INTRODUCTION — The hepatitis D virus (HDV, also called the delta virus) is a defective pathogen that requires the presence of the hepatitis B virus (HBV) for infection. The HD virion is composed of an outer lipoprotein envelope made of the surface antigen of the HBV (HBsAg) and an inner ribonucleoprotein structure in which the HDV genome resides. The HDV genome consists of a single stranded RNA which is folded as a rod-like structure through internal base-pairing (figure 1). It is complexed with the only HDV-encoded antigen, the HDAg [1].
HDAg can elicit a specific immune response in the infected host, consisting of antibodies of the IgM and IgG class (anti-HDV). In HDV infected individuals, the timing of appearance and level of HDV RNA, HDAg, and anti-HDV in serum allow the three HDV-related clinical entities to be discriminated:
●Acute HBV/HDV coinfection
●Acute HDV superinfection of a chronic HBV carrier
●Chronic HDV infection
This topic will review the diagnosis of HDV infection. The pathogenesis, epidemiology, clinical manifestations and treatment of HDV are discussed separately. (See "Pathogenesis, epidemiology, natural history, and clinical manifestations of hepatitis D virus infection" and "Treatment and prevention of hepatitis D virus infection".)
DIAGNOSIS OF HDV INFECTION — Due to the dependence of hepatitis D virus (HDV) on hepatitis B virus (HBV), the presence of hepatitis B surface antigen (HBsAg) is necessary for the diagnosis of HDV infection. The additional presence of IgM antibody to hepatitis B core antigen (IgM anti-HBc) is necessary for the diagnosis of acute HBV/HDV coinfection (table 1).
Detection of serum HDV RNA — HDV RNA can be detected in serum by reverse transcriptase-polymerase chain reaction (RT-PCR)-based assays (table 1).
RT-PCR assays are very sensitive and may detect viral loads as low as 10 genomes per mL [2-6]. However, the extensive sequence heterogeneity of different HDV isolates makes it difficult to choose suitable primers for the amplification of HDV RNA since only a few conserved regions exist in the HDV genome. In addition, the secondary and tertiary structure of the HDV RNA may hamper efficient amplification, even of highly conserved regions [5]. The best efficiency is obtained by amplifying the C-terminal half of the HDV antigen (HDAg)-encoding region [5]. Although automated assays are commercially available to allow quantification of HDV RNA in the serum of patients with HDV infection before and during treatment [7,8], none of them have been approved for clinical use by the US Food and Drug Administration.
HDV genotyping can be performed by direct sequencing [9-12], but this test is not routinely recommended since the clinical relevance of HDV genotyping is uncertain. Furthermore, the phylogenetic classification of the various HDV genotypes is in debate [13]. (See "Pathogenesis, epidemiology, natural history, and clinical manifestations of hepatitis D virus infection", section on 'Natural history of chronic hepatitis D'.)
Sequence analysis of HDV RNA can also be used to detect common sources of infection, such as after intrafamilial or perinatal transmission [14,15]. However, it is unclear if HDV RNA levels in the blood can be used as a prognostic factor to determine disease progression [16].
Detection of anti-HDV antibody — Total (IgM and IgG) anti-HDV antibodies can be detected by enzyme immunoassays (EIAs) or radioimmunoassays (RIAs) (table 1).
●Total anti-HDV antibody usually appears after four weeks of acute infection in acute hepatitis D (figure 2). As a result, its clinical value is limited unless repeated testing is performed [17,18]. Nevertheless, a well-documented anti-HDV seroconversion may be the only way to diagnose acute HDV infection in the absence of other markers of HDV infection.
●High-titer anti-HDV of the IgG class is present in chronic HDV infection. It correlates well with ongoing HDV replication and may help in differentiating current from past HDV infection [19].
●Anti-HDV of the IgM class can be detected by EIAs or RIAs, but these assays are not available for clinical use in the United States [20]. IgM anti-HDV is transient and delayed if the course of acute hepatitis D is self-limited, but it may be the only serum marker of acute HDV infection [17]. In patients who progress to chronic HDV infection, which is usually the case in those with HDV superinfection, IgM anti-HDV is brisk and long-lasting. It should be remembered, however, that differentiating between HBV/HDV coinfection and HDV superinfection in an HBV carrier relies mainly on the detection of high-titer IgM anti-HBc in patients with coinfection.
IgM anti-HDV is present in high titers during chronic HDV infection, and the titers correlate with the level of HDV replication and severity of liver disease [20], although HDV replication is best assessed by quantifying HDV RNA in serum. IgM anti-HDV gradually disappears from serum in patients who have persistent remission after interferon therapy and following liver transplantation [21].
Detection of serum HDAg — Serum HDV antigen (HDAg) can be detected by EIA or RIA. However, these assays are rarely used and are not approved for clinical diagnosis in the United States.
●In acute HDV infection, serum HDAg appears early, but its detection by EIA is short-lived and may require repeated testing (figure 2) [17,18,22,23], except in immunocompromised individuals [24].
●In chronic HDV infection, anti-HDV is present in high titers, but HDAg cannot be detected by EIA since it is complexed with anti-HDV. In this setting, serum HDAg is best detected by immunoblot assay [25], which is more sensitive [26]; however, this assay is technically difficult and time- and labor-consuming.
Tissue markers of HDV infection — Both HDAg and HDV RNA can be detected in liver tissues routinely processed for histopathologic evaluation.
●HDAg can be detected by direct immunofluorescence or immunohistochemistry. Although initially proposed as the "gold" standard for diagnosis of current HDV infection [27], as many as 50 percent of liver biopsy specimens from patients who have been infected for 10 or more years may be negative for HDAg, suggesting that the levels of HDV replication may decrease with time [6,28]. In patients who are negative for HDAg, the diagnosis of current HDV infection has to rely on the detection of HDV RNA or high-titer anti-HDV antibodies in the serum. (See 'Detection of serum HDV RNA' above and 'Detection of anti-HDV antibody' above.)
●HDV RNA can be detected by in-situ hybridization. However, the techniques involved are time-consuming and tedious; they are not recommended for clinical use [29].
SUMMARY AND RECOMMENDATIONS — Due to the dependence of hepatitis D virus (HDV) on hepatitis B virus (HBV), the diagnosis of HDV cannot be made in the absence of markers of HBV infection. Screening for HDV infection should be considered in all hepatitis B surface antigen (HBsAg)-positive persons (table 1). (See 'Acute HBV infection' below and 'Acute hepatitis of undetermined origin in a chronic HBV carrier' below and 'HBsAg-positive chronic liver disease' below.)
Acute HBV infection — In patients with acute HBV infection, testing for HDV coinfection should be performed in those who have risk factors for HDV (intravenous drug users and patients from endemic countries) or those who present with unusually severe or protracted hepatitis.
Patients should be positive for HBsAg and have high titer IgM anti-HBc. However, markers of HBV replication may precede or follow those of HDV. In addition, occasional patients have already seroconverted to anti-HBs if they present during the second phase of biphasic hepatitis. These patients should still be positive for high-titer IgM anti-HBc (table 1).
Serum HDV RNA and/or HDV antigen (HDAg) are usually positive at presentation. If these assays are not available, repeated testing for anti-HDV (total or IgM) should be performed to document anti-HDV seroconversion.
Acute hepatitis of undetermined origin in a chronic HBV carrier — Tests for HDV should be obtained to rule out acute HDV superinfection in this setting. Since HDV superinfection may occur in previously unrecognized chronic HBV carriers, distinguishing between this condition and acute HBV/HDV coinfection can be difficult. This distinction is important because of the differences in prognosis (table 1). (See "Pathogenesis, epidemiology, natural history, and clinical manifestations of hepatitis D virus infection".)
●HBsAg is present in both situations, but IgM anti-HBc should be negative in acute HDV superinfection (figure 3).
●The diagnosis is made more difficult since HDV superinfection may cause transient suppression of HBV replication, resulting in very low and, rarely, undetectable levels of HBsAg.
●As in patients with acute HBV/HDV coinfection, patients with acute HDV superinfection are usually positive for HDV RNA and/or HDAg in serum at the time of presentation. However, in contrast to acute coinfection, acute HDV superinfection is characterized by persistent detection of HDV RNA in serum and rapidly increasing titers of anti-HDV (total and IgM).
HBsAg-positive chronic liver disease — Screening for HDV should be considered in all HBsAg-positive patients to rule out coexistent chronic HDV infection. This is best achieved by testing for total anti-HDV antibody. The presence of active HDV infection should be confirmed by testing for serum HDV RNA. Quantification of serum HDV RNA is important in evaluating the need for and efficacy of antiviral therapy. (See "Treatment and prevention of hepatitis D virus infection", section on 'Treatment of chronic hepatitis D'.)
In patients with chronic HDV infection, serum HBV DNA is usually present at low levels, but may be undetectable. In addition, patients are often hepatitis B e antigen (HBeAg) negative and hepatitis B e antibody (anti-HBe) positive.
When and how to test for HDV infection — A one-time screening for HDV infection should be considered in all persons with HBV infection. In particular, testing should be performed in patients who emigrated from countries with a high prevalence of HDV infection (eg, Eastern European, Mediterranean, and Central Asian countries) and in those with a history of injection drug use.
Patients should be tested initially for total anti-HDV. When possible, the diagnosis should be confirmed by reverse transcriptase-polymerase chain reaction (RT-PCR) assays for HDV RNA in serum. Total anti-HDV should be detected in all patients with chronic HDV infection; however, a negative test for total anti-HDV does not necessarily exclude a diagnosis of acute HBV/HDV coinfection (table 1). In addition, HDAg may not be detectable in all liver biopsies in patients with chronic HDV infection, particularly during the late stages. (See 'Diagnosis of HDV infection' above.)
