Beatriz Meurer Moreira

ANTIMICROBIAL RESISTANCE IN STAPHYLOCOCCI: Epidemiology, Molecular Mechanisms, and Clinical Relevance

Staphylococcal infections continue to pose important clinical problems in children and adults. Antibiotic resistance among the staphylococci has rendered therapy of these infections a therapeutic challenge. Despite early, uniform susceptibility to penicillin, staphylococci acquired a gene elaborating beta-lactamase that rendered penicillin inactive and that is borne by nearly all clinical isolates. Penicillinase-resistant beta-lactams, such as methicillin, were introduced in the early 1960s, but resistance to them has become an increasing concern. The mechanism of the so-called methicillin resistance is complex. Moreover, once confined to the ecology of hospitals and other institutions, a recent increase in community-acquired methicillin-resistant S. aureus infections has been observed. Glycopeptides, until now the only uniformly reliable therapeutic modality, have been increasingly used for therapy of staphylococcal infections. The recent recognition of clinical isolates with reduced susceptibility to glycopeptides is of concern.

Identification of Clinical Isolates of Indole-Positive and Indole-Negative Klebsiella spp.

ABSTRACT Biochemical methods employed to classify bacterial species have limitations and may have contributed to the taxonomic complexity recently reported for the genus Klebsiella. The objective of the present study was to apply a simple biochemical test panel to classify a collection of human Klebsiella isolates. We found that with only three additional tests, it is possible to place most isolates in a defined species. Analysis of a 512-bp sequence of the rpoB gene was used as the reference. A total of 16 conventional and 4 supplementary tests were used to evaluate 122 recent isolates identified as Klebsiella from 120 patients, isolated at the clinical laboratory of a university hospital in Minas Gerais, Brazil. Of these, 102 (84%) isolates were identified as Klebsiella pneumoniae or Klebsiella variicola, 19 (15%) as Klebsiella oxytoca, and 1 (1%) as Raoultella planticola. Enterobacterial repetitive intergenic consensus-PCR typing revealed a diversity of genotypes. rpoB gene sequencing confirmed the phenotypic identification and detected five K. variicola isolates among the K. pneumoniae/K. variicola group. Three additional tests that include growth at 10°C and histamine and d-melezitose assimilation should be considered essential tests for the typing of Klebsiella isolates.

Risk Factors for Acquisition of Multidrug-Resistant Pseudomonas aeruginosa Producing SPM Metallo-β-Lactamase

ABSTRACT To evaluate risk factors for colonization or infection due to multidrug-resistant Pseudomonas aeruginosa (MDRPa) carrying the blaSPM gene (SPM-MRDPa) among hospitalized patients, we undertook a case control study at a 480-bed, tertiary-care university hospital. Two different case definitions were used. In the first definition, a case patient (SPM case patient) was defined as a patient who had at least one isolate of SPM-MDRPa (14 patients). In the second, a case patient (non-SPM case patient) was defined as a patient who had at least one isolate of non-SPM-MDRPa (18 patients). For each case patient, we selected two controls, defined as a patient colonized and/or infected by a non-MDRPa isolate during the same study period and with the closest duration of hospitalization until the isolation of P. aeruginosa as cases. The use of quinolones was the single independent predictor of colonization and/or infection by blaSPM MDRPa (odds ratrio [OR] = 14.70, 95% confidence interval [95% CI] = 1.70 to 127.34, P = 0.01), whereas the use of cefepime was the single predictor of colonization and/or infection by non-blaSPM MDRPa (OR = 8.50, 95% CI = 1.51 to 47.96, P = 0.01). The main risk factor for MDRPa was a history of antibiotics usage. Stratification of risk factor analysis by a precise mechanism of resistance led us to identify a specific antibiotic, a quinolone, as a predictor for SPM-MDRPa.

Clonal composition of Escherichia coli causing community-acquired urinary tract infections in the State of Rio de Janeiro, Brazil.

Recent studies from North America and Europe have demonstrated community-wide clonal spread of uropathogenic Escherichia coli (UPEC). To investigate if a similar pattern of spread occurs in Brazil, we characterized UPEC from women with community-acquired urinary tract infection (UTI) in Rio de Janeiro. E. coli isolates from women with UTI in one public outpatient clinic were evaluated for antibiotic susceptibility, E. coli phylogenetic grouping, enterobacterial repetitive intergenic consensus (ERIC) 2 PCR and pulsed-field gel electrophoresis fingerprinting, and multilocus sequence typing. From March 2005 to November 2006, 344 patients were studied. Of these, 186 (54%) had confirmed UTI, 118 (63.4%) of which were caused by E. coli. More than 50% of these isolates were resistant to ampicillin and trimethoprim/sulfamethoxazole. Of these, 96 (81%) belonged to 19 ERIC2 clonal groups. The largest group included 15 isolates, all belonging to multilocus sequence typing group ST69 and phylogenetic group D; they had pulsed-field gel electrophoresis patterns sharing at least 89% similarity compared with the CgA reference strain ATCC BAA-457. CgA strains have been found to be widespread in the United States in the early 2000s. Clonal group E. coli strains accounted for a large proportion (52%) of all UTIs and 82% of the trimethoprim/sulfamethoxazole-resistant E. coli UTIs. Thus, as in North America and Europe, UPECs that cause UTI in Rio de Janeiro also show clonal distribution, and a substantial proportion of drug-resistant UTI is caused by a small set of genetically related E. coli strains.

Antimicrobial Resistance in Staphylococci

Staphylococci have developed a variety of strategies for dealing with the presence of antibiotics encountered in clinical environments. Resistance to beta-lactams and other antimicrobial agents has been accomplished by a diverse array of molecular mechanisms. Options available to treat infections caused by staphylococci resistant to methicillin are limited, and the next generation of antibiotics to be introduced, should glycopeptide resistance become an important clinical problem, is not yet on the horizon.

Use of fimH Single-Nucleotide Polymorphisms for Strain Typing of Clinical Isolates of Escherichia coli for Epidemiologic Investigation

ABSTRACT Strain typing methods that compare electrophoresis banding patterns are commonly used but are difficult to standardize and poorly portable. Multilocus sequence typing (MLST) is a sequence-based alternative, but it is not practical for large-scale epidemiological studies. In the present study, the usefulness of fimH single-nucleotide polymorphisms (SNPs) for Escherichia coli typing was explored. fimH SNPs were determined for 345 E. coli clinical isolates (including 3 reference strains) and compared to PCR-based ECOR (E. coli reference collection) phylogrouping. The fimH gene could be amplified for 316 (92%) of the 345 isolates. fimH SNP analysis found 46 distinct terminal groups in the nucleotide sequence-based phylogenetic tree (fimH types). A subset of the E. coli isolates (162 clinical isolates and the 3 reference strains) were compared by fimH type, PCR phylogroup, and MLST. These isolates fell into 27 fimH types and 18 MLST clonal complexes (CCs) that contained 2 to 28 isolates per complex. The combination of PCR phylogroup and fimH type corresponded to a single CC for 113 (68%) isolates and 2 or 3 CCs for the other 52 (32%) isolates. We propose that the combination of PCR phylogrouping and fimH SNP analysis may be a useful method to type a large collection of clinical E. coli isolates for epidemiologic studies.

Imported and Intensive Care Unit-Born Acinetobacter baumannii Clonal Complexes: One-Year Prospective Cohort Study in Intensive Care Patients

The main objective of this study was to assess the frequency and possible sources of colonization and infection by Acinetobacter in the intensive care unit (ICU) of a university hospital in Rio de Janeiro, Brazil, and characterize the isolates for relatedness to internationally and locally disseminated lineages. Patients consecutively admitted to the ICU from April 2007 to April 2008 were screened for colonization and infection. Species were identified by rpoB sequencing. The presence of acquired and intrinsic carbapenemase genes was assessed by polymerase chain reaction (PCR). Strains were typed by random amplification of polymorphic DNA (RAPD)-PCR, pulsed-field gel electrophoresis, and multilocus sequence typing (MLST) using the schemes hosted at the University of Oxford (UO) and Institut Pasteur (IP). Of 234 patients, 98 (42%) had at least one specimen positive for the Acinetobacter isolate, and 24 (10%) had infection. A total of 22 (92%) infections were caused by Acinetobacter baumannii and one each (4%) by Acinetobacter nosocomialis and Acinetobacter berezinae. A. baumannii isolates from 60 patients belonged to RAPD types that corresponded to MLST clonal complexes (CCs) 109/1 (UO/IP scheme, known as International Clone I), CC 110/110 (UO/IP), CC 113/79 (UO/IP), and CC 104/15 (UO/IP). Most CCs were carbapenem resistant and carried the bla(OXA-23)-like gene. Strains were introduced by patients transferred from other wards of the same hospital (11 patients, 18%) or acquired from cross-transmission within the ICU (49 patients, 82%). A. nosocomialis lineage sequence type 260 colonized 10% of the whole study population. A. baumannii have become established in this hospital as a part of a global epidemic of successful clones. Once introduced into the hospital, such clones have become entrenched among patients in the ICU.

Rapid Induction of High-Level Carbapenem Resistance in Heteroresistant KPC-Producing Klebsiella pneumoniae

ABSTRACT Enterobacteriaceae strains producing the Klebsiella pneumoniae carbapenemase (KPC) have disseminated worldwide, causing an urgent threat to public health. KPC-producing strains often exhibit low-level carbapenem resistance, which may be missed by automated clinical detection systems. In this study, eight Klebsiella pneumoniae strains with heterogeneous resistance to imipenem were used to elucidate the factors leading from imipenem susceptibility to high-level resistance as defined by clinical laboratory testing standards. Time-kill analysis with an inoculum as low as 3 × 106 CFU/ml and concentrations of imipenem 8- and 16-fold higher than the MIC resulted in the initial killing of 99.9% of the population. However, full recovery of the population occurred by 20 h of incubation in the same drug concentrations. Population profiles showed that recovery was mediated by a heteroresistant subpopulation at a frequency of 2 × 10−7 to 3 × 10−6. Samples selected 2 h after exposure to imipenem were as susceptible as the unexposed parental strain and produced the major outer membrane porin OmpK36. However, between 4 to 8 h after exposure, OmpK36 became absent, and the imipenem MIC increased at least 32-fold. Individual colonies isolated from cultures after 20 h of exposure revealed both susceptible and resistant subpopulations. Once induced, however, the high-level imipenem resistance was maintained, and OmpK36 remained unexpressed even without continued carbapenem exposure. This study demonstrates the essential coordination between blaKPC and ompK36 expression mediating high-level imipenem resistance from a population of bacteria that initially exhibits a carbapenem-susceptibility phenotype.

Association of Class 1 and 2 Integrons with Multidrug-Resistant Acinetobacter baumannii International Clones and Acinetobacter nosocomialis Isolates

ABSTRACT The Acinetobacter baumannii clonal complex 113/79 (CC113/79) and class 2 integrons predominate in Latin America; a relationship between these characteristics was explored. The presence of integrases was determined in successive hospital Acinetobacter isolates (163 A. baumannii isolates and 72 Acinetobacter nosocomialis isolates). Most isolates had integrons, but class 1 and 2 integrons were present significantly more often in CC109/1 and CC113/79, respectively. The high prevalence of CC113/79 in Latin America may account for the predominance of class 2 integrons.

Case-Crossover Study of Burkholderia cepacia Complex Bloodstream Infection Associated with Contaminated Intravenous Bromopride

OBJECTIVEnTo investigate an outbreak of healthcare-associated Burkholderia cepacia complex (BCC) primary bloodstream infections (BCC-BSI).nnnDESIGN AND SETTINGnCase-crossover study in a public hospital, a university hospital and a private hospital in Rio de Janeiro, Brazil, from March 2006 to May 2006.nnnPATIENTSnTwenty-five patients with BCC-BSI.nnnDESIGNnAfter determining the date BCC-BSI symptoms started for each patient, 3 time intervals of data collection were defined, each one with a duration of 3 days: the case period, starting just before BCC-BSI symptoms onset; the control period, starting 6 days before BCC-BSI symptoms onset; and the washout period, comprising the 3 days between the case period and the control period. Exposures evaluated were intravascular solutions and invasive devices and procedures. Potential risk factors were identified by using the McNemar chi(2) adjusted test. Cultures of samples of potentially contaminated solutions were performed. BCC strain typing was performed by pulsed-field gel electrophoresis using SpeI.nnnRESULTSnThe statistical analysis revealed that the use of bromopride and dipyrone was associated with BCC-BSI. A total of 21 clinical isolates from 17 (68%) of the 25 patients and an isolate obtained from the bromopride vial were available for strain typing. Six pulsotypes were detected. A predominant pulsotype (A) accounted for 11 isolates obtained from 11 patients (65%) in the 3 study hospitals.nnnCONCLUSIONnOur investigation, using a case-crossover design, of an outbreak of BCC-BSI infections concluded it was polyclonal but likely caused by infusion of contaminated bromopride. The epidemiological finding was validated by microbiological analysis. After recall of contaminated bromopride vials by the manufacturer, the outbreak was controlled.