Espectrometría de masas matrix-assisted laser desorption ionization time-of-flight vs. metodología convencional en la identificación de Candida no-albicans / Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry vs conventional methods in the identification of Candida non-albicans

Introducción La infección por Candida se ha convertido en un importante problema de salud en todo el mundo. La epidemiología de la candidemia se ha modificado considerablemente por la emergencia de las especies de Candida no-albicans. Esta variación tiene especial importancia en la elección de la profilaxis y tratamiento empírico. Los métodos bioquímicos y los basados en biología molecular presentan limitaciones para la identificación correcta de las especies de Candida. El objetivo de este estudio es demostrar la capacidad del sistema de espectrometría de masas MALDI-TOF para la identificación de estas especies y compararlo con la tecnología utilizada en la actualidad. Métodos Se incluyeron todos los aislados recogidos durante dos años (n=73) de Candida no-albicans procedentes de muestras invasivas. La identificación se realizó mediante los sistemas Vitek-2 YST y API CAUX. Las identificaciones del MALDI-TOF se hicieron con el sistema Axima Confidence (Shimadzu Corporation), utilizando el software Shimadzu Launchpad y la base de datos SARAMIS (AnagnosTec GmbH). Las discrepancias se resolvieron mediante PCR multiplex LightCycler SeptiFast, PCR específica de C. glabrata y digestión enzimática con BanI del fragmento SADH en los aislados de C. parapsilosis. Resultados De los 73 aislados de Candida no-albicans, los métodos bioquímicos identificaron de forma concluyente 67 a nivel de especie y 6 a nivel de género. El sistema MALDI-TOF obtuvo identificaciones a nivel de especie en todas ellas. La correlación en la especie de todos los aislados estudiados fue del 85,07%, llegando al 94,52% si se estudia la correlación entre la identificación obtenida mediante métodos bioquímicos junto con los métodos empleados para el análisis de las discrepancias. En los aislados de C. parapsilosis, el sistema MALDI-TOF obtuvo una identificación de C. orthopsilosis en tres de ellos, confirmándose por digestión con BanI del fragmento SADH (..) (AU)

Introduction: Candida infection has become a major health problem worldwide. The epidemiology of Candidaemia has substantially changed by the emergence of the species Candida non-albicans. This variation is particularly important in the choice of prophylaxis and empirical treatment. The methods based on biochemical and molecular biology have limitations for the correct identification of Candida species. The aim of this study is to demonstrate the ability of the MALDI-TOF mass spectrometry for the identification of these species and compare it with the technology used today. Methods: We included all isolates collected over 2 years (n=73) of Candida non-albicans from non-invasive samples. The identification was carried out by Vitek-2 systems YST and API CAUX. The MALDI-TO Fidentifications were made with Confidence Axima system (Shimadzu Corporation) using the Shimadzu Launchpad software and database SARAMIS (AnagnosTec GmbH). Discrepancies were resolved by Septi-Fast Light Cycler multiplex PCR, specific PCR C. glabrata and enzymatic digestion with BanI SADH fragment in isolates of C. parapsilosis. Results: Of the 73 isolates of Candida non-albicans, the biochemical methods conclusively identified 67 to species level and 6 at the genus level. The MALDI-TOF system obtained identifications at the species level in all cases. The correlation in the species of all isolates studied was 85.07%, reaching 94.52% when the correlation was made between the identification obtained by biochemical methods and the methods for the analysis of the discrepancies. In isolates of C. parapsilosis, MALDI-TOF system obtained an identification (..) (AU)

[Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry vs conventional methods in the identification of Candida non-albicans]. / Espectrometría de masas matrix-assisted laser desorption ionization time-of-flight vs. metodología convencional en la identificación de Candida no-albicans.

INTRODUCTION: Candida infection has become a major health problem worldwide. The epidemiology of Candidaemia has substantially changed by the emergence of the species Candida non-albicans. This variation is particularly important in the choice of prophylaxis and empirical treatment. The methods based on biochemical and molecular biology have limitations for the correct identification of Candida species. The aim of this study is to demonstrate the ability of the MALDI-TOF mass spectrometry for the identification of these species and compare it with the technology used today. METHODS: We included all isolates collected over 2 years (n=73) of Candida non-albicans from non-invasive samples. The identification was carried out by Vitek-2 systems YST and API CAUX. The MALDI-TOF identifications were made with Confidence Axima system (Shimadzu Corporation) using the Shimadzu Launchpad software and database SARAMIS (AnagnosTec GmbH). Discrepancies were resolved by SeptiFast LightCycler multiplex PCR, specific PCR C. glabrata and enzymatic digestion with BanI SADH fragment in isolates of C. parapsilosis. RESULTS: Of the 73 isolates of Candida non-albicans, the biochemical methods conclusively identified 67 to species level and 6 at the genus level. The MALDI-TOF system obtained identifications at the species level in all cases. The correlation in the species of all isolates studied was 85.07%, reaching 94.52% when the correlation was made between the identification obtained by biochemical methods and the methods for the analysis of the discrepancies. In isolates of C. parapsilosis, MALDI-TOF system obtained an identification of C. orthopsilosis. In 3 of them it was confirmed by digestion with BanI SADH fragment. CONCLUSION: This study has demonstrated the use of mass spectrometry (MALDI-TOF system) to provide the microbiology laboratory with greater efficiency and reliability to identify isolates of Candida non-albicans to species level. It also shows its potential usefulness in identifying related species, such as C. parapsilosis, metapsilosis and orthopsilosis.

Emergence in Spain of a multidrug-resistant Enterobacter cloacae clinical isolate producing SFO-1 extended-spectrum beta-lactamase.

Between February 2006 and October 2009, 38 patients in different wards at the A Coruña University Hospital (northwest Spain) were either infected with or colonized by an epidemic, multidrug-resistant (MDR), and extended-spectrum-ß-lactamase (ESBL)-producing strain of Enterobacter cloacae (EbSF), which was susceptible only to carbapenems. Semiautomated repetitive extragenic palindromic sequence-based PCR (rep-PCR) and pulsed-field gel electrophoresis (PFGE) analysis revealed that all of the E. cloacae isolates belonged to the same clone. Cloning and sequencing enabled the detection of the SFO-1 ESBL in the epidemic strain and the description of its genetic environment. The presence of the ampR gene was detected upstream of bla(SFO-1), and two complete sequences of IS26 surrounding ampR and ampA were detected. These IS26 sequences are bordered by complete left and right inverted repeats (IRL and IRR, respectively), which suggested that they were functional. The whole segment flanked by two IS26 copies may be considered a putative large composite transposon. A gene coding for aminoglycoside acetyltransferase (gentamicin resistance gene [aac3]) was found downstream of the 3' IS26. Despite the implementation of strict infection control measures, strain EbSF spread through different areas of the hospital. A case-control study was performed to assess risk factors for EbSF acquisition. A multivariate analysis revealed that the prior administration of ß-lactam antibiotics, chronic renal failure, tracheostomy, and prior hospitalization were statistically associated with SFO-1-producing E. cloacae acquisition. This study describes for the first time an outbreak in which an SFO-1-producing E. cloacae strain was involved. Note that so far, this ß-lactamase has previously been isolated in only a single case of E. cloacae infection in Japan.