Quality Control of the Qualitative Real Time PCR Method for the Detection of Aspergillus DNA in Clinical Samples

Introduction: The culture of clinical samples is positive in only 50% of the aspergillosis cases. In most neutropenic fever patients, anti-fungal treatment is generally commenced based on empirical considerations and reduces the growth rates in microbiological cultures. Polymerase chain reaction (PCR) screening for circulating DNA is not related to treatment in order for the DNA to be detected under antifungal effect. We evaluated the utility of qualitative real time PCR assay for the detection of Aspergillus DNA. Materials and Methods: DNA isolation was performed without a commercial system. The presence of DNA from the samples was detected using TaqMan PCR targeting fungal 28S rDNA gene. Diagnostic approach of the assay was evaluated by using different samples. Sixty samples including blood samples contaminated with Aspergillus conidia, colony suspensions, tissue and blood samples of experimentally infected animals, and patient samples obtained from suspected cases were subjected to testing. Results: False negative results were obtained from tissue samples of experimentally infected animals. When the results were analysed, 4 (40%) of the tissue samples, 3 (30%) of the blood samples, 2 (20%) of the simulated blood samples, and 1 (10%) of the patient samples failed to achieve the same level of detection. Colony suspensions were found to be all positive (100%). Conclusion: The efficiency of the Aspergillus PCR is limited in tissue samples, but sufficient in blood samples, and should be qualified to exclude false negativity. Clinical studies with large number of samples should be initiated to integrate PCR in the diagnostic armentarium of aspergillosis.