An increased diversity of HCV isolates were characterized among 393 patients with liver disease in China representing six genotypes, 12 subtypes, and two novel genotype 6 variants

Abstract

Background: We have recently determined HCV isolates among volunteer blood donors and IDUs in southern China and revealed the genotype distribution patterns not only different between the two studied cohorts but also from what we have sampled in 2002. A changed pattern could have also occurred among patients with liver disease.

Materials and methods: Both E1 and NS5B sequences of HCV were characterized among 393 patients with liver disease followed by phylogenetic analysis.

Results: Six HCV genotypes, 12 subtypes (1b: 65.9%, 6a: 17.1%, 2a: 7.4%, 3a: 3.6%, 3b: in 3.3%, 6e: 0.76%, and 1a, 1c, 2b, 2f, 4d, and 5a each 0.25%), and two novel genotype 6 variants were classified, showing the greatest complexity of HCV hitherto found in China. Although the predominance of 1b followed by 6a is largely consistent with what we have sampled in 2002, the identification of single isolates of 1c, 2f, 4d, 5a, and two novel HCV-6 variants were first reported. Excluding 4d from a European visitor, all the others were from Chinese patients. Since the 6a proportion (17.1%, 67/393) was unexpectedly lower than what we have recently detected among blood donors (34.8%, 82/236) and IDUs (51.5%, 70/136), further statistical analyses were conducted. Comparison of the mean ages showed that among the 393 patients, those infected with 1b were significantly (6.7 years) older than those with 6a, while the 393 patients as a whole were significantly older than the 236 blood donors (8.4 years) and 136 IDUs (12.6 years) we have recently reported. Explanations are that younger individuals had higher proportions of 6a infections while patients with liver disease could have acquired their infections earlier than volunteer blood donors and IDUs.

Conclusion: Among 393 patients with liver disease, a great diversity in HCV was detected, which reflects a constantly changing pattern of HCV genotypes in China over time.

Figure 1

Two circular ML trees reconstructed for the 393 partial E1 (A) and NS5B (B) region sequences, corresponding to the nucleotide numbering of 869-1289 and 8276-8615, respectively, in the H77 genome. Subtype designations are given at the internal nodes and bootstrap values shown in percentages. A scale in the upper middle of each tree measures 0.1 nucleotide substitutions per site. Initially, a large number of reference sequences were included for genotyping the 393 isolates. However, to reduce the taxa number shown in the trees, all the reference sequences are removed after genotyping.

Figure 2

ML trees reconstructed for the 259 subtype 1b isolates using (A) E1 and (B) NS5B sequences. The 1a sequence M62321 is used as an outlier group. In each tree, two rectangles highlight the classification of A and B clusters. The scale bar at the bottom of each tree represents 0.02 nucleotide substitutions per site.

Figure 3

ML trees reconstructed for the 67 subtype 6a isolates using (A) E1 and (B) NS5B sequences. In each tree, three rectangles highlight the classification of I, II, and III clusters. The 6b sequence D84262 was initially used as an outlier group. However, it was removed from the figure after the 6a sequences were rooted.

Figure 4

ML trees reconstructed for the 67 isolates of other HCV genotypes/subtypes using (A) E1 and (B) NS5B region sequences. Subtype designations are given at the internal nodes and bootstrap supports were shown in percentages.