Hepatitis C Workup

Updated: Mar 28, 2016
  • Author: Vinod K Dhawan, MD, FACP, FRCPC, FIDSA; Chief Editor: BS Anand, MD  more...
  • Print

Approach Considerations

Serologic screening for hepatitis C virus (HCV) involves an enzyme immunoassay (EIA). These assays are 97% specific but cannot distinguish acute from chronic infection. A rapid antibody test for HCV is available. The recombinant immunoblot assay is used to confirm HCV infection.

A meta-analysis comparing point-of-care screening tests (POCTs) with rapid diagnostic tests (RDTs) indicated that POCTs are highly accurate for diagnosing hepatitis C. [35, 36] POCTs do not require special equipment or electricity and are more robust than RDTs at high temperatures; thus, they may enable expanded screening.

Health care personnel who sustain a needle-stick injury involving an HCV-infected patient should undergo polymerase chain reaction (PCR) testing for HCV immediately and then every 2 months for 6 months. If HCV infection is diagnosed, therapy can be instituted.

Other baseline studies include the following:

  • Complete blood cell count (CBC) with differential
  • Liver function tests, including alanine aminotransferase (ALT) level
  • Thyroid function studies
  • HCV genotyping as an aid for guiding treatment
  • Quantitative HCV RNA assay
  • Screening tests for coinfection with HIV or hepatitis B virus (HBV)
  • Screening for alcohol abuse, drug abuse, and/or depression

The CBC demonstrates thrombocytopenia in approximately 10% of patients. Low thyroxine levels are found in approximately 10% of patients, as well. Stress testing may be necessary in appropriate patients. An ophthalmologic examination may also be necessary.

A diagnostic algorithm for the evaluation of hepatitis C is shown in the following image.

Diagnostic algorithm for hepatitis C virus infecti Diagnostic algorithm for hepatitis C virus infection.

In August 2012, the Centers for Disease Control and Prevention (CDC) expanded their existing, risk-based testing guidelines to recommend a 1-time blood test for hepatitis C virus (HCV) infection in baby boomers—the generation born between 1945 and 1965, who account for approximately three fourths of all chronic HCV infections in the United States—without prior ascertainment of HCV risk (see Recommendations for the Identification of Chronic Hepatitis C Virus Infection Among Persons Born During 1945–1965). [31]

One-time HCV testing in this population could identify nearly 808,600 additional people with chronic infection. All individuals identified with HCV should be screened and/or managed for alcohol abuse, followed by referral to preventive and/or treatment services, as appropriate. [31]

In June 2013, The US Preventive Services Task Force also updated its 2004 HCV screening and treatment recommendations, advocating a 1-time screening for all persons born between 1945 and 1965. The new recommendation arose from the fact that a lack of universal blood screening for the virus prior to 1992 placed persons born between the mid-1940s and mid-1960s at increased risk of exposure to HCV. [37, 38, 39] Screening for HCV in the emergency department (ED) has been found to be feasible, albeit costly. [40, 41]


Hepatitis C Antibody Test

Anti-HCV EIAs include second- and third-generation EIAs. These assays are 97% specific but cannot distinguish acute from chronic infection. The most recent third-generation EIA detects antibodies against core protein and nonstructural proteins 3, 4, and 5 and can yield positive results an average of 8 weeks after the onset of infection.

False-negative results for the presence of HCV antibody can occur in persons with compromised immune systems, such as those with HIV type 1 infection, renal failure, or HCV-associated essential mixed cryoglobulinemia. False-positive EIA results can occur; the likelihood of a false-positive result is greater in persons without risk factors and in those without signs of liver disease, such as blood donors or health care workers.

The FDA has approved OraQuick HCV Rapid Antibody Test, which can be used for persons at risk for hepatitis or for those with signs or symptoms of hepatitis. The test strip can be used with a sample collected from a fingerstick or venipuncture whole blood. [42]


Recombinant Immunoblot Assay

The recombinant immunoblot assay is used to confirm HCV infection. A positive immunoblot assay result is defined as the detection of antibodies against 2 or more antigens and an indeterminate assay result defined as the detection of antibodies against a single antigen.

A positive immunoblot assay result followed by 2 or more instances of undetectable HCV RNA suggests HCV infection has resolved. A positive anti-HCV immunoassay result followed by a negative immunoblot assay result represents a false-positive immunoassay, and no further testing is required.

The recombinant immunoblot assay has limited usefulness in clinical practice.


Qualitative and Quantitative Assays for HCV RNA

Qualitative assays can be used to test for HCV RNA. HCV RNA can be detected in blood using amplification techniques such as PCR or transcription-mediated amplification (TMA). The FDA has approved the following 2 PCR-based tests for qualitative HCV RNA detection:

  • Cobas Amplicor Hepatitis C Virus Test, version 2.0 (Roche Molecular Systems): PCR with a lower limit of detection of 50 IU/mL
  • Versant HCV RNA Qualitative Assay (Bayer HealthCare): TMA with a lower limit of detection of 9.6 IU/mL

Quantitative assays ascertain HCV RNA quantity in blood, using signal amplification (branched DNA [bDNA] assay) or target amplification techniques (PCR, TMA). Reverse transcriptase PCR (RT-PCR) is more sensitive than bDNA testing. The HCV RNA level in blood helps predict the likelihood of a response to treatment, and the change in HCV RNA level can also be used to monitor response.

The same quantitative test should be used throughout therapy to avoid confusion, and results should be reported in international units (IU) to standardize data. The only FDA-approved quantitative test is Versant HCV RNA, version 3.0 (Bayer HealthCare). It is based on bDNA technology and has a dynamic range of 615-7,700,000 IU/mL.


HCV Genotyping

Genotyping is helpful for predicting the likelihood of response and duration of treatment. Patients with genotypes 1 and 4 are generally treated longer than genotypes.

Genotyping can be performed by direct sequence analysis, reverse hybridization to genotype-specific oligonucleotide probes, or restriction fragment length polymorphisms (RFLPs).

In June 2013, the FDA approved the Abbott RealTime HCV Genotype II test, which, by analyzing a sample of an infected patient’s blood plasma or serum, can differentiate HCV genotypes 1, 1a, 1b, 2, 3, 4, and 5. This new test is approved for use in adult, non-immunocompromised patients with known chronic HCV infection but has not been approved for diagnostic use or as a screening test for HCV genetic material. FDA approval was based partly on a comparison of the test's accuracy with that of a validated gene-sequencing method. [43, 44]

Two other genotype tests are available, although neither has been approved by the FDA. They are as follows:

  • Trugene HCV 5'NC Genotyping kit (Visible Genetics; Toronto, Canada): Based on direct sequencing followed by comparison with a reference sequence database
  • Line Probe Assay (Inno LiPA HCVII, Innogenetics; Ghent, Belgium): Based on reverse hybridization of PCR amplicon on a nitrocellulose strip coated with genotype-specific oligonucleotide probes

In addition to HCV genotype, a growing body of research indicates that patient genetics play a role in response to treatment. The single-nucleotide polymorphism (SNP) rs12979860, located near the IL28B gene on chromosome 19, which encodes type III interferon, is associated with more than a 2-fold difference in the rate of sustained virologic response to antiviral treatment with pegylated interferon and ribavirin. This SNP can be detected by PCR and is an independent predictor of sustained virologic response regardless of HCV genotype. [45]


Serologic Testing

Della Rossa et al reported that cryoglobulins are found in as many as 50% of persons with HCV infection. [46] HCV is the primary cause of essential mixed cryoglobulinemia (ie, type 2 cryoglobulinemia); as many as 90% of affected persons have HCV viremia. Cryoprecipitates usually contain large amounts of HCV antigens and antibodies. Vasculitis, arterial hypertension, purpura, lichen planus, arthralgias, and low thyroxine levels were associated with titers positive for cryoglobulin.

Other common serologic findings in patients with chronic HCV infection include one or more of the following:

  • Antinuclear antibody (ANA; 41%)
  • Rheumatoid factor (38%)
  • Anticardiolipin antibody (27%)
  • Antithyroid antibody (13%)
  • Anti–smooth muscle antibody (9%)

Liver Biopsy

Liver biopsy is not considered mandatory before the initiation of treatment, but it may be helpful for assessing the activity and severity of HCV-related liver disease. However, some experts recommend biopsy only in the following situations:

  • The diagnosis is uncertain
  • Other coinfections or disease may be present
  • The patient being considered for treatment has normal liver enzyme levels and no extrahepatic manifestations
  • The patient is immunocompromised

Histologic Findings

Lymphocytic infiltration, moderate degrees of inflammation and necrosis, and portal or bridging fibrosis are noted. Regenerative nodules are seen in patients with cirrhosis. Some patients also may have findings indicative of HCC.

Most pathologists give separate measurements of disease activity (grade) and fibrosis (stage). Many scoring systems are used, including the Ishak (6-point scale) and the Knodell histologic activity index (18-point score); both are useful for assessing improvements in histologic findings in studies but are impractical for clinical use because of interobserver disagreement.

The METAVIR score was developed by the French METAVIR Cooperative Study Group and reported by Bedossa and Poynard in 1996; it is frequently used in European trials. This score consists of a 3-point activity scale and 4-point fibrosis score, with good agreement among pathologists. In the United States, many pathologists use a scale described by Batts and Ludwig in 1995, which consists of an activity grade (0-4) and a fibrosis stage (0-4).

Noninvasive methods of assessing hepatic fibrosis are in development. Current serum assays are directed at measuring breakdown products of extracellular matrix constituents (eg, glycoproteins, propeptides) and their regulatory enzymes (eg, lysyl oxidase, lysyl hydroxylase, propyl hydroxylase).


Radiologic Studies

A liver stiffness test (FibroScan) is available as a noninvasive method of staging liver disease in persons with chronic hepatitis C. Obesity, female sex, operator experience, and age older than 52 may give invalid results. Falsely high estimates of liver fibrosis have also been reported with acute inflammation and recent food intake.

On December 17, 2014, the FDA gave marketing approval for the Hepatiq radiologic image processing system. [47, 48] The software application uses quantitative analysis of nuclear medicine liver-spleen images to determine the severity of liver disease and to predict clinical outcomes. [48] The developer noted that Hepatiq "automates the Quantitative Liver Spleen Scan (QLSS) that has been proven to be an accurate predictor of clinical outcomes in the recently concluded HALT-C [ epatitis C ntiviral ong-term reatment against irrhosis] trial." [48] The HALT-C trial was a multicenter, randomized controlled study that evaluated whether long-term interferon would suppress HCV, prevent progression to cirrhosis, prevent liver cancer, and reduce the need for liver transplantation. [49]