Differential prevalence and geographic distribution of hepatitis C virus genotypes in acute and chronic hepatitis C patients in Vietnam

Abstract

Background: The highest burden of disease from hepatitis C virus (HCV) is found in Southeast Asia, but our understanding of the epidemiology of infection in many heavily burdened countries is still limited. In particular, there is relatively little data on acute HCV infection, the outcome of which can be influenced by both viral and host genetics which differ within the region. We studied HCV genotype and IL28B gene polymorphism in a cohort of acute HCV-infected patients in Southern Vietnam alongside two other cohorts of chronic HCV-infected patients to better understand the epidemiology of HCV infection locally and inform the development of programs for therapy with the increasing availability of directly acting antiviral therapy (DAAs).

Methods: We analysed plasma samples from patients with acute and chronic HCV infection, including chronic HCV mono-infection and chronic Human Immunodeficiency Virus (HIV)-HCV coinfection, who enrolled in four epidemiological or clinical research studies. HCV infection was confirmed with RNA testing. The 5' UTR, core and NSB5 regions of HCV RNA positive samples were sequenced, and the genotype and subtype of the viral strains were determined. Host DNA from all HCV positive patients and age- and sex-matched non-HCV-infected control individuals were analysed for IL28B single nucleotide polymorphism (SNP) (rs12979860 and rs8099917). Geolocation of the patients were mapped using QGIS.

Results: 355 HCV antibody positive patients were analysed; 54.6% (194/355) and 46.4% (161/355) were acute and chronic infections, respectively. 50.4% (81/161) and 49.6.4% (80/161) of chronic infections had HCV mono-infection and HIV-HCV coinfection, respectively. 88.7% (315/355) and 10.1% (36/355) of the patients were from southern and central regions of Vietnam, respectively. 92.4% (328/355) of patients were HCV RNA positive, including 86.1% (167/194) acute and 100% (161/161) chronic infections. Genotype could be determined in 98.4% (322/328) patients. Genotypes 1 (56.5%; 182/322) and 6 (33.9%; 109/322) predominated. Genotype 1 including genotype 1a was significantly higher in HIV-HCV coinfected patients compared to acute HCV patients [43.8% (35/80) versus 20.5% (33/167)], (p = <0.001), while genotype 6 was significantly higher in chronic HCV mono-infected patients [(44.4% (36/81) versus 20.0% (16/80)] (p = < 0.004) compared to HIV-HCV coinfected patients. The prevalence of IL28B SNP (rs12979860) homozygous CC was 86.46% (83/96) in control individuals and was significantly higher in acutely-infected compared to chronically-infected patients [93.2 (82/88) versus 76.1% (35/46)] (p = < 0.005).

Conclusion: HCV genotype 6 is highly prevalent in Vietnam and the high prevalence in treatment naïve chronic HCV patients may results from poor spontaneous clearance of acute HCV infection with genotype 6.

Fig 1. A midpoint rooted tree showing the relationship between the Vietnamese HCV (n = 322) sequences with 43 reference sequences downloaded from the GenBank database (all genotypes and selected subtypes).

Tree was constructed using Neighbor-Joining method available in Genious software using GTR+G+I nucleotide substitution model with 1000 bootstrapping replicates. Bootstrap values greater than 70% is shown on the branch nodes. Acute, chronic mono-infection and HIV-HCV coinfection is presented by suffix A (in black text), suffix C (in blue text) and suffix H (in purple text) followed by the patient ID and sequence region respectively. The reference genomes are presented as subtype followed by Gene Bank accession number and country of origin. The reference genomes are highlighted with a filled dark circle. The scale bar indicates the number of nucleotide substitution. Red, purple, green, and brown color is used for genotype 1, 2, 3, and 6. (a) Analysis was based on a 377 bp HCV NS5B gene (position 8259 to 8636 relative to GenBank sequence AY051292). Sequences with discordant genotype between NS5B and 5 UTR, and NS5B and core is marked with * and ** respectively. (b) Analysis was based on a 464 bp core sequence (position 288 to 752) relative to GenBank sequence AY051292). Sequences which have discordant genotype between core and 5’UTR with NS5B is marked with ***.

Fig 2. Geographical distribution of HCV genotypes in different sub regions of Vietnam.

Sub regions are presented by different colors. Number of isolates and proportion of different genotypes in each sub region are presented in blue (genotype 1), green (genotype 2), red (genotype 3) and purple (genotype 6) colors.

Fig 3. The prevalence of IL28B SNP (rs12979860 and rs8099917) in 171 patients.

88 acute, 46 chronic HCV mono-infection and 37 HIV-HCV coinfection and 96 age (±5 years) and sex matched non hepatitis control patients. The percent prevalence of SNPs is presented in Y axis and the patient groups and controls in X axis. P value was calculated using chi square and fisher’s exact test as appropriate. (A) The prevalence CC, CT and TT for rs8099917. The proportion of CC, CT and TT alleles are presented in green, orange and blue color bars respectively. P value was calculated between control and all groups (acute HCV, chronic HCV mono-infection and HIV-HCV coinfection) individually and acute HCV and chronic HCV mono-infection group. (B) The prevalence TT, T/G and G/G for rs12979860. The proportion of TT, GT and GG alleles are presented in green, orange and blue color bars respectively. P value was calculated between control and all groups (acute HCV, chronic HCV mono-infection and HIV-HCV coinfection) individually and acute HCV and chronic HCV mono-infection group.