Using the GoFlowChip to understand SARS-CoV-2 infection of the gastrointestinal mucosa of humans and bats

Summary The emerging severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a highly infectioushuman respiratory tract pathogen that has the capacity to infect many different organ systems, including thegastrointestinal (GI) tract. The overall goal of this project is to elucidate the role of the GI mucosa in SARS-CoV-2 infection and transmission. Specifically, we seek to identify mucosal immune mechanisms involved inCOVID-19 pathogenesis and compare epithelial responses to SARS-CoV-2 in the gut of humans and bats.Investigating bat cells is important, because bats are considered the original hosts for SARS-CoV-2, but do notdevelop overt disease upon infection. Our recently developed GOFlowChip platform, which integrates 3-dimensional GI organoids and immune cells on a millifluidic chip with luminal and basolateral flow capacity, isideally suited to probe the mechanisms involved in SARS-CoV-2 infection of the gut mucosa. Here, wepropose to utilize the GOFlowChip to elucidate mechanisms of viral infection, replication and spread involvedin SARS-CoV-2 infection of the GI tract. Specifically, we seek to (1) Define how SARS-CoV-2 infection of theGI epithelium contributes to viral spread. (2) Determine to what extent GI mucosal immune mechanismsregulate SARS-CoV-2 infection and spreading in the GI tract. (3) Compare GI epithelial responses to SARS-CoV-2 between bats and humans. For Specific Aim 1, we will optimize the GOFlowChip for use under BSL-3laboratory conditions, followed by SARS-CoV-2 infection experiments that will elucidate susceptibility and viralreplication dynamics in different gut compartments. Specific Aim 2 will leverage our organoid-mononuclearphagocyte (MNP) co-culture system to elucidate whether MNP-dependent transport mechanisms impact viralinvasion of the GI mucosa, and we will screen SARS-CoV-2 reactive patient sera for their ability to modulategut infection. For Specific Aim 3, we will establish organoid lines from bat GI tissues in order to compareepithelial responses to SARS-CoV-2 in bats and humans using RNA sequencing in order to understanddifferences in viral pathogenicity between humans and bats. The proposed work is directly integrated with ourexisting project, because we will use out GOFlowChip to investigate how SARS-CoV-2 interacts with epithelialcells and immune system components to cross the GI barrier. Our research is technologically innovative,because we are using gut organoid-immune cell co-cultures in a fully contained tissue chip design to study aBSL-3 level pathogen. Our project is conceptually innovative, because it is the first to compare gut organoidsfrom Jamaican fruit bats to human organoids for their ability to sustain SARS-CoV-2 infection and to mountcellular antiviral responses. The proposed research is significant because it will yield crucial information on therole of the GI mucosa in COVID-19 pathogenesis that may inform future strategies to identify viral carriers,prevent viral transmission, and design novel antiviral treatments and vaccines.