Optimisation of a single-primer sequence-independent amplification (SP-SIA) assay: detection of previously undetectable norovirus strains associated with outbreaks of gastroenteritis

The introduction of molecular diagnostic methods for investigation of gastroenteritis has significantly reduced the diagnostic gap. However, approximately 25% of cases of gastroenteritis remain undiagnosed even after screening for bacteria, parasites and viruses using the most sensitive PCR and RT-PCR methods available. In recent years, it has become apparent that viruses are responsible for the majority of outbreaks of gastroenteritis. In this study, a panel of samples from outbreaks of gastroenteritis for which no aetiological agent had been identified was selected for investigation by random amplification molecular methods. An algorithm for virus purification and concentration was developed followed by a single-primer sequence-independent amplification method. These methods resulted in the identification of viruses in 5 out of 51 previously negative outbreaks. Noroviruses undetectable using two available broadly reactive diagnostic methods were detected in 4 of these outbreaks.