Abstract: Objective: To establish a one-step reverse transcriptase droplet digital polymerase chain reaction (RT-ddPCR) assay for the detection of rotavirus (RV) group A strains. Methods: A set of specific primers and probes was selected for RT-ddPCR. The specificity of this assay was evaluated with RNA samples of other foodborne viruses. A reference material containing the target fragment of RV group A strains was prepared and diluted into several gradients for use in evaluation of the limit of quantitation (LOQ), accuracy, repeatability and reproducibility. Artificially positive samples were prepared by adding the diluted positive fetal sample to different food matrices. Viruses were separated and concentrated from the artificially positive samples, and viral RNA was extracted sufficiently. The effect of food matrices on the quantitative performance of RT-ddPCR was investigated. Results: The RT-ddPCR assay displayed a high accuracy, sensitivity, repeatability and reproducibility. Its LOQ ranged from 58 000.00 to 5.80 copies/μL. Statistically significant disparities (P < 0.05) in the detectable amount and recovery of virus among different food matrices were discovered at each contamination level. Conclusion: RT-ddPCR could quantitatively and accurately detect RNA of RV group A strains. However, the quantitative results were sufficiently affected by the type of food matrixes. The load of RV group A strains in food could be calculated from the recovery rate.

Key words: rotavirus group a strain, one-step reverse transcriptase droplet digital polymerase chain reaction, quantitative detection, food matrix