The gut virome in non-alcoholic fatty liver disease: distinct changes in Phapecoctavirus composition in a human microbiota associated animal model

Abstract

Background and aims: There is increasing knowledge on the role of the gut microbiome in the development of non-alcoholic fatty liver disease (NAFLD), but the role of the gut viral community in disease development remains poorly understood. We aimed to discover gut virome related to NAFLD development via a human-microbiotaassociated (HMA) rodent model of NAFLD. Method: Human fecal donors were recruited and classified into obese NAFLD, non-obese NAFLD and non-obese healthy controls. Liver steatosis was assessed by vibration-controlled transient elastography (Fibroscan, Echosens, Paris). The HMA-NAFLD rodent modelwasestablishedbyhigh-fatdiet andtransplantationofhuman fecal microbiota (FMT) for 12 weeks. The profile of the gut virome of both human donors and rodents was assessed by shotgun metagenomic sequencing (Illumina NovaSeq 6000, US). Results: Human donors with NAFLD had a significantly higher controlled attenuation parameter measurement when compared to healthy controls (median 324 [IQR 305.5–355.75] vs. 182.5 [179.25199.25] dB/m, p <0.001). Based on Bray-Curtis distance and PERMANOVA testing, human NAFLD donors had a distinct gut DNA viromecomparedwithhealthycontrols(p=0.04). Analysis via linear discriminate analysis (LDA) effect size method identified the Phapecoctavirus genus as a potential biomarker (LDA score 3.80), indicating a lower median relative abundance of Phapecoctavirus in NAFLDdonors whencomparedtohealthy donors (3.54% [IQR 2.93%3.98%] vs. 4.15% [3.94%-4.94%], p=0.038). As a putative host for Phapecoctavirus, the abundance of Escherichia coli was negatively associated with Phapecoctavirus (rho= −0.51, p=0.012). In the animal model, mice receiving FMT from obese NAFLD (FMT-ON) displayed a more severe NAFLD phenotype than those receiving FMT from healthycontrol (FMT-HC) (liver triglyceride: median 65.35 [IQR 62.32–73.33] vs. 48.7 [38.61–54.44] mg/g liver tissue, p=0.015; blood triglyceride: 207.89 [187.87–216.94] vs.110.05 [102.16–120.76] mg/dL,p<0.001). Phapecoctavirus genus was similarly identifiedas a potential fecal biomarker when comparingFMT-ONvs.FMT-HC(LDA score 3.19). It wasthe only viral genusthat distinguished NAFLD from control in both human donors and mouse recipients. FMT-ON mice with more severe NAFLD had a significantly lower level of fecal PhapecoctavirusthanFMT-HCmice(median2.47%[IQR2.39%-2.75%] vs. 2.82% [2.6%-3.14%], p =0.038). Conclusion: We reported the association of reduced fecal Phapecoctavirus with NAFLD in both human donors and HMA-mice recipients. Further interventional studies in animal and human are warrantedtodetermineitspotentialcausalityandfunctioninNAFLD.