Volume 123, Issue 5, Pages 1485-1493 (November 2002) Evolution of hepatitis C viral quasispecies after liver transplantation  Andre C. Lyra, *, Xiaofeng Fan, *, Dorothy M. Lang, ‡, Karina Yusim, ‡, Sanjay Ramrakhiani, *, Elizabeth M. Brunt, §, Bette Korber, ‡, Alan S. Perelson, ‡, Adrian M. Di Bisceglie, *  Gastroenterology  Volume 123, Issue 5, Pages 1485-1493 (November 2002) DOI: 10.1053/gast.2002.36546 Copyright © 2002 American Gastroenterological Association Terms and Conditions

Fig. 1 RNA levels before and after OLT in 4 patients with mild and 7 patients with severe recurrent hepatitis C. RNA was measured between 12 and 80 months after OLT (median, 14 months) in (A) the mild group, and between 3.4 and 80 months in (B) the severe group (median, 27 months). Gastroenterology 2002 123, 1485-1493DOI: (10.1053/gast.2002.36546) Copyright © 2002 American Gastroenterological Association Terms and Conditions

Fig. 2 Intrasample HVR1 diversity of all samples included in the study. Patients 1–4 had mild recurrent hepatitis C, whereas patients 5–11 had severe recurrent hepatitis C. The x-axis represents serum and/or histologic follow-up in months of each individual patient; thus the scale is different in each graph. Negative numbers represent time before transplant. The y-axis represents the intrasample diversity at each time point. The stage of fibrosis is also noted at each time point. The closed circle and open triangle symbols correspond to intrasample genetic distance and nonsynonymous substitutions per nonsynonymous site, respectively. Gastroenterology 2002 123, 1485-1493DOI: (10.1053/gast.2002.36546) Copyright © 2002 American Gastroenterological Association Terms and Conditions

Fig. 3 Intrasample HVR1 diversity. (A and B) Mild group. Comparison between mean group values from before (analysis of 4 samples, 40 clones) and from all time points after OLT (8 samples, 80 clones) showed a significant increase (P = 0.014 for genetic distance, P = 0.038 for nonsynonymous substitutions, and P = 0.003 for synonymous substitutions in the HVR1). In the flanking regions, these same values were P = 0.029 for genetic distance, P = 0.556 for nonsynonymous substitutions, and P = 0.021 for synonymous substitutions. (C and D) Severe group. Comparison between before (7 samples, 70 clones) and after (11 samples, 110 clones) OLT showed P = 0.718 for genetic distance, P = 0.806 for nonsynonymous substitutions, and P = 0.628 for synonymous substitutions in the HVR1 and P = 0.054 for genetic distance, P = 0.101 for nonsynonymous substitutions, and P = 0.037 for synonymous substitutions in the flanking regions. Gastroenterology 2002 123, 1485-1493DOI: (10.1053/gast.2002.36546) Copyright © 2002 American Gastroenterological Association Terms and Conditions

Fig. 4 Correlation of amino acid complexity and stage of fibrosis after OLT. All pairs of serum samples and liver histology (n = 19) obtained at the same time from all 11 patients were analyzed (r = −0.543; P = 0.016). Samples after retransplantation were not included in the analysis. Gastroenterology 2002 123, 1485-1493DOI: (10.1053/gast.2002.36546) Copyright © 2002 American Gastroenterological Association Terms and Conditions

Fig. 5 Overall intersample diversity/year between time point 0 and time points after OLT from the mild (8 time points) and severe (11 time points) groups. (A) HVR1. Genetic distance was 131 × 10−3 and 105 × 10−3 (P = 0.016), nonsynonymous substitutions per nonsynonymous site were 124 × 10−3 and 132 × 10−3 (P = 0.503), and synonymous substitutions were 146 × 10−3 and 37 × 10−3 (P < 0.001) in the mild and severe groups, respectively, for each comparison. (B) Flanking regions. Genetic distance was 31 × 10−3 and 21 × 10−3 (P < 0.001), nonsynonymous substitutions per nonsynonymous site were 12 × 10− and 9 × 10−3 (P = 0.002), and synonymous substitutions were 82 × 10−3 and 64 × 10−3 (P = 0.002) in the mild and severe groups, respectively, for each comparison. Gastroenterology 2002 123, 1485-1493DOI: (10.1053/gast.2002.36546) Copyright © 2002 American Gastroenterological Association Terms and Conditions

Fig. 6 Neighbor-joining trees of 462-bp nucletide sequences including HVR1. Trees were constructed with the distance matrix calculated using the Kimura 2-parameter method in the MEGA program. (A) Tree of patient 4 from the mild group. The circle, triangle, and square symbols correspond, respectively, to samples from 15 months before OLT (control), time point 0, and 12 months after OLT. (B) Tree of patient 6 from the severe group. The circle, triangle, and square symbols correspond, respectively, to samples from 22 months before OLT (control), at time point 0, and from 12 months after OLT. Gastroenterology 2002 123, 1485-1493DOI: (10.1053/gast.2002.36546) Copyright © 2002 American Gastroenterological Association Terms and Conditions

Fig. 7 Rate of amino acid change over time from all patients in HVR1 and flanking regions. The x-axis represents time of follow-up in months; negative numbers represent time before OLT. The y-axis represents the average amino acid change. The rate of amino acid change is most variable in the first 3 years after OLT and appears to become constant after 36 months. Gastroenterology 2002 123, 1485-1493DOI: (10.1053/gast.2002.36546) Copyright © 2002 American Gastroenterological Association Terms and Conditions