Detection, Identification, and Distribution of Fungi in Bronchoalveolar Lavage Specimens by Use of Multilocus PCR Coupled with Electrospray Ionization/Mass Spectrometry

Abstract

dAs pulmonary fungal infections continue to increase due to an increasing number of immunocompromised patients, rapid detection and accurate identification of these fungal pathogens are critical. A broad fungal assay was developed by incorporating broad-range multilocus PCR amplification and electrospray ionization/mass spectrometry (PCR/ESI-MS) to detect and identify fungal organisms directly from clinical specimens. The aims of this study were to evaluate the performance of PCR/ESI-MS for detection, identification, and determination of the distribution of fungal organisms in bronchoalveolar lavage (

Broad-Range Direct Detection and Identification of Fungi by Use of the PLEX-ID PCR-Electrospray Ionization Mass Spectrometry (ESI-MS) System

“…Our results are comparable to those of a study performed by Shin et al that examined 691 nonduplicate BAL fluid specimens by using the Broad Fungal assay on the PLEX-ID platform (12).…”

“…Previous published studies with the PLEX-ID system have used specimens sent to the manufacturer's facility for analysis by the personnel there (12). Thus, this study is the first to assess the performance of the PLEX-ID system in a clinical mycology laboratory.…”

“…PCRs were assembled by adding 10 l of extracted nucleic acid to each well of the 96-well microtiter Broad Fungal assay plate by using the Corbett Robotics CAS 1200 (QUIagility, Qiagen, Valencia, CA) fluid handling robotic platform. The Broad Fungal assay targets and primers have been previously described (12). The plates were sealed with PLEX-ID PCR seals (Abbott Molecular, Des Plaines, IL) and loaded onto an Eppendorf Mastercycler ProS Thermal Cycler (Eppendorf, Hamburg, Germany).…”

Clinical chorioamnionitis at term: the amniotic fluid fatty acyl lipidome

“…Briefly, DNA was extracted from 300 µl of amniotic fluid using a method that combines beadbeating cell lysis with a magnetic-bead-based extraction method (38,39). The extracted DNA was amplified by the previously described broad bacteria and candida detection assay, according to the manufacturer's instructions.…”

Broad-Spectrum Biosensor Capable of Detecting and Identifying Diverse Bacterial and Candida Species in Blood

“…Genome preparation and PCR. The PCR/ESI-MS system used in this study included automated sample lysis, nucleic acid extraction, PCR, PCR cleanup (desalting), mass spectrometry, and bioinformatic data analysis components, as previously described (Ibis Biosciences, Carlsbad, CA) (13,14). Strict separation of PCR/sample set-up areas and amplification/analysis areas (personnel/equipment movement and airflow) was employed to prevent backflow of amplicons from the thermocycler and mass spectrometer components, according to Clinical and Laboratory Standards Institute (CLSI) guidelines for open PCR processes.…”