Polymerase Chain Reaction (PCR) Assays for Hepatitis B Virus

Reliable Estimator of Infectivity
HBV is now considered a more serious health threat than HIV/AIDS in developing countries, as its "infectivity" is as high as 33 per cent, against the HIV "infectivity" of 0.3 per cent. Mortality in case of HBV-related diseases is much higher. India is sitting on a Hepatitis time bomb that is silently ticking away with an estimated 40 million Indians carrying the Hepatitis-B virus which is 200 times more infectious than AIDS and another 20 million hosting the even more deadly Hepatitis C virus. The incidence among pregnant mothers is 4.22 percent. Antenatal transmission is 73.8 per cent against 33 per cent in case of HIV (Indian Express, 1999).

Hepatitis B is a silent killer and carriers of the virus are at a risk of developing fatal cirrhosis and liver cancer.

Serological Diagnosis
The routine laboratory testing for the hepatitis B virus includes assays for multiple antibodies (anti-HBsAg, anti-HBc IgM, anti-HBc IgG, anti-HBe) and viral proteins (HBsAg, HBeAg).

Disadvantages Of Serological Assays
The above mentioned array of markers determine whether the infection is present and, when combined with routine "liver function test", help to evaluate the stage of disease (i.e acute, chronic active, or resolved infection)

The most significant weakness of the standard serological test from the clinical perspective, however, is that they provide no reliable measure of viral replication or viremia (two indices that should be related) in a patient known to have been infected with HBV. In certain settings, however, these measures fall short and molecular assays for the presence of HBV DNA have been used to supplement the testing algorithm.

The presence of hepatitis B e antigen (HBeAg) is often taken as a marker of active viral replication, viremia, and infectivity (Robinson, 1995). Conversely, the presence of anti-HbeAg is a marker of viral quiescence and non-infectivity. It is now recognized, however, that the correlation between these clinical and laboratory parameters is unreliable and that, in fact, a majority of anti-HBeAg negative individuals have ongoing active viral replication and viremia that can be detected by a sensitive PCR-based assay (Brechot, 1993 and Zaaijer et al., 1994). This apparent discrepancy is explained in part by the discovery that HBV can develop a mutation in the pre-core region of the gene encoding the pre-core/core protein, which prevent the secretion of HBeAg but does not disrupt viral replication. The clinical significance of this distinction is that the HBeAg-negative patients who have detectable HBV DNA in blood have, in general, more severe hepatitis and a worse response to interferon therapy than do DNA negative individuals (Carman et al., 1992 and Omata et al., 1991).

AURO-HBV-PCR (QUALITATIVE)
Detection of HBV DNA is the most reliable estimator of infectivity and viral replication.

AURO-HBV-PCR is a nested PCR where primers are used to amplify the core and pre-core regions of Hepatitis B Virus genome.

Clinical Significance of PCR based detection of HBV
Detection of HBV-DNA is the most reliable estimator of infectivity and viral replication. The results of many studies indicate that the serum level of HBV DNA provides a more accurate assessment of the amount of circulating virus.

	Fulminant hepatitis, an uncommon presentation of HBV infection, which may pose a diagnostic challenge, since the serological response may be incomplete and viral proteins, may be undetectable in up to one-half of patients (Wright et al., 1999).
	Chronic hepatitis of unknown etiology in a patient with risk factors for HBV infection: In some studies up to 50% of the cases are related to HBV infection (Lai et al., 1989). Some new studies show that up to 50% of the cases are related to HBV infection (Lai et al., 1989).
	Unexplained post-transfusion hepatitis, in which up to 20% of cases may be related to HBV infection.
	Liver transplant recipients, where pre-existent seropositivity or immunoprophylaxis with anti-HBsAg-specific immunoglobulin interferes with the diagnosis of post-transplant re-infection; detection of HBV DNA may precede the detection of HBsAg or elevated liver function tests by several months.

	Brechot C. 1993. Polymerase chain reaction for the diagnosis of viral hepatitis B and C. Gut Suppl: S39-S44
	Carman WF, Thomas HC. 1992. Genetic variation in hepatitis B virus. Gastroenterology 102: 711-719
	Indian Express. March 12, 1999
	Lai ME, Farci P. Figus A, Balestrieri A ,Arnone M, Vygas GN. 1989. Hepatitis B virus DNA in the serum of Sardinian blood donars negative for the hepatitis B surface antigen. Blood 73: 17-19.
	Omata M, Ehata T, Yokosuka O,Hosoda K,Ohto M. 1991. Mutations in the precore region of hepatitis B virus in patients with fulminant and severe hepatitis. N Engl J Med 324: 1699-1704
	Robinson WS. Hepatitis B virus and hepatitis D virus. In: Mandell GL, Bennet JE, Dolin R,eds. Mandell, Douglas, and Bennett's Principle and Practice of Infectious Diseases. 4th ed. New York: Churchill Livingstone, 1995: 1406-1439.
	Wright TL, Mamisk D, Combs C, Kim M, Donegan E, Ferrell L, Lake J, Roberts J , Ascher N. 1992. Hepatitis B virus and apparent fulminant non-A, non-B hepatitis. Lancet 339: 952-955.
	Zaaijer HL, Ter Borg F, Cuypers HTM, Hermus MCAH, Lelie PN. 1994. Comparision of methods for detection of hepatitis B virus DNA. J Clin Microbiol 32: 2088-2091.