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

Abstract

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.

Genetic Characterization of Hepatitis B Virus in Peripheral Blood Leukocytes: Evidence for Selection and Compartmentalization of Viral Variants with the Immune Escape G145R Mutation

“…However, until now, studies of serum HBV isolates from different parts of India could not detect this subgenotype in the sera of the Indian population (1,4,16,20,34). Nevertheless, for the first time we reveal that subgenotype Ae/A2 sequences remain present in the leukocytes in our population.…”

A case-control study for differences among hepatitis B virus infections of genotypes A (subtypes Aa and Ae) and D †

“…A novel method for confirmation of genotype A and specific determination of HBV/Ae consisted of two PCR cycles with heminested primers followed by restriction fragment length polymorphism with the restriction site specific for HBV/Ae. 32 The first-round PCR was performed with a sense primer in a 96-well cycler. The second-round PCR was performed twice separately with different sets of primers.…”

Influence of genotypes and precore mutations on fulminant or chronic outcome of acute hepatitis B virus infection

“…21 Subgenotypes of HBV/A designated Ae prevalent in Europe and Aa frequent in Africa as well as Asia,9 which corresponds to subgroup AЈ originally reported by Bowyer et al, 22 were determined by PCR restriction fragment length polymorphism (RFLP) involving nucleotide conversions in an immediate upstream of the precore region that are specific for each of them. 16,23 HBV/Bj (Japanese type) lacking the recombination with C over the precore region and the core gene and Ba (Asian type) with the recombination were determined by its absence or presence on HBV DNA sequences, as well as RFLP based on specific nucleotide substitutions, after the methods described previously. 15,24 Subgenotypes of HBV/C, Cs (Southeast Asian type) found only in Southeast Asia, including Vietnam, Myanmar, Thailand, Laos, Bangladesh, Hong Kong, and Southern China, and Ce (East Asian type), found in Far East Asia, including Japan, Korea, and Northern China, were determined by the PCR-RFLP method described previously.…”