title: Real-time PCR on the first galactomannan-positive serum sample for diagnosing invasive aspergillosis in liver transplant recipients.
abstract: Invasive aspergillosis (IA) is a life-threatening complication of liver transplantation. Detection of circulating galactomannan (GM) in serum samples is a method to improve the microbiological diagnosis in patients at risk for IA. However, the assay is hampered by false-positive results. The search for circulating Aspergillus DNA in the first GM-positive sample could improve the specificity of the test. Among 484 liver transplant recipients followed in a single center over 4 years, 25 patients had at least 1 GM-positive serum sample. The threshold of GM positivity was a ratio >or=1. These 25 patients were classified by the clinicians as probable IA (n=11), possible IA (n=2), and no IA (n=12) using the EORTC/MSG criteria with blinding to the polymerase chain reaction (PCR) results. After 1 mL aliquots of the first GM-positive serum sample were thawed, 2 independent DNA extractions were performed using the MagNA Pure Compact apparatus. Real-time amplification targeted at Aspergillus fumigatus mitochondrial DNA was performed on 10 microL of the final eluate in duplicate in the 2 independent DNA extractions using a LightCycler instrument. A sample was considered positive when the crossing point was <or=43 cycles in at least 2 out of the 4 replicates. Among the 13 probable or possible IA, 8 patients were PCR positive. The other 12 patients who had no IA were all PCR negative. Our data suggest that a concomitant real-time PCR performed on the first GM-positive sample improves the specificity of the first GM-positive assay result.
authors.affiliation: Assistance Publique-Hôpitaux de Paris, CHU Henri Mondor, Service de Parasitologie-Mycologie, Créteil, France. botterel@univ-paris12.fr
authors.forename: F
authors.initials: F
authors.lastname: Botterel
authors.forename: C
authors.initials: C
authors.lastname: Farrugia
authors.forename: P
authors.initials: P
authors.lastname: Ichai
authors.forename: J-M
authors.initials: JM
authors.lastname: Costa
authors.forename: F
authors.initials: F
authors.lastname: Saliba
authors.forename: S
authors.initials: S
authors.lastname: Bretagne
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D001228:Aspergillosis / Q000097:blood / Q000175:diagnosis* / Q000209:etiology; D001230:Aspergillus / Q000235:genetics / Q000302:isolation & purification*; D017931:DNA Primers; D004271:DNA, Fungal / Q000097:blood*; D005260:Female; D005690:Galactose / Q000031:analogs & derivatives; D006801:Humans; D016031:Liver Transplantation / Q000009:adverse effects*; D008297:Male; D008351:Mannans / Q000097:blood*; D008875:Middle Aged; D009894:Opportunistic Infections / Q000097:blood / Q000175:diagnosis* / Q000209:etiology; D016133:Polymerase Chain Reaction; D012189:Retrospective Studies; D012680:Sensitivity and Specificity; D055815:Young Adult
chemical_list: D017931:DNA Primers; D004271:DNA, Fungal; D008351:Mannans; C012990:galactomannan; D005690:Galactose