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Journal of Virology, July 2004, p. 7575-7581, Vol. 78, No. 14
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.14.7575-7581.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Novel Hepatitis B Virus Genotype A Subtyping Assay That Distinguishes Subtype Aa from Ae and Its Application in Epidemiological Studies

Izumi Hasegawa,1,2 Yasuhito Tanaka,1 Anna Kramvis,3 Takanobu Kato,1 Fuminaka Sugauchi,2 Subrat K. Acharya,4 Etsuro Orito,2 Ryuzo Ueda,2 Michael C. Kew,3 and Masashi Mizokami1*

Department of Clinical Molecular Informative Medicine,1 Department of Internal Medicine and Molecular Science, Nagoya City University Graduate School of Medical Sciences, Nagoya, Japan,2 MRC/CANSA/University Molecular Hepatology Research Unit, Department of Medicine, University of the Witwatersrand, Johannesburg, South Africa,3 Sub Bagian Hepatologi, Department of Gastroenterology, All India Institute of Medical Sciences, New Delhi, India4

Received 3 December 2003/ Accepted 22 March 2004

The eight genotypes of hepatitis B virus (HBV) have different geographical distributions, virological characteristics, and clinical manifestations. A unique subtype of HBV genotype A (HBV/A) was reported in sub-Saharan Africa, raising the possibility that patients infected with this subtype (HBV/Aa ["a" for African and Asian]) may have different clinical outcomes than other HBV/A isolates (HBV/Ae ["e" for European]). Comparison between 30 HBV/Aa and 30 HBV/Ae isolates indicated that almost all HBV/Ae isolates had G at nucleotide (nt) 1809 and C at nt 1812, whereas HBV/Aa isolates had T1809/T1812. Taking advantage of these two single nucleotide polymorphisms (SNPs), a novel subtype-specific PCR assay in the X/precore/core region was developed. This assay was combined with a restriction fragment length polymorphism assay using BglII in a different region (nt 1984 to 1989), which has a SNP distinguishing HBV/Aa from HBV/Ae, resulting in 100% specificity for the combined assay. Application of the subtyping assay using sera from 109 paid donors in the United States indicated significantly different distributions of HBV/A subtypes among races; African-Americans, Caucasians, and Hispanics had HBV/Ae, whereas Asians had mainly HBV/Aa, suggesting that the HBV/Aa isolates may have been imported by recent immigration from Asia. In conclusion, the specificity and sensitivity of the combined subtyping assay were confirmed, and its usefulness was demonstrated in a practical context.

* Corresponding author. Mailing address: Department of Clinical Molecular Informative Medicine, Nagoya City University Graduate School of Medical Sciences, Kawasumi 1, Mizuho, Nagoya 467-8601, Japan. Phone: 81-52-853-8292. Fax: 81-52-842-0021. E-mail: mizokami{at}med.nagoya-cu.ac.jp.

Journal of Virology, July 2004, p. 7575-7581, Vol. 78, No. 14
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.14.7575-7581.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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