Hepatitis C virus: Prevention, screening, and interpretation of assays
ABSTRACTPatients at risk of hepatitis C virus (HCV) infection should be screened for it so that they can be treated and potentially cured, or can at least avoid transmitting the disease to others. The authors describe why and how to screen for HCV and how to interpret the test results.
KEY POINTS
- Patients who should be screened include intravenous drug abusers, people infected with human immunodeficiency virus, patients with unexplained elevated alanine aminotransferase levels, infants born to infected mothers, and people with infected sexual partners.
- Patients at risk of HCV infection should be tested for anti-HCV antibody using an enzyme immunoassay (EIA).
- Positive results on anti-HCV EIA testing should be confirmed with an assay for HCV RNA.
- HCV genotyping can help predict the response to therapy. Genotypes 2 or 3 are more likely to respond to therapy than genotype 1.
CONFIRMATORY TESTING WITH ASSAYS FOR HCV RNA
As stated above, a positive result on an anti-HCV EIA needs to be confirmed with an assay for HCV RNA, of which there are two types, ie, qualitative and quantitative.
Each involves trade-offs. Qualitative assays are more sensitive and detect more cases, but they provide no information about the amount of virus (viral load). Quantitative assays are less sensitive, so a negative result does not completely exclude hepatitis C, although they can still can detect 95% of cases. They do, however, measure the viral load.
Therefore, the type of test to use depends on the patient’s risk profile, the goals of testing, and the setting in which future care will be provided. The primary objective when a patient has a positive EIA test is to determine whether he or she has ongoing infection, a goal most expeditiously achieved using a qualitative assay. However, since a quantitative assay can detect the vast majority of cases of active HCV infection, many clinicians select this as the test of first choice when the probability of HCV is high (eg, in a patient with risk factors and abnormal liver tests). If the pretest probability is low, a qualitative assay is the better choice.
Many commercial assays are available for detecting (qualitative assays) or measuring (quantitative assays) HCV RNA.
Qualitative HCV RNA assays
The approved qualitative assays are:
- Amplicor HCV Test, version 2.0 (Roche Molecular Diagnostics, Pleasanton, CA)
- Cobas Amplicor HCV Test, version 2.0 (Roche Molecular Diagnostics)
- Ampliscreen (Roche Molecular Diagnostics)
- Versant HCV RNA Qualitative Assay (Siemens Healthcare Diagnostics, Deerfield, IL)
- Procleix HIV-1/HCV Assay (Chiron, Emeryville, CA).
Quantitative HCV RNA assays
The approved quantitative assays are:
- Amplicor HCV Monitor (Roche Molecular Diagnostics)
- Cobas Amplicor HCV Monitor, version 2.0 (Roche Molecular Diagnostics)
- Versant HCV RNA 3.0 Assay (bDNA) (Siemens Healthcare Diagnostics)
- Cobas Taqman HCV Test (Roche Molecular Diagnostics).
Quantitative tests use target amplification with PCR, transcription-mediated amplification (TMA), or a signal amplification technique such as a branched DNA (bDNA) assay. The sensitivity varies for different types of amplification. TMA assays appear to be the most sensitive for detecting HCV RNA.
The latest innovation is real-time PCR, which shortens the typical time for PCR processing from 1.5 hours to 35 minutes. It may also detect relapsed HCV infection earlier than regular PCR. With the recent availability of real-time PCR assays, which have sensitivities of 10 to 50 IU/mL, many experts feel there is no longer a need for qualitative assays.74 In fact, many laboratories no longer offer qualitative testing. The Cleveland Clinic laboratory has recently stopped offering this test.
Because RNA testing is widely available, the recombinant immunoblot assay (RIBA) has become obsolete in diagnosing HCV infection, except in special circumstances. Currently, the primary purpose of RIBA testing is to distinguish between resolved HCV infection (EIA-positive, HCV RNA-negative, RIBA-positive) and a false-positive EIA (EIA-positive, HCV RNA-negative, RIBA-negative).
In summary, patients suspected of having acute or chronic HCV infection should first be tested for anti-HCV. Subsequently, HCV RNA testing should be performed in:
- Patients with a positive anti-HCV test
- Patients for whom antiviral treatment is being considered (using a sensitive quantitative assay)
- Patients with unexplained liver disease whose anti-HCV test is negative and who are immunocompromised or suspected of having acute HCV infection.
Significance of the HCV viral load
The significance of the HCV viral load is widely misunderstood. The amount of virus in the blood does not correlate with symptoms, histologic liver injury, or the stage or aggressiveness of disease. Its sole importance is in relation to therapy.
The HCV viral load, measured before treatment, helps predict the likelihood of a treatment response: the lower the pretreatment viral load, the more likely that the patient will respond to current HCV therapies.
Additionally, the pretreatment viral load serves as a baseline for comparison with subsequent measurements during treatment. Patients with HCV genotype 1 who do not achieve more than a 2-log (99%) reduction in viral load by the 12th week of treatment (an early virologic response) have a low response rate, and treatment should generally be stopped, given its cost and side effects.76 However, measuring the viral load to detect an early virologic response is less helpful in patients with HCV genotype 2 or 3 infection, since these patients require only 24 weeks of therapy and most of them clear the virus by week 12 and respond to therapy.
Additionally, patients with genotype 2 or 3 and those with a viral load of less than 600,000 IU/mL have been found to achieve higher rates of sustained virologic response.15 A sustained virologic response is defined as the absence of HCV RNA 24 weeks after stopping treatment and is now considered to be the best predictor of long-term treatment response. A sustained virologic response is generally regarded as a “virologic cure.”
HCV GENOTYPE AFFECTS SUCCESS AND DURATION OF TREATMENT
HCV has at least six major genotypes.1,3–6 Several genotypes are subclassified as “a” or “b” (ie, genotype 1a or 1b); however, these distinctions are of little clinical use.
In the laboratory, HCV genotypes are identified by restriction fragment length polymorphism, by direct sequence analysis, or by reverse hybridization. Once the HCV genotype has been identified, there is no need to repeat the test.
Different genotypes are more common in some areas of the world than in others. Genotype 1 is the one most common in the United States (accounting for 70% to 75% of cases), followed by genotypes 2 and 3 (25%–30%). Genotype 4 is most common in Egypt and the Arabian peninsula.
HCV genotyping is important because it can help predict the likelihood of a response to treatment and in planning the dose and duration of therapy.77 For example, treatment with pegylated interferon plus ribavirin is predicted to work approximately 50% of the time for people with genotype 1, but 80% to 90% of the time for people with genotypes 2 or 3.15–17,78 Additionally, patients with genotype 1 need 12 months of therapy to achieve maximum benefit, whereas those with genotypes 2 and 3 require treatment for only 6 months to achieve maximum benefit.
