures to oxacillin, vancomycin, tigecycline, linezolid, daptomycin, and quinupristin/dalfopristin over a period of 12 years. S. epidermidis and S. haemolyticus isolated from blood cultures of inpatients, attended at a teaching hospital, were analyzed for the presence of the mecA gene and by SCCmec typing. The minimum inhibitory concentration (MIC) values of tigecycline, linezolid, daptomycin, quinupristin/dalfopristin, and vancomycin were determined. Isolates exhibiting vancomycin MICs of ≥2 μg/ml were typed by pulsed-field gel electrophoresis (PFGE). The rate of mecA positivity was 92.9% and 100% in S. epidermidis and S. haemolyticus, respectively. The most frequent SCCmec types were type III (53.2%) in S. epidermidis and type I (32.1%) in S. haemolyticus. All isolates were susceptible to linezolid and daptomycin, but 7.1% of S. haemolyticus and 2.3% of S. epidermidis isolates were resistant to tigecycline, and 1.2% each of S. haemolyticus and S. epidermidis were resistant and intermediately resistant to quinupristin/dalfopristin, respectively. S. epidermidis exhibited higher vancomycin MICs (40% with MIC of ≥2 μg/ml). Clonal typing of strains with vancomycin MIC of ≥2 μg/ml revealed the presence of different PFGE types of S. epidermidis and S. haemolyticus over a period of up to 4 years (2002-2004, 2005-2008, 2006-2009, 2010-2011). Despite the observation of a high prevalence of mecA, the clinical strains were fully susceptible to vancomycin and to the new drugs linezolid, daptomycin, tigecycline, and quinupristin/dalfopristin. The PFGE types with vancomycin MIC of ≥2 μg/ml exhibited a great diversity of SCCmec cassettes, demonstrating that S. epidermidis and S. haemolyticus may easily acquire these resistance-conferring genetic elements.