Polymerase Chain Reaction (PCR)-Based Tests

Abstract

PCR to aid in the diagnosis of invasive aspergillosis has been in use for over 15 years but little or no agreement over methodology has limited both widespread clinical use and large scale multi-centre evaluation. Each stage of the assay needs to be evaluated, including specimen type, extraction protocol, PCR amplification and result interpretation. The technical laboratory procedures are the least complicated to assess, although may be specimen-specific and recently efforts have been made to standardise methodology when testing blood specimens. The specimen type influences all downstream applications and will limit assay performance, particularly if the specimen is combined with unsuitable extraction processes. Advances in our knowledge of the disease process and host response may elucidate the optimal specimen but this will vary with disease type and the underlying condition of the host. Indeed, the underlying status of the patient may restrict the use of PCR if testing intrusive specimens is favoured. Large scale clinical evaluations using multiple specimen types and/or the testing of animal models will allow us to define the correct specimen type(s), optimise methodology accordingly and define guidelines for confident result interpretation. Only then will Aspergillus PCR gain widespread acceptance and use. This chapter describes the complexities related to each stage of Aspergillus PCR and analyses PCR performance based on specimen type, extraction protocol and PCR amplification to explicate what we currently know and reveal what research still needs to be performed.