To the Editor: Previous attempts to document vertical transmission of hepatitis C virus (HCV) were accomplished by detecting antibodies to HCV and HCV RNA in the infants/children born to mothers with HCV infection (1-5). For the high-risk infants, documentation of HCV transmission using these approaches is difficult due to the exposure to multiple potential sources of HCV, which include blood-product transfusion and surgery, in addition to vertical transmission. HCV is also characterized by its heterogeneity and circulates as a spectrum of quasispecies. Accordingly, the HCV genomic sequences in the donor and the recipient may be different. We recently proposed that the transmission of HCV can be confirmed by molecular evolutionary analysis, which is based on the estimation of genomic divergence (6,7). Here, we report the use of this approach to document the vertical transmission of HCV.
A 8-lb boy was born by vaginal delivery in January 1992. At 8 weeks, he developed jaundice. Serum liver biochemistry revealed marked cholestasis (total bilirubin 132 mol/L, AST 249 IU/L, ALT 143 IU/L, alkaline phosphatase 107 IU/L). The child had a normal α1-antitrypsin level and was seronegative for both human immunodeficiency and hepatitis B virus. Serum was reactive to two HCV antigens by RIBA-2. Ultrasound revealed absent gallbladder, and HIDA scan did not image the gallbladder or biliary tree. Liver biopsy showed features compatible with biliary atresia. The child underwent liver transplantation in August 1992. The child's mother was 35-year-old white woman who was also found to have an elevated serum liver biochemistry (ALT 68 and AST 65 IU/L) and seropositive for antibody to HCV by EIA-2. Serum from the mother and the child (before liver transplantation) as well as breast milk from the mother were available for study.
An attempt was made to detect HCV RNA in the breast milk (including using 1 ml of breast milk for RNA extraction, adding HCV RNA to the RNA extract as a positive control to account for the possibility of having inhibitory substances for the amplification, and repeating the experiments four times), and it was consistently negative for HCV RNA. Both mother and child were positive for HCV RNA and both were determined to have HCV genotype 2b based on genotype-specific primers for RT-PCR based on the HCV core region (8). Since genotype 2b is relatively uncommon in the United States, there is a good chance that the child acquired HCV from his mother (9). To confirm this claim, the HCV core region from both mother and child was amplified, cloned, and seven clones from the mother's isolate and four clones from the baby's isolate were sequenced. The sequences were aligned, the number of nucleotide substitutions estimated by the six-parameter method, and a phylogenetic tree was constructed by the neighbor-joining method (Fig. 1) using the computer software ODEN (version 1.1). The clones from the mother (89) were divided into two groups, 89-1,2,3,7 and 89-4,5,6 within the subtype 2b, and the latter group was very close in evolutionary distance from the clones isolated from the child (90-1,2,3,4,5). In addition, the clones from the child were clustered within the nest containing one group of the isolated clones from the mother (89-4,5,6) (8), and the nucleotide diversity of the mother's clones was also more variable than that of the baby's, supporting the direction of transmission being from mother to child (10).
This report documents the transmission of HCV from mother to child using molecular evolutionary analysis technique. This method is especially useful for the documentation of HCV transmission in high-risk groups (10).
Robert G. Gish; *Kenneth L. Cox
Departments of Transplantation and * Pediatrics
California Pacific Medical Center
San Francisco, California, U.S.A.
Masashi Mizokami; Tomoyoshi Ohno
Second Department of Medicine
Nagoya City University Medical School
Johnson Y. N. Lau
Section of Hepatobiliary Diseases
Department of Medicine
University of Florida
Gainesville, Florida, U.S.A.
1. Lam JPH, McOmish F, Burns SM, Yap PL, Mok JYQ, Simmonds P. Infrequent vertical transmission of hepatitis C virus. J Infect Dis
2. Reinus JF, Leikin EL, Alter HJ, et al. Failure to detect vertical transmission of hepatitis C virus. Ann Intern Med
3. Wejstål R, Widell A, Månsson A-S, Hermodsson S, Norkrans G. Mother-to-infant transmission of hepatitis C virus. Ann Intern Med
4. Inoue Y, Takeuchi K, Chou WH, et al. Silent mother-to-child transmission of hepatitis C virus through two generations determined by comparative nucleotide sequence analysis of viral cDNA. J Infect Dis
5. Ohto H, Terazawa S, Sasaki N, et al. Transmission of hepatitis C virus from mothers to infants. N Engl J Med
6. Susuki K, Mizokami M, Lau JYN, et al. Confirmation of hepatitis C virus transmission through needlestick accident by molecular evolutionary analysis. J Infect Dis
7. Mizokami M, Gobojori T, Lau JYN. Molecular evolutionary medicine—its application to the study of hepatitis C virus. Gastroenterology
8. Okamoto H, Sugiyama Y, Okada S, et al. Typing hepatitis C virus by polymerase chain reaction with type-specific primers: application to clinical surveys and tracing infection sources. J Gen Virol
9. Lau JYN, Mizokami M, Kolberg JA, et al. Application of six hepatitis C virus genotyping systems to sera from chronic hepatitis C patients in the United States. J Infect Dis
10. Weiner AJ, Thaler MM, Crawford K, et al. A unique, predominant hepatitis C virus variant found in an infant born to a mother with multiple variants. J Virol