Screening for resistant gram-negative microorganisms to guide empiric therapy of subsequent infection.

OBJECTIVE: To define the potential of resistant gram-negative colonization surveillance to predict etiology of subsequent infection and improve adequacy of empiric antimicrobial treatment. DESIGN: Retrospective cohort study. SETTING: A mixed medical-surgical six-bed intensive care unit (ICU), from November 2003 to December 2006. PATIENTS: All patients having at least one episode of ventilator-associated pneumonia (VAP) or bloodstream infection (BSI) caused by a resistant gram-negative pathogen during the study period. INTERVENTIONS: Colonization surveillance of the respiratory tract and gastrointestinal tract was systematically performed in all ICU patients. Tracheal aspirates were obtained twice weekly and rectal swabs once weekly. Both tracheal and rectal samples were cultured in antibiotic-enriched media (containing ceftazidime, ciprofloxacin, imipenem or piperacillin/tazobactam), to focus on resistant gram-negative pathogen isolation. MEASUREMENTS AND RESULTS: Colonization concordance between resistant, gram-negative pathogens of infectious episodes and previous, recent (

Colistin susceptibility testing by Etest and disk diffusion methods.

The accuracy of disk susceptibility methods for colistin against 778 bacterial pathogens was evaluated in comparison with Etest using interpretive criteria available from the Clinical and Laboratory Standards Institute (CLSI). Colistin exhibited excellent activity against Acinetobacter baumannii and Escherichia coli isolates (minimum inhibitory concentration for 90% of the organisms (MIC(90))=0.5 mg/L), whilst it was less active both against Enterobacter spp. and Klebsiella pneumoniae (MIC for 50% of the organisms (MIC(50))=0.5 mg/L, MIC(90)=16 mg/L). Colistin also showed good activity against Pseudomonas aeruginosa (MIC(90)=2 mg/L, MIC(50)=1 mg/L) but poor activity against Stenotrophomonas maltophilia (MIC(50)=8 mg/L, MIC(90)=128 mg/L). Only 0.8% of minor errors were observed between the studied methods for P. aeruginosa isolates when the CLSI criteria were applied. All A. baumannii isolates with a zone diameter < or =12 mm were resistant and those with a zone diameter > or =14 mm were susceptible according to MIC breakpoints established by the CLSI. Among nine isolates exhibiting a zone diameter of 13 mm, one was resistant to colistin (MIC=8 mg/L) and eight isolates were susceptible (MIC=0.5 mg/L). Applying a MIC breakpoint of < or =2 mg/L for susceptibility in Enterobacteriaceae, all isolates with a zone diameter > or =14 mm were susceptible, whilst all isolates with a zone diameter < or =11 mm were resistant. Among isolates with zone diameters of 12-13 mm, 59% were characterised as susceptible. Major errors were observed only in K. pneumoniae isolates at a rate of 0.8%. The poor agar diffusion characteristics of colistin limit the predictive accuracy of the disk diffusion test and consequently values of 12-13 mm should be confirmed with MIC determination by Etest or broth dilution method.

Epidemiology and molecular characterisation of metallo-ß-lactamase-producing Enterobacteriaceae in a university hospital Intensive Care Unit in Greece.

The molecular epidemiology of VIM-producing Enterobacteriaceae isolated at the beginning of an epidemic in the Intensive Care Unit (ICU) of a university hospital in Athens, Greece, was studied. All Gram-negative organisms isolated from March 2004 to November 2005 positive for metallo-ß-lactamase (MBL) production were submitted to polymerase chain reaction (PCR) and sequencing, to repetitive sequence-based PCR (Rep-PCR) for molecular typing, and to S1 nuclease digestion for plasmid DNA characterisation. Conjugation experiments and isoelectric focusing were performed to identify co-existing ß-lactamases. Amongst 23 patients, 12 suffered one or more clinical infections. Eighty-two isolates representing one isolate per clone, source and ICU patient were studied, including Klebsiella pneumoniae (77), Enterobacter cloacae (2), Citrobacter freundii (1) and Pseudomonas aeruginosa (2). High clonal diversity was detected amongst the K. pneumoniae, with 10 distinct clones identified. Conjugation was successful in 54.5% of K. pneumoniae, and five different-sized plasmids were detected. All K. pneumoniae and both E. cloacae isolates shared the same bla(VIM-1)-containing class 1 integron structure also carrying aacA7, dhfrI and aadA1 gene cassettes. The C. freundii isolate carried a different integron that included bla(VIM-1) and aac(6')-IIc. Both P. aeruginosa isolates were positive for bla(VIM-2). It was not possible to identify specific clones with the potential to cause clinical infections. In conclusion, a multiclonal cluster of MBL-producers was responsible for the first cases of colonisation and/or infection in the ICU. A single integron structure, common in Greek hospitals, efficiently disseminated between clones and species, suggesting that the epidemic was mainly the result of successful horizontal transfer of mobile genetic material rather than the result of horizontal transfer of one or a few clones.

Early changes of procalcitonin may advise about prognosis and appropriateness of antimicrobial therapy in sepsis.

PURPOSE: The objective of this study is to define if early changes of procalcitonin (PCT) may inform about prognosis and appropriateness of administered therapy in sepsis. METHODS: A prospective multicenter observational study was conducted in 289 patients. Blood samples were drawn on day 1, that is, within less than 24 hours from advent of signs of sepsis, and on days 3, 7, and 10. Procalcitonin was estimated in serum by the ultrasensitive Kryptor assay (BRAHMS GmbH, Hennigsdorf, Germany). Patients were divided into the following 2 groups according to the type of change of PCT: group 1, where PCT on day 3 was decreased by more than 30% or was below 0.25 ng/mL, and group 2, where PCT on day 3 was either increased above 0.25 ng/mL or decreased less than 30%. RESULTS: Death occurred in 12.3% of patients of group 1 and in 29.9% of those of group 2 (P < .0001). Odds ratio for death of patients of group 1 was 0.328. Odds ratio for the administration of inappropriate antimicrobials of patients of group 2 was 2.519 (P = .003). CONCLUSIONS: Changes of serum PCT within the first 48 hours reflect the benefit or not of the administered antimicrobial therapy. Serial PCT measurements should be used in clinical practice to guide administration of appropriate antimicrobials.

Colistin-resistant isolates of Klebsiella pneumoniae emerging in intensive care unit patients: first report of a multiclonal cluster.

OBJECTIVES: Infections due to multidrug-resistant (MDR) Gram-negative pathogens in the ICU have prompted the use of colistin, an antibiotic forgotten for decades. The aim of this retrospective observational study was to record and present the emergence of colistin-resistant Klebsiella pneumoniae (CRKB) in a Greek ICU. METHODS: In a new university tertiary hospital, the first patients admitted in the ICU were already colonized or infected with MDR pathogens, and this led to frequent colistin use as part of empirical or microbiologically documented therapy. Colistin resistance was defined as MIC >4 mg/L by the Etest method. All CRKB isolated in surveillance cultures or clinical specimens in the ICU during the period 2004-5 were recorded along with patients' characteristics. RESULTS: Eighteen CRKB were isolated from 13 patients over a 16 month period, representing either colonizing or infective isolates. Patients' mean age was 70 years, with a mean APACHE II score at admission of 22. They all had a long hospitalization (median 69 days) and a long administration of colistin (median 27 days). Colistin-resistant isolates were implicated as pathogens in two bacteraemias, a ventilator-associated pneumonia and two soft tissue infections. Repetitive extragenic palindromic PCR identified six distinct clones, and horizontal transmission was also documented. CONCLUSIONS: Selective pressure due to extensive or inadequate colistin use may lead to the emergence of colistin resistance among K. pneumoniae isolates, jeopardizing treatment options in the ICU, potentially increasing morbidity and mortality of critically ill patients and necessitating prudent use of colistin.

Molecular characterization of an Escherichia coli clinical isolate that produces both metallo-beta-lactamase VIM-2 and extended-spectrum beta-lactamase GES-7: identification of the In8 integron carrying the blaVIM-2 gene.

OBJECTIVES: We have previously reported the isolation of a VIM-2-producing Escherichia coli clinical isolate and the coexistence of blaVIM-2 and blaGES-7 in the same isolate. The aim of this study was to elucidate the genetic environment of these genes. METHODS: PCR and sequence analysis were used to identify and analyse the blaVIM-containing integron. The location of the blaVIM-2 and blaGES-7 genes was identified by hybridization experiments on I-CeuI-digested genomic DNA after PFGE. RESULTS: Only the blaVIM-2 gene has been found on a class I integron, designated In8 (1474 bp). Hybridization with the blaVIM-2, the blaGES-7 and the 16S-23S rRNA probes confirmed the chromosomal location of both genes. The blaVIM-2 probe hybridized with a 710 kb fragment whereas the blaGES-7 probe hybridized with a 144 kb fragment of genomic DNA of the E. coli isolate. CONCLUSIONS: This report provides evidence for the chromosomal location of both blaVIM-2 and blaGES-7 genes carried by an E. coli clinical isolate. In8, containing blaVIM-2, presents an emerging threat of carbapenem resistance among E. coli isolates in Greece.

In vitro activity of tigecycline against multiple-drug-resistant, including pan-resistant, gram-negative and gram-positive clinical isolates from Greek hospitals.

The in vitro activities of tigecycline and selected antimicrobials were evaluated against a variety of multiple-drug-resistant clinical isolates, including extended-spectrum beta-lactamase- and/or metallo-beta-lactamase-producing gram-negative strains, colistin-resistant strains, vancomycin- and/or linezolid-resistant enterococci, and methicillin-resistant Staphylococcus aureus (MRSA). Tigecycline showed excellent activity against a collection of difficult-to-treat pathogens currently encountered in the hospital setting.