Coagulase-negative staphylococci strains resistant to oxacillin isolated from neonatal blood cultures.

Coagulase-negative staphylococci (CoNS) are the microorganisms most frequently isolated from clinical samples and are commonly found in neonatal blood cultures. Oxacillin is an alternative treatment of choice for CoNS infections; however, resistance to oxacillin can have a substantial impact on healthcare by adversely affecting morbidity and mortality. The objective of this study was to detect and characterise oxacillin-resistant CoNS strains in blood cultures of newborns hospitalised at the neonatal ward of the University Hospital of the Faculty of Medicine of Botucatu. One hundred CoNS strains were isolated and the mecA gene was detected in 69 of the CoNS strains, including 73.2% of Staphylococcus epidermidis strains, 85.7% of Staphylococcus haemolyticus strains, 28.6% of Staphylococcus hominis strains and 50% of Staphylococcus lugdunensis strains. Among these oxacillin-resistant CoNS strains, staphylococcal cassette chromosome mec (SCCmec) type I was identified in 24.6%, type II in 4.3%, type III in 56.5% and type IV in 14.5% of the strains. The data revealed an increase in the percentage of CoNS strains isolated from blood cultures from 1991-2009. Furthermore, a predominant SCCmec profile of the oxacillin-resistant CoNS strains isolated from neonatal intensive care units was identified with a prevalence of SCCmec types found in hospital-acquired strains.

Evaluation of discrepancies between oxacillin and cefoxitin susceptibility in coagulase-negative staphylococci.

The Clinical and Laboratory Standards Institute (CLSI) recommends testing coagulase-negative staphylococci (CoNS) strains to determine resistance against oxacillin by testing for mecA, PBP2a, or with cefoxitin disk. However, discrepant results of resistance to oxacillin and susceptibility to cefoxitin were found. In this study, we aimed to investigate the oxacillin resistance and cefoxitin susceptibility of CoNS in Taiwan. Of 9,017 strains collected from 2005 to 2010, 131 (1.5%) of the isolates were oxacillin-resistant and cefoxitin-susceptible. Species identification was carried out using the Vitek 2 system or 16S ribosomal RNA sequencing. Oxacillin minimum inhibitory concentrations (MICs) were examined by the agar dilution method. The presence of mecA and the activity of ß-lactamase were performed by polymerase chain reaction (PCR) and Cefinase disks, respectively. Overall, 33% (43/129) of the strains carried mecA and 43% (37/86) of mecA-negative isolates tested positive for ß-lactamase. The remaining 49 isolates were negative for both mecA and ß-lactamase, and were mainly Staphylococcus cohnii ssp. urealyticus and S. saprophyticus (oxacillin MICs 0.5-2 µg/ml) obtained from bloodstream and urinary tract infections. Our study suggests that incorrect reporting can be found in CoNS using cefoxitin disk alone to determine the susceptibility to oxacillin, and the strains should be further tested for oxacillin MICs and detection of the mecA gene or ß-lactamase activity.

Pulsed-field gel electrophoretic analysis and some characteristics of Staphylococcus aureus isolated from retail foods and human hands.

This study investigates whether there is a predominant Staphylococcus aureus strain in retail foods and healthy human hands, and examines the relationship between pulsed-field gel electrophoresis (PFGE) banding patterns and the S. aureus characteristics of staphylococcal enterotoxin (SE) type, coagulase type, and ß-lactamase activity. Ninety-four strains of S. aureus isolated from retail foods and healthy human hands were analyzed by PFGE. Several strains isolated from the same shop or a chain store showed identical patterns, indicating that the origins of these strains were identical. After excluding these strains showing identical patterns, 54 strains were used for the PFGE analysis. No spread of a particular clone in the environment surrounding the food was apparent. The PFGE analysis of these 54 strains was classified in 6 lineages (L1-L6). There was no relationship between the PFGE banding pattern and coagulase type or SE type. Eleven (84.6%) of the 13 isolates in PFGE banding pattern L5 did not produce ß-lactamase, suggesting that the production of ß-lactamase influenced a specific PFGE banding pattern.

Reduction in coagulase-negative staphylococcus infection rates in the NICU using evidence-based research.

Coagulase-negative Staphylococcus (CoNS) bloodstream infection is the most common cause of sepsis in the NICU and can lead to significant morbidity and mortality. There is evidence that hand hygiene using an alcohol-based gel and wearing gloves during patient care, management of central and peripheral intravenous lines using the Centers for Disease Control and Prevention (CDC) guidelines, and a closed medication administration system can reduce the incidence to CoNS sepsis in the (NICU). To successfully apply the evidence and decrease the CoNS infection rate, a systematic process is necessary. One approach to process change that significantly reduced the CoNS infection rate in a health care system with two Level III NICUs included using system thinking; working within a multidisciplinary team; using evidence to revise, develop, and implement policies and procedures; developing staff education programs; and monitoring and providing feedback to all staff members.

Molecular characterization of high-level mupirocin resistance in methicillin-resistant staphylococci isolated from companion animals.

High-level mupirocin resistance (HLMR) is determined by the plasmid-located ileS2 gene flanked by two copies of the insertion sequence 257 (IS257). The molecular epidemiology of high-level mupirocin-resistant isolates could be assessed by the determination of their IS257-ileS2 spacer regions conformation. In this study, 188 isolates of methicillin-resistant staphylococci were subjected to the detection of HLMR, and analysis of the conformation of the IS257-ileS2 spacer regions. Mupirocin resistance was detected in five (2,6%) isolates, among which two were recognized as Staphylococcus pseudintermedius, two as Staphylococcus haemolyticus, and one as Staphylococcus aureus. High-level mupirocin resistance was revealed by the agar disk diffusion method, and MIC values, and was confirmed by the detection of the ileS2 gene. The conformations of the IS257-ileS2 spacer regions were homologous in two S. haemolyticus strains tested. The remaining three isolates showed diverse IS257-ileS2 conformations. The results of this study indicate that HLMR occasionally occurs in staphylococci isolated from companion animals. The heterogeneity and the homogeneity of the IS257-ileS2 spacer regions confirm that the ileS2 gene spread among staphylococci of animal origin by the transfer of different as well as the same plasmids. Surveillance of the occurrence of mupirocin resistance and molecular characterization of resistant isolates are strongly recommended due to the possibility of plasmid-located resistance gene transfer between staphylococci.

Decreased susceptibility to teicoplanin and vancomycin in coagulase-negative Staphylococci isolated from orthopedic-device-associated infections.

We studied 315 coagulase-negative Staphylococcus strains recovered prospectively during 240 surgical procedures (206 subjects) from proven or suspected device-associated bone and joint infections. Sixteen strains (5.1%) had decreased susceptibility to glycopeptides: 15 (12 S. epidermidis strains, 2 S. capitis strains, and 1 S. haemolyticus strain) to teicoplanin alone (MIC of 16 mg/liter, n = 9; MIC of 32 mg/liter, n = 6) and one (S. epidermidis) to both teicoplanin and vancomycin (MIC, 16 and 8 mg/liter, respectively). Decreased susceptibility to teicoplanin was more prevalent in "infecting" strains (i.e., strains recovered from >/=2 distinct intraoperative samples) than in "contaminants" (i.e., strains not fulfilling this criterion) (8.1% [12/149] versus 2.4% [4/166], respectively [P = 0.022]). One hundred percent (13/13) of S. epidermidis strains with decreased susceptibility to teicoplanin were resistant to methicillin (versus 112/173 [64.7%] for S. epidermidis strains susceptible to teicoplanin; P = 0.021).

In vitro antimicrobial synergy testing of coagulase-negative staphylococci isolated from prosthetic joint infections using Etest and with a focus on rifampicin and linezolid.

In recent years, coagulase-negative staphylococci (CoNS) have been increasingly recognised as causative agents of various infections, especially in immunocompromised patients and related to implanted foreign body materials. CoNS, and especially Staphylococcus epidermidis, transform into a stationary growth phase and produce biofilm when involved in a foreign body infection, making them difficult to eradicate with antimicrobials. Rifampicin has the ability to penetrate biofilm, but resistance may develop rapidly. To reduce the emergence of resistance, rifampicin should be combined with additional antimicrobials, of which several different ones have been proposed, including the relatively new class of antimicrobials, oxazolidinones, represented by linezolid. Thirty-seven CoNS isolates from patients with prosthetic joint infection were investigated by synergy testing using Etest. Nine antimicrobial combinations, based on either rifampicin or linezolid, were tested. For 16 (43%) of the isolates, a synergistic (n = 5), additive (n = 14) and/or antagonistic (n = 11) effect were identified. In conclusion, Etest is an objective and easily performed in vitro method for antimicrobial synergy testing. However, each isolate requires testing for the specific combination considered for treatment.

Staphylococcal cassette chromosome mec (SCC mec) in methicillin-resistant coagulase-negative staphylococci. A review and the experience in a tertiary-care setting.

Coagulase-negative staphylococci (CNS) are increasingly recognized to cause clinically significant infections, with S. epidermidis often cited as the third most common cause of nosocomial sepsis. Among CNS, there is a high prevalence of methicillin resistance associated with staphylococcal cassette chromosome (SCCmec) elements. Although identical SCCmec types can exist in S. aureus and CNS, some novel classes of SCCmec may be unique to CNS. Differences in the accuracy of identification of CNS species and use of non-standardized methods for the detection of methicillin resistance have led to confusing data in the literature. In addition to the review of SCCmec in CNS, in this paper we report a 2-year surveillance of methicillin-resistant CNS in a tertiary-care hospital in Guadalajara, Mexico.

Enterotoxigenicity and Antibiotic Resistance of Coagulase-Negative Staphylococci Isolated from Raw Buffalo and Cow Milk.

Staphylococcal food poisoning is considered to be one of the most common foodborne illnesses worldwide. Because milk is rich in nutrients and its neutral pH, it leads to the growth of various bacteria. To date, the correlation between enterotoxigenic potential in Staphylococcus species and antimicrobial resistance (AMR), using bioinformatics analysis in buffalo and cow raw milk and the possible health risks from these bacteria, has not been examined in Egypt. A total of 42 Staphylococcus isolates representing 12 coagulase-positive staphylococci (Staphylococcus aureus and Staphylococcus intermedius) and 30 coagulase-negative staphylococci (Staphylococcus capitis, Staphylococcus xylosus, Staphylococcus carnosus, Staphylococcus saccharolyticus, and Staphylococcus auricularis) were isolated. An assay of the antimicrobial resistance phenotypes indicated low resistance against vancomycin (9.5%). The blaZ gene was associated with penicillin G and methicillin resistance and not with sulbactam + ampicillin. The presence of the gene ermB presented the correlation with erythromycin resistance and tetK with tetracycline resistance (correlation index: 0.57 and 0.49, respectively), despite the absence of the same behavior for ermC and tetM, respectively. Interestingly, the gene mecA was not correlated with resistance to methicillin or any other ß-lactam. Correlation showed that slime-producing isolates had more resistance to antibiotics than those of nonslime producers. The multiple correlations between antibiotic resistance phenotypes and resistance genes indicate a complex nature of resistance in Staphylococcus species. The antimicrobial resistance could potentially spread to the community and thus, the resistance of Staphylococcus species to various antibiotics does not depend only on the use of a single antimicrobial, but also extends to other unrelated classes of antimicrobials.

Management of the catheter in documented catheter-related coagulase-negative staphylococcal bacteremia: remove or retain?

BACKGROUND: Studies and guidelines recommending the retention of the central venous catheter (CVC) in patients with coagulase-negative staphylococcal bacteremia were based on loose definitions of bacteremia and/or did not evaluate the risk of recurrence. In this study, we used strict definitions of coagulase-negative staphylococcal bacteremia to determine the impact of CVC retention on response to and recurrence of infection. METHODS: During the period from July 2005 through December 2007, we retrospectively evaluated 188 patients with coagulase-negative staphylococcal bacteremia. Bacteremia was defined using the strict Centers for Disease Control and Prevention criteria of 2 positive blood culture results. Catheter-related bacteremia was confirmed by differential quantitative blood cultures (>or=3:1) or time to positivity (>2 h). RESULTS: Resolution of infection within 48 h after commencement of antimicrobial therapy was not influenced by CVC removal or exchange versus retention and occurred in 175 patients (93%). Multiple logistic regression analysis showed that infection was 7.0 times (95% confidence interval [CI], 1.5-32.6 times) more likely to fail to resolve in patients with an intensive care unit stay prior to infection ( P = .013 ) and 3.8 times (95% CI, 1.1-13.3 times) more likely to fail to resolve in patients who had other concurrent sites of infection (P = .041 ). Duration of therapy did not affect recurrence. Multiple logistic regression analysis revealed that patients with catheter retention were 6.6 times (95% CI, 1.8-23.9 times) more likely to have a recurrence than were those whose catheter was removed or exchanged (P = .004). CONCLUSIONS: CVC retention does not have an impact on the resolution of coagulase-negative staphylococcal bacteremia but is a significant risk factor of recurrence.

Comparison of the BD Phoenix system with the cefoxitin disk diffusion test for detection of methicillin resistance in Staphylococcus aureus and coagulase-negative staphylococci.

The BD Phoenix system was compared to the cefoxitin disk diffusion test for detection of methicillin (meticillin) resistance in 1,066 Staphylococcus aureus and 1,121 coagulase-negative staphylococcus (CoNS) clinical isolates. The sensitivity for Phoenix was 100%. The specificities were 99.86% for S. aureus and 88.4% for CoNS.

Differentiating culture samples representing coagulase-negative staphylococcal bacteremia from those representing contamination by use of time-to-positivity and quantitative blood culture methods.

Differentiating true coagulase-negative staphylococcal infection from contamination has an important impact on therapeutic implications. Time to positivity reflects bacterial density and may help in the interpretation of blood cultures. We retrospectively reviewed the records of 272 patients from June 2005 to January 2008 for clinical characteristics, microbiological data, and therapeutic outcome. Four groups were identified. The first three groups, as follows, included patients with one positive quantitative blood culture: the low-colony-count group (<10 CFU/ml), the moderate-colony-count group (30 to 100 CFU/ml), and the high-colony-count group (>100 CFU/ml). The control group included patients with two positive quantitative blood cultures and definite coagulase-negative staphylococcal bloodstream infection. The high-colony-count group had shorter time to positivity (< or = 16 h) than did the low-colony-count group (P < 0.0001). The low-colony-count group had a significantly longer time to positivity, >20 h (P = 0.001), than did the moderate-colony-count group. Even though antibiotics were not provided in 71% of cases and central venous catheter was retained in 83%, the low-colony-count group had a favorable outcome, suggesting that <10 CFU/ml represents contamination. The high-colony-count group, similar to the positive control group, required antibiotics in 81% of cases and central venous catheter removal in 51% (P = 0.001). A time to positivity of < or = 16 h reflects high-grade bacteremia with CFU of >100. Similar to the positive control group, these patients required an active therapeutic approach. A time to positivity of >20 h indicates possible contamination with a CFU of <10, and active therapy may not be required.

Characterization of methicillin-resistant Staphylococcus aureus and methicillin-resistant coagulase-negative Staphylococcus spp. isolated from US West Coast public marine beaches.

OBJECTIVES: The aim of this study was to isolate and characterize methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant coagulase-negative Staphylococcus spp. (MRCoNS) from marine water and intertidal beach sand from public beaches in Washington State, USA. METHODS: Fifty-one staphylococci from Washington State beaches were characterized using antimicrobial susceptibility testing, carriage of acquired tetracycline and/or macrolide resistance genes, staphylococcal cassette chromosome mec (SCCmec) typing, the BBL Crystal Gram-Positive ID System and/or 16S rRNA sequencing, coagulase test and multilocus sequence typing (MLST) for MRSA. RESULTS: Five multidrug-resistant MRSA SCCmec type I, of which three were MLST type ST45, one ST59 and one a new MLST type, ST1405, plus one susceptible non-typeable (NT) MRSA ST30 were characterized. Thirty-three MRCoNS isolates, representing 21 strains from 9 Staphylococcus spp., carried a range of SCCmec types [I (2), II (6), III (3), V (2), I/II (1) and NT (7)] and varied in their antibiotic susceptibility to other antibiotic classes and carriage of acquired tetracycline/macrolide resistance gene(s). MRSA and MRCoNS donors co-transferred tet(M) and erm(A) genes to an Enterococcus faecalis recipient at a frequency of 10(-8). CONCLUSIONS: This is the first report of MRSA and MRCoNS isolated from marine water and intertidal beach sand. The MLST types and antibiotic carriage of five MRSA isolates were similar to hospital MRSA isolates rather than US community-acquired MRSA isolates. Our results suggest that public marine beaches may be a reservoir for transmission of MRSA to beach visitors as well as an ecosystem for exchange of antibiotic resistance genes among staphylococci and related genera.

Effectiveness of antibiotics and antiseptics on coagulase-negative staphylococci for the decontamination of bone allografts.

Bone allografts retrieved from multi-organ donors can be decontaminated with minimally aggressive methods. Therefore, we evaluated the efficacy of antibiotics and antiseptics in the decontamination of bone fragments actively contaminated with coagulase-negative staphylococci. Gentamicin (512/1,024 microg/mL), rifampicin (400/1,000 microg/mL), chlorhexidine in alcohol and chlorhexidine soap were tested with different contact times and temperatures and a delay in starting decontamination. Gentamicin-susceptible strains dried on bone could be removed by gentamicin 512 microg/mL after 19 h of contact, while strains not dried on bone could be eliminated by soaking bone for 60 min in gentamicin 512 microg/mL. Rifampicin-susceptible strains could be eliminated by soaking bone for 60 min in rifampicin 1,000 microg/mL. In none of the experimental conditions could gentamicin/rifampicin-resistant staphylococci be eliminated. Antiseptics could not eliminate staphylococci from bone. Different antibiotics need different protocols in order to decontaminate bone allografts.

The role of virulence factors in the outcome of staphylococcal peritonitis in CAPD patients.

BACKGROUND: Peritonitis continues to be the most frequent cause of peritoneal dialysis (PD) failure, with an important impact on patient mortality. Gram-positive cocci such as Staphylococcus epidermidis, other coagulase-negative staphylococci (CoNS), and Staphylococcus aureus are the most frequent etiological agents of PD-associated peritonitis worldwide. The objective of the present study was to compare peritonitis caused by S. aureus and CoNS and to evaluate the factors influencing outcome. METHODS: Records of 86 new episodes of staphylococcal peritonitis that occurred between 1996 and 2000 in the Dialysis unit of a single university hospital were studied (35 due to S. aureus, 24 to S. epidermidis and 27 to other CoNS). The production of slime, lipase, lecithinase, nuclease (DNAse), thermonuclease (TNAse), alpha- and beta-hemolysin, enterotoxins (SEA, SEB, SEC, SED) and toxic shock syndrome toxin-1 (TSST-1) was studied in S. aureus and CoNS. Antimicrobial susceptibility was evaluated based on the minimal inhibitory concentration determined by the E-test. Outcome predictors were evaluated by two logistic regression models. RESULTS: The oxacillin susceptibility rate was 85.7% for S. aureus, 41.6% for S. epidermidis, and 51.8% for other CoNS (p = 0.001). Production of toxins and enzymes, except for enterotoxin A and alpha-hemolysin, was associated with S. aureus episodes (p < 0.001), whereas slime production was positive in 23.5% of CoNS and 8.6% of S. aureus strains (p = 0.0047). The first model did not include enzymes and toxins due to their association with S. aureus. The odds of resolution were 9.5 times higher for S. epidermidis than for S. aureus (p = 0.02) episodes, and were similar for S. epidermidis and other CoNS (p = 0.8). The resolution odds were 68 times higher for non-slime producers (p = 0.001) and were not influenced by oxacillin resistance among vancomycin-treated cases (p = 0.89). In the second model, the resolution rate was similar for S. aureus and S. epidermidis (p = 0.70), and slime (p = 0.001) and alpha-hemolysin (p = 0.04) production were independent predictors of non-resolution. CONCLUSION: Bacterial species and virulence factors rather than antibiotic resistance influence the outcome of staphylococcal peritonitis.

Diversity of staphylococcal cassette chromosome in coagulase-negative staphylococci from animal sources.

AIMS: To type the staphylococcal cassette chromosome (SCC) in coagulase-negative staphylococci (CoNS) from animal sources. METHODS AND RESULTS: A total of 92 CoNS isolates recovered from farm animals was analysed. The top three staphylococcal species were Staphylococcus lentus (34), S. sciuri (31), and S. xylosus (13). The presence of the cassette chromosome recombinase (ccr) genes ccrA1, ccrB1, ccrA2, ccrB2, ccrA3, ccrB3 and ccrC, the mec regulatory genes mecI and mecR1, and Tn554 was used to differentiate the SCC. A total of 60 of the 92 isolates were methicillin resistant. Among the 60 methicillin-resistant Staphylococcus spp. isolates, SCCmec (mecA-carrying SCC) types I, III, IV and V were identified in 24 isolates based on the combinations of the ccr genes and the mec regulatory genes, with type III being predominant. The single S. epidermidis carried SCCmec type IV. SCC type III was also identified in two of 32 methicillin-susceptible isolates. Identical SCCmec types were present in different species of CoNS. Pulsed-field gel electrophoresis (PFGE) generated 64 patterns out of 81 PFGE typeable isolates. Indistinguishable clones were detected in animals from different farms. CONCLUSIONS: Heterogeneous SCC existed in CoNS of diverse genetic background. Both clonal transmission of methicillin-resistant CoNS and horizontal transfer of SCCmec occurred in the animal production environment. SIGNIFICANCE AND IMPACT OF THE STUDY: This study adds to our knowledge of SCCmec type and the diversity of SCC in CoNS.

Phenotypic and genotypic methods for the detection of methicillin resistant coagulase negative staphylococci: a comparative study.

CoNS are the major cause of nosocomial infection in last decade and methicillin resistant CoNS has emerged as a major clinical problem. The present study was to compare different phenotypic methods with genotypic method PCR, for the detection of methicillin resistance in CoNS. 100 CoNS isolates from different samples were studied for the detection of mecA gene. PCR was considered as "gold standard". Oxacillin and cefoxitin antibiotics were used for different phenotypic tests (DD, Agar dilution and MHOX). The sensitivities of oxacillin and cefoxitin disks for all CoNS were found to be 92.30% and 88.46% respectively and the specificities were 87.5% and 100% respectively. The sensitivities of the agar dilution test for oxacillin and cefoxitin were 86.53% and 80.76%, respectively, where as the specificities were 79.16% and 85.41%, respectively. The sensitivity of MHOX was observed to be 96.16% and specificity 72.91%. Cefoxitin D.D and oxacillin AD methods could be used as initial test for the determination of methicillin resistance in CoNS isolates. The result of MHOX shows that it could be the best single method for the evaluation of oxacillin resistance mediated by the mecA gene for all CoNS species.

Key role for clumping factor B in Staphylococcus aureus nasal colonization of humans.

BACKGROUND: Staphylococcus aureus permanently colonizes the vestibulum nasi of one-fifth of the human population, which is a risk factor for autoinfection. The precise mechanisms whereby S. aureus colonizes the nose are still unknown. The staphylococcal cell-wall protein clumping factor B (ClfB) promotes adhesion to squamous epithelial cells in vitro and might be a physiologically relevant colonization factor. METHODS AND FINDINGS: We define the role of the staphylococcal cytokeratin-binding protein ClfB in the colonization process by artificial inoculation of human volunteers with a wild-type strain and its single locus ClfB knock-out mutant. The wild-type strain adhered to immobilized recombinant human cytokeratin 10 (CK10) in a dose-dependent manner, whereas the ClfB(-) mutant did not. The wild-type strain, when grown to the stationary phase in a poor growth medium, adhered better to CK10, than when the same strain was grown in a nutrient-rich environment. Nasal cultures show that the mutant strain is eliminated from the nares significantly faster than the wild-type strain, with a median of 3 +/- 1 d versus 7 +/- 4 d (p = 0.006). Furthermore, the wild-type strain was still present in the nares of 3/16 volunteers at the end of follow-up, and the mutant strain was not. CONCLUSIONS: The human colonization model, in combination with in vitro data, shows that the ClfB protein is a major determinant of nasal-persistent S. aureus carriage and is a candidate target molecule for decolonization strategies.

Identification of coagulase-negative Staphylococci by using the BD phoenix system in the low-inoculum mode.

The new "low-inoculum" mode of the Phoenix system was evaluated to identify clinical coagulase-negative staphylococci. API ID32 Staph panels were used as comparators, and discrepancies were resolved by 16S rRNA and tuf gene analysis. The system correctly identified 90.5% of isolates, with a mean time of 10.2 h. Accuracy was satisfactory for Staphylococcus epidermidis, S. saprophyticus, and S. haemolyticus.