Objective: To establish a method for simultaneous detection of norovirus (NV), rotavirus (RV), astrovirus (AV) and hepatitis A virus (HAV) by multiplex reverse transcription-polymerase chain reaction (RT-PCR).
Methods: Specific primers of the four viruses were designed based on the high conserved sequences, the reaction system and conditions optimized and the specificity and sensitivity confirmed. The method was then applied to detect the four viruses in clinical samples.
Results: The steady detection limits were 100 pg/ml for hepatitis A virus, 50 pg/ml for rotavirus, norovirus and astrovirus respectively. When the developed method was used to detect clinical fecal samples, 62 (48.44%) were identified as rotavirus, 8 (6.25%) as norovirus, 11 (8.59%) as astrovirus and 4 (3.12%) as hepatitis A virus in a total of 128 samples.
Conclusion: Data from our study showed that multiplex RT-PCR system could be used to simultaneously detect the four viruses in routine monitoring and risk assessment in disease outbreaks with high specificity and sensitivity.