We have developed a method based on broad-range RT-PCR followed by electrospray ionization mass spectrometry (RT-PCR/ ESI-MS) for rapid and accurate detection of influenza virus, subspecies characterization, and early identification of genetic changes in circulating viruses. This method has previously been applied to detection of other pathogens in human clinical samples [19, 20, 21, 22] , but it has unique capabilities and advantages for influenza surveillance. Here, we show how a highthroughput assay incorporating eight parallel RT-PCR reactions followed by ESI-MS analysis can be used to simultaneously survey for all species of influenza viruses, provide clade-level resolution, identify mixed viral populations in the same sample, detect reassortants, and facilitate monitoring of viral evolution, all integral components of broad influenza surveillance.