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PubMed2000

Correlation between the resistance genotype determined by multiplex PCR assays and the antibiotic susceptibility patterns of Staphylococcus aureus and Staphylococcus epidermidis.

Clinical isolates of Staphylococcus aureus (a total of 206) and S. epidermidis (a total of 188) from various countries were tested with multiplex PCR assays to detect clinically relevant antibiotic resistance genes associated with staphylococci. The targeted genes are implicated in resistance to oxacillin (mecA), gentamicin ¿aac(6')-aph(2"), and erythromycin (ermA, ermB, ermC, and msrA). We found a nearly perfect correlation between genotypic and phenotypic analysis for most of these 394 strains, showing the following correlations: 98% for oxacillin resistance, 100% for gentamicin resistance, and 98.5% for erythromycin resistance. The discrepant results were (i) eight strains found to be positive by PCR for mecA or ermC but susceptible to the corresponding antibiotic based on disk diffusion and (ii) six strains of S. aureus found to be negative by PCR for mecA or for the four erythromycin resistance genes targeted but resistant to the corresponding antibiotic. In order to demonstrate in vitro that the eight susceptible strains harboring the resistance gene may become resistant, we subcultured the susceptible strains on media with increasing gradients of the antibiotic. We were able to select cells demonstrating a resistant phenotype for all of these eight strains carrying the resistance gene based on disk diffusion and MIC determinations. The four oxacillin-resistant strains negative for mecA were PCR positive for blaZ and had the phenotype of beta-lactamase hyperproducers, which could explain their borderline oxacillin resistance phenotype. The erythromycin resistance for the two strains found to be negative by PCR is probably associated with a novel mechanism. This study reiterates the usefulness of DNA-based assays for the detection of antibiotic resistance genes associated with staphylococcal infections.

Anti-Bacterial Agents↗
PubMed1986

Evaluation of laboratory tests for detection of methicillin-resistant Staphylococcus aureus and Staphylococcus epidermidis.

Few studies evaluating susceptibility testing of methicillin-resistant staphylococci have included isolates of Staphylococcus epidermidis, a known pathogen in many types of serious infections. We tested 175 S. epidermidis and 95 Staphylococcus aureus isolates to determine the most sensitive procedures for detecting methicillin-resistant staphylococci. Reference procedures included agar dilution with methicillin and 4% NaCl in the agar and broth microdilution with methicillin and 2% NaCl in cation-supplemented Mueller-Hinton broth. After 24 h of incubation, the results from both methods correlated well and were within 1 log2 dilution for all isolates tested. Only one-half of all resistant isolates (92 of 183) were detected at 18 h by using the standard disk diffusion technique with 5-micrograms methicillin disks, and even fewer were detected with 10-micrograms methicillin disks and newly recommended zone-size criteria. However, the standard disk diffusion method with 4% NaCl in the agar increased the sensitivity and specificity for identification of the proper phenotype to greater than 92%. The spread plate and new spot techniques, both using agar with 4% NaCl, were also sensitive methods. Of 47 S. epidermidis isolates tested against oxacillin, 6 (13%) were oxacillin susceptible but methicillin resistant. Two automated systems, the Automicrobic system (Vitek Systems) and MicroScan (American MicroScan), as well as two broth screening systems available from Remel and Austin Biological Laboratories, failed to detect several resistant isolates, depending on the species.

Culture Media↗
PubMed2006

Antimicrobial susceptibility of Staphylococcus aureus and Staphylococcus epidermidis biofilms isolated from infected total hip arthroplasty cases.

BACKGROUND: It is difficult to treat implant-related infections because bacteria form biofilms on implants. Biofilms protect the organisms from the host's immune system and prevent penetration of antimicrobial agents. We investigated the susceptibility of six strains isolated from infected total hip arthroplasty (THA) cases to six antimicrobial agents. METHODS: Six Staphylococcus aureus and S. epidermidis strains isolated from infected THA cases and S. aureus ACTT 25923, positive strains of biofilms, were used in this study. Biofilms were developed on 96-well titer plates. After 7 days the nutrient solution was aspirated, and wells were rinsed three times with sterile water. A series of doubling dilutions of each antimicrobial agent (16, 32, 64, 128, 256, 512, and 1024 microg/ml) in trypticase soy broth were added to the wells. The plates were incubated at 37 degrees C. After 24 h, the contents were aspirated and wells were rinsed three times with sterile water. The surfaces of wells were vigorously scraped with a sterile swab and applied to sheep blood agar plates at 37 degrees C in an aerobic chamber for 24 h. The minimum bactericidal concentrations (MBCs) were determined as the lowest concentration of antibiotic resulting in 100% reduction of viable cells compared to the antibiotic-free control. Biofilms were produced on stainless steel washers, and the time-course changes were observed by scanning electron microscopy (SEM). RESULTS: Planktonic bacteria were sensitive to the antimicrobial agents, but biofilm bacteria were markedly resistant. SEM revealed that biofilms grew larger with time. CONCLUSIONS: The MBCs of antimicrobial agents for biofilms bacteria were higher than those of planktonic bacteria. Therefore, it is suggested that implant-related infection is difficult to treat with antimicrobial agents alone.

Adult↗
PubMed2001

In vitro synergy between cefepime and vancomycin against methicillin-susceptible and -resistant Staphylococcus aureus and Staphylococcus epidermidis.

The in vitro activity of cefepime combined with vancomycin was assessed by the chequerboard method against 35 clinical isolates of methicillin-susceptible (MSSA, n = 8) or -resistant (MRSA, n = 10) Staphylococcus aureus and methicillin-susceptible (MSSE, n = 9) or -resistant (MRSE, n = 8) Staphylococcus epidermidis and S. aureus ATCC 25923 (MSSA). The combination was synergic against 16 isolates and additive/indifferent against 20. For 10 of the clinical isolates (two MSSA, three MRSA, two MSSE, three MRSE) and the reference strain, the interaction of cefepime and vancomycin was also determined by the time-kill method. Except for one MRSA isolate, synergic killing was demonstrated with clinically achievable concentrations of vancomycin (0.5-1 mg/L) and cefepime (methicillin-susceptible isolates: 0.5-1 mg/L; methicillin-resistant isolates: 2-64 mg/L).

Anti-Bacterial Agents↗
PubMed2005

Prevalence and characterization of Staphylococcus aureus and enterotoxigenic Staphylococcus aureus in retail raw chicken meat throughout Japan.

A total of 444 samples of raw chicken meat (thighs, breasts, wings, livers, gizzards, hearts and ovaries) that retailed at 145 different supermarkets in 47 prefectures in Japan were examined for contamination with Staphylococcus aureus in association with its enterotoxigenicity. S. aureus was isolated from 292 (65.8%) of the samples, and from 131 of the 145 supermarkets. There was no significant difference in the detection rate of S. aureus according to the type of meat examined. About 80% of 714 isolates belonged to the poultry (57.1%) and human biotypes (22.1%). Seventy-eight (21.7%) of 360 isolates were enterotoxigenic and isolated from 78 samples in 53 supermarkets in 31 prefectures. Staphylococcal enterotoxins (SEs) produced were SEB (50 isolates), SEA (14), SEC (8), SED (2), SEA+SEB (2), and SEA+SEC (2). Most of the enterotoxigenic isolates belonged to the human and poultry biotypes, coagulase type VII, VIII or IV, and were lysed by phages of group III. Identical SE types, biotypes, coagulase types and pulsed-field gel electrophoresis (PFGE) patterns were shown in isolates from different types of meat at the same supermarket and from samples taken from different supermarkets in the same prefectures or in isolates from samples obtained from several different prefectures. Among the 50 SEB-producing isolates, 27 yielded three similar PFGE patterns that differed by only a few fragments, suggesting that they were closely related genetically. The three patterns were found in isolates of samples that retailed at 17 supermarkets in 11 prefectures, indicating that they may be disseminated among raw chicken meat in Japan.

Animals↗
PubMedDate not supplied

[Antibacterial activity of clindamycin and lincomycin in broth, serum, and in combination with polymorphonuclear leukocytes against Staphylococcus aureus and Staphylococcus epidermidis].

We investigated the antibacterial activity of clindamycin and lincomycin at 1/4 X minimum inhibitory concentration (MIC), 1 X MIC and 4 X MIC against a serum-resistant Staphylococcus aureus and a serum-resistant Staphylococcus epidermidis strain in broth, in serum with and without the presence of leukocytes and in Hank's medium in combination with leukocytes alone. Against both test strains, lincomycin in broth and serum was similarly effective, whereas against S. aureus clindamycin in broth was somewhat more active. In the combined test mixture of serum with leukocytes, even a 1/4 X MIC of clindamycin or lincomycin markedly improved leukocyte killing of S. aureus, whereas both compounds could not further enhance the marked leukocyte killing of S. epidermidis, even at inhibitory concentrations. In Hank's medium with leukocytes alone, clindamycin and lincomycin had at the most only a bacteriostatic effect against both test strains.

Blood↗
PubMed1962

Studies on the interactions between components of Staphylococcus aureus and Staphylococcus bacteriophage.

The cell walls of Staphylococcus aureus are capable of inactivating S. aureus bacteriophage. Furthermore, the cell walls isolated from S. aureus of a given phage type inactivate a variety of different staphylococcal bacteriophages. Under the conditions employed neither the isolated mucopeptide nor teichoic acid components of the cell walls act as bacteriophage receptor.

Bacteriophages↗
PubMed2006

Biofilm-forming ability profiling of Staphylococcus aureus and Staphylococcus epidermidis mastitis isolates.

Biofilm-forming ability has been increasingly recognized as an important virulence factor in Staphylococci, facilitating their persistence in the host, evading its defences and allowing bacterial survival at high antimicrobial concentrations. Staphylococcus aureus remains a major pathogen of chronic mastitis, but in the last years Staphylococcus epidermidis has emerged as a relevant mastitis pathogen. The present work aimed at the evaluation of the biofilm-forming ability of Staphylococci field isolates from bovine subclinical mastitis and at the development of a fluorescent in situ hybridisation (FISH) protocol that would allow the direct observation of biofilm formation in milk samples. The analysis of phenotypic expression in Congo Red Agar (CRA) and by FISH, showed that 37.5% of the S. aureus isolates produced biofilm, while by optical density measurement only 18.75% isolates revealed this phenotype. The results showed a fair agreement according to the kappa coefficient test (kappa = 0.259). Regarding S. epidermidis mastitis isolates, 37.5% revealed the ability to produce biofilm, but only four isolates were positive by all methods. This agreement was moderate (kappa = 0.467). The application of FISH to artificially contaminated milk samples allowed the direct observation of biofilm production by 37.5% isolates, showing total agreement with the CRA results. This method better mimics the in vivo conditions, especially in terms of the presence of calcium and iron, which in high concentrations, respectively, are known to inhibit or induce biofilm production.

Animals↗
PubMed1976

Susceptibility of Staphylococcus aureus and Staphylococcus epidermidis to 65 antibiotics.

The susceptibilities of 36 recent isolates of Staphylococcus aureus and 35 recent isolates of Staphylococcus epidermidis were determined against each of 65 antimicrobial agents and against two of them in combination. Rifampin was the most active of all the agents tested against both S. aureus and S. epidermidis. Among the penicillins, cloxacillin, dicloxacillin, and nafcillin were most active, although benzylpenicillin and phenoxymethyl penicillin were more active against susceptible strains. Cephaloridine was the most active of the cephalosporins, and sisomicin was the most active aminoglycoside. Minocycline was more active than the other tetracycline analogues tested. Among the macrolide-lincomycin compounds in clinical use, clindamycin was more active, and lincomycin was less active than erythromycin. The synergy of trimethoprim-sulfamethoxazole was more striking against S. aureus than against S. epidermidis. The median minimal inhibitory concentrations of the penicillins, cephalosporins, and aminoglycosides were lower against S. aureus, whereas the minimal inhibitory concentrations of the tetracyclines were lower against S. epidermidis.

Anti-Bacterial Agents↗
PubMed2003

Lysostaphin disrupts Staphylococcus aureus and Staphylococcus epidermidis biofilms on artificial surfaces.

Staphylococci often form biofilms, sessile communities of microcolonies encased in an extracellular matrix that adhere to biomedical implants or damaged tissue. Infections associated with biofilms are difficult to treat, and it is estimated that sessile bacteria in biofilms are 1,000 to 1,500 times more resistant to antibiotics than their planktonic counterparts. This antibiotic resistance of biofilms often leads to the failure of conventional antibiotic therapy and necessitates the removal of infected devices. Lysostaphin is a glycylglycine endopeptidase which specifically cleaves the pentaglycine cross bridges found in the staphylococcal peptidoglycan. Lysostaphin kills Staphylococcus aureus within minutes (MIC at which 90% of the strains are inhibited [MIC(90)], 0.001 to 0.064 microg/ml) and is also effective against Staphylococcus epidermidis at higher concentrations (MIC(90), 12.5 to 64 microg/ml). The activity of lysostaphin against staphylococci present in biofilms compared to those of other antibiotics was, however, never explored. Surprisingly, lysostaphin not only killed S. aureus in biofilms but also disrupted the extracellular matrix of S. aureus biofilms in vitro on plastic and glass surfaces at concentrations as low as 1 microg/ml. Scanning electron microscopy confirmed that lysostaphin eradicated both the sessile cells and the extracellular matrix of the biofilm. This disruption of S. aureus biofilms was specific for lysostaphin-sensitive S. aureus, as biofilms of lysostaphin-resistant S. aureus were not affected. High concentrations of oxacillin (400 microg/ml), vancomycin (800 microg/ml), and clindamycin (800 microg/ml) had no effect on the established S. aureus biofilms in this system, even after 24 h. Higher concentrations of lysostaphin also disrupted S. epidermidis biofilms.

Anti-Bacterial Agents↗
PubMed1981

Antibiotic susceptibility patterns of Staphylococcus aureus and Staphylococcus epidermidis.

The susceptibilities of 100 clinically significant isolates of Staphylococcus aureus and 45 clinically significant isolates of Staphylococcus epidermidis against various antibiotics were determined by the agar dilution method. Rifampin was the most active of all agents tested against both S. aureus and S. epidermidis. Among the beta-lactam antibiotics, penicillin G was least active, but had the lowest MIC in range. All strains of S. aureus were susceptible to cloxacillin and cephalothin at concentrations of 2 and 4 mcg/ml, respectively. Cloxacillin was less active against S. epidermidis, while cephalothin was more active. Minocycline was more active than doxycycline and tetracycline. Chloramphenicol was not only almost as poor in activity against S. aureus as tetracycline, but also the least active of all agents tested against S. epidermidis. Gentamicin was the most active aminoglycoside. Erythromycin, lincomycin and clindamycin showed the bimodal pattern of susceptibility. These results suggest the marked variability in antibiotic susceptibility of staphylococci.

Anti-Bacterial Agents↗
PubMed1994

Immunological recognition of fibronectin-binding proteins of Staphylococcus aureus and Staphylococcus capitis, strain LK 499.

Antibodies to fibronectin-binding proteins (FnBPs) of Staphylococcus aureus, including binding domain of FnBPA, the D region, or the A-C regions of FnBPB were produced in rabbits and mice. These antibodies were used to characterize cell-associated FnBPs of S. aureus strain Cowan I, S. aureus strain U320 and a coagulase-negative Staphylococcus capitis strain LK499 as well as extracellular FnBPs in culture supernatants of the strain U320. FnBPs of S. aureus were predominantly FnBPA, while FnBPB was hardly detected on the cells or in culture supernatant of these S. aureus strains. Moreover, S. capitis strain LK499 possessed different FnBP(s) compared to S. aureus because the antibodies to S. aureus FnBPs did not recognize FnBP(s) on S. capitis.

Adhesins, Bacterial↗
PubMed1983

Intramammary inoculation of the dairy cow with Staphylococcus aureus and Staphylococcus epidermidis during the nonlactating period.

The susceptibility of nonlactating mammary glands to intramammary inoculation with staphylococci was determined. Susceptibility was determined by inoculation of bacteria through the teat canal directly into the gland sinus. In experiment 1, Staphylococcus aureus (Newbould 305, ATCC 29740) was inoculated into glands at various intervals throughout the nonlactating period. During the first month, 35 of 52 inoculated glands became infected (67.3%). During the month before parturition when 37 glands were inoculated, 36 became infected (97.3%). In experiment 2, Staphylococcus epidermidis (A-44) was inoculated into mammary glands. During the first month of the nonlactating period, 35 glands were inoculated and only 6 glands became infected (17%). During the month before parturition, 36 of 38 inoculated glands became infected (94%). It appears that the mammary gland is susceptible to staphylococcal infection during the nonlactating period, because all glands became infected when they were inoculated during the month before parturition.

Animals↗
PubMed1990

Comparison of the staphylocoagulase activities of Staphylococcus aureus and Staphylococcus intermedius on Chromozym-TH.

The staphylocoagulases of Staphylococcus intermedius (39 strains from clinical samples from dogs and 1 strain from a pigeon) and Staphylococcus aureus (40 strains from nine different animal species) were compared by using the chromogenic methods of Engels et al. (W. Engels, M. Kemps, and C. P. A. van Boven, J. Clin. Microbiol. 14:496-500, 1981). S. intermedius produced staphylocoagulase which resembled that of S. aureus in its rate and method of action on prothrombin, but S. intermedius produced it in lesser amounts. Therefore, chromogenic substrate tests such as Chromozym-TH (Boehringer GmbH, Mannheim, Federal Republic of Germany) (as described by Engels et al.) can be used for the detection of staphylocoagulase in both of these species. However, to detect accurately the presence of S. intermedius staphylocoagulase by this method, preconcentration of the extracellular proteins or an extension of the reaction time of the test would be required. The method described by Engels et al. was designed specifically for clinical laboratories that handle human samples. Under those circumstances the method could be regarded as specific, enabling an identification of S. aureus only. However, as both S. aureus and S. intermedius give positive results in this test, in veterinary diagnostic laboratories, heavy reliance must not be placed on this test for determination of organisms to the species level.

Animals↗
PubMed1988

Problems with rapid agglutination methods for identification of Staphylococcus aureus when Staphylococcus saprophyticus is being tested.

Six rapid agglutination tests for identification of Staphylococcus aureus were evaluated by using 62 strains of S. aureus, 63 strains of S. saprophyticus, and 67 strains of other coagulase-negative staphylococci. S. saprophyticus was responsible for 19 of 26 false-positive results and 20 uninterpretable reactions. Thus, urinary staphylococcal isolates that are positive by rapid agglutination tests may require other confirmatory tests for the identification of possible S. saprophyticus.

Hemagglutination Tests↗
PubMed2004

Development of a PCR test to differentiate between Staphylococcus aureus and Staphylococcus intermedius.

The presence of Staphylococcus intermedius in food remains unclear because routine laboratory analysis does not discriminate between S. intermedius and Staphylococcus aureus, a major cause of food poisoning. Both species share many phenotypic characteristics, including coagulase and thermonuclease production. In both species, some strains can produce enterotoxin and therefore can be the cause of food poisoning outbreaks. Although the ID32 Staph System (bioMérieux, SA, Marcy l'Etoile, France), based on a miniaturized phenotypic characterization, gives satisfactory results for discriminating between these two species, some rapid molecular PCR-based methods have been developed to identify S. aureus specifically, but they do not identify S. intermedius. Here, we developed a rapid, accurate, and discriminative multiplex PCR method that targets species-specific sequences in the nuc gene, which encodes thermonuclease in the two species. The test includes an internal positive control that targets a highly conserved region of 16S ribosomal RNA gene (rDNA). A total of 116 strains were used to validate our test. The test gave no signal on the following Staphylococcus species: S. epidermidis, S. chromogenes, S. hyicus, S. warneri, S. xylosus, S. lentus, and S. sciuri. It allowed a 100% successful discrimination between S. aureus (44 strains tested) and S. intermedius (57 strains) isolated from different origins.

Animals↗
PubMed1983

Construction and characterization of plasmid vectors for cloning in Staphylococcus aureus and Staphylococcus carnosus.

Several plasmid vectors for cloning in Staphylococcus aureus and S. carnosus have been constructed and characterized. The chimeric plasmids are composed of parts of the following parental plasmids: The chloramphenicol-resistance plasmid, pC194, the tetracycline-resistance plasmid, pMK148, and the erythromycin-resistance plasmid, pE12. All the chimeric plasmids confer two selectable antibiotic-resistance markers on host cells. Insertional inactivation of the various antibiotic-resistance markers occurred at the BclI site of pE12, and the Sau96- or AvaII-site of pMK148; only a slight inactivation of the chloramphenicol-resistance marker occurred at the HaeIII-site of pC194. The chimeric plasmids pCT20 and pCE10 are both stable in S. aureus and S. carnosus. In addition, the hybrid plasmids of pCT20 and pCE10, containing lambda-DNA fragments in various restriction sites between 0.4 and 1.2 kb, are stably maintained. The inserted lambda-DNA fragments appear unchanged.

Chloramphenicol↗