Nationwide German multicenter study on prevalence of antibiotic resistance in staphylococcal bloodstream isolates and comparative in vitro activities of quinupristin-dalfopristin

Antibiotic-resistant gram-positive bacteria have become an increasing problem in the last two decades. In order to evaluate the prevalence of antibiotic resistance in staphylococcal bloodstream isolates in Germany, 2,042 staphylococci collected in 21 tertiary-care hospitals were investigated during a 3-year period (March 1996 to March 1999). Altogether, 1,448 S. aureus isolates and 594 coagulase-negative staphylococci (CoNS) that comprised 13 different species were included. Furthermore, the antistaphylococcal activities of quinupristin-dalfopristin were compared with those of eight other compounds by the broth microdilution method. The rates of oxacillin resistance in Staphylococcus aureus, S. epidermidis, S. haemolyticus, and other CoNS were 13.5, 69, 90, and 34%, respectively. In oxacillin-resistant strains high rates of resistance (up to 100%) to erythromycin, clindamycin, ciprofloxacin, and gentamicin were also observed. However, no strain appeared to be resistant to vancomycin or quinupristin-dalfopristin. The streptogramin combination exhibited excellent in vitro activity against all staphylococcal species tested, regardless of the patterns of resistance to other drug classes. In terms of MICs at which 90% of the isolates are inhibited, quinupristin-dalfopristin was 2 times more active against S. aureus isolates, 4 to 16 times more active against S. haemolyticus, and 8 to 32 times more active against S. epidermidis than vancomycin or teicoplanin.     

In vitro activity of antimicrobial agents against oxacillin resistant staphylococci with special reference to Staphylococcus haemolyticus

One hundred and sixty seven isolates of staphylococci isolated from the inpatients of a tertiary care referral hospital in South India were speciated and activity of oxacillin, glycopeptides, linezolid and quinupristin/dalfopristin against these isolates was tested by broth microdilution method. Of the 114 coagulase negative staphylococci (CoNS), 49.1 % were S. haemolyticus, isolated predominantly from urine (64.6%), while the rest belonged to 11 other species. More than half the isolates of S. aureus (52.8%) and 68.4% of the CoNS were oxacillin resistant. All the strains were uniformly susceptible to vancomycin, linezolid and quinupristin/dalfopristin; but 25.6% isolates of S. haemolyticus showed reduced susceptibility to teicoplanin (MIC: 8-16 mg/L). Our study demonstrates the high prevalence of oxacillin resistance among hospital isolates of S. aureus and CoNS in India. Vancomycin, along with the newer agents like linezolid and quinupristin/dalfopristin remains the drug of choice for treating multi drug resistant staphylococcal infections.     

Molecular Epidemiology of Oxacillin-resistant Staphylococcus aureus and in vitro Activity of Glycopeptides against Staphylococcus species

Oxacillin resistant Staphylococcus aureus and coagulase negative Staphylococcus species (MRSA and MRSCoN)have become major pathogens in nosocomial infections.The first MRSA isolate in the world was identified in En gland in 1961 [1]. Since that tim…     

Vancomycin resistance in Staphylococcus haemolyticus causing colonization and bloodstream infection.

The increase in the incidence of infections due to beta-lactam-resistant coagulase-negative staphylococci has resulted in expanded use of vancomycin for such infections. Despite this, coagulase-negative staphylococci have remained susceptible to vancomycin in recent years. This report describes a strain of Staphylococcus haemolyticus with increased resistance to vancomycin (MIC, 8.0 to 16 micrograms/ml). S. haemolyticus was initially isolated from a patient with acute leukemia and neutropenia in surveillance throat and stool cultures. The microdilution vancomycin MICs for these isolates were 1.0 to 2.0 micrograms/ml. Subsequent S. haemolyticus isolates from the bloodstream and tracheal aspirate occurred in the setting of prolonged empirical vancomycin therapy. MICs for these isolates were 8.0 to 16 micrograms/ml. Further vancomycin resistance (MIC, 32 micrograms/ml) could be selected for in vitro in all four isolates. Restriction endonuclease analysis of plasmid DNA indicated that the isolates were very closely related and likely to be of the same strain. We conclude that colonization with a vancomycin-susceptible strain of S. haemolyticus was subsequently linked to a nosocomial bloodstream infection with an apparently identical strain with intermediate levels of vancomycin resistance. Prolonged empirical vancomycin therapy was temporally associated with this episode.     

Prevalence of isolates with reduced glycopeptide susceptibility in orthopedic device-related infections due to methicillin-resistant Staphylococcus aureus

We evaluated, by an improved susceptibility testing method, the prevalence and significance of low-level glycopeptide resistance in methicillin-resistant Staphylococcus aureus (MRSA) isolates, which belonged to a previously described, retrospective cohort of patients treated for orthopedic device-related infections (ODRI) at the Geneva University Hospital between 2000 and 2008. Fifty-seven individual or multiple isolates were retrieved from 41 ODRI patients for glycopeptide susceptibility and clonality studies, including 20 patients with prosthetic joint (PJ) and 21 with osteosynthesis (OS) MRSA infections. Low-level glycopeptide resistance was detected by elevated teicoplanin or/and vancomycin minimum inhibitory concentrations (MICs ≥4?mg/L), as determined by a previously validated combination of macrodilution and agar dilution assays of improved sensitivity. MRSA isolates with elevated teicoplanin MICs were detected in 20/41 (49?%) ODRI patients at the onset or during the course of glycopeptide therapy, namely, in 10 of 20 patients with PJ and 10 of 21 patients with OS infections. Only one isolate developed a concomitant increase in vancomycin MIC during therapy. 13/20 (65?%) glycopeptide-intermediate S. aureus (GISA)-infected patients, including 7/10 (70?%) with PJ and 6/10 (60?%) with OS, experienced treatment failure. In contrast, therapy failed in only 5/21 (24?%) ODRI patients with non-GISA isolates (p?=?0.012), including 2/10 (20?%) with PJ and 3/11 (27?%) with OS infections. The emergence of low-level teicoplanin resistance could not be explained by teicoplanin administration, since only four patients received teicoplanin. The evaluation of low-level teicoplanin resistance may improve the detection of GISA isolates. Further studies are warranted to evaluate the impact of low-level teicoplanin resistance on the outcome of glycopeptide therapy.     

Nosocomial infection by Staphylococcus haemolyticus and typing methods for epidemiological study.

A patient with chronic myelogenous leukemia became colonized with a Staphylococcus haemolyticus strain and experienced a septic episode caused by this strain during a cytostatic course. The strain was multiply resistant to antibiotics; the MIC and MBC of vancomycin were 2 and 4 mg/liter, and the MIC and MBC of teicoplanin were 4 and 16 mg/liter, respectively. We performed a surveillance study on the carriage of S. haemolyticus in medical and nursing staff of the hospital ward where the patient was treated. S. haemolyticus was isolated from 18 sites on 12 of the 39 people tested. A number of typing methods were performed in order to investigate the possible relationships among the isolates. Methods used were immunoblotting of staphylococcal peptides, plasmid analysis, restriction fragment length polymorphism of chromosomal DNA, and pulsed-field gel electrophoresis of total DNA. Compared with the immunoblotting technique, the molecular methods were more discriminative. The strain colonizing the patient showed a consistent pattern by all typing methods during isolation. When the immunoblot technique was used, similar patterns were found with isolates from hospital staff and isolates from unrelated sources. With the molecular techniques, no evidence of a local spread of the patient's strain was found. However, plasmid profiles and restriction fragment length polymorphism and pulsed-field gel electrophoresis patterns showed that S. haemolyticus isolates collected from hospital ward personnel were related, which was not the case with isolates collected from unrelated sources. Restriction fragment length polymorphism analysis was more discriminative when IS431 was used as a DNA probe instead of a probe based on the 16S rRNA gene. S. haemolyticus, as in this case, may develop resistance to vancomycin and teicoplanin. These antibiotics are considered the last-resort drugs for the therapy of nosocomial gram-positive infections. Thus, local spread of staphylococci resistant to these drugs is an important problem, which should be prevented by strict hygienic measures and antibiotic policy.     

Antimicrobial resistance in coagulase-negative staphylococci

Patterns of resistance to antimicrobial agents were studied in 193 strains of coagulase-negative staphylococci isolated from hospital patients. Strains isolated from patients with malignant disease were significantly more often resistant to sulphonamide, trimethoprim, gentamicin and methicillin than were strains from other sources. Susceptibility to various beta-lactam antibiotics and aminoglycosides was investigated in members of the two most frequent species: Staphylococcus epidermidis and S. haemolyticus. S. haemolyticus strains were not only more often resistant to methicillin than S. epidermidis strains (respectively 81% and 17%) but they were more highly resistant (mean MICs respectively 85 and 19 mg/L). Methicillin-resistant S. haemolyticus strains were highly resistant to nine other beta-lactam antibiotics, whereas methicillin-resistant S. epidermidis strains showed both lower levels and a narrower spectrum of cross-resistance. Resistance to methicillin in members of both species was "heterogeneous", i.e., only a minority of cells in a culture showed significant resistance. Almost all gentamicin-resistant strains were sensitive to netilmicin and amikacin; rifampicin, vancomycin and teicoplanin were also highly active in vitro.     

Comparative activity of ceftobiprole against coagulase-negative staphylococci from the BSAC Bacteraemia Surveillance Programme, 2013–2015

Coagulase-negative staphylococci (CoNS) are a significant cause of bacteraemia, the treatment of which is becoming increasingly complex due to the emergence of multidrug-resistant strains. This study aimed to evaluate the in vitro activity of ceftobiprole, an advanced-generation cephalosporin, as compared with other antimicrobial agents against CoNS from patients with bacteraemia. As part of the British Society for Antimicrobial Chemotherapy (BSAC) Bacteraemia Surveillance Programme, 650 blood isolates of CoNS were obtained from patients with bacteraemia at 74 centres throughout the UK and Ireland for the years 2013–2015. Minimum inhibitory concentrations (MICs) of ceftobiprole and other antimicrobial agents were determined using the BSAC agar dilution method. Susceptibility was assessed by European Committee on Antimicrobial Susceptibility Testing (EUCAST) criteria. The majority of the isolates (63.2%) were Staphylococcus epidermidis. Overall, methicillin resistance, as determined by oxacillin susceptibility testing, was observed in 64.2% of isolates. The MIC_50/90 of ceftobiprole was 1/2 mg/L, and 100% of CoNS isolates were inhibited at the EUCAST ceftobiprole non-species-specific pharmacokinetic/pharmacodynamic breakpoint of 4 mg/L. Only one isolate was resistant to vancomycin. Overall rates of resistance to ciprofloxacin, clindamycin, erythromycin and teicoplanin were 50.5, 25.1, 68.2 and 20.9%, respectively. In S. epidermidis, resistance to oxacillin was associated with increased resistance to other antimicrobials. Ceftobiprole demonstrated in vitro activity against all CoNS species isolated from patients with bacteraemia and was active against species resistant to other antistaphylococcal antimicrobials. The collection of clinical data regarding the efficacy of ceftobiprole in treating CoNS bacteraemia is warranted.     

Prevalence, molecular epidemiology, and clinical significance of heterogeneous glycopeptide-intermediate Staphylococcus aureus in liver transplant recipients

We investigated the prevalence, molecular epidemiology, and clinical significance of heterogeneous glycopeptide-intermediate Staphylococcus aureus (hGISA) isolates in 48 liver transplant recipients infected or colonized with methicillin-resistant S. aureus over a 5-year period. Strains were screened for hGISA on Mueller-Hinton agar containing 5 mg of teicoplanin per liter. Heterogeneous glycopeptide resistance was confirmed by the E-test method with a dense inoculum and a simplified method of population analysis. hGISA strains were found in 13 (27%) of the 48 patients studied. Eleven of the 13 strains shared a common multiresistant phenotype with homogeneous methicillin resistance and gentamicin resistance, and they were closely related according to the results of pulsed-field gel electrophoresis. Only 2 of the 13 patients infected or colonized with hGISA strains had previously received glycopeptide therapy. Most patients were successfully treated with vancomycin, but one patient who failed to respond to vancomycin subsequently died. These results suggest that the high prevalence of hGISA among our patients was due to the clonal spread of a multiresistant strain.     

Glycopeptide resistance in coagulase-negative staphylococci isolated in blood cultures from patients with hematological malignancies during three decades

The aim of this study was to determine if there was a long-term increase in glycopeptide minimum inhibitory concentration (MIC) values, MIC creep, among bloodstream isolates of Staphylococcus epidermidis and S. haemolyticus isolated from patients with hematological malignancies. We conducted a retrospective single-center study where all positive blood cultures of S. epidermidis (n = 387) and S. haemolyticus (n = 19) isolated from patients with hematological malignancies during three decades, 1980 to 2009, were re-evaluated for the presence of reduced susceptibility to vancomycin and teicoplanin. Three different methods for the detection of reduced susceptibility to glycopeptides were used; standard Etest, macromethod Etest, and glycopeptide resistance detection (GRD) Etest. The median MIC value for vancomycin was 2 mg/L. MIC values for vancomycin and teicoplanin did not show any statistically significant increase during the study period. The presence of heterogeneously glycopeptide-intermediate staphylococci (hGIS) was analyzed among 405 coagulase-negative staphylococci (CoNS) isolates. hGIS were found in 31–45% of the CoNS isolates by the macromethod Etest and in 53–67% by the GRD Etest during the three decades. In conclusion, we did not observe any long-term glycopeptide MIC creep determined by the standard Etest, although a high and increasing proportion of heterogeneous vancomycin resistance was observed.     

Distribution and expression of macrolide resistance genes in coagulase-negative staphylococci

In total, 494 isolates of coagulase-negative staphylococci (CoNS) were identified to the species level by biochemical tests and sodA sequencing. Erythromycin resistance phenotypes were determined and specific resistance genes were identified by PCR. The prevalence of erythromycin resistance varied widely among staphylococcal species, from 0% in Staphylococcus lugdunensis to almost 90% in Staphylococcus haemolyticus. Most (63%) erythromycin-resistant isolates carried constitutively expressed erm(C) as the sole resistance determinant, with the notable exception of Staphylococcus hominis subsp. hominis, which carried inducible erm(C). The erm(A) and erm(B) determinants were comparatively rare. The msr(A) gene was carried by 20-30% of all erythromycin-resistant isolates, with little variation among species, and was combined in 16.7% of isolates with mph(C), a resistance gene of unknown clinical relevance found previously in isolates of veterinary origin. No erythromycin resistance that could not be attributed to the genes investigated was detected. It was concluded that the presence of methylases cannot be assumed in CoNS isolates that appear erythromycin-resistant and clindamycin-susceptible; thus, methods that detect the export mechanism should be used with clinically significant isolates to indicate whether use of clindamycin may be effective. In Staphylococcus epidermidis and S. haemolyticus, 46% and 66%, respectively, of erythromycin-resistant, clindamycin-susceptible isolates were susceptible to clindamycin therapy.     

PCR-based DNA fingerprinting of Staphylococcus haemolyticus to investigate nosocomial infections

Objective: To apply PCR-based DNA fingerprinting in a clinical microbiology laboratory to investigate nosocomial infections with Staphylococcus haemolyticus.
Method: DNA fingerprints were generated by PCR on 99 S. haemolyticus isolates using different primer combinations based on ERIC, REP or arbitrarily chosen simple repeat sequences.
Results: Primer combinations REP1+(GTC)₆ and ERIC1+ERIC2 had sufficient discrimatory power and were chosen to analyze the clinical isolates. DNA fingerprint patterns from strains isolated from the patients nursed in the same hospital ward in the period 1991–94 were approximately 90% similar to each other. One staff member, sampled in 1991, carried a strain with a similar fingerprint.
Conclusions: PCR based DNA fingerprinting is a suitable method to perform in a clinical laboratory. An S. haemolyticus strain appeared to be endemic in the hospital ward and had most probably been transmitted from patient to patient. S. haemolyticus may carry glycopeptide resistance and needs attention as a causative agent of nosocomial infections.     

The VISA/GISA problem: therapeutic implications

The emergence of vancomycin intermediate resistant Staphylococcus aureus (VISA) isolates in Japan, USA, France, Hong Kong and Korea among methicillin-resistant Staphylococcus aureus (MRSA) clinical isolates, is of great concern. Vancomycin has been the drug of choice for the treatment of multiresistant MRSA infections in the last three decades, but the management of invasive MRSA infections will become a serious problem if VISA strains become widespread. VISA isolates reported to date have a vancomycin MIC of 8 mg/L, and were isolated from patients with underlying diseases whose long-term vancomycin treatment apparently failed. Since many VISA isolates also have been resistant to teicoplanin, the term glycopeptide-intermediate S. aureus (GISA) is more appropriate. The frequency of GISA isolates appears to be extremely low; to date, only 10 GISA infections have been reported worldwide. However, heterogeneous resistance to glycopeptides (h-GISA) have been reported in Japan, Europe and Thailand. These h-GISA strains showed vancomycin MICs ranging from 1 to 4 mg/L, but had subpopulations that could grow on agar plates containing 4–8 mg/L, which may represent the first step in the development of GISA strains. Although GISA isolates have shown resistance to many antimicrobials, all GISA isolates remain susceptible to co-trimoxazole and some of them to other common antimicrobials. Currently, there are no recommended therapy guidelines for GISA infections, although in recent studies, several new drugs have shown promising activity against GISA strains. In addition, synergy between glycopeptides and β-lactams against GISA strains was observed in some in vivo and in vitro studies. Specific MRSA/GISA control programs, rational antibiotic policies, including the reduction of glycopeptide use, and rapid laboratory detection of GISA and h-GISA strains are the key measures in preventing the spread of these strains.     

Glycopeptide Resistance in Coagulase-Negative Staphylococci

 Coagulase-negative staphylococci (CNS) were the first organisms in which acquired glycopeptide resistance was recognized. Ever since the early reports, it has been apparent that resistance to teicoplanin is more common than that to vancomycin and that resistance occurs mostly in species such as Staphylococcus haemolyticus and Staphylococcus epidermidis. The minimum inhibitory concentrations (MICs) of teicoplanin for CNS usually fall over a wide range, and, especially in some methicillin-resistant isolates of the two above-mentioned species, they can reach and even exceed the resistance breakpoint, whereas vancomycin MICs tend to remain more stable over a narrower range within the limits of susceptibility. CNS strains intermediately susceptible and even resistant not only to teicoplanin but also to vancomycin have, however, been isolated, most frequently from patients subjected to prolonged glycopeptide treatment. Laboratory detection of glycopeptide-resistant CNS may be problematic, mainly because susceptibility tests, particularly those for teicoplanin, are influenced by various technical factors, and agar diffusion tests may yield false susceptibility data. In studies with experimental glycopeptides, some molecules have exhibited improved in vitro activity compared with teicoplanin and vancomycin, but these encouraging microbiological findings have not usually been followed by in vivo trials. Stepwise and single-step exposure to teicoplanin and vancomycin has allowed stable clones for which glycopeptide MICs are increased to be obtained from susceptible CNS strains, particularly strains of Staphylococcus haemolyticus and Staphylococcus epidermidis. In these studies, resistance to teicoplanin was generally easier to obtain than resistance to vancomycin, and the levels of teicoplanin resistance were higher. Population studies have demonstrated the usually heterogeneous nature of glycopeptide resistance in CNS. Although glycopeptide-resistant CNS have been shown to differ in several features from their glycopeptide-susceptible counterparts, the exact mechanism of staphylococcal glycopeptide resistance remains unknown.     

Coagulase negative Staphylococci as causative agents of urinary tract infections-prevalence and resistance status in IGIMS,Patna

Previously considered solely as the laboratory contaminants and normal flora of skin in man, coagulase negative Staphylococci are now a major cause of nosocomial and opportunistic infections. This study was conducted at IGIMS, Patna to find out the coagulase negative Staphylococcus isolates from urine and their antimicrobial resistance. In a period of ten months, the relative frequency of main coagulase negative Staphylococci were as follows--Staphylococcus epidermidis-45.90%, Staphylococcus saprophyticus 34% and Staphylococcus haemolyticus-8.50%. others were 11.60%. most of the Staphylococcus saprophyticus was isolated from young female patients suffering from uncomplicated acute cystitis and Staphylococcus epidermidis was mainly from patients with indwelling catheters and complicated cases. Staphylococcus saprophyticus showed the highest sensitivities to all the antimicrobials whereas Staphylococcus haemolyticus had the highest resistance rates. 66.6% of staphylococcus epidermidis and 60% of staphylococcus haemolyticus were resistant to oxacillin, whereas only 10% of staphylococcus saprophyticus resistant and 90% were sensitive to it. As is in the emerging state, vancomycin resistance was very less but in future it may cause a major problem to treat these cases. So this area needs further exploration.     

In vitro activity of teicoplanin against gram-positive cocci

The glycopeptide susceptibility of 443 clinical isolates of gram-positive cocci collected from nine general hospitals in 1996 was determined according to the recommendations of the CA-SFM (the Antibiogram Committee of the French Society for Microbiology). In total, 234 isolates of Staphylococcus aureus, 84 isolates of coagulase-negative staphylococci (CNS), 98 enterococci and 27 streptococci were collected. The mecA gene confirming resistance to methicillin was found in 42.7% of S. aureus isolates and 51.2% of CNS isolates. No resistance to teicoplanin and vancomycin was found in S. aureus but four isolates of CNS had an MIC of teicoplanin > or = 8 mg/L. All isolates of Enterococcus faecalis tested were susceptible to both glycopeptides. This study confirms that teicoplanin has a very good in vitro activity against gram-positive cocci, isolated in France from nosocomial infections.     

Nosocomial spread of a Staphylococcus capitis strain with heteroresistance to vancomycin in a neonatal intensive care unit

A premature infant in a neonatal intensive care unit (NICU) developed a bloodstream infection caused by coagulase-negative staphylococci (CoNS) sensitive to vancomycin. The infection persisted for 3 weeks, despite therapy with vancomycin and replacement of all intravenous catheters. The neonate died due to necrotizing enterocolitis which developed during the ongoing sepsis. We screened this strain and 216 other strains of CoNS from cultures of blood obtained from neonates between 1997 and 2000 for heteroresistance to vancomycin. Forty-eight isolates, including the strain that caused ongoing sepsis, proved heteroresistant. All isolates were identified as Staphylococcus capitis and were identical, just as their resistant stable subcolonies were, when they were genetically fingerprinted by amplified-fragment length polymorphism analysis. The heteroresistant phenotype of this endemic strain was confirmed by population analysis. We conclude that heteroresistance to vancomycin occurs in S. capitis and might be the cause of therapeutic failures in NICUs. Moreover, heteroresistant strains can become endemic in such units.     

Clonal spread of Staphylococcus aureus heterogeneously resistant to vancomycin in a university hospital in Korea

Since vancomycin-intermediate Staphylococcus aureus (VISA) was first reported in Japan in 1997, there has been great concern that heterogeneous vancomycin-intermediate S. aureus (hetero-VISA) is the putative precursor of VISA. To investigate the prevalence, clinical significance, and molecular epidemiology of S. aureus with reduced susceptibility to vancomycin, all consecutive isolates of S. aureus isolated from clinical specimens from December 1998 to August 1999 at Asan Medical Center were screened for VISA and hetero-VISA by using brain heart infusion agar containing 4 microg of vancomycin/ml. Screen-positive isolates were confirmed by susceptibility testing and population analysis of subpopulations with reduced susceptibility to vancomycin. The isolates confirmed as hetero-VISA were typed by pulsed-field gel electrophoresis (PFGE). Medical records were reviewed to evaluate the clinical significance and risk factors for the acquisition of hetero-VISA. Of the 4,483 isolates that were tested, 53 were screen positive; no VISA was detected, but 24 isolates (0.54%) from 22 patients were hetero-VISA. All but two strains appeared to be clones of the Korean VISA strain, AMC11094, in the PFGE analysis. A total of 18 patients were in intensive care units, and 16 underwent major surgeries during the same admission. Only 10 of the 22 patients had previous methicillin-resistant S. aureus infections and 11 had previous vancomycin or teicoplanin therapy. Only 7 of the 22 patients from whom hetero-VISA strains were isolated were infected, and the remaining 15 patients were colonized. All seven infected patients were successfully treated with vancomycin. These results suggest that hetero-VISA can be treated with vancomycin, but the spread of hetero-VISA clonal to VISA is of concern, since many believe that VISA can arise from hetero-VISA, although this phenomenon was not observed in this study.     

StaphPlex system for rapid and simultaneous identification of antibiotic resistance determinants and Panton-Valentine leukocidin detection of staphylococci from positive blood cultures

Phenotypic methods take several days for identification and antimicrobial susceptibility testing of staphylococcal isolates after gram-positive cocci in clusters (GPCC) are observed in positive blood cultures. We developed and validated a StaphPlex system that amplifies and detects 18 gene targets simultaneously in 1 reaction for species-level identification of staphylococci, detection of genes encoding Panton-Valentine leukocidin (PVL), and antimicrobial resistance determinants of staphylococci. The StaphPlex system was compared to phenotypic methods for organism identification and antimicrobial resistance detection for positive blood culture specimens in which GPCC were observed. Among a total of 360 GPCC specimens, 273 (75.8%), 37 (10.3%), 37 (10.3%), 1 (0.3%), 3 (0.8%), and 9 (2.5%) were identified by StaphPlex as coagulase-negative Staphylococcus (CoNS), methicillin-resistant Staphylococcus aureus (MRSA), methicillin-susceptible S. aureus (MSSA), or mixed infections of CoNS and MRSA, CoNS and MSSA, or nonstaphylococci, respectively, with an overall accuracy of 91.7%. The 277 CoNS-containing specimens were further identified to the species level as containing 203 (73.3%) Staphylococcus epidermidis isolates, 10 (3.6%) Staphylococcus haemolyticus isolates, 27 (9.7%) Staphylococcus hominis isolates, 1 (0.4%) Staphylococcus lugdunensis isolate, and 36 (13.0%) other CoNS isolates, with an overall accuracy of 80.1% compared to an API STAPH test and CDC reference identification. Numerous very major errors were noticed when detection of aacA, ermA, ermC, tetM, and tetK was used to predict in vitro antimicrobial resistance, but relatively few major errors were observed when the absence of these genes was used to predict susceptibility. The StaphPlex system demonstrated 100% sensitivity and specificity, ranging from 95.5% to 100.0% when used for staphylococcal cassette chromosome mec typing and PVL detection. StaphPlex provides simultaneous staphylococcal identification and detection of PVL and antimicrobial resistance determinants within 5 h, significantly shortening the time needed for phenotypic identification and antimicrobial susceptibility testing.     

Persistent strains of coagulase-negative staphylococci in a neonatal intensive care unit: virulence factors and invasiveness

Coagulase-negative staphylococci (CoNS) are the major cause of nosocomial bacteraemia in neonates. The aim of this study was to investigate whether persistent strains of CoNS possess specific bacterial characteristics as compared with sporadic non-cluster isolates. In total, 180 blood culture isolates (95 contaminants and 85 invasive isolates) obtained from a single neonatal unit over a 12-year period were studied. Pulsed-field gel electrophoresis (PFGE) identified 87 persistent CoNS strains (endemic clones). The two largest PFGE clusters belonged to a single clonal complex according to multilocus sequence typing. Patients colonised or infected with endemic clones were of lower gestational age than those infected with non-cluster strains. One Staphylococcus haemolyticus cluster appeared to selectively colonise and infect the most extreme pre-term infants. Endemic clones were characterised by high levels of antibiotic resistance and biofilm formation. All 51 isolates belonging to the two largest PFGE clusters were ica operon-positive. Genes encoding Staphylococcus epidermidis surface protein B and the production of phenol-soluble modulins (PSMs) were also more prevalent among endemic clones than among non-cluster strains. However, endemic clones were not more prevalent among invasive isolates than among contaminants. These findings indicate that multiple selective factors, including antibiotic resistance, biofilm formation, surface proteins with adhesive properties, and PSMs regulated by agr, increase the ability of CoNS to persist in a hospital environment. It may be more prudent, when searching for new therapeutic targets, to focus on ubiquitous components of CoNS instead of putative virulence factors that do not clearly contribute to increased invasive capacity.