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Clostridium difficile infection (CDI) has become the main cause of diarrhea-related diseases in domestic (China) inpatients. High-sensitivity and high-specificity detection methods for CDI must be applied clinically for CDI supervisory control. In this paper, we introduce a detection method for C. difficile and Norovirus based on real-time PCR. We developed and optimized a primer–probe for Norovirus targets tcdA and tcdB with remarkably increased detection sensitivity. We then used this method in an integrated cassette, and found increased detection efficiency for Norovirus standards in the cassette compared to C. difficile samples. These results provide a basis for further exploration of automatic testing system design.