Rapid detection of Hepatitis E virus RNA by reverse transcription-polymerase chain reaction using universal oligonucleotide primers

A rapid reverse transcription-polymerase chain reaction (RT-PCR) procedure for the detection of Hepatitis E virus (HEV) RNA in serum is described. Tot al nucleic acids are extracted from a small volume of human serum and rever se transcribed using random hexamers. An aliquot of cDNA is then utilized i n nested PCR employing degenerate HEV consensus primers. These primers are designed to sequences conserved between the Burma, Mexico, and US HEV strai ns, generating amplicons within each of the three open reading frames. Reac tions are analyzed by agarose gel electrophoresis and sampler showing an et hidium bromide stained band of the appropriate size in the first and second amplification, or in the second amplification only, are designated as posi tive. This protocol allows for the rapid and sensitive detection of HEV inf ection in human serum. (C) 1999 Elsevier Science B.V. All rights reserved.