title: Volume dependency for culture of fungi from respiratory secretions and increased sensitivity of Aspergillus quantitative PCR.
abstract: Diagnosis of aspergillosis is often difficult. We compared fungal yields from respiratory specimens using the Health Protection Agency standard culture method (BSOP57), a higher volume undiluted culture method Mycology Reference Centre Manchester (MRCM) and Aspergillus quantitative real time polymerase chain reaction (qPCR). Sputum, bronchial aspirate and bronchoalveolar lavage (BAL) samples (total 23) were collected from aspergillosis patients. One fraction of all samples was cultured using the MRCM method, one BSOP57 and one was used for qPCR. The recovery rate for fungi was significantly higher by MRCM (87%) than by BSOP57 (8.7%) from all 23 specimens. Sputum samples were 44% positive by MRCM compared to no fungi isolated (0%) by BSOP57. Bronchial aspirates were 75% positive by MRCM and 0% by BSOP57. BAL samples were positive in 20% by MRCM and 10% by BSOP57. qPCR was always more sensitive than culture (95.6%) from all samples. In general, over 100 mould colonies (81 Aspergillus fumigatus) were grown using the MRCM method compared with only one colony from BSOP57. This study provides a reference point for standardisation of respiratory sample processing in diagnostic laboratories. Culture from higher volume undiluted respiratory specimens has a much higher yield for Aspergillus than BSOP57. qPCR is much more sensitive than culture and the current UK method requires revision.
authors.affiliation: Institute of Inflammation and Repair, Education and Research Centre, Manchester Academic Health Science Centre (MAHSC), University of Manchester, Manchester, UK; Mycology Reference Centre, Education and Research Centre, University Hospital of South Manchester (UHSM), Manchester, UK.
authors.forename: Marcin G
authors.initials: MG
authors.lastname: Fraczek
authors.forename: Marie B
authors.initials: MB
authors.lastname: Kirwan
authors.forename: Caroline B
authors.initials: CB
authors.lastname: Moore
authors.forename: Julie
authors.initials: J
authors.lastname: Morris
authors.forename: David W
authors.initials: DW
authors.lastname: Denning
authors.forename: Malcolm D
authors.initials: MD
authors.lastname: Richardson
mesh_terms: D001228:Aspergillosis / Q000175:diagnosis* / Q000382:microbiology; D001230:Aspergillus / Q000235:genetics / Q000302:isolation & purification*; D012634:Bodily Secretions / Q000382:microbiology; D001992:Bronchoalveolar Lavage Fluid / Q000382:microbiology; D006801:Humans; D008828:Microbiological Techniques / Q000379:methods*; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013048:Specimen Handling / Q000379:methods*; D006113:United Kingdom
keywords: Aspergillus; bronchoalveolar lavage; culture; qPCR; sputum