Long-term Expansion of Porcine Intestinal Organoids serves as an Ex Vivo Model for Swine Enteric Coronavirus infection

We designed a long-term culture system for porcine intestinal organoids from intestinal crypt or single Lgr5+ stem cells by combining previously defined insights in the growth requirements of intestinal epithelium of human and mouse. We showed that long-term cultured swine intestinal organoids were expanded in vitro more than six months at least and maintained the potential to differentiate into different types of cells. These organoids were successfully infected with porcine enteric coronavirus including porcine epidemic diarrhea virus (PEDV) and transmissible gastroenteritis virus (TGEV). RNA-seq analysis showed that robust induction of transcripts associated with antiviral signaling in response to enteric coronavrius infection, including a number of interferon-stimulated genes and cytokines. Moreover, gene set enrichment analysis indicated that PEDV infection could suppress immune response in organoids. This 3D intestinal organoid model offers a long-term, renewable resource for investigating porcine intestinal infections with a variety of pathogens. Overall design: To better understanding how porcine intestinal organoids respond to porcine coronavirus infection, we performed RNA sequencing (RNA-seq) experiments using porcine intestinal organoids infected with 0.1 MOI of PEDV, TGEV or MOCK infected and evaluated the transcript landscape of porcine small intestinal organoids at 48 hrs after viral challenges. Total RNAs were isolated and sequencing libraries were prepared. The libraries were sequenced using standard methods for paired-end 150-bp reads, on an Illumina Novaseq 6000 sequencer.