The development of multicolour real-time PCR cyclers and "ready-to-use" commercial multiplex real-time PCR kits has also made it possible to combine several assays within a single tube. Major advantages of multiplexing include a reduced sample requirement, which is especially important when sample material is scarce [182, 183] , and the ability to combine assays with an internal control system. However, it is important to optimize these assays in order to limit competitive interactions that may significantly impact upon assay sensitivity. The combined properties of high sensitivity and specificity, low contamination risk, and speed have made real-time PCR technology a highly attractive alternative to tissue culture-or immunoassaybased methods for diagnosing many infectious diseases [184] . Clinical diagnostic applications and the use of realtime PCRs are growing exponentially, and real-time PCR is rapidly replacing conventional PCR and other established diagnostic methods such as antigen-ELISA and cell culture isolation.