Comparison of mean somatic hypermutation (SHM) frequency in IGH VDJ sequences, estimated in the BCR immunoglobulin gene repertoires of 22 HCV-infected patients and 7 HD, across 4 B-cell subsets. Solid points represent outliers. As the 2 biological replicates for each B-cell subset were very similar, the mean of the 2 values was taken for the calculation. Only the IGHV genes of productive IGH VDJ rearrangements were considered for mutation analysis. The mean somatic mutation frequency estimated for each B-cell subset is for the healthy individuals: naive = 0.16%, CD5⁺ = 0.10%, CS memory = 7.76%, NCSM = 4.77%, and for the HCV⁺ patients: naive = 0.41%, CD5⁺ = 0.17%, CS memory = 7.43%, NCSM = 4.51%. For naive B cells, the P value (.003) indicates a significantly different mean SHM frequency between HCV patients and HD. The 2 outliers among naive and CD5⁺ B cells of HCV⁺ patients are for both B-cell subsets patients 11 and 12. In these cell samples, the higher average mutation load is due to a substantial number of rearrangements with mutation frequencies typical for memory B cells. Hence, the higher mutation load of these samples is likely due to contamination of the naive and CD5⁺ B cells by memory B cells, or in these patients, some NCSM B cells had reduced CD27 and increased CD23 expression, so that they were indistinguishable from the naive and CD5⁺ B cells in the sorting procedure. CD5⁺, CD5⁺ mature B cells.