Mixed viral infection identified using heteroduplex mobility analysis (HMA)

It is now recognised that mixed viral infection, or infection of an individ ual with two or more distinct strains of a single viral species, often occu rs particularly with RNA viruses. Current methods for detection of mixed in fection normally involve genotyping or cloning and DNA sequencing. These me thods are not always accurate or sensitive at detecting mixed infection and cannot be used for large numbers of samples. Furthermore subsequent sequen ce determination of the coinfecting viruses is labour intensive. This paper describes a simple, generic method based upon PCR and heteroduplex mobilit y analysis (HMA) that can be used to rapidly determine mixed infection with two strains of the same virus. The utility of this method is illustrated w ith hepatitis C virus (HCV) and TT virus (TTV) as examples. PCR-HMA detecte d mixed infection in 3 (8%) of 38 sera from intravenous drug users (IVDU) a nd 28 (30%) of 70 TTV-positive sera from Australia, China, and Vietnam. HMA can also be used to screen recombinant colonies to identify the sequences of the coinfecting viruses. The methods described here could be applied to analyse any PCR product containing two or more divergent sequences, whether derived from viruses, bacteria, or eukaryotic organisms. (C) 2000 Academic Press.