Newer methods such as PCR are being investigated in order to improve the diagnosis of invasive
aspergillosis. One of the major obstacles to using PCR to diagnose
aspergillosis is a reliable, simple method for extraction of the
fungal DNA. The presence of a complex, sturdy cell wall that is resistant to lysis impairs extraction of the
DNA by conventional methods employed for bacteria. Numerous
fungal DNA extraction protocols have been described in the literature. However, these methods are time-consuming, require a high level of skill and may not be suitable for use as a routine diagnostic technique. Here, a number of extraction methods were compared: a freeze-thaw method, a freeze-boil method,
enzyme extraction and a bead-beating method using Mini-BeadBeater-8. The quality and quantity of the
DNA extracted was compared using real-time PCR. It was found that the use of a bead-beating method followed by extraction with AL
buffer (Qiagen) was the most successful extraction technique, giving the greatest yield of
DNA, and was also the least time-consuming method assessed.