Publications (35)169.87 Total impact
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Article: Enhancing resistance to cephalosporins in class C beta-lactamases: impact of Gly214Glu in CMY-2.
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ABSTRACT: The biochemical properties of CMY-32, a class C enzyme possessing a single-amino acid substitution in the Omega loop (Gly214Glu), were compared to those of the parent enzyme, CMY-2, a widespread class C beta-lactamase. In parallel with our microbiological characterization, the Gly214Glu substitution in CMY-32 reduced catalytic efficiency (k(cat)/K(m)) by 50-70% against "good" substrates (i.e., cephalothin) while increasing k(cat)/K(m) against "poor" substrates (i.e., cefotaxime). Additionally, CMY-32 was more susceptible to inactivation by sulfone beta-lactamase inhibitors (i.e., sulbactam and tazobactam) than CMY-2. Timed electrospray ionization mass spectrometry (ESI-MS) analysis of the reaction of CMY-2 and CMY-32 with different substrates and inhibitors suggested that both beta-lactamases formed similar intermediates during catalysis and inactivation. We next showed that the carbapenems (imipenem, meropenem, and doripenem) form long-lived acyl-enzyme intermediates and present evidence that there is beta-lactamase-catalyzed elimination of the C(6) hydroxyethyl substituent. Furthermore, we discovered that the monobactam aztreonam and BAL29880, a new beta-lactamase inhibitor of the monobactam class, inactivate CMY-2 and CMY-32 by forming an acyl-enzyme intermediate that undergoes elimination of SO(3)(2-). Molecular modeling and dynamics simulations suggest that the Omega loop is more constrained in CMY-32 than CMY-2. Our model also proposes that Gln120 adopts a novel conformation in the active site while new interactions form between Glu214 and Tyr221, thus explaining the increased level of cefotaxime hydrolysis. When it is docked in the active site, we observe that BAL29880 exploits contacts with highly conserved residues Lys67 and Asn152 in CMY-2 and CMY-32. These findings highlight (i) the impact of single-amino acid substitutions on protein evolution in clinically important AmpC enzymes and (ii) the novel insights into the mechanisms by which carbapenems and monobactams interact with CMY-2 and CMY-32 beta-lactamases.Biochemistry 11/2009; 49(5):1014-23. · 3.42 Impact Factor -
Article: Extended-spectrum and CMY-type beta-lactamase-producing Escherichia coli in clinical samples and retail meat from Pittsburgh, USA and Seville, Spain.
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ABSTRACT: Infections due to Escherichia coli producing extended-spectrum beta-lactamase (ESBL) or CMY-type beta-lactamase (CMY) are increasingly observed in non-hospitalized patients. The origin of these organisms is uncertain, but retail meat contaminated with E. coli may be a source. In the present study, clinical information and strains collected from patients infected or colonized with ESBL-producing and CMY-producing E. coli at hospitals in Pittsburgh, USA and Seville, Spain were investigated. Retail meat purchased in these cities was also studied for the presence of these organisms. Twenty-five and 79 clinical cases with ESBL-producing E. coli and 22 cases and one case with CMY-producing E. coli were identified in Pittsburgh and Seville, respectively. Among them all, community-acquired and healthcare-associated cases together constituted 60% of the cases in Pittsburgh and 73% in Seville. Community-acquired cases were more common in Seville than in Pittsburgh (49% vs. 13%; p <0.001). ESBL-producing and CMY-producing E. coli isolates were commonly recovered from the local retail meat. In particular, 67% (8/12) of retail chickens in Seville and 85% (17/20) of those in Pittsburgh contained ESBL-producing and CMY-producing E. coli isolates, respectively. Among the ESBL-producing isolates, CTX-M and SHV were the most common ESBL types in both clinical and meat isolates. Approximately half of the ESBL-producing and CMY-producing E. coli isolates from meat belonged to phylogenetic groups associated with virulent extra-intestinal infections in humans. Community and healthcare environments are now significant reservoirs of ESBL-producing and CMY-producing E. coli. Retail meat is a potential source of these organisms.Clinical Microbiology and Infection 08/2009; 16(1):33-8. · 4.54 Impact Factor -
Article: Molecular epidemiology of CTX-M-producing Escherichia coli isolates at a tertiary medical center in western Pennsylvania.
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ABSTRACT: A combination of phenotypic and genotypic methods was used to investigate 70 unique Escherichia coli clinical isolates identified as producing extended-spectrum beta-lactamases (ESBLs) at a medical center in Pittsburgh, PA, between 2007 and 2008. Fifty-seven isolates (81%) produced CTX-M-type ESBLs, among which CTX-M-15 was predominant (n = 46). Isolates producing CTX-M-2, -9, -14, and -65 were also identified. One CTX-M-producing isolate coproduced CMY-2 cephalosporinase. Ten isolates (14%) produced SHV-type ESBLs, either SHV-5 or SHV-7. Two isolates produced only CMY-2 or -32. Pulsed-field gel electrophoresis revealed the presence of two major clusters of CTX-M-15-producing E. coli isolates, one in phylotype B2 (n = 15) and the other in phylotype A (n = 19). Of four phylotype B2 isolates that were able to transfer the bla(CTX-M-15)-carrying plasmids, three showed fingerprints related (>60%) to those of plasmids from phylotype A isolates. In phylotype B2, all CTX-M-15-producing isolates, as well as three isolates producing CTX-M-14, two producing SHV-5, and one producing SHV-7, belonged to the international epidemic clone defined by serotype O25:H4 and sequence type 131. The plasmids from eight of nine CTX-M-15-producing E. coli isolates of phylotype A that were examined were highly related to each other and were also found in two isolates belonging to phylotype D, suggesting horizontal transfer of this bla(CTX-M-15)-carrying plasmid between phylotypes. Our findings underscore the need to further investigate the epidemiology and virulence of CTX-M-producing E. coli in the United States.Antimicrobial Agents and Chemotherapy 08/2009; 53(11):4733-9. · 4.84 Impact Factor -
Article: Reduced susceptibility to cefepime among Escherichia coli clinical isolates producing novel variants of CMY-2 beta-lactamase.
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ABSTRACT: Here we describe three Escherichia coli clinical isolates with reduced susceptibility to cefepime. Sequencing of the bla(CMY) genes revealed two novel variants (CMY-33 and -44) with two- to four-amino-acid deletions in the H-10 helix. The deletions were responsible for 12- to 24-fold increases in the MICs of cefepime.Antimicrobial Agents and Chemotherapy 06/2009; 53(7):3159-61. · 4.84 Impact Factor -
Article: Klebsiella pneumoniae carbapenemase-producing enterobacteriaceae, northeast florida
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ABSTRACT: BACKGROUND: Since 2001 there have been several reported outbreaks due to carbapenem-resistant Klebsiella pneumoniae (Kp), particularly in the northeastern states. METHODS: Carbapenemase-producing Enterobacteriaceae from healthcare facilities in Northeast Florida were phenotypically identified and confirmed using PCR amplification and sequencing of the blaKPC gene. RESULTS: Results from PFGE analysis of these isolates demonstrated possible horizontal spread from two possible "outbreak" strains during the study period. CONCLUSIONS: We present the first published cluster of Kp and Escherichia coli (Ec) cases in Florida carrying the KPC-2 or KPC-3 gene.Southern medical journal 06/2009; · 0.92 Impact Factor -
Article: Activity of temocillin against KPC-producing Klebsiella pneumoniae and Escherichia coli.
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ABSTRACT: Temocillin, a 6-a-methoxy derivative of ticarcillin, is currently approved for treatment of infections due to Enterobacteriaceae in Belgium and the UK. ...Antimicrobial Agents and Chemotherapy 04/2009; 53(6):2700-1. · 4.84 Impact Factor -
Article: Clinical features and molecular epidemiology of CMY-type beta-lactamase-producing Escherichia coli.
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ABSTRACT: Knowledge of the clinical features of infections caused by Escherichia coli strains that produce plasmid-mediated AmpC beta-lactamase is limited. Of the several groups of plasmid-mediated AmpC beta-lactamases, CMY-type beta-lactamase is the most common in the United States. We prospectively identified patients infected or colonized with E. coli strains that produce CMY-type beta-lactamase, and we collected clinical data over a 7-month period. A retrospective cohort study was performed to identify features associated with these cases. Patients with extended-spectrum beta-lactamase-producing E. coli were used as a control group. Pulsed-field gel electrophoresis, plasmid analysis, and phylogenetic typing were performed. Twenty-two patients with infection or colonization due to CMY-type beta-lactamase-producing E. coli and 25 patients with infection or colonization due to extended-spectrum beta-lactamase-producing E. coli were identified. The demographic characteristics of the patients were similar in both cohorts. Patients with CMY-type beta-lactamase-producing E. coli were significantly more likely to have symptomatic infection than were patients with extended-spectrum beta-lactamase-producing E. coli (P = .028). The CMY-type beta-lactamase was identified as CMY-2 or its variants. Ninety-four percent of the CMY-type beta-lactamase-producing isolates belonged to E. coli phylogenetic groups B2 and D, which are associated with virulence. Many of the isolates shared similar plasmid profiles, whereas the pulsed-field gel electrophoresis profiles were diverse. Co-resistance to non-beta-lactam antimicrobials was common. In Pittsburgh, Pennsylvania, CMY-type beta-lactamase-producing E. coli strains are almost as common as extended-spectrum beta-lactamase-producing E. coli strains, and they cause symptomatic infection in the majority of cases.Clinical Infectious Diseases 03/2009; 48(6):739-44. · 9.15 Impact Factor -
Article: Simple disk-based method for detection of Klebsiella pneumoniae carbapenemase-type beta-lactamase by use of a boronic acid compound.
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ABSTRACT: A disk potentiation method using carbapenems as substrates and 3-aminophenyl boronic acid as an inhibitor was evaluated for the detection of Klebsiella pneumoniae carbapenemase (KPC)-type beta-lactamases. When combined with nonsusceptibility to ertapenem, the method was easy to perform and reliably differentiated isolates producing KPC-type beta-lactamases from those producing other types of beta-lactamases.Journal of clinical microbiology 11/2008; 46(12):4083-6. · 4.16 Impact Factor -
Article: Genetic basis of multidrug resistance in Acinetobacter baumannii clinical isolates at a tertiary medical center in Pennsylvania.
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ABSTRACT: A total of 49 unique clinical isolates of multidrug-resistant (MDR) Acinetobacter baumannii identified at a tertiary medical center in Pittsburgh, Pennsylvania, between August 2006 and September 2007 were studied for the genetic basis of their MDR phenotype. Approximately half of all A. baumannii clinical isolates identified during this period qualified as MDR, defined by nonsusceptibility to three or more of the antimicrobials routinely tested in the clinical microbiology laboratory. Among the MDR isolates, 18.4% were resistant to imipenem. The frequencies of resistance to amikacin and ciprofloxacin were high at 36.7% and 95.9%, respectively. None of the isolates was resistant to colistin or tigecycline. The presence of the carbapenemase gene bla(OXA-23) and the 16S rRNA methylase gene armA predicted high-level resistance to imipenem and amikacin, respectively. bla(OXA-23) was preceded by insertion sequence ISAba1, which likely provided a potent promoter activity for the expression of the carbapenemase gene. The structure of the transposon defined by ISAba1 differed from those reported in Europe, suggesting that ISAba1-mediated acquisition of bla(OXA-23) may occur as an independent event. Typical substitutions in the quinolone resistance-determining regions of the gyrA and parC genes were observed in the ciprofloxacin-resistant isolates. Plasmid-mediated quinolone resistance genes, including the qnr genes, were not identified. Fifty-nine percent of the MDR isolates belonged to a single clonal group over the course of the study period, as demonstrated by pulsed-field gel electrophoresis.Antimicrobial Agents and Chemotherapy 09/2008; 52(11):3837-43. · 4.84 Impact Factor -
Article: Genetic environment of 16S rRNA methylase gene rmtD.
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ABSTRACT: The genetic environment of the 16S rRNA methylase gene rmtD was investigated. rmtD was flanked by a novel ISCR motif located downstream of class I integron In163 in the original Pseudomonas aeruginosa strain. rmtD found in Klebsiella pneumoniae appeared to have been mobilized from P. aeruginosa by an IS26-mediated event.Antimicrobial Agents and Chemotherapy 07/2008; 52(6):2270-2. · 4.84 Impact Factor -
Article: High prevalence of CTX-M-15-producing Klebsiella pneumoniae among inpatients and outpatients with urinary tract infection in Southern India.
Journal of Antimicrobial Chemotherapy 07/2008; 61(6):1393-4. · 5.07 Impact Factor -
Article: Multiclonal outbreak of Klebsiella pneumoniae producing extended-spectrum beta-lactamase CTX-M-2 and novel variant CTX-M-59 in a neonatal intensive care unit in Brazil.
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ABSTRACT: An outbreak of cephalosporin-resistant Klebsiella pneumoniae occurred in a neonatal intensive care unit in São Paulo, Brazil. Of the 10 pulsotypes identified during the outbreak and follow-up periods, nine produced CTX-M-2 or its new variant CTX-M-59 and one produced SHV-5. bla(CTX-M-2/59) genes were located on closely related plasmids that were transferable.Antimicrobial Agents and Chemotherapy 06/2008; 52(5):1790-3. · 4.84 Impact Factor -
Article: Escherichia coli isolate coproducing 16S rRNA Methylase and CTX-M-type extended-spectrum beta-lactamase isolated from an outpatient in the United States.
Antimicrobial Agents and Chemotherapy 04/2008; 52(3):1204-5. · 4.84 Impact Factor -
Article: 16S ribosomal RNA methylase RmtD produced by Klebsiella pneumoniae in Brazil.
Journal of Antimicrobial Chemotherapy 04/2008; 61(3):746-7. · 5.07 Impact Factor -
Article: Activity of temocillin against KPC-producing Klebsiella pneumoniae and Escherichi coli
Top co-authors
Top Journals
Institutions
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2008–2011
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University of Pittsburgh
- Division of Infectious Diseases
Pittsburgh, PA, USA
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2009
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University of Queensland
Brisbane, Queensland, Australia -
Baptist Medical Center, Jacksonville
Jacksonville, FL, USA
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2008–2009
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Instituto Adolfo Lutz
São Paulo, Estado de Sao Paulo, Brazil
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