Scientists from Erasmus University in the Netherlands have designed and evaluated a set of molecular assays that they say are a sensitive and good alternative for conventional diagnostic methods and can produce results in one day without the need for additional equipment. The results were recently published in The Journal of Molecular Diagnostics.
The investigators report on a set of real-time polymerase chain reaction (RT-PCR) assays for quantification and subtyping of human influenza A and B viruses, as well as four assays for detecting drug resistant mutations.
The influenza quantification assay was used to check for virus positivity and to obtain virus particle counts for all analyzed samples. Influenza A viruses were then subtyped and tested for presence of oseltamivir resistance mutations using the resistance RT-PCR assays. In total, 129 respiratory specimens tested positive for influenza A and 60 for influenza B virus. One sample tested positive for both virus types.
“RT-PCR based assays have become the standard in most diagnostic laboratories worldwide in recent years,” comments lead investigator Martin Schutten, PhD, Head of the Clinical Virology Unit at the Erasmus University, Rotterdam, the Netherlands. “The assays described here cover all currently circulating human influenza viruses and can detect major resistance mutations to oseltamivir. By introducing external quantification and internal standards, longitudinal assay performance can be monitored carefully and a virus particle count can be assigned to an analyzed sample.
The scientists state that the algorithm can generate useful data to assist in the management of individual influenza virus infected patients and to evaluate clinical trials. Information regarding influenza virus type and subtype, viral load and antiviral susceptibility can be obtained within one working day. “These assays are a powerful tool for the clinical management of influenza virus infected patients,” he concludes,” adds Schutten.
Although infection from H7N9, the new potential pandemic Influenza strain, or H5N1, a continuing pandemic threat since 1997, can be identified by exclusion (positive in the Influenza matrix RT-PCR but negative in RT-PCR typing), development of rapid typing RT-PCR for these potential pandemic viruses may be useful in complementing the existing set.