HCV subtype characterization among injection drug users: implication for a crucial role of Zhenjiang in HCV transmission in China

Abstract

Background: HCV transmission is closely associated with drug-trafficking routes in China. However, the transmission route of HCV in Eastern China remains unclear. Here, we investigate the role of Zhenjiang city of Jiangsu province, an important transportation hub linking Shanghai with other regions of China, in HCV transmission.

Methodology/principal findings: A total of 141 whole blood samples were collected from injection drug users (IDUs) in Zhenjiang and then tested for HCV infection. Of them, 115 HCV positive plasmas were subjected to RNA extraction, RT-PCR amplification, and sequencing. The subtype characterization and the evolutionary origin of HCV strains circulating in Zhenjiang were determined using polygenetic or phylogeographic analyses. Seven HCV subtypes 1b, 2a, 3a, 3b, 6a, 6e and 6n were detected among Zhenjiang IDUs, showing a complex HCV epidemic. The most predominant subtypes were 3a (38%) and 1b (26.8%). Among these subtypes, subtypes 3b, 6n and 6e originated from Southwestern China (i.e., Yunnan and/or Guangxi), subtypes 2a and 6a from Southern China (i.e., Guangdong), subtype 1b from Central (i.e., Henan) and Northwestern (i.e., Xinjiang) China, and subtype 3a from Southwestern (i.e., Yunnan) and Northwestern (i.e., Xinjiang) China. From Zhenjiang, subtypes 1b and 2a were further spread to Eastern (i.e., Shanghai) and Northern (i.e., Beijing) China, respectively.

Conclusions/significance: The mixing of seven HCV subtypes in Zhenjiang from all quarters of China indicates that as an important middle station, Zhenjiang plays a crucial role in HCV transmission, just as it is important in population migration between other regions of China and Eastern China.

Figure 1. Phylogenetic tree of NS5B fragments of HCV strains isolated from Zhenjiang.

The sequences correspond to nucleotides 8262–9312 in HCV H77 genome (NC-004102). Only bootstrap values >75% are shown. The sequences from Zhenjiang IDUs are labeled by red solid triangles.

Figure 2. Comparison of HCV subtype distributions among IDUs between Zhenjiang and other regions of China.

Figure 3. The NS5B phylogenetic relationship of HCV strains isolated from Zhenjiang IDUs with those from other regions of China.

Please see Figure 6 for the location of other regions in China and Figure 1 for other details. (A) The whole phylogenetic tree. (B) The sub-tree of HCV subtype 1b strains. (C) The sub-tree of HCV subtype 3a strains.

Figure 4. Maximum clade credibility tree of the HCV 1b sequences.

The tree was reconstructed based on the genomic region (H77 1341–1835 nt) located in C/E2. Ancestral geographic states were reconstructed using Bayesian phylogeographic inference framework implemented in the BEAST v1.5.4 package. The tree branches are colored according to their respective geographical locations. The time scale is shown at the bottom.

Figure 5. Maximum clade credibility tree of the HCV 3a sequences.

The tree was reconstructed based on the genomic region (H77 1356–1826 nt) located in C/E2. Please see Figure 4 for other details.

Figure 6. Potential migration routes of several HCV subtypes in China.

The arrows represent the potential migration routes of HCV. XJ: Xinjiang; LN: Liaoning; SD: Shandong; HN: Henan; JS: Jiangsu; SH: Shanghai; YN: Yunnan; GX: Guangxi; GD: Guangdong; HK: Hong Kong. Because few corresponding sequences from Taiwan are available for phylogenetic analyses, this Figure does not include Taiwan.