Bile acids accumulate norovirus-like particles and enhance binding to and entry into human enteric epithelial cells

Human norovirus (HuNoV) is a leading cause of acute viral gastroenteritis, but despite high impact on public health and health care, the mechanisms of viral attachment to and entry into target cells are not yet fully understood. It is well known that body fluids such as blood can transmit unrelated viruses, but recent reports also indicate that saliva and bile contribute to transmission of HuNoV. For example, human bile acids increase cell surface ceramide levels in human enteroids, which improves norovirus entry into cells resulting in enhanced replication. Bile acids can also interact directly with the norovirus capsid, but it is not known whether bile or other gastrointestinal body fluids directly affect HuNoV attachment to host cells. In this study, we investigated the effects of patient-derived gastric juice, pancreatic juice, and bile on HuNoV GII.4 virus-like particle (VLP) attachment to and entry into a human duodenal cell line, HuTu-80. We show that while gastric juice and pancreatic juice do not affect viral attachment or entry, bile - in particular hydrophobic bile acids - significantly enhance cellular attachment and subsequent entry of GII.4 VLPs into cells. In addition, we show that hydrophobic bile acids induce accumulation of viral particles in the vicinity of cells. Our results suggest the presence of a new en masse infection mechanism, where bile acids aggregate virions, and allow direct and more efficient attachment to and entry into target cells. ImportanceViruses transmitted by the fecal-oral route encounter secreted host factors in gastrointestinal fluids. Some host factors can be exploited by the virus to facilitate infection. Human bile acids indirectly promote norovirus entry into and infection of human enteroids, but the direct effect of bile acids on attachment and uptake, along with the impact of other gastrointestinal fluids, remain unknown. Here, we investigated the direct effects of human body fluids on cellular attachment of norovirus VLPs. We show that human bile and hydrophobic bile acids induce an accumulation of norovirus VLPs, which is associated with significantly enhanced attachment and entry into human duodenal cell lines. These results highlight the differential effects of gastrointestinal body fluids on viral attachment and entry, while providing useful information into the complex HuNoV-host interactions that facilitate infection.