Evaluación de los riesgos del proceso

Diagnostic tests for HCV infection are divided into serologic assays for antibodies and molecular tests for viral particles. Screening assays based on antibody

detection have markedly reduced the risk of transfusion-related infection, and once a person seroconverts, they usually remain positive for antibodies. However, recent data indicate that the level of HCV antibodies decreases gradually over time in the few patients in whom infection spontaneously resolves (Takaki et al., 2000).

Diagnostic tests: ELISA

Anti-HCV antibody is detected by ELISA. The third-generation test is usually very sensitive and very specific. The currently used third-generation ELISAs contain core protein as well as non-structural proteins 3, 4 and 5 and can detect antibodies 4 to 10 weeks after infection. If false positive or false negative results are suspected, the best test for confirmation of HCV infection is HCV RNA PCR. In low-risk

populations, the test misses only 0.5 to 1 % of cases. False negative tests can occur in persons with compromised immunity, such as HIV-1 infection, patients with renal failure, and those with HCV-associated essential mixed cryoglobulinaemia. Anti- HCV antibody is still detectable during and after treatment, whatever the response, and should not be retested.

Diagnostic tests: PCR amplification

In the past few years, assays based on the molecular detection of HCV RNA have been introduced. These tests can be categorised as qualitative and quantitative. Samples to be tested should be separated and frozen within three hours of

phlebotomy. Qualitative HCV RNA tests are based on the PCR technique and have a lower limit of detection of less than 100 copies of HCV RNA per ml of serum. These are the tests of choice for the confirmation of viraemia and the assessment of treatment response. Testing for HCV RNA is a reliable way of demonstrating HCV infection and is the most specific test of infection.

A qualitative PCR assay is particularly useful when: transaminases are normal; several causes of liver disease are possible (i.e. alcohol consumption); in

immunosuppressed patients (i.e. after transplantation, in HIV coinfected patients); and in acute hepatitis C before occurrence of antibodies.

Diagnostic tests: genotype and serotype

There are six genotypes of hepatitis C and more than 50 subtypes. Knowing the genotype or serotype (genotype-specific antibodies) is helpful when choosing the interferon–ribavirin treatment duration. Response rates to treatment are around 88 % for genotypes 2 and 3, and around 48 % for genotypes 1, 4, 5 and 6. Genotypes do not change during the course of infection and must not be tested again. Serotyping (1, 2, etc.) is cheaper than genotyping but does not allow assessment of the subtype, which is only determined by genotype (1a,1b, 2a, 2b, etc.). Knowing the subtypes (i.e. 1a versus 1b) is currently not clinically helpful and therefore serotyping could be more cost-effective. There is no relationship between the severity of the disease (fibrosis stage) and genotypes.

Diagnostic tests: quantification of HCV RNA in serum

Methods for measuring the level of virus in serum used quantitative PCR and a branched DNA (bDNA) test (Pawlotsky et al., 2000). In the more recent studies, the median of viral load ranged from 2 to 4 million copies/ml (‘Superquant’ assay, National Genetics Institute, Los Angeles, California). Knowing the viral load is helpful for the choice of interferon–ribavirin treatment duration. Patients with a high initial viral load have higher relapse rates and benefit more from a 48-week treatment regimen than patients with a lower viral load. In contrast to HIV

infection, viral load does not correlate with the severity of hepatitis (fibrosis progression).

Recently, an effort was made to define clinically relevant HCV RNA loads in standardised international units (IU) for use in routine clinical and research

applications based on standardised quantitative assays validated with appropriate calibrated panels (Pawlotsky et al., 2000). Two HCV RNA quantitative assays have already been assessed: the ‘Superquant’ assay, for which possibly relevant

thresholds were established; and the semi-automated ‘Cobas Amplicor HCV Monitor’ assay version 2.0 (Cobas v2.0, Roche Molecular Systems, Pleasanton, California), which measures HCV RNA loads in IU/ml. A value of 2 000 000 copies/ml (6.3 log10 _{copies/ml) with ‘Superquant’ was converted to nearly}

800 000 IU/ml (5.9 log10_{IU/ml), and 3 500 000 copies/ml (6.5 log}10_copies/ml)

to nearly 1 300 000 IU/ml (6.1 log10 _{IU/ml). To simplify diagnosis, we recommend}

a decision threshold of 1 000 000 IU/ml (6.0 log10_{IU/ml) to tailor the interferon-}

alpha/ribavirin treatment duration. Diagnostic tests: liver biopsy

PCR HCV RNA testing can diagnose hepatitis C infection. Biopsy is generally recommended for the initial assessment of persons with chronic HCV infection (Consensus statement, 1999). Biopsy is necessary for staging the severity of disease (fibrosis stage) and grading the amount of necrosis and inflammation (Metavir Cooperative Study Group, 1994; Bedossa and Poynard, 1996). Biopsy is also helpful in ruling out other causes of liver disease such as alcoholic features, non-alcoholic steatohepatitis, autoimmune hepatitis, medication-induced,

coinfection with HBV, HIV or iron overload.

Liver biopsy is helpful before treating a patient, as an aid to the choice and duration of therapy. Biopsy is usually not helpful when cirrhosis is clinically or biologically obvious. Liver biopsy is usually performed by the intercostal route. In the case of clotting disorders, the transjugular route is used.

Complications of liver biopsy

From nine large-scale observations, gathering 98 445 cases of liver biopsy, the incidence of severe adverse events was 3.1 per 1 000 (95 % confidence interval: 2.8–3.5) with a 0.3 per 1 000 mortality (95 % confidence interval: 0.2–0.5) (Poynard et al., 2000a). Factors associated with severe adverse events and mortality were cirrhosis, age of the patient, and the presence of liver cancer. Biochemical markers of liver fibrosis and activity

In the next decades, liver biopsy indications should decrease because of the validation of serum markers (Poynard et al., 2000b, 2002a; Imbert-Bismut et al., 2001). We recently made a prospective assessment of the predictive value of a combination of six simple serum biochemical markers for the diagnosis of significant fibrosis (ranging from few septa to cirrhosis) and necroinflammatory activity (Imbert-Bismut et al., 2001). From these results, we suggested that biochemical markers could lead to a significant reduction in the number of liver biopsies performed in patients with chronic Figure 1: Longitudinal assessment of biochemical markers of liver fibrosis (fibrosis

index) according to virological response. Adapted with permission (Poynard et al., 2002a)

0.00 0.10 0.20 0.30 0.40 0.50 Baseline 6 12 24 Fibrosis index Non- responders Sustained Relapsers

hepatitis C. Several studies confirmed the diagnostic value of these fibrosis and activity indices in different multicentre populations including longitudinal assessment with two liver biopsies (Poynard et al., 2002a, 2003) (Figure 1) and in patients coinfected with HIV (Myers, 2003).