Browsing by Author "Olivares, Jorge"
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Publication Ceftazidime/avibactam resistance is associated with PER-3-producing ST309 lineage in Chilean clinical isolates of non-carbapenemase producing Pseudomonas aeruginosa(2024) Soto, Katherine; Alcalde, Manuel; Ugalde, Juan; Olivares, Jorge; Quiroz, Valeria; Brito, Bárbara; Rivas Jiménez, Lina María; Munita, Jose M.; García, Patricia; Wozniak, AnielaIntroduction: 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 β-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 β-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 β-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 β-lactams.Item Comparación del número de especies detectadas por los métodos de diagnóstico para DEMODEX SPP. en pacientes con rosácea de Clínica Alemana de Vitacura, año 2022(2023) Di Gennaro, Angelo; Olivares, Jorge; Olivares, TomásDemodex spp. es un ectoparásito microscópico que forma parte de la microbiota de la piel del ser humano, sin embargo, su presencia se ha relacionado a un rol patógeno en la Rosácea, que es una enfermedad inflamatoria cutánea crónica que se presenta con flushing, eritema persistente, telangiectasias, pápulas y pústulas en la zona facial. Para su diagnóstico, se pueden utilizar dos métodos, la Biopsia de Superficie Cutánea Estandarizada y el Examen Microscópico Directo, útiles para tomar muestras de piel en las que se pueda observar este ectoparásito, pero se desconoce, actualmente, cuál de ellos detecta mayor cantidad de ácaros en pacientes con Rosácea. Objetivo: Determinar cuál método detecta significativamente una mayor cantidad de ácaros en pacientes con Rosácea de la Clínica Alemana de Vitacura, Santiago de Chile. Metodología: Se aplicaron los métodos de Biopsia de Superficie Cutánea Estandarizada y Examen Microscópico Directo en 30 pacientes diagnosticados con Rosácea, para luego hacer el conteo de ácaros al microscopio, que fue registrado posteriormente en una base de datos, obteniendo así el promedio de los ácaros pesquisados con ambos métodos y realizando un análisis estadístico. Resultado: El método de Biopsia de Superficie Cutánea Estandarizada detecta significativamente mayor cantidad de ácaros que el método de Examen Microscópico Directo en pacientes con Rosácea. Conclusión: el método de Biopsia de Superficie Cutánea Estandarizada posee mayor detección de Demodex spp. en pacientes con Rosácea, lo que influye en la positividad y en la obtención del tratamiento oportuno, por ende, es importante implementarlo en el diagnóstico.Publication Genetic characterization of clinically relevant class 1 integrons carried by multidrug resistant bacteria (MDRB) isolated from the gut microbiota of highly antibiotic treated Salmo salar(2022) Vásquez, Felipe; Higuera, Sebastián; Parás, Juan; Cortés, Jimena; Opazo, Andrés; Ugalde, Juan; Alcalde, Manuel; Olivares, JorgeObjectives: The main objective of this study was the genetic characterization of clinically relevant class 1 integrons carried by multidrug resistant bacteria isolated from the intestinal microbiota of aquaculture salmon treated with high concentrations of antibiotics. Methods: In 82 multidrug resistant bacterial isolates, the prevalence of both the conserved elements of the integrons, qacEΔ1 and sul1 genes, and the variable region (VR) was determined. Further, whole genome sequencing and complete genetic analysis was performed in VR-positive isolates. Results: Despite the fact that 100% of the bacterial isolates presented the intI1 gene, only 12.3% carried the qacEΔ1 and sul1 genes and only two (2.4%) presented a VR with gene cassettes. In the Pseudomonas baetica 25P2F9 isolate, a VR carrying aac(6')31, qacH, and blaOXA-2 gene cassettes was described, whereas the VR of Aeromonas salmonicida 30PB8 isolate showed a dfrA14 gene cassette. The array of gene cassettes found in the Pseudomonas isolate appears with high frequency in clinically relevant pathogens such as Pseudomonas aeruginosa or Escherichia coli. Additionally, it was possible to determine that these integrons are contained in plasmids and coul be easily transferred. Resistome analysis demonstrated that both isolates carried a great diversity of antibiotic resistance genes, including many β-lactamases. Even in the Aeromonas isolate a new oxacillin-hydrolyzing beta-lactamase gene was described (blaOXA-956). Conclusion: The presence of multidrug resistant bacteria and clinically relevant genetic elements in the salmon intestinal microbiota make the aquaculture a hotspot in the phenomenon of antibiotic resistance; therefore, the control of antibiotics used in this activity is a key point to avoid its escalation.Item Real-world Performance of Susceptibility Testing for Ceftolozane/Tazobactam Against Non-Carbapenemase-Producing Carbapenem-Resistant Pseudomonas aeruginosa(2021) Rivas, Lina; Alcalde-Rico, Manuel; Martínez, José R.; Moreno, Victoria; Rojas, Pamela; Wozniak, Aniela; García, Patricia; Olivares, Jorge; Miller, William R.; Arias, Cesar A.; Khan, Ayesha; Munita, JoséCeftolozane/tazbactam (C/T) is a potent anti-pseudomonal agent that has clinical utility against infections caused by non-carbapenemase producing carbapenem-resistant P. aeruginosa (non-CP-CR-PA). Accurate, precise and reliable antimicrobial susceptibility testing (AST) is crucial to guide clinical decisions. However, studies assessing the performance of different AST methods against non-CP-CR-PA- (the main clinical niche for C/T), are lacking. Here, we evaluated performance of gradient strips (Etest and MIC test strip (MTS), and disk diffusion (DD) using CLSI breakpoints. Additionally, we assessed the performance of DD using EUCAST breakpoints. For all susceptibility tests, we used a collection of 97 non-CP-CR-PA clinical isolates recovered from 11 Chilean hospitals. Both gradient strips and DD had acceptable performance when using CLSI breakpoints, yielding a categorical agreement (CA) of >90% and 92%, respectively. In contrast, DD using EUCAST breakpoints performed sub-optimally (CA 81%). MTS yielded a higher essential agreement (EA, >90%) than Etest (84%). Importantly, the performance of all methods varied significantly when the isolates were stratified by their degree of susceptibility to other anti-pseudomonal β-lactams. All methods had 100% CA when testing isolates that were pan-susceptible to all β-lactams (Pan-β-S). However, the CA markedly decreased when testing isolates resistant to all β-lactams (Pan-β-R). Indeed, the CA was 81% for Etest (6 errors), 78% for MTS (7 errors) and 78% and 56% for DD when using CLSI (7 errors) or EUCAST breakpoints (14 errors), respectively. Our results suggest that all manual AST methods have strikingly decreased performance in the context of Pan-β-R P. aeruginosa with potentially major clinical implications.