Clostridium perfringens is a Gram-positive anaerobic bacterium that causes a variety of toxin-mediated diseases including human clostridial myonecrosis or gas gangrene. To obtain an insight into host-pathogen interactions in clostridial myonecrosis we carried out comparative RNA-seq analysis, measuring the expression of both bacterial and murine genes in a murine C. perfringens infection model. We compared the in vivo bacterial expression levels obtained from the wild-type strain with expression levels from its equivalent in vitro culture. The results showed that in the wild-type strain 32% (888) of thegenes were differentially expressed in vivo (FDR <0.01, fold change >4). Of these genes, 53% were up-regulated and 57% were down-regulated. Unexpectedly, the genes encoding the key toxins alpha-toxin and perfringolysin O were expressed at a lower relative level in the early stages of infection compared to the equivalent in vitro culture.The gene encoding an MprF-like flippase was highly up-regulated (227x) in vivo. In another bacterium this flippase is critical for membrane phospholipid layer assembly and provides protection from host cationic antimicrobial peptides, but its function in C. perfringens is unknown. Analysis of the murine transcriptome from the wild-type infection indicated that 270 genes were up-regulated compared to the transcriptome from control mock-infected mice. KEGG pathway analysis of these genes revealed an enrichment of Toll-like receptor (TLR) and Nod-like receptor (NLR) signaling components. Real-time PCR validation suggested that TLR2 and NLRP3 inflammasome signalling was activated in myconecrosis infections. In conclusion, we report the first successful expression profiling of both bacterial and host genes during an active C. perfringens infection. These studies have identified other bacterial genes that may be involved in the disease process and have provided novel insights into host-pathogen interactions.