Acinetobacter baumannii isolates from pets and horses in Switzerland: molecular characterization and clinical data

Objectives We investigated whether Acinetobacter baumannii isolates of veterinary origin shared common molecular characteristics with those described in humans. Methods Nineteen A. baumannii isolates collected in pets and horses were analysed. Clonality was studied using repetitive extragenic palindromic PCR (rep-PCR) and multilocus sequence typing (MLST). PCR and DNA sequencing for various β-lactamase, aminoglycoside-modifying enzyme, gyrA and parC, ISAba1 and IS1133, adeR and adeS of the AdeABC efflux pump, carO porin and class 1/2/3 integron genes were performed. Results Two main clones [A (n = 8) and B (n = 9)] were observed by rep-PCR. MLST indicated that clone A contained isolates of sequence type (ST) ST12 (international clone II) and clone B contained isolates of ST15 (international clone I). Two isolates of ST10 and ST20 were also noted. Seventeen isolates were resistant to gentamicin, 12 to ciprofloxacin and 3 to carbapenems. Isolates of ST12 carried blaOXA-66, blaADC-25, blaTEM-1, aacC2 and IS1133. Strains of ST15 possessed blaOXA-69, blaADC-11, blaTEM-1 and a class 1 integron carrying aacC1 and aadA1. ISAba1 was found upstream of blaADC (one ST10 and one ST12) and/or blaOXA-66 (seven ST12). Twelve isolates of different STs contained the substitutions Ser83Leu in GyrA and Ser80Leu or Glu84Lys in ParC. Significant disruptions of CarO porin and overexpressed efflux pumps were not observed. The majority of infections were hospital acquired and in animals with predisposing conditions for infection. Conclusions STs and the molecular background of resistance observed in our collection have been frequently described in A. baumannii detected in human patients. Animals should be considered as a potential reservoir of multidrug-resistant A. baumanni

NDM-5 and OXA-181 Beta-Lactamases, a Significant Threat Continues To Spread in the Americas

ABSTRACT Among Gram-negative bacteria, carbapenem-resistant infections pose a serious and life-threatening challenge. Here, the CRACKLE network reports a sentinel detection and characterization of a carbapenem-resistant Klebsiella pneumoniae ST147 isolate harboring bla NDM-5 and bla OXA-181 from a young man who underwent abdominal surgery in India. bla NDM-5 was located on an IncFII plasmid of ≈90 kb, whereas bla OXA-181 was chromosomally encoded. Resistome and genome analysis demonstrated multiple copies of the transposable element IS 26 and a “hot-spot region” in the IncFII plasmid

Analysis of antibiotic resistance genes in multidrug-resistant acinetobacter sp. isolates from military and civilian patients treated at the Walter Reed Army Medical Center

Military medical facilities treating patients injured in Iraq and Afghanistan have identified a large number of multidrug-resistant (MDR) Acinetobacter baumannii isolates. In order to anticipate the impact of these pathogens on patient care, we analyzed the antibiotic resistance genes responsible for the MDR phenotype in Acinetobacter sp. isolates collected from patients at the Walter Reed Army Medical Center (WRAMC). Susceptibility testing, PCR amplification of the genetic determinants of resistance, and clonality were determined. Seventy-five unique patient isolates were included in this study: 53% were from bloodstream infections, 89% were resistant to at least three classes of antibiotics, and 15% were resistant to all nine antibiotics tested. Thirty-seven percent of the isolates were recovered from patients nosocomially infected or colonized at the WRAMC. Sixteen unique resistance genes or gene families and four mobile genetic elements were detected. In addition, this is the first report of blaOXA-58-like and blaPER-like genes in the U.S. MDR A. baumannii isolates with at least eight identified resistance determinants were recovered from 49 of the 75 patients. Molecular typing revealed multiple clones, with eight major clonal types being nosocomially acquired and with more than 60% of the isolates being related to three pan-European types. This report gives a “snapshot” of the complex genetic background responsible for antimicrobial resistance in Acinetobacter spp. from the WRAMC. Identifying genes associated with the MDR phenotype and defining patterns of transmission serve as a starting point for devising strategies to limit the clinical impact of these serious infections. © 2006, American Society for Microbiolog

Colistin Resistance in Carbapenem-Resistant Klebsiella pneumoniae: Laboratory Detection and Impact on Mortality

Background: Polymyxins including colistin are an important "last-line" treatment for infections caused by carbapenem-resistant Klebsiella pneumoniae (CRKp). Increasing use of colistin has led to resistance to this cationic antimicrobial peptide. Methods: A cohort nested within the Consortium on Resistance against Carbapenems in Klebsiella pneumoniae (CRACKLE) was constructed of patients with infection, or colonization with CRKp isolates tested for colistin susceptibility during the study period of December, 2011 to October, 2014. Reference colistin resistance determination as performed by broth macrodilution was compared to results from clinical microbiology laboratories (Etest) and to polymyxin resistance testing. Each patient was included once, at the time of their first colistin-tested CRKp positive culture. Time to 30-day in-hospital all-cause mortality was evaluated by Kaplan-Meier curves and Cox proportional hazard modeling. Results: In 246 patients with CRKp, 13% possessed ColR CRKp. ColR was underestimated by Etest (very major error rate = 35%, major error rate = 0.4%). A variety of rep-PCR strain types were encountered in both the ColS and the ColR groups. Carbapenem resistance was mediated primarily by blaKPC-2 (46%) and blaKPC-3 (50%). ColR was associated with increased hazard for in-hospital mortality (aHR 3.48; 95% confidence interval, 1.73-6.57; P < .001). The plasmid-associated ColR genes, mcr-1 and mcr-2 were not detected in any of the ColR CRKp. Conclusions: In this cohort, 13% of patients with CRKp presented with ColR CRKp. The apparent polyclonal nature of the isolates suggests de novo emergence of ColR in this cohort as the primary factor driving ColR. Importantly, mortality was increased in patients with ColR isolates

Population Structure of KPC-Producing Klebsiella pneumoniae Isolates from Midwestern U.S. Hospitals

ABSTRACT Genome sequencing of carbapenem-resistant Klebsiella pneumoniae isolates from regional U.S. hospitals was used to characterize strain diversity and the bla KPC genetic context. A phylogeny based on core single-nucleotide variants (SNVs) supports a division of sequence type 258 (ST258) into two distinct groups. The primary differences between the groups are in the capsular polysaccharide locus ( cps ) and their plasmid contents. A strict association between clade and KPC variant was found. The bla KPC gene was found on variants of two plasmid backbones. This study indicates that highly similar K. pneumoniae subpopulations coexist within the same hospitals over time

A Prospective Observational Study of the Epidemiology, Management, and Outcomes of Skin and Soft Tissue Infections Due to Carbapenem-Resistant Enterobacteriaceae

Abstract Background: This study was performed to characterize the epidemiology, management, and outcomes of skin and soft tissue infection (SSTI) and colonization due to carbapenem-resistant Enterobacteriaceae (CRE). Methods: Patients from the Consortium on Resistance Against Carbapenem in Klebsiella and Other Enterobacteriaceae (CRACKLE-1) from December 24, 2011 to October 1, 2014 with wound cultures positive for CRE were included in the study. Predictors of surgical intervention were analyzed. Molecular typing of isolates was performed using repetitive extragenic palindromic polymerase chain reaction (PCR). Carbapenemase genes were detected using PCR. Results: One hundred forty-two patients were included: 62 had SSTI (44%) and 56% were colonized. Mean age was 61 years, and 48% were male: median Charlson score was 3 (interquartile range, 1–5). Forty-eight percent of patients were admitted from long-term care facilities (LTCFs), and 31% were from the community. Two strain types (ST258A and ST258B) were identified (73% of 45 tested). Carbapenemase genes were detected in 40 of 45 isolates (blaKPC-3 [47%], blaKPC-2 [42%]). Sixty-eight patients (48%) underwent surgical intervention, 63% of whom had SSTI. Patients admitted from LTCFs were less likely to undergo surgical intervention (odds ratio [OR], 0.36; 95% confidence interval [CI], 0.18–0.71). In multivariable analysis, among patients with SSTI, those admitted from LTCFs were less likely to undergo debridement (OR, 0.18; 95% CI, 0.04–0.93). Conclusions: Patients admitted from LTCFs with CRE SSTI were less likely to undergo surgical intervention. Sixteen percent of the patients died, and approximately 50% of survivors required more intensive care upon discharge. These findings suggest a unique, impactful syndrome within the CRE infection spectrum. Further studies are needed to assess the role of surgical debridement in management of CRE-SSTI, particularly among LTCF residents

Longitudinal Analysis of the Temporal Evolution of Acinetobacter baumannii Strains in Ohio, USA, by Using Rapid Automated Typing Methods

Genotyping methods are essential to understand the transmission dynamics of Acinetobacter baumannii. We examined the representative genotypes of A. baumannii at different time periods in select locations in Ohio, using two rapid automated typing methods: PCR coupled with electrospray ionization mass spectrometry (PCR/ESI-MS), a form of multi-locus sequence typing (MLST), and repetitive-sequence-based-PCR (rep-PCR). Our analysis included 122 isolates from 4 referral hospital systems, in 2 urban areas of Ohio. These isolates were associated with outbreaks at 3 different time periods (1996, 2000 and 2005–2007). Type assignments of PCR/ESI-MS and rep-PCR were compared to each other and to worldwide (WW) clone types. The discriminatory power of each method was determined using the Simpson's index of diversity (DI). We observed that PCR/ESI-MS sequence type (ST) 14, corresponding to WW clone 3, predominated in 1996, whereas ST 12 and 14 co-existed in the intermediate period (2000) and ST 10 and 12, belonging to WW clone 2, predominated more recently in 2007. The shift from WW clone 3 to WW clone 2 was accompanied by an increase in carbapenem resistance. The DI was approximately 0.74 for PCR/ESI-MS, 0.88 for rep-PCR and 0.90 for the combination of both typing methods. We conclude that combining rapid automated typing methods such as PCR/ESI-MS and rep-PCR serves to optimally characterize the regional molecular epidemiology of A. baumannii. Our data also sheds light on the changing sequence types in an 11 year period in Northeast Ohio

Evaluation of Ceftazidime and NXL104 in Two Murine Models of Infection Due to KPC-Producing Klebsiella pneumoniae▿

We evaluated the efficacy of NXL104, a novel β-lactamase inhibitor, in combination with ceftazidime (CAZ) in two murine infection models (septicemia and thigh infection). We chose two KPC-producing Klebsiella pneumoniae strains (VA-361 and VA-406) showing MICs of CAZ of ≥256 μg/ml. Septicemia was induced by the intraperitoneal injection of KPC-producing K. pneumoniae followed 30 min later by a single subcutaneous treatment with CAZ alone or CAZ-NXL104 in ratios of 2:1, 4:1, 8:1, and 16:1. In this model, the median effective doses for 50% (ED50) of the animals for CAZ alone versus VA-361 and VA-406 were 1,578 and 709 mg/kg of body weight, respectively. When combined with NXL104 at 2:1, 4:1, 8:1, and 16:1 ratios, the CAZ ED50s for VA-361 and VA-406 were reduced to 8.1 and 3.5 mg/kg, 15.1 and 3.8 mg/kg, 16.9 and 7.2 mg/kg, and 29.5 and 12.1 mg/kg, respectively. For thigh infection, neutropenia was induced by the intraperitoneal injection of cyclophosphamide at days −4 and −1 preinfection. Infection was established by the intramuscular injection of KPC-producing K. pneumoniae into the right thigh. Mice were treated 1.5 h postinfection with either CAZ alone or CAZ-NXL104 at constant ratios of 4:1. When thighs were removed at 24 h postinfection, a >2-log CFU reduction was observed for mice treated with CAZ-NXL104 at doses of ≥128:32 mg/kg. In contrast, CAZ doses of ≥1,024 mg/kg were unable to reduce the numbers of CFU. Despite resistance to CAZ and possessing a complex β-lactamase background, NXL104 combined with CAZ proved to be very effective in murine models of infection due to contemporary highly resistant KPC-producing K. pneumoniae isolates