Real-time PCR on the First Galactomannan-positive Serum Sample for Diagnosing Invasive Aspergillosis in Liver Transplant Recipients

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.