Frericks, N. | Search bioRxiv

bioRxiv2026

Yellow fever vaccine propagation in primary human hepatocytes triggers antiviral and cytolytic responses

Yellow fever virus (YFV) infection can cause severe-to-fatal liver damage in humans, while immunization with the attenuated 17D vaccine strain has an excellent safety record, priming protective host immunity in the absence of pathology. To investigate virus-host correlates associated with these differential clinical outcomes, we combined YFV genome-level evolutionary analyses with investigations of vaccine and virulent strain hepatotropism. Evolutionary analyses confirmed purifying selection is the dominant force driving global YFV genome divergence, with functional constraints associated with the YFV transmission cycle selecting against vaccine attenuating mutations in virulent strains. In immune deficient hepatoma cells, 17D exhibited enhanced early propagation, spreading and apoptosis induction when compared to virulent strains. Ex vivo infections performed in primary human hepatocytes (PHH) from multiple donors confirmed robust propagation of both 17D and virulent YFV strains. RNA-sequencing revealed consistent and shared induction of IFNB and IFNL1-4, modulating overlapping gene programs associated with antiviral responses, immunity, chemotaxis and inflammation, cell-death, metabolic reprogramming and protein translation. Subtle differences in virion production kinetics and the magnitude and tempo of PHH transcriptional responses were observed. Antiviral responses to 17D were activated earlier while responses to virulent YFV were delayed but enhanced, mirroring virus propagation kinetics. More broadly, cellular responses to YFV infection are likely dominated by paracrine IFN signalling, with enriched LRP1 expression coupled with impaired IFN production in PHH contributing to YFVs robust hepatotropism. In summary, we demonstrate comparable propagation kinetics and PHH transcriptional responses to vaccine and virulent YFV strains, highlighting impaired hepatotropism is not a correlate of vaccine attenuation. These data imply that unknown barriers restrict liver access and associated organ pathology upon vaccination with 17D.

microbiology↗

bioRxiv2023

Hepatitis C virus cell culture adaptive mutations enhance cell culture propagation by multiple mechanisms but boost antiviral responses in primary human hepatocytes

Hepatitis C virus (HCV) infection progresses to chronicity in the majority of infected individuals. Its high intra-host genetic variability enables HCV to evade the continuous selection pressure exerted by the host, contributing to persistent infection. Utilizing a cell culture adapted HCV population (p100pop) which exhibits increased replicative capacity in various liver cell lines, this study investigated virus and host determinants which underlie enhanced viral fitness. Characterization of a panel of molecular p100 clones revealed that cell culture adaptive mutations optimize a range of virus-host interactions, resulting in expanded cell tropism, altered dependence on the cellular co-factor micro-RNA 122 and increased rates of virus spread. On the host side, comparative transcriptional profiling of hepatoma cells infected either with p100pop or its progenitor virus revealed that enhanced replicative fitness correlated with activation of endoplasmic reticulum stress signaling and the unfolded protein response. In contrast, infection of primary human hepatocytes with p100pop led to a mild attenuation of virion production which correlated with a greater induction of cell-intrinsic antiviral defense responses. In summary, long-term passage experiments in cells where selective pressure from innate immunity is lacking improves multiple virus-host interactions, enhancing HCV replicative fitness. However, this study further indicates that HCV has evolved to replicate at low levels in primary human hepatocytes to minimize innate immune activation, highlighting that an optimal balance between replicative fitness and innate immune induction is key to establishing persistence. Author SummaryHCV infection remains a global health burden with 58 million people currently chronically infected. However, the detailed molecular mechanisms which underly persistence are incompletely defined. We utilized a long-term cell culture adapted HCV, exhibiting enhanced replicative fitness in different human liver cell lines, in order to identify molecular principles by which HCV optimizes its replication fitness. Our experimental data revealed that cell culture adaptive mutations confer changes in the host response and usage of various host factors. The latter allows functional flexibility at different stages of the viral replication cycle. However, increased replicative fitness resulted in an increased activation of the innate immune system, which likely poses boundary for functional variation in authentic hepatocytes, explaining the observed attenuation of the adapted virus population in primary hepatocytes.

molecular biology↗