Linezolid-resistant Staphylococci Research Articles

Characterization of linezolid- and methicillin-resistant coagulase-negative staphylococci in a tertiary hospital in China

BackgroundRecently, linezolid-resistant staphylococci have become an emerging problem worldwide. Understanding the mechanisms of resistance, molecular epidemiology and transmission of linezolid-resistant CoNS in hospitals is very important.MethodsThe antimicrobial susceptibilities of all isolates were determined by the microdilution method. The resistance mechanisms and molecular characteristics of the strains were determined using whole-genome sequencing and PCR.ResultsAll the strains were resistant to oxacillin and carried the mecA gene; 13 patients (36.1%) had prior linezolid exposure. Most S. epidermidis and S. hominis isolates were ST22 and ST1, respectively. MLST typing and evolutionary analysis indicated most linezolid-resistant CoNS strains were genetically related. In this study, we revealed that distinct CoNS strains have different mechanisms of linezolid resistance. Among ST22-type S. epidermidis, acquisition of the T2504A and C2534T mutations in the V domain of the 23 S rRNA gene, as well as mutations in the ribosomal proteins L3 (L101V, G152D, and D159Y) and L4 (N158S), were linked to the development of linezolid resistance. In S. cohnii isolates, cfr, S158Y and D159Y mutations in the ribosomal protein L3 were detected. Additionally, emergence of the G2576T mutation and the cfr gene were major causes of linezolid resistance in S. hominis isolates. The cfr gene, G2576T and C2104T mutations, M156T change in L3 protein, and I188S change in L4 protein were found in S. capitis isolates.ConclusionThe emergence of linezolid-resistant CoNS in the environment is concerning because it involves clonal dissemination and frequently coexists with various drug resistance mechanisms.

CHROMAgar™ LIN-R as an efficient screening tool to assess the prevalence of linezolid-resistant enterococci in German hospital patients-a multicentre study approach, 2021-2022.

In recent years, an increasing number of linezolid-resistant enterococci (LRE) was recognized at the German National Reference Centre (NRC) for Enterococci. National guidelines on infection prevention recommend screening for LRE in epidemiologically linked hospital settings without referring to a reliable and rapid diagnostic method. Since 2020, CHROMAgar™ provide a chromogenic linezolid screening agar, LIN-R, suitable to simultaneously screen for linezolid-resistant staphylococci and enterococci. To assess the applicability of CHROMAgar™ LIN-R in clinical settings for detecting LRE directly from patient material and to infer prevalence rates of LRE amongst German hospital patients. During the 3-month trial period, clinical samples were plated on CHROMAgar™ LIN-R. Antimicrobial susceptibility testing was performed using VITEK2 or disc diffusion. At the NRC, linezolid resistance was determined by broth microdilution, multiplex-PCR for cfr/optrA/poxtA and by a restriction-based assay for 23S rDNA mutations. The 12 participating study sites used 13 963 CHROMAgar™ LIN-R plates during the study period. Of 442 presumptive LRE, 192 were confirmed by phenotypic methods. Of these, 161 were received by the NRC and 121 (75%) were verified as LRE. Most of LR-E. faecium 53/81 (65%) exhibited a 23S rRNA gene mutation as the sole resistance-mediating mechanism, whereas optrA constituted the dominant resistance trait in LR-E. faecalis [39/40 (98%)]. Prevalence of LRE across sites was estimated as 1% (ranging 0.18%-3.7% between sites). CHROMAgar™ LIN-R represents a simple and efficient LRE screening tool in hospital settings. A high proportion of false-positive results demands validation of linezolid resistance by a reference method.

Distribution and Antibiotic Resistance Analysis of Blood Culture Pathogens in a Tertiary Care Hospital in China in the Past Four Years.

This study aimed to investigate the antibiotic resistance patterns and clinical distribution of blood culture-positive isolates at Suining Central Hospital between 2018 and 2021. The findings of this study can provide a basis for ensuring rational and effective use of antibiotic therapy in clinical settings. This retrospective study analyzed the data of 3660 non-repeating strains that tested positive for clinical blood culture, collected from the microbiology laboratory of Suining Central Hospital between January 2018 and December 2021. The identification of bacterial species and their antibiotic resistance patterns were analyzed. The study found that 76.7% of the bacterial strains identified were Gram-negative bacteria, while 23.3% were Gram-positive bacteria. Escherichia coli (44.8%), Klebsiella spp. (19.2%), Staphylococcus aureus (9.2%), Enterococcus spp. (5.3%), and Enterobacter spp. were the top five bacterial ratios observed. These bacteria were detected most frequently in the Digestion Center, intensive care unit (ICU), Neurology Center, Urology Department, and Hematology Department. Among the Staphylococcus spp., methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant coagulase-negative staphylococci (MRCNS) were detected at rates of 39.3% and 71.8%, respectively. However, no vancomycin- or linezolid-resistant staphylococci were identified. Enterococcus faecalis showed higher susceptibility to most antibiotic than Enterococcus faecium, except for tetracycline. The resistance rates of E. coli and Klebsiella spp. to meropenem and imipenem were low, but the resistance rates for other antibiotic were above 40%. The results of this study show a rising incidence of bacterial antibiotic resistance in positive blood culture specimens at Suining Central Hospital. Clinicians should carefully consider the importance of blood culture antibiotic susceptibility testing to ensure effective treatment. The Department of Microbiology at Suining Central Hospital should regularly analyze the distribution of pathogenic bacteria and antibiotic resistance in blood cultures to ensure the most effective treatment possible.

Evaluation of CHROMagar™ LIN-R for the Screening of Linezolid Resistant Staphylococci from Positive Blood Cultures and Nasal Swab Screening Samples.

The increasing number of nosocomial pathogens with resistances towards last resort antibiotics, like linezolid for gram positive bacteria, leads to a pressing need for screening and, consequently, suitable screening media. Some national guidelines on infection prevention (e.g., in Germany) have already recommended screening for linezolid-resistant bacteria, despite an accurate screening medium that was not available yet. In this study, we analyzed the performance and reliability of the first commercial chromogenic medium, CHOMagar™ LIN-R, for screening of linezolid-resistant gram-positive isolates. Thirty-four pure bacterial cultures, 18 positive blood cultures, and 358 nasal swab screening samples were tested. This medium efficiently detected linezolid-resistant S. epidermidis isolates from pure bacterial cultures and from positive blood cultures with a high sensitivity (100%) and specificity (100%). Among the 358 nasal swab screening samples prospectively tested, 10.9% were cultured with linezolid-resistant isolates (mostly S. epidermidis). Of note, slight growth was observed for 7.5% samples with linezolid-susceptible isolates of S. epidermidis (n = 1), S. aureus (n = 1), Enterococcus faecalis (n = 4), Lactobacillus spp. (n = 3), gram negatives (n = 18). Moreover, few Candida spp. also cultured on this medium (1.4%).

Linezolid-Resistant Staphylococcus haemolyticus: Emergence of G2447U and C2534U Mutations at the Domain V of 23Sr RNA Gene

Background: Linezolid an oxazolidinone drug available in both parenteral and oral formulations has emerged as a novel alternative to vancomycin and other second-generation drugs for the treatment of infections from gram-positive cocci. Clinical isolates of linezolid-resistant staphylococci and enterococci were reported. Since then, linezolid-resistant strains have become an increasing problem worldwide. The most frequently reported mechanisms of linezolid resistance include the mutation in 23S ribosomal nucleic acid and presence of cfr gene. Methicillin-resistant coagulase-negative staphylococci (MR-CoNS) and vancomycin-resistant Enterococcus (VRE) have become a worrisome clinical problem. Objective: Therefore, we aimed to determine the distribution of linezolid-resistant strains in an inpatient setting of a tertiary-care hospital in India and to evaluate the resistance mechanisms among these isolates. In addition, the clonal diversity of the isolates was determined by pulsed-field gel electrophoresis (PFGE). Methods: The distribution, clonal diversity, and resistance mechanism of linezolid resistant-Staphylococcus haemolyticus (LRSH) strains were determined. The isolates were identified by MALDI-TOF. The mechanism of resistance was determined by sequence analysis of the domain V of 23SrRNA and screening for cfr gene. Clonal relatedness was defined by PFGE. Results: In total, 13 LRSH isolates were recovered from pus specimens. The 13 LRSH strains isolated had an linezolid MIC of 256 g/mL. Sequencing results revealed G2576T mutations in 7 (53.8%), G2447U in 4 (30.7%) and C2534U in 1 (7.6%) isolate of S. haemolyticus. One isolate of S. haemolyticus showed 2 simultaneous mutations (G2576T and G2447U) in the domain V region of 23Sr RNA gene. PFGE of the LR-SH isolates revealed the presence of 11 clones. Of the 11 clones, clones I and II had 2 isolates each. Isolates of clone I exhibited a band pattern identical with the previous isolates of LRSH isolated from the orthopedic unit. Similarly, isolates of clone II also shared the same band pattern with the previous LRSH isolates from the dermatology unit of our center. Conclusions: This study highlights the importance of continuous monitoring of vigilance of linezolid resistance in staphylococci. Rationalizing the use of linezolid and implementing methods to control the spread of hospital clones is of paramount importance to prevent further dissemination of these strains.Funding: NoneDisclosures: None

Antibiogram of staphylococcal isolates with special reference to inducible clindamycin resistance

Introduction and Objectives: Expression of inducible clindamycin resistance (iMLSB) by Staphylococcal isolates limits the effective use of clindamycin resulting in treatment failure. The present study was aimed to estimate the prevalence of iMLSB. Materials and Methods: Fifty Staphylococci isolated from various clinical specimens were identified by standard microbiological methods and were subjected to antibiotic susceptibility testing by Kirby-Bauer disc diffusion method and D test. Methicillin resistance was detected by cefoxitin disc diffusion method. Results: Staphylococcus aureus constituted 62% and co-agulase negative Staphylococci 38% of total Staphylococcal isolates. Methicillin resistance was noted in 27(54%) of isolates. Seventeen (34%) isolates were sensitive to both erythromycin and clindamycin. Among the resistant phenotypes, constitutive clindamycin resistance (cMLSB), Macrolide Streptogramin B resistance (MS) phenotype and iMLSB was found in 16 (32%), 10 (20%) and in 7(14%) isolates respectively. Majority of the Staphylococcal isolates showed resistance to penicillin (93.6%) followed by ciprofloxacin (79.6%) and erythromycin (66%). Linezolid resistance was observed in 8% of isolates. None of the isolates were found resistant to vancomycin. Interpretation and Conclusion: Routine testing for iMLSB for all clinical isolates of Staphylococci should be carried out to prevent failure of clindamycin therapy. Isolation of linezolid resistant Staphylococci is an alarming sign and a matter of concern. Keywords: D test, Inducible clindamycin resistance, Methicillin resistance, Staphylococci.

Five-Year Summary of In Vitro Activity and Resistance Mechanisms of Linezolid against Clinically Important Gram-Positive Cocci in the United States from the LEADER Surveillance Program (2011 to 2015).

This report describes linezolid susceptibility testing results for 6,741 Gram-positive pathogens from 60 U.S. sites collected during 2015 for the LEADER Program. In addition, the report summarizes linezolid in vitro activity, resistance mechanisms, and molecular typing obtained for 2011 to 2015. During 2015, linezolid showed potent activity in testing against Staphylococcus aureus, inhibiting >99.9% of 3,031 isolates at ≤2 µg/ml. Similarly, linezolid showed coverage against 99.2% of coagulase-negative staphylococci, 99.7% of enterococci, and 100.0% of Streptococcus pneumoniae, virdans group, and beta-hemolytic streptococcus isolates tested. The overall linezolid resistance rate remained a modest <1% from 2011 to 2015. Staphylococci, especially Staphylococcus epidermidis, showed a range of linezolid resistance mechanisms. Increased annual trends for the presence of cfr among Staphylococcus aureus isolates were not observed, but 64.3% (9/14) of the isolates with decreased susceptibility (MIC, ≥4 µg/ml) to linezolid carried this transferrable gene (2011 to 2015). The cfr gene was detected in 21.9% (7/32) of linezolid-resistant staphylococci other than S. aureus from 2011 to 2015. The optrA gene was noted in half (2/4) of the population of linezolid-nonsusceptible Enterococcus faecalis isolates from 2011 to 2015, while linezolid-nonsusceptible Enterococcus faecium isolates showed alterations predominantly (16/16) in the 23S rRNA gene (G2576T). This report confirms a long record of linezolid activity against Gram-positive isolates in the United States since regulatory approval in 2000 and reports the oxazolidinones evolving resistance mechanisms.

Oritavancin Activity Tested against Molecularly Characterized Staphylococci and Enterococci Displaying Elevated Linezolid MIC Results.

Oritavancin (MIC50/90, 0.03/0.06 to 0.12 μg/ml) had potent activity against linezolid-resistant staphylococci, as well as Enterococcus faecalis and Enterococcus faecium (oritavancin MIC50/90, 0.015/0.12 μg/ml against both species). All linezolid-resistant isolates were inhibited by oritavancin at ≤0.12 μg/ml. These results confirmed the absence of cross-resistance between linezolid and oritavancin in staphylococci and enterococci.

In vitro activity of tedizolid against linezolid-resistant staphylococci and enterococci

The tedizolid MIC of 27 clinical isolates of linezolid-resistant staphylococci and enterococci was determined. Tedizolid MICs were ≥1μg/mL and were 4- to 32-fold lower than those of linezolid. Linezolid resistance mechanisms included G2576T 23S rRNA gene and rplC and rplD mutations.

Linezolid-resistant clinical isolates of meticillin-resistant coagulase-negative staphylococci and Enterococcus faecium from China

Seventeen meticillin-resistant coagulase-negative staphylococci (MRCoNS), including ten Staphylococcus capitis, four Staphylococcus cohnii, two Staphylococcus haemolyticus and one Staphylococcus sciuri, and an Enterococcus faecium isolate with various levels of linezolid resistance were isolated from intensive care units in a Chinese hospital. PFGE indicated that the four S. cohnii isolates belonged to a clonal strain, and that nine of the S. capitis isolates were indistinguishable (clone A1) and the other one was closely related (clone A2). A G2576T mutation was identified in domain V of the 23S rRNA gene in the E. faecium isolate. Besides the G2576T mutation, a novel C2104T mutation was detected in the nine clone A1 S. capitis isolates. The cfr gene was detected in all the staphylococci except an S. sciuri isolate, whose 23S rRNA gene contained the G2576T mutation. There was a clonal dissemination of linezolid-resistant MRCoNS in intensive care units of our hospital, and this is the first report, to our knowledge, of linezolid-resistant staphylococci and enterococci in China.

In vitro activity of tedizolid (TR-700) against linezolid-resistant staphylococci

To compare the activity of tedizolid (formally known as torezolid and TR-700) with that of 15 agents against a collection of linezolid-resistant staphylococci (164 coagulase-negative staphylococci and 5 Staphylococcus aureus). Antimicrobial susceptibility tests were performed using the broth microdilution method following the recommendations of the CLSI. All isolates were susceptible to vancomycin and tigecycline. Based on the MIC(90) values, the potency of tedizolid against coagulase-negative staphylococci was >16-fold greater than that of linezolid. Tedizolid retained activity against most of the linezolid-resistant staphylococci tested, including multidrug-resistant isolates with elevated linezolid MICs (32 to >128 mg/L). Of the isolates, 79.2% and 31.4% were inhibited by tedizolid at ≤ 4 mg/L and ≤ 2 mg/L, respectively. The results of this study confirm the activity of tedizolid against linezolid-resistant staphylococci. This new oxazolidinone could have an important role as a potential therapeutic agent against multidrug-resistant staphylococci.

Diseminación nosocomial de Staphylococcus hominis resistente al linezolid en dos hospitales de Mallorca

Since March 2008, several linezolid and teicoplanin-resistant Staphylococcus hominis (S. hominis) isolates have been recovered from patients admitted to the two major hospitals on the island of Majorca, Spain. For this reason, a study was conducted to determine the molecular epidemiology of these isolates and the mechanism of linezolid resistance. The molecular epidemiology study was performed by pulsed-field gel electrophoresis (PFGE) analysis, after digestion with ApaI. Linezolid resistance mechanisms were evaluated by PCR amplification of a fragment of the domain V of the 23S rRNA gene (followed by sequencing) and cfr gene. From March 2008 to February 2009, 15 linezolid and teicoplanin-resistant S. hominis isolates were recovered from 14 patients. All of them, except one, were hospitalised in the intensive care units of either of the two institutions. Isolates were obtained mainly from blood cultures (9). The majority of infected patients (12 of 15 infectious episodes, 80.0%) had received courses of linezolid prior to detection of the resistant isolate. PFGE analysis revealed the presence of a unique clone among linezolid resistant S. hominis isolates. The G2576T mutation was detected in all the linezolid resistant strains. None of the resistant isolates showed a positive PCR for the cfr gene. All of the isolates were also resistant to penicillin, oxacillin, trimethoprim-sulfamethoxazole, ciprofloxacin, levofloxacin, and tobramicin; whereas all of them were susceptible to erythromycin, tetracycline, gentamicin, and daptomycin. The MIC of vancomycin was 4μg/ml for all the strains. The detection of linezolid resistant Staphylococci highlights the need to rationalise the use of linezolid, and maintain an active surveillance of its resistance to preserve the clinical usefulness of this antimicrobial.

Evaluation of the activity of fusidic acid tested against contemporary Gram-positive clinical isolates from the USA and Canada

We evaluated the antimicrobial activity of fusidic acid (CEM-102) against 1140 clinical strains of Gram-positive bacteria obtained from patients with bacteraemia or skin and skin-structure infections collected in more than 30 medical centres in the USA and Canada over a 10-year period (1997–2006). Fusidic acid was very active against meticillin-susceptible Staphylococcus aureus (MSSA), meticillin-resistant S. aureus (MRSA) and coagulase-negative staphylococci (CoNS), with MIC 90 values (minimum inhibitory concentration encompassing 90% of isolates tested) at 0.12 μg/mL for US strains of MSSA, MRSA and CoNS and 0.25 μg/mL for Canadian strains of MSSA and MRSA. A progressive increase in fusidic acid resistance was observed among Canadian strains of S. aureus (12.2% in 2005–2006) and among Canadian strains of CoNS. In contrast, no fusidic acid resistance was detected among US S. aureus strains and only 1.5% among CoNS. Fusidic acid was equally active against community-acquired MRSA and linezolid-resistant staphylococci. Fusidic acid exhibited equal or greater potency against staphylococci compared with vancomycin, daptomycin and linezolid.

Polyphyletic Emergence of Linezolid-Resistant Staphylococci in the United States

Since the year 2000, linezolid has been used in the United States to treat infections caused by antimicrobial-resistant Gram-positive cocci. At present, linezolid-resistant (Linr) Staphylococcus aureus and Staphylococcus epidermidis strains are rare and the diversity of their genetic backgrounds is unknown. We performed sequence-based strain typing and resistance gene characterization of 46 Linr isolates that were collected from local and national sources between the years 2004 and 2007. Resistance was found to occur in at least three clonal complexes (CCs; lineages) of S. aureus and in at least four subclusters of a predominant, phylogenetically unstable CC of S. epidermidis. New candidate resistance mutations in 23S rRNA and the L4 riboprotein were identified among the S. epidermidis isolates. These findings suggest that linezolid resistance has emerged independently in multiple clones of S. aureus and with a variety of ribosomal mutations in multiple clones of S. epidermidis.

Reduced biofilm production associated with increasing linezolid MICs among linezolid-resistant staphylococci

Sir, To follow-up our previous finding of reduced polystyrene adherence accompanying the acquisition of linezolid resistance in a clinical bloodstream isolate of methicillin-resistant Staphylococcus aureus (MRSA), we evaluated the polystyrene adherence properties of linezolid-resistant staphylococci obtained from our institution as well as isolates obtained from multiple medical centres throughout the USA collected as part of the LEADER programme. Eleven coagulase-negative staphylococci from one medical centre identified by the Microscan susceptibility method (10 Staphylococcus epidermidis and 1 Staphylococcus haemolyticus) and 50 staphylococci identified from the LEADER programme (33 S. epidermidis, 7 S. aureus, 2 Staphylococcus caprae, 2 Staphylococcus capitis, 2 Staphylococcus warnerii, 1 Staphylococcus cohnii, 1 S. haemolyticus, 1 Staphylococcus saprophyticus and 1 non-speciated) with linezolid MICs .4 mg/L were available for susceptibility testing by broth microdilution methods of the CLSI. The biofilm formation phenotype was assayed by polystyrene adherence assays as previously described. After approval was obtained from the internal review board for research, chart review of 13 patients at our hospital (11 cases whose isolates were used in the analysis plus 2 additional cases whose isolates were not available) with linezolid-resistant coagulase-negative staphylococci isolated from blood cultures was performed to determine patient age, sex, hospital ward, exposure to linezolid, hospital day at time of isolation, patient co-morbidities and whether the blood culture isolate fulfilled CDC criteria for catheter-related bloodstream infection. The patients had a median age of 60 years (range 18–78), 62% of the patients were male and the patients were hospitalized for a median of 37 days (range 6–217) at the time of isolation of the linezolid-resistant coagulase-negative staphylococci. Fifty-four percent received prior linezolid during the hospitalization but none of the patients was on linezolid at the time cultures were obtained. Of the 13 patients, 2 isolates (15%) were deemed clinically significant by fulfilling CDC criteria for catheter-related bloodstream infection. PFGE demonstrated a predominant clonal type, two closely related types and three unique clonal types. Regression tree modelling revealed a dichotomous inverse relationship between polystyrene adherence and linezolid MIC for the 11 isolates from a single medical centre [P1⁄40.02, Kruskal–Wallis analysis of variance (ANOVA)]. This relationship was examined for reproducibility among the 50 isolates from the LEADER programme (Figure 1). The break occurred at a linezolid MIC of 32 mg/L, above which isolates showed significantly reduced polystyrene adherence (P1⁄40.004, Kruskal– Wallis ANOVA). Evaluation of the subset of 43 LEADER coagulase-negative staphylococci (excluding the 7 S. aureus) demonstrated consistency of this relationship (P1⁄40.02, Kruskal–Wallis ANOVA). Linezolid resistance has until recently remained relatively uncommon among staphylococci. A recent report of a 4.4% prevalence of linezolid resistance among coagulase-negative staphylococci at a tertiary medical centre raises concern. In this report, we identified 13 patients from whom these isolates were obtained from blood cultures, although only 2 (15%) represented clinically relevant isolates as aetiologies in catheter-associated bacteraemia. The majority of patients (54%) had received linezolid during the hospitalization, lower than the 76% seen in the prior report. This suggests that linezolid-resistant staphylococci may reside as part of the indigenous nosocomial flora that can subsequently colonize hospitalized patients. Further studies are required to examine the point prevalence of such isolates at individual centres and whether linezolid utilization can impact this prevalence. An evaluation of 11 linezolid-resistant coagulase-negative staphylococci from a single institution and 50 linezolid-resistant staphylococci acquired from multiple medical centres around the USA demonstrated a dichotomous inverse relationship between linezolid MIC and polystyrene adherence. A linezolid MIC 32 mg/L was associated with reduced polystyrene adherence when compared with linezolid-resistant staphylococci at the lower end of the resistant MIC range (MIC 8–16 mg/L). This finding may represent another example where a specific virulence property of bacteria may be attenuated by the acquisition of antimicrobial resistance, although the mechanism behind this observation is unknown and not immediately obvious. This relationship may provide at least a partial explanation as to why linezolid-resistant staphylococci remain relatively rare among clinical isolates. This study provides a preliminary assessment of the epidemiological, microbiological and physiological features of linezolid resistance in staphylococci. A more thorough epidemiological evaluation and prevalence study of linezolid resistance among coagulase-negative staphylococci in hospitals is warranted. Studies evaluating the genetic background of these strains and their mechanisms of linezolid resistance are currently in progress in our laboratories. Because of potential future implications on the treatment of staphylococcal infections mediated Journal of Antimicrobial Chemotherapy doi:10.1093/jac/dkp324

Linezolid-ResistantStaphylococcus cohnii,Greece

To the Editor: Since 2003, linezolid has typically been used to treat infections caused by multidrug-resistant gram-positive cocci such as vancomycin-resistant Enterococcus faecium and methicillin-resistant Staphylococcus aureus (1). In Greece, a major problem is nosocomial dissemination of vancomycin-resistant enterococci. Use of linezolid for the treatment of such infections led to the emergence of linezolid–vancomycin resistant E. faecium; however, linezolid resistance of staphylococci is still relatively low in this country (2). We describe an outbreak caused by a linezolid-resistant S. cohnii in an intensive care unit (ICU) in Greece. From July through October 2007, nonrepetitive coagulase-negative staphylococci that exhibited resistance to linezolid were isolated from blood cultures from 4 separate patients hospitalized in the ICU at Sismanoglion General Hospital of Athens, a 450-bed tertiary care hospital. The ICU is a 10-bed, level II unit, comprising 2 rooms with 1 bed each and 2 rooms with 4 beds each. Each isolate was recovered in 2 of 2 blood culture sets per patient, indicating true bacteremia. The demographic and clinical information for the patients is described in the Table. The mean duration of time preceding linezolid therapy was 22 days. Table Clinical characteristics of 4 patients from whom linezolid-resistant Staphylococcus cohnii was isolated, Greece, 2007* Isolates were first identified to the species level by using an API Staph system (bioMerieux, la Balme les Grottes, France) and a molecular method based on the tuf gene followed by sequencing analysis (3). Susceptibility testing for various antimicrobial agents was performed by disk diffusion and using Clinical Laboratory Standards Institute criteria; susceptibilities were interpreted according to Institute guidelines (4). In addition, MICs to oxacillin, vancomycin, teicoplanin, quinupristin-dalfopristin, linezolid, daptomycin, and tigecycline were determined by Etest (AB Biodisk, Solna, Sweden) according to manufacturer’s instructions. Resistance genes mecA, vat, vga, erm, aac(6´)-Ie+aph(2″), ant(4′)-Ia, and aph(3′)-IIIa, as markers for resistance to β-lactams, dalfopristin, macrolides, and aminoglycosides, were identified by PCR as previously reported (5,6). The presence of G2576T in domain V of the 23S rRNA, which is mainly associated with linezolid resistance in clinical isolates, was detected by using PCR and digestion of the product with NheI (2). The number of mutated versus nonmutated alleles was determined as described by Pillai et al. (7). In addition, isolates were examined for the presence of the cfr gene, which was found to be correlated with linezolid resistance in some coagulase-negative staphylococci and for mutations of ribosomal protein L4, L22 genes (8,9). Clonality of isolates was assessed by pulsed-field gel electrophoresis (PFGE) after digestion of chromosomal DNA with SmaI (2). The molecular method identified the isolates as S. cohnii subsp. ureolyticus. The API Staph system has correctly identified 2 of them; the remaining 2 isolates were falsely characterized as S. xylosus. According to disc diffusion test results, the isolates were resistant to cefoxitin, oxacillin, penicillin, rifampin, quinupristin-dalfopristin, erythromycin, clindamycin, fusidic acid, tobramycin, gentamicin, and linezolid. MICs were linezolid 32, oxacillin 256, quinupristin-dalfopristin 8, vancomycin 2, teicoplanin 2, tigecycline 0.125, and daptomycin 0.5 mg/L. Molecular methods detected the following resistance genes: mecA, ermA, aac(6´)-Ie+aph(2″), and aph(3´)-IIIa. The isolates, despite their resistance to streptogramins, were negative for vat and vgaA genes. In addition, all isolates carried the G2576T mutation and had 4 of 5 mutated alleles. No isolate carried the cfr gene or any mutation on ribosomal protein L4 and L22 genes. PFGE results indicated that all isolates were clonally related, belonging to the same clone. Outbreaks caused by linezolid-resistant staphylococci are rare worldwide (10); in Sismanoglion Hospital, before the outbreak period, no linezolid-resistant staphylococci and enterococci had been isolated. However, in the ICU, a statistically significant increase in the use of linezolid was observed in 2004 and in 2007 (0.58 vs. 1.34 defined daily doses/100 patient-days, respectively); heavy use of linezolid may have created substantial selection pressure in favor of the linezolid-resistant isolates. The 4 patients were treated in the same room by the same personnel; thus, a potential explanation for this outbreak is patient-to-patient transmission of linezolid-resistant strains on the hands of healthcare personnel. However, cultures of ICU personnel (nasal cavity and hands) grew only methicillin-resistant S. aureus and methicilllin-resistant S. epidermidis. In addition, environmental samples taken from the beds and the equipment of these patients were negative for S. cohnii. Strict control measures were taken (e.g., isolation of infected patients, increased environmental cleaning, and reinforcement of proper glove and gown use and hand disinfection with alcohol gel), and the outbreak strain was not recovered from other patients in the ICU or in other departments of the hospital after the initial outbreak. In conclusion, to avoid spread of staphylococcal resistance in ICUs, measures such as hand hygiene and adequate central venous catheter handling should be taken, and policies regarding antimicrobial drug use must be applied.

Spectrum and antibiotic resistance of uropathogens from hospitalised patients with urinary tract infections: 1994–2005

From 1994–2005, all uropathogens cultured from the urine of hospitalised urological patients were identified and their sensitivity was tested against the most important antibiotics for the treatment of urinary tract infection (UTI). Duplicate isolates were eliminated. The following results were obtained: (i) there was no general trend of increase in resistance; (ii) certain uropathogens developed resistance to some antibiotics; (iii) vancomycin- or linezolid-resistant staphylococci or enterococci did not play a role; (iv) the lowest overall rates of resistance were found with piperacillin/tazobactam; and (v) ciprofloxacin and trimethoprim/sulfamethoxazole showed the next favourable overall activity. Adequate initial antibiotic therapy is critical in the treatment of severe UTI. Therefore, ongoing surveillance of antibiotic resistance must be performed in every institution. Additionally, it reflects antibiotic and hospital infection policies in a defined department or institution.

Potential utility of a peptide deformylase inhibitor (NVP PDF-713) against oxazolidinone-resistant or streptogramin-resistant Gram-positive organism isolates.

To evaluate the potency of a novel peptide deformylase inhibitor, NVP PDF-713, against Gram-positive organisms having resistances to linezolid or quinupristin/dalfopristin. A total of 45 strains from three genera (six species groups) were tested by reference broth microdilution methods. The mechanism of resistance to the oxazolidinone was determined by sequencing of the gene encoding the ribosomal target. NVP PDF-713 retained activity against linezolid-resistant staphylococci (MIC range 0.25-2 mg/L), Streptococcus oralis (MIC 0.5 mg/L), Enterococcus faecalis (MIC range 2-4 mg/L) and Enterococcus faecium (MIC range 0.5-4 mg/L). Quinupristin/dalfopristin-resistant E. faecium (MIC range 1-2 mg/L) and staphylococci (MIC range 0.12-2 mg/L) were also inhibited by NVP PDF-713. Many (10 of 13 strains) of the linezolid-resistant enterococci were resistant to vancomycin and these clinical strains had a G2576U ribosomal target mutation. NVP PDF-713 appears to be a promising clinical candidate among the peptide deformylase inhibitors for the treatment of infections caused by Gram-positive organisms that possess resistances to oxazolidinones or streptogramin combinations.