Ceftazidime/avibactam resistance is associated with PER-3-producing ST309 lineage in Chilean clinical isolates of non-carbapenemase producing Pseudomonas aeruginosa.

Introduction: Ceftazidime/avibactam (CZA) is indicated against multidrug-resistant Pseudomonas aeruginosa, particularly those that are carbapenem resistant. CZA resistance in P. aeruginosa producing PER, a class A extended-spectrum ß-lactamase, has been well documented in vitro. However, data regarding clinical isolates are scarce. Our aim was to analyze the contribution of PER to CZA resistance in non-carbapenemase-producing P. aeruginosa clinical isolates that were ceftazidime and/or carbapenem non-susceptible. Methods: Antimicrobial susceptibility was determined through agar dilution and broth microdilution, while bla PER gene was screened through PCR. All PER-positive isolates and five PER-negative isolates were analyzed through Whole Genome Sequencing. The mutational resistome associated to CZA resistance was determined through sequence analysis of genes coding for PBPs 1b, 3 and 4, MexAB-OprM regulators MexZ, MexR, NalC and NalD, AmpC regulators AmpD and AmpR, and OprD porin. Loss of bla PER-3 gene was induced in a PER-positive isolate by successive passages at 43°C without antibiotics. Results: Twenty-six of 287 isolates studied (9.1%) were CZA-resistant. Thirteen of 26 CZA-resistant isolates (50%) carried bla PER. One isolate carried bla PER but was CZA-susceptible. PER-producing isolates had significantly higher MICs for CZA, amikacin, gentamicin, ceftazidime, meropenem and ciprofloxacin than non-PER-producing isolates. All PER-producing isolates were ST309 and their bla PER-3 gene was associated to ISCR1, an insertion sequence known to mobilize adjacent DNA. PER-negative isolates were classified as ST41, ST235 (two isolates), ST395 and ST253. PER-negative isolates carried genes for narrow-spectrum ß-lactamases and the mutational resistome showed that all isolates had one major alteration in at least one of the genes analyzed. Loss of bla PER-3 gene restored susceptibility to CZA, ceftolozane/tazobactam and other ß-lactamsin the in vitro evolved isolate. Discussion: PER-3-producing ST309 P. aeruginosa is a successful multidrug-resistant clone with blaPER-3 gene implicated in resistance to CZA and other ß-lactams.

Mechanical and Antimicrobial Polyethylene Composites with CaO Nanoparticles.

Low-density polyethylene composites containing different sizes of calcium oxide (CaO) nanoparticles were obtained by melt mixing. The CaO nanoparticles were synthesized by either the sol-gel or sonication methods, obtaining two different sizes: ca. 55 nm and 25 nm. These nanoparticles were used either as-synthesized or were modified organically on the surface with oleic acid (Mod-CaO), at concentrations of 3, 5, and 10 wt% in the polymer. The Mod-CaO nanoparticles of 25 nm can act as nucleating agents, increasing the polymer's crystallinity. The Young's Modulus increased with the Mod-CaO nanoparticles, rendering higher reinforcement effects with an increase as high as 36%. The reduction in Escherichia coli bacteria in the nanocomposites increased with the amount of CaO nanoparticles, the size reduction, and the surface modification. The highest antimicrobial behavior was found in the composites with a Mod-CaO of 25 nm, presenting a reduction of 99.99%. This strong antimicrobial effect can be associated with the release of the Ca2+ from the composites, as studied for the composite with 10 wt% nanoparticles. The ion release was dependent on the size of the nanoparticles and their surface modification. These findings show that CaO nanoparticles are an excellent alternative as an antimicrobial filler in polymer nanocomposites to be applied for food packaging or medical devices.

A Multicenter Study To Evaluate Ceftaroline Breakpoints: Performance in an Area with High Prevalence of Methicillin-Resistant Staphylococcus aureus Sequence Type 5 Lineage.

Ceftaroline (CPT) is a broad-spectrum agent with potent activity against methicillin-resistant Staphylococcus aureus (MRSA). The sequence type 5 (ST5) Chilean-Cordobés clone, associated with CPT nonsusceptibility, is dominant in Chile, a region with high rates of MRSA infections. Here, we assessed the in vitro activity of CPT against a collection of MRSA isolates collected between 1999 and 2018 from nine hospitals (n = 320) and community settings (n = 41) in Santiago, Chile, and evaluated performance across testing methodologies. We found that our hospital-associated isolates exhibited higher CPT MIC distributions (MIC50 and MIC90 of 2 mg/liter) than the community isolates (MIC50 and MIC90 of 0.5 mg/liter), a finding that was consistent across time and independent of the culture source. High proportions (64%) of isolates were CPT nonsusceptible despite the absence of CPT use in Chile. Across methodologies, the Etest underestimated the MIC relative to the gold standard broth microdilution (BMD) test (MIC50 and MIC90 of 1 and 1.5 mg/liter, respectively). There was low (∼51%) categorical agreement (CA) between Etest and BMD results across CLSI and EUCAST breakpoints. The recent revision of CLSI guidelines abolished "very major error" (VME) from the previous guidelines (81%), which perform similarly to the EUCAST guidelines. The level of concordance between CLSI and EUCAST for BMD testing and Etest was >95%. Disk diffusion performed poorly relative to BMD under CLSI (CA, 55%) and EUCAST (CA, 36%) guidelines. Comparison of EUCAST to CLSI for disk diffusion (with EUCAST used as the reference) showed low agreement (CA, 25%; VME, 70%). In summary, CPT-nonsusceptible MRSA are dominant in clinical settings in Chile. Our results provide data to support the reevaluation of CPT breakpoints and to improve agreement across methodologies and agencies.

Detección de serinocarbapenemasas de clase A y otros mecanismos de resistencia enzimática a β-lactámicos en cepas de enterobacterias con susceptibilidad disminuida a carbapenémicos, aisladas de pacientes de un hospital universitario de Santiago, Chile / Detection of serinocarbapenemases Class A and other mechanisms of enzymatic resistance to β-lactams in enterobacteria strains with diminished susceptibility to carbapenems isolated of patients in a university hospital

Background: The emergence of carbapenemase mediated resistance in Enterobacteriaceae has a strong clinical impact. This study aimed to do a genotypic and phenotypic characterization of the enzymatic resistance to β-lactams in clinical Enterobacteriaceae isolates with decreased susceptibility to carbapenems in a university medical center in Santiago. Methods: During April-September 2010 at Hospital Clinico Universidad de Chile, 23 isolates of carbapenem non susceptible Enterobacteriaceae were collected. We used PCR for the detection of class A carbapenemases (SME, IMI, NMC, GES and KPC) and the modified Hodge with the boronic acid test to phenotypically assess the presence of serine-carbapenemases. To assess extended spectrum β-lactamases (ESBLs) the CLSI phenotypic tests were performed. Metallo-β-lactamases (MBL) and AmpC were assessed with commercial tablets. Results: 18/23 were Klebsiellapneumoniae and 5/23 strains were Enterobacter cloacae. All PCR to class A carbapenemases were negative. 3/23 strains (all E. cloacae), were positive to the Hodge modified test and 1/23, a K.pneumoniae, was positive to the boronic acid test. ESBLs were detected in 14/23 os the strains and AmpC in 5/23. No MBL was detected. Conclusion: No class A serine-carbapenemasa was detected. The decreased susceptibility to carbapenems is probably explained by the β-lactamase activity and due to porin loss.

Introducción: La emergencia de resistencia a β-lactámicos por carbapenemasas en enterobacterias tiene gran importancia clínica. El objetivo de este estudio fue caracterizar genotípica y fenotípicamente la resistencia enzimática a β-lactámicos en enterobacterias con susceptibilidad disminuida a carbapenémicos, en cepas aisladas de pacientes de un hospital universitario de Santiago. Metodología: Durante abril-septiembre 2010, en el Hospital Clínico de la Universidad de Chile se recolectaron 23 aislados. Se detectaron serinocarbapenemasas clase A (SME, IMI, NMC, GES y KPC) mediante RPC. Se empleó el test de Hodge modificado y acido fenil-borónico (APB) para la detección fenotípica de serinocarbapenemasas. Se detectó la presencia de β-lactamasas de espectro extendido según CLSI y AmpC y MBL mediante tabletas comerciales. Resultados: 18 cepas (78,26%) correspondieron a Klebsiella pneumoniae y 5 cepas (21,74%) a Enterobacter cloacae. Todas las RPC para serinocarbapenemasas fueron negativas, en tanto, el test de Hodge fue positivo para 3/23 cepas, todas E. cloacae. Una cepa de K. pneumoniae fue positiva para APB. Se detectó BLEE en 14/23 cepas, AmpC en 5/23 cepas y no se detectó MBL. Conclusiones: En las cepas estudiadas no se detectaron serinocarbapenemasas clase A. Probablemente los mecanismos que explican la susceptibilidad disminuida a carbapenémicos, involucran resistencia enzimática, combinados con cambios en la permeabilidad de la membrana bacteriana.

[Detection of serinocarbapenemases Class A and other mechanisms of enzymatic resistance to ß-lactams in enterobacteria strains with diminished susceptibility to carbapenems isolated of patients in a university hospital]. / Detección de serinocarbapenemasas de clase A y otros mecanismos de resistencia enzimática a ß-lactámicos en cepas de enterobacterias con susceptibilidad disminuida a carbapenémicos, aisladas de pacientes de un hospital universitario de Santiago, Chile.

BACKGROUND: The emergence of carbapenemase mediated resistance in Enterobacteriaceae has a strong clinical impact. This study aimed to do a genotypic and phenotypic characterization of the enzymatic resistance to ß-lactams in clinical Enterobacteriaceae isolates with decreased susceptibility to carbapenems in a university medical center in Santiago. METHODS: During April-September 2010 at Hospital Clinico Universidad de Chile, 23 isolates of carbapenem non susceptible Enterobacteriaceae were collected. We used PCR for the detection of class A carbapenemases (SME, IMI, NMC, GES and KPC) and the modified Hodge with the boronic acid test to phenotypically assess the presence of serine-carbapenemases. To assess extended spectrum ß-lactamases (ESBLs) the CLSI phenotypic tests were performed. Metallo-ß-lactamases (MBL) and AmpC were assessed with commercial tablets. RESULTS: 18/23 were Klebsiellapneumoniae and 5/23 strains were Enterobacter cloacae. All PCR to class A carbapenemases were negative. 3/23 strains (all E. cloacae), were positive to the Hodge modified test and 1/23, a K.pneumoniae, was positive to the boronic acid test. ESBLs were detected in 14/23 os the strains and AmpC in 5/23. No MBL was detected. CONCLUSION: No class A serine-carbapenemasa was detected. The decreased susceptibility to carbapenems is probably explained by the ß-lactamase activity and due to porin loss.