Identification of genetic variants of hepatitis A virus.
Although detection of hepatitis A virus (HAV) has been greatly aided by the development of polymerase chain reaction (PCR) technology, identification of genetic variants requires sequencing PCR products, which necessarily limits the length of the HAV genome (typically 2%) that can be analyzed. From a regulatory standpoint, identification of the specific strain detected by PCR is a prerequisite not only to overrule contamination of test samples in the diagnostic laboratory, but also to possibly locate the origin of the virus detected by PCR. We explored alternatives to sequencing PCR products to achieve these goals. The findings indicate that restriction fragment length polymorphism (RFLP) analysis of PCR products from two noncontiguous regions of the HAV genome encompassing 765 nucleotides (approximately 10% of the genome) by the restriction endonucleases HinfI and AluI, which cut frequently within the HAV genome, can distinguish the common tissue culture adapted strains of HAV from stool isolates. The resolution can be greatly enhanced by combining single strand conformation polymorphism (SSCP) analysis with restriction enzyme digestion, when most of the seventeen strains analyzed could be identified.
Goswami
B B
BB
Food and Drug Administration, Center for Food Safety and Applied Nutrition, Washington, D.C. 20204, USA.
Burkhardt
W
W
Cebula
T A
TA
D004279:DNA, Viral / Q000737:chemistry / Q000302:isolation & purification; D014644:Genetic Variation; D006507:Hepatovirus / Q000737:chemistry / Q000235:genetics*; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length
D004279:DNA, Viral