Federico Perez

Global Challenge of Multidrug-Resistant Acinetobacter baumannii

We may soon be facing the end of the “antibiotic era.” The initial and seemingly unstoppable success of antibiotics, the fruit of human ingenuity, has been countered by an escalation of resistance mechanisms in bacteria. This crisis has been described as an “unwinnable war” (www.wellcome.org). The statistics compiled as a result of surveillance efforts illustrate the emergence of many genera of bacteria that are resistant to all antibiotics (57, 60). The genus Acinetobacter epitomizes this trend and deserves close attention. Acinetobacter spp. display mechanisms of resistance to all existing antibiotic classes as well as a prodigious capacity to acquire new determinants of resistance (7). The increasing recovery in the clinic of multidrug-resistant (MDR) Acinetobacter baumannii is a frightening reality (112). This review summarizes the worldwide emergence of antibiotic-resistant A. baumannii as a nosocomial pathogen and focuses on its mechanisms of resistance against selected antibiotics. It concludes with a summary of current strategies in the treatment of MDR A. baumannii and offers perspectives on the control of this global public health threat.

Characterization of blaKPC-containing Klebsiella pneumoniae isolates detected in different institutions in the Eastern USA

BACKGROUND The emergence of bla(KPC)-containing Klebsiella pneumoniae (KPC-Kp) isolates is attracting significant attention. Outbreaks in the Eastern USA have created serious treatment and infection control problems. A comparative multi-institutional analysis of these strains has not yet been performed. METHODS We analysed 42 KPC-Kp recovered during 2006-07 from five institutions located in the Eastern USA. Antimicrobial susceptibility tests, analytical isoelectric focusing (aIEF), PCR and sequencing of bla genes, PFGE and rep-PCR were performed. Results By in vitro testing, KPC-Kp isolates were highly resistant to all non-carbapenem beta-lactams (MIC(90)s >or= 128 mg/L). Among carbapenems, MIC(50/90)s were 4/64 mg/L for imipenem and meropenem, 4/32 mg/L for doripenem and 8/128 for ertapenem. Combinations of clavulanate or tazobactam with a carbapenem or cefepime did not significantly lower the MIC values. Genetic analysis revealed that the isolates possessed the following bla genes: bla(KPC-2) (59.5%), bla(KPC-3) (40.5%), bla(TEM-1) (90.5%), bla(SHV-11) (95.2%) and bla(SHV-12) (50.0%). aIEF of crude beta-lactamase extracts from these strains supported our findings, showing beta-lactamases at pIs of 5.4, 7.6 and 8.2. The mean number of beta-lactamases was 3.5 (range 3-5). PFGE demonstrated that 32 (76.2%) isolates were clonally related (type A). Type A KPC-Kp isolates (20 bla(KPC-2) and 12 bla(KPC-3)) were detected in each of the five institutions. rep-PCR showed patterns consistent with PFGE. CONCLUSIONS We demonstrated the complex beta-lactamase background of KPC-Kp isolates that are emerging in multiple centres in the Eastern USA. The prevalence of a single dominant clone suggests that interstate transmission has occurred.

Carbapenem-resistant Acinetobacter baumannii and Klebsiella pneumoniae across a hospital system: impact of post-acute care facilities on dissemination

BACKGROUND Resistance to carbapenems among Acinetobacter baumannii and Klebsiella pneumoniae presents a serious therapeutic and infection control challenge. We describe the epidemiology and genetic basis of carbapenem resistance in A. baumannii and K. pneumoniae in a six-hospital healthcare system in Northeast Ohio. METHODS Clinical isolates of A. baumannii and K. pneumoniae distributed across the healthcare system were collected from April 2007 to April 2008. Antimicrobial susceptibility testing was performed followed by molecular analysis of carbapenemase genes. Genetic relatedness of isolates was established with repetitive sequence-based PCR (rep-PCR), multilocus PCR followed by electrospray ionization mass spectrometry (PCR/ESI-MS) and PFGE. Clinical characteristics and outcomes of patients were reviewed. RESULTS Among 39 isolates of A. baumannii, two predominant genotypes related to European clone II were found. Eighteen isolates contained bla(OXA-23), and four isolates possessed bla(OXA-24/40). Among 29 K. pneumoniae isolates with decreased susceptibility to carbapenems, two distinct genotypes containing bla(KPC-2) or bla(KPC-3) were found. Patients with carbapenem-resistant A. baumannii and K. pneumoniae were elderly, possessed multiple co-morbidities, were frequently admitted from and discharged to post-acute care facilities, and experienced prolonged hospital stays (up to 25 days) with a high mortality rate (up to 35%). CONCLUSION In this outbreak of carbapenem-resistant A. baumannii and K. pneumoniae across a healthcare system, we illustrate the important role post-acute care facilities play in the dissemination of multidrug-resistant phenotypes.

Outbreak of Colistin-Resistant, Carbapenem-Resistant Klebsiella pneumoniae in Metropolitan Detroit, Michigan

ABSTRACT Carbapenem-resistant Klebsiella pneumoniae has spread worldwide and throughout the United States. Colistin is used extensively to treat infections with this organism. We describe a cluster of colistin-resistant, carbapenem-resistant K. pneumoniae infection cases involving three institutions in Detroit, MI. A cluster of five cases of colistin-resistant, carbapenem-resistant K. pneumoniae was identified at Detroit Medical Center (DMC) from 27 July to 22 August 2009. Epidemiologic data were collected, and transmission opportunities were analyzed. Isolates were genotyped by using pulsed-field gel electrophoresis and repetitive extragenic palindromic PCR. Data regarding the use of colistin were obtained from pharmacy records. The index case of colistin-resistant, carbapenem-resistant K. pneumoniae was followed 20 days later by four additional cases occurring in a 6-day interval. All of the patients, at some point, had stayed at one particular institution. The mean number of opportunities for transmission between patients was 2.3 ± 0.5, and each patient had at least one opportunity for transmission with one of the other patients. Compared to 60 colistin-susceptible, carbapenem-resistant K. pneumoniae controls isolated in the previous year at DMC, case patients were significantly older (P = 0.05) and the carbapenem-resistant K. pneumoniae organisms isolated from them displayed much higher MICs to imipenem (P < 0.001). Colistin use was not enhanced in the months preceding the outbreak. Genotyping revealed two closely related clones. This report of a colistin-resistant, carbapenem-resistant K. pneumoniae outbreak is strongly linked to patient-to-patient transmission. Controlling the spread and novel emergence of bacteria with this phenotype is of paramount importance.

Emergence of blaKPC-containing Klebsiella pneumoniae in a long-term acute care hospital: a new challenge to our healthcare system

OBJECTIVES To characterize isolates of Klebsiella pneumoniae producing KPC carbapenemase (KPC-Kp) associated with an outbreak in a long-term acute care hospital (LTACH) in South Florida. METHODS During 21 March to 20 April 2008, 241 K. pneumoniae isolates detected at Integrated Regional Laboratories (Ft. Lauderdale, FL) for which the ertapenem MICs were > or =4 mg/L were studied. PCR, cloning and sequence analysis were used to detect bla(KPC) and to characterize the beta-lactamase and outer membrane proteins (Omps). The expression level of KPC enzymes was studied by immunoblotting. Genetic relatedness of isolates was investigated with rep-PCR and PFGE. Clinical records of patients were investigated. RESULTS Seven KPC-Kp strains were isolated from different patients located at a single LTACH, with a further three isolates being recovered from patients at different hospitals. All KPC-Kp isolates in patients from the LTACH and from one hospital patient were genetically related and shared PFGE patterns that clustered with known sequence type (ST) 258 strains. These strains were highly resistant to carbapenems (MICs > or = 32 mg/L) due to an increased level of KPC expression and loss of Omps. Rectal colonization was documented in all LTACH patients with KPC-Kp isolates. Treatment failures were common (crude mortality rate of 69%). Active surveillance and enhanced infection control practices terminated the KPC-Kp outbreak. CONCLUSIONS The detection of KPC-Kp in an LTACH represents a serious infection control and therapeutic challenge in a new clinical setting. The speed at which the epidemic of KPC-Kp is spreading in our healthcare system mandates urgent action.

In Vitro Activity of Fosfomycin Against blaKPC-containing Klebsiella pneumoniae Isolates including those nonsusceptible to tigecycline and/or colistin

ABSTRACT In vitro activity of fosfomycin was evaluated against 68 blaKPC-possessing Klebsiella pneumoniae (KpKPC) isolates, including 23 tigecycline- and/or colistin-nonsusceptible strains. By agar dilution, 93% of the overall KpKPC were susceptible (MIC50/90 of 16/64 μg/ml, respectively). The subgroup of 23 tigecycline- and/or colistin-nonsusceptible strains showed susceptibility rates of 87% (MIC50/90 of 32/128 μg/ml, respectively). Notably, 5 out of 6 extremely drug-resistant (tigecycline and colistin nonsusceptible) KpKPC were susceptible to fosfomycin. Compared to agar dilution, disk diffusion was more accurate than Etest.

Identification of a New Allelic Variant of the Acinetobacter baumannii Cephalosporinase, ADC-7 β-Lactamase: Defining a Unique Family of Class C Enzymes

ABSTRACT Acinetobacter spp. are emerging as opportunistic hospital pathogens that demonstrate resistance to many classes of antibiotics. In a metropolitan hospital in Cleveland, a clinical isolate of Acinetobacter baumannii that tested resistant to cefepime and ceftazidime (MIC = 32 μg/ml) was identified. Herein, we sought to determine the molecular basis for the extended-spectrum-cephalosporin resistance. Using analytical isoelectric focusing, a β-lactamase with a pI of ≥9.2 was detected. PCR amplification with specific A. baumannii cephalosporinase primers yielded a 1,152-bp product which, when sequenced, identified a novel 383-amino-acid class C enzyme. Expressed in Escherichia coli DH10B, this β-lactamase demonstrated greater resistance against ceftazidime and cefotaxime than cefepime (4.0 μg/ml versus 0.06 μg/ml). The kinetic characteristics of this β-lactamase were similar to other cephalosporinases found in Acinetobacter spp. In addition, this cephalosporinase was inhibited by meropenem, imipenem, ertapenem, and sulopenem (Ki < 40 μM). The amino acid compositions of this novel enzyme and other class C β-lactamases thus far described for A. baumannii, Acinetobacter genomic species 3, and Oligella urethralis in Europe and South Africa suggest that this cephalosporinase defines a unique family of class C enzymes. We propose a uniform designation for this family of cephalosporinases (Acinetobacter-derived cephalosporinases [ADC]) found in Acinetobacter spp. and identify this enzyme as ADC-7 β-lactamase. The coalescence of Acinetobacter ampC β-lactamases into a single common ancestor and the substantial phylogenetic distance separating them from other ampC genes support the logical value of developing a system of nomenclature for these Acinetobacter cephalosporinase genes.

New Insights into Dissemination and Variation of the Health Care-Associated Pathogen Acinetobacter baumannii from Genomic Analysis

ABSTRACT Acinetobacter baumannii is a globally important nosocomial pathogen characterized by an increasing incidence of multidrug resistance. Routes of dissemination and gene flow among health care facilities are poorly resolved and are important for understanding the epidemiology of A. baumannii, minimizing disease transmission, and improving patient outcomes. We used whole-genome sequencing to assess diversity and genome dynamics in 49 isolates from one United States hospital system during one year from 2007 to 2008. Core single-nucleotide-variant-based phylogenetic analysis revealed multiple founder strains and multiple independent strains recovered from the same patient yet was insufficient to fully resolve strain relationships, where gene content and insertion sequence patterns added additional discriminatory power. Gene content comparisons illustrated extensive and redundant antibiotic resistance gene carriage and direct evidence of gene transfer, recombination, gene loss, and mutation. Evidence of barriers to gene flow among hospital components was not found, suggesting complex mixing of strains and a large reservoir of A. baumannii strains capable of colonizing patients. IMPORTANCE Genome sequencing was used to characterize multidrug-resistant Acinetobacter baumannii strains from one United States hospital system during a 1-year period to better understand how A. baumannii strains that cause infection are related to one another. Extensive variation in gene content was found, even among strains that were very closely related phylogenetically and epidemiologically. Several mechanisms contributed to this diversity, including transfer of mobile genetic elements, mobilization of insertion sequences, insertion sequence-mediated deletions, and genome-wide homologous recombination. Variation in gene content, however, lacked clear spatial or temporal patterns, suggesting a diverse pool of circulating strains with considerable interaction between strains and hospital locations. Widespread genetic variation among strains from the same hospital and even the same patient, particularly involving antibiotic resistance genes, reinforces the need for molecular diagnostic testing and genomic analysis to determine resistance profiles, rather than a reliance primarily on strain typing and antimicrobial resistance phenotypes for epidemiological studies. Genome sequencing was used to characterize multidrug-resistant Acinetobacter baumannii strains from one United States hospital system during a 1-year period to better understand how A. baumannii strains that cause infection are related to one another. Extensive variation in gene content was found, even among strains that were very closely related phylogenetically and epidemiologically. Several mechanisms contributed to this diversity, including transfer of mobile genetic elements, mobilization of insertion sequences, insertion sequence-mediated deletions, and genome-wide homologous recombination. Variation in gene content, however, lacked clear spatial or temporal patterns, suggesting a diverse pool of circulating strains with considerable interaction between strains and hospital locations. Widespread genetic variation among strains from the same hospital and even the same patient, particularly involving antibiotic resistance genes, reinforces the need for molecular diagnostic testing and genomic analysis to determine resistance profiles, rather than a reliance primarily on strain typing and antimicrobial resistance phenotypes for epidemiological studies.

Presence of Plasmid-Mediated Quinolone Resistance in Klebsiella pneumoniae Isolates Possessing blaKPC in the United States

ABSTRACT The presence of plasmid-mediated quinolone resistance genes [i.e., qnrA, qnrB, qnrS, aac(6′)-Ib-cr, and qepA] was evaluated among 42 blaKPC-containing Klebsiella pneumoniae isolates collected in the eastern United States. One isolate carried the blaKPC-3 and qnrB19 genes on the same conjugative plasmid, whereas another carried the blaKPC-3 and qnrA1 genes on separate plasmids.

Carbapenem-resistant Enterobacteriaceae: a menace to our most vulnerable patients.

The emergence of carbapenem-resistant Enterobacteriaceae (CRE) highlights the importance of effective antibiotics to maintain the safety of our health care system. Clinicians will encounter CRE as a cause of difficult-to-treat and often fatal infections in hospitalized patients. We review the mechanisms of carbapenem resistance, the dissemination and clinical impact of these resistant organisms, and challenges to their detection, treatment, and control. This public health crisis demands redefined and refocused efforts in the diagnosis, treatment, and control of infections in hospitalized patients.