Understanding Human Astrovirus from Pathogenesis to Treatment

While human astroviruses (HAstV) were discovered nearly 45 years ago, these small positive-sense RNA viruses remain critically understudied. These studies provide fundamental new research on astrovirus pathogenesis and disruption of the gut epithelium by induction of epithelial-mesenchymal transition (EMT) following astrovirus infection. Here we characterize HAstV-induced EMT as an upregulation of SNAI1 and VIM with a down regulation of CDH1 and OCLN, loss of cell-cell junctions most notably at 18 hours post-infection (hpi), and loss of cellular polarity by 24 hpi. While active transforming growth factor- (TGF-) increases during HAstV infection, inhibition of TGF- signaling does not hinder EMT induction. However, HAstV-induced EMT does require active viral replication. These are among the first studies describing the induction of EMT by a non-oncogenic virus and provides an exciting opportunity to understand EMT induction independent of cancer. Our findings likely extend beyond astrovirus to other viruses and may shed light on novel ways pathogens can circumvent the barriers meant to protect against them. Crossing these barriers can lead to systemic and even fatal infections. Astroviruses can be especially problematic in immunocompromised individuals and infants where the virus has been associated with necrotizing enterocolitis, severe and persistent diarrhea, and even encephalitis and meningitis. Using our novel tools and models, we demonstrate that the FDA-approved broad-spectrum anti-infective drug nitazoxanide (NTZ) blocks astrovirus replication in vitro with a 50% effective concentration (EC50) of approximately 1.47μM. It can be administered up to 8 hours post-infection and is effective against multiple human astrovirus serotypes including clinical isolates. Most importantly, NTZ reduces viral shed in vivo, exhibiting its potential as a future clinical therapeutic. Overall, these studies will further our understanding of astrovirus pathogenesis leading to the development of therapeutic options for vulnerable populations

Human Astroviruses: A Tale of Two Strains.

Since the 1970s, eight closely related serotypes of classical human astroviruses (HAstV) have been associated with gastrointestinal illness worldwide. In the late 2000s, three genetically unique human astrovirus clades, VA1-VA3, VA2-VA4, and MLB, were described. While the exact disease associated with these clades remains to be defined, VA1 has been associated with central nervous system infections. The discovery that VA1 could be grown in cell culture, supports exciting new studies aimed at understanding viral pathogenesis. Given the association of VA1 with often lethal CNS infections, we tested its susceptibility to the antimicrobial drug, nitazoxanide (NTZ), which we showed could inhibit classical HAstV infections. Our studies demonstrate that NTZ inhibited VA1 replication in Caco2 cells even when added at 12 h post-infection, which is later than in HAstV-1 infection. These data led us to further probe VA1 replication kinetics and cellular responses to infection in Caco-2 cells in comparison to the well-studied HAstV-1 strain. Overall, our studies highlight that VA1 replicates more slowly than HAstV-1 and elicits significantly different cellular responses, including the inability to disrupt cellular junctions and barrier permeability

Human Astroviruses: A Tale of Two Strains

Since the 1970s, eight closely related serotypes of classical human astroviruses (HAstV) have been associated with gastrointestinal illness worldwide. In the late 2000s, three genetically unique human astrovirus clades, VA1-VA3, VA2-VA4, and MLB, were described. While the exact disease associated with these clades remains to be defined, VA1 has been associated with central nervous system infections. The discovery that VA1 could be grown in cell culture, supports exciting new studies aimed at understanding viral pathogenesis. Given the association of VA1 with often lethal CNS infections, we tested its susceptibility to the antimicrobial drug, nitazoxanide (NTZ), which we showed could inhibit classical HAstV infections. Our studies demonstrate that NTZ inhibited VA1 replication in Caco2 cells even when added at 12 h post-infection, which is later than in HAstV-1 infection. These data led us to further probe VA1 replication kinetics and cellular responses to infection in Caco-2 cells in comparison to the well-studied HAstV-1 strain. Overall, our studies highlight that VA1 replicates more slowly than HAstV-1 and elicits significantly different cellular responses, including the inability to disrupt cellular junctions and barrier permeability

Astrovirus Pathogenesis

Astroviruses are a major cause of diarrhea in the young, elderly, and the immunocompromised. Since the discovery of human astrovirus type 1 (HAstV-1) in 1975, the family Astroviridae has expanded to include two more human clades and numerous mammalian and avian-specific genotypes. Despite this, there is still little known about pathogenesis. The following review highlights the current knowledge of astrovirus pathogenesis, and outlines the critical steps needed to further astrovirus research, including the development of animal models of cell culture systems

Astrovirus Pathogenesis

Astroviruses are a major cause of diarrhea in the young, elderly, and the immunocompromised. Since the discovery of human astrovirus type 1 (HAstV-1) in 1975, the family Astroviridae has expanded to include two more human clades and numerous mammalian and avian-specific genotypes. Despite this, there is still little known about pathogenesis. The following review highlights the current knowledge of astrovirus pathogenesis, and outlines the critical steps needed to further astrovirus research, including the development of animal models of cell culture systems

Astrovirus replication is dependent on induction of double-membrane vesicles through a PI3K-dependent, LC3-independent pathway.

Human astrovirus is a positive-sense, single-stranded RNA virus. Astrovirus infection causes gastrointestinal symptoms and can lead to encephalitis in immunocompromised patients. Positive-strand RNA viruses typically utilize host intracellular membranes to form replication organelles, which are potential antiviral targets. Many of these replication organelles are double-membrane vesicles (DMVs). Here, we show that astrovirus infection leads to an increase in DMV formation through a replication-dependent mechanism that requires some early components of the autophagy machinery. Results indicate that the upstream class III phosphatidylinositol 3-kinase (PI3K) complex, but not LC3 conjugation machinery, is utilized in DMV formation. Both chemical and genetic inhibition of the PI3K complex lead to significant reduction in DMVs, as well as viral replication. Elucidating the role of autophagy machinery in DMV formation during astrovirus infection reveals a potential target for therapeutic intervention for immunocompromised patients. IMPORTANCE These studies provide critical new evidence that astrovirus replication requires formation of double-membrane vesicles, which utilize class III phosphatidylinositol 3-kinase (PI3K), but not LC3 conjugation autophagy machinery, for biogenesis. These results are consistent with replication mechanisms for other positive-sense RNA viruses suggesting that targeting PI3K could be a promising therapeutic option for not only astrovirus, but other positive-sense RNA virus infections

Indoleamine 2,3-dioxygenase 1 regulates cell permissivity to astrovirus infection

Astroviruses cause a spectrum of diseases spanning asymptomatic infections to severe diarrhea, but little is understood about their pathogenesis. We previously determined that small intestinal goblet cells were the main cell type infected by murine astrovirus-1. Here, we focused on the host immune response to infection and inadvertently discovered a role for indoleamine 2,3-dioxygenase 1 (Ido1), a host tryptophan catabolizing enzyme, in the cellular tropism of murine and human astroviruses. We identified that Ido1 expression was highly enriched among infected goblet cells, and spatially corresponded to the zonation of infection. Because Ido1 can act as a negative regulator of inflammation, we hypothesized it could dampen host antiviral responses. Despite robust interferon signaling in goblet cells, as well as tuft cell and enterocyte bystanders, we observed delayed cytokine induction and suppressed levels of fecal lipocalin-2. Although we found Ido-/- animals were more resistant to infection, this was not associated with fewer goblet cells nor could it be rescued by knocking out interferon responses, suggesting that IDO1 instead regulates cell permissivity. We characterized IDO1-/- Caco-2 cells and observed significantly reduced human astrovirus-1 infection. Together this study highlights a role for Ido1 in astrovirus infection and epithelial cell maturation