A visualizable hepatitis A virus and hepatitis C virus coinfection model in vitro: coexistence of two hepatic viruses under limited competition in viral RNA synthesis

Hepatitis A virus (HAV) and hepatitis C virus (HCV) coinfection in patients usually leads to HCV suppression or clearance. Whether this suppression/clearance is caused by direct virus-virus interaction or indirect interactions is still unknown. Here, we present a robust and visualizable HAV/HCV coinfection model for investigating their interactions in vitro. We find that HAV super infects HCV-persistently-infected Huh-7.5.1 cells without obvious virus-virus exclusion and vice versa, while there is a mild reciprocal viral interference in coinfection. Through single-cell scale confocal microscopy analysis and treating co-infected cells with rNTPs or antivirals e.g. Sofosbuvir, Simeprevir, and Ledipasvir, we find that HAV and HCV exploit cellular machinery in a compatible manner, but with reciprocal competition for rNTPs in their RNA synthesis. In conclusion, our findings reveal the absence of direct HAV-HCV interaction but the presence of indirect interaction, which may be due to limited competition in viral RNA synthesis. Therefore, we propose that suppression/clearance of HCV in HAV/HCV coinfected patients is probably due to indirect interactions e.g. viral competition or immunological interactions.\n\nAuthor summaryVirus-virus interactions could usually be categorized in three aspects: (1) direct interaction induced by interaction of viral gene or gene products; (2) indirect interaction mediated by immune response or (3) host environmental alteration. Hepatitis A virus (HAV) and hepatitis C virus (HCV) are two important causatives of human hepatitis. In patients, their coinfection often induces HCV suppression/clearance, and sometimes may lead to fulminant hepatitis. In this study, we find that a limited reciprocal viral interference occurs in HAV/HCV coinfection in vitro, which results from a viral competition of rNTPs in viral RNA synthesis. Our study provides a new paradigm of studying virus-virus interactions in single-cell scale. To our knowledge, we identify the rNTPs competition in virus coinfection for the first time by experiment, though several mathematical modelings have predicted the resource competition in virus coinfection.