Adenoviral load diagnostics by quantitative polymerase chain reaction: techniques and application.
Human adenoviruses (HAdV) can cause fatal complications such as disseminated disease especially in a post-transplant setting. With conventional methods, disseminated HAdV disease could only be diagnosed with delay. Quantification of the HAdV load by real-time PCR in peripheral blood promised to solve this diagnostic dilemma. Here we review the development, applications and significance of quantitative HAdV PCR. The high genetic divergence of the 56 HAdV types was a major obstacle for developing a quantitative HAdV PCR covering all types. Several protocols focused either on a few, probably predominating types or tried to detect all known HAdV types by using a bundle of assays or a few multiplexed PCRs. Alternatively, generic quantitative real-time HAdV PCR protocols using primer and probe consensus sequences have been designed, providing considerable reduction of costs and hands-on time. Application of HAdV load testing by several studies on stem cell transplant (SCT) recipients indicated that rapidly increasing HAdV blood loads as well as high HAdV DNAemia (e.g. >10(4) copies/ml) are predictive for disseminated HAdV disease although a universal threshold value has not yet been established. HAdV load testing has been implemented for systematic screening of SCT patients permitting early diagnosis, pre-emptive treatment initiation and monitoring of antiviral therapy. However, further investigations are required to validate proposed virus load thresholds. Moreover, other applications of quantitative HAdV PCR, such as the diagnosis of localized HAdV disease, the analysis of environmental samples and monitoring of gene therapy with adenoviral vectors will be addressed in this review. Copyright © 2011 John Wiley & Sons, Ltd.