Assay Sheet

Test ID

1300 Hepatitis C Genotyping (HCVG)

CPT Code

87902 

Clinical Utility

Hepatitis C Genotyping can be used to help predict the outcome of therapy and to influence the choice of therapeutic drugs. Clinical outcome depends on the genotype of the infection, pretreatment viral load, and whether or not liver cirrhosis is present. Genotypes 1a and 1b have the poorest clinical outcomes with sustained viral response of only 46% with combination antiviral therapy. The ViraCor HCVG assay detects genotypes 1-6 and can detect dual genotype infections.

Procedure

Extraction of nucleic acid from plasma; reverse transcription of the target RNA to generate complementary DNA. Detection of hepatitis C genotypes 1 through 6 occurs by utilizing primers and probes specific to the different HCV genotypes with real-time PCR.
Note: 1300 Hepatitis C Genotyping must be ordered with 1200 Hepatitis C (HCV) Real-time RNA qPCR. HCV Genotyping is performed following confirmation of adequate viral load to obtain a genotyping result. If the HCV viral load is <1,000 IU/ml, genotyping may not be successful.

Specimens

Whole Blood: 7-10 ml submitted in EDTA, ACD Solution A, or PPT sterile tube.

Minimum specimen requirement is 2 ml plasma.

• Separate plasma from cells within 4 hours of collection by centrifuge at 1000 xg for 10-15 minutes. Do not clarify by filtration or further centrifugation.
• Ship ambient or frozen.

Specificity

Detects all 6 HCV genotypes. The primers and probes used in this assay will distinguish HCV genotypes 1a, 1b, 2a, 2b, 3, 4, 5, and 6.

Hepatitis C Genotyping Assay Range

Genotypes: 1a, 1b, 2a, 2b, 3, 4, 5, or 6

Turnaround Time

Next day, following quantitation result

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PCR tests are performed pursuant to a license agreement with Roche Molecular Systems, Inc.
This assay was developed and the performance characteristics were determined at ViraCor Laboratories. This test is performed in a CLIA certified laboratory. FDA approval is not required for the performance of this test.
AS15B-0108

Pathogen Overview

ABOUT THE VIRUS

Hepatitis C virus (HCV) is a member of the Flaviviridae family. It is a single-stranded, positive-sense RNA virus, which is spherical and lipid-enveloped. HCV was discovered in the late 1980s, and is considered to be one of the leading causes of liver disease in the United States. Six major genotypes have been identified to date, 1-6, and more than 50 subtypes. Genotype 1 is the most common strain in the United States, accounting for approximately 75% of all infections. This is unfortunate, as genotype 1 is the least likely to respond to treatment. The genotypes differ by 31-34% in their nucleotide sequences; subtypes differ by 20-23% based on their full-length genomic sequence comparisons. This extensive genetic heterogeneity, as well as the propensity for mutation, has hindered vaccine development.

CLINICAL MANIFESTATIONS

HCV is considered to be responsible for most cases of hepatitis since the hepatitis B (HBV) vaccine came into widespread use. The most common risk factors for HCV infection include a blood transfusion before 1992, when testing methods had not yet been developed to screen out HCV-tainted blood, IV drug abuse or contact with contaminated blood, such as tattoos or needlestick injuries. To a lesser extent, infants born to an infected mother and sex with an infected partner are also risk factors. However, an estimated 20% of patients infected with HCV do not have identifiable risk factors.

According to the CDC website, as of late 2006, approximately 170 million people worldwide are infected with HCV; 4.1 million are Americans. About 3.4 million new infections occur worldwide each year.

HCV infection is generally asymptomatic during the acute phase, making diagnosis of the infection during the acute phase very unusual. In rare cases, acute hepatitis is accompanied by jaundice, malaise, weakness, and anorexia. More rarely, fulminant hepatitis may develop in the acute phase. It is estimated that 74-86% of individuals develop persistent viremia, which subsequently leads to chronic infection and possible cirrhosis or hepatocellular carcinoma. The development of persistent infection in most individuals is probably due to a poor T-lymphocyte response and the high propensity of the virus to mutate.

The time interval between infection and development of chronic liver disease can range from 20 to over 30 years. Once a chronic infection is established, its clinical progression can be accelerated by such things as alcohol consumption, coinfection with HIV-1 (human immunodeficiency virus) or HBV, being of the male sex, African American, or of an older age at the time of infection. A typical presentation of chronic HCV infection involves a relapsing remitting infection with recurrent bouts of hepatitis, marked by fluctuation in serum AST/ALTs. Specific symptoms are related to liver dysfunction, and involve jaundice, ascites, or GI bleeding. These are typically seen only in patients with disease that is far advanced. Depending on the patient, AST/ALT levels may even be normal during periods of remission, so the absence of ALT abnormalities does not automatically preclude the presence of HCV infection.

End stage liver disease caused by chronic HCV infection has become a primary cause of liver transplantations in Western countries. Epidemiological studies have shown the association between chronic HCV infections and hepatocellular carcinoma, which is known to result in death in approximately 33% of patients with HCV cirrhosis.

LABORATORY DIAGNOSIS

Most individuals with HCV are diagnosed after a routine physical examination shows elevated liver enzymes. HCV is then diagnosed by two antibody tests. First, an ELISA is performed, if positive, it is often confirmed by a RIBA test. A positive antibody test means the patient has been exposed to HCV and has a 70% chance of developing chronic hepatitis C. Since HCV cannot be grown in the clinical laboratory, molecular testing is needed to confirm the presence of the virus. Tests for HCV infection have typically included both serologic assays to determine serostatus and molecular tests to monitor the viral burden (viral load) in individuals. Any patient undergoing treatment for HCV infection will have their viral load monitored regularly by quantitative real-time PCR testing. Real-time PCR testing is highly sensitive and specific, making it the ideal assay to monitor a patient’s response to antiviral therapies. The ViraCor HCV quantitative real-time PCR assay has an unusually wide assay range, with a 5 IU/ml lower limit of detection, which makes it particularly helpful to the physician in ascertaining viral clearance.

TREATMENT

Therapy with pegylated interferon and ribavarin has become the standard of care, although not all individuals will clear the virus, particularly those infected with HCV genotype 1. If the patient does not show at least a 2-log10 drop in viral load 12 weeks after initiating therapy, the likelihood of a sustained response is quite low and therapy should be discontinued.

Liver transplantation is the only available treatment option for individuals who have developed decompensated HCV-related cirrhosis. Liver transplantation is also indicated for some patients who have early stage hepatocellular carcinoma. Despite the fact that recurrence of HCV is almost inevitable in these individuals, their 1 and 5 year survival rates are similar to those of patients with other common indications for liver transplantation.

Selected References

Knipe D, Howley P. Fields Virology. 5th ed. Philadelphia, PA: Lippincott Williams & Wilkins; 2006.

Lauer GM, Walker BD. Hepatitis C virus infection. N Engl J Med. 2001;(345):41-52.

Lemon SM, Brown EA. Hepatitis C virus. In: Mandell GL, Bennett JE, Dolin, eds. Mandell, Douglas and Bennett’s Principles and Practice of Infectious Diseases. Vol 2. 4th ed. New York, NY: Churchill Livingstone; 1995:1474-1486.

Pirsch JD, Becker BN, Becker YT, Odorico JS, Chin LT, et al. HCV-related kidney disease after kidney transplantation. American Transplant Congress; 2003. Abstract 576.
Rodriguez-Luna H, Vargas H, De Petris G, Byrne T, Moss A, et al. Hepatitis C virus recurrence in living donor transplantation vs. cadaveric liver transplantation. American Transplant Congress; 2003. Abstract 34.

Zigneno AL, Gragnani L, Di Pietro E, Solazzo V, Puliti S, et al. HCV infection, malignancy, and liver transplantation. Transplant Proc. 2003;(35):1031-1033.

PAO-06-0707 PCR tests are performed pursuant to a license agreement with Roche Molecular Systems, Inc.

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Collection & Shipping

Turnaround Time: Within 24 hours of specimen receipt at ViraCor

PCR tests are performed pursuant to a license agreement with Roche Molecular Systems Inc.
ImmuKnow is a registered trademark of Cylex Incorporated.
Respiratory Viral Panel is a product of Luminex Corporation.

Abstracts & Publications

Herrine SK. Approach to the patient with chronic Hepatitis C virus infection. Ann Intern Med. 2002;(136):747-757.

Chronic hepatitis C virus (HCV) infection is common and often asymptomatic. Antibodies against HCV are a highly sensitive marker of infection. Molecular testing for HCV is used to confirm a positive result on antibody testing and to provide prognostic information for treatment; however, quantitative HCV RNA does not correlate with disease severity or risk for progression. Chronic HCV infection is most frequently associated with remote or current intravenous drug use and blood transfusion before 1992, although as many as 20% of infected patients have no identifiable risk factor. In an estimated 15% to 20% of persons infected with HCV, the infection progresses to cirrhosis; alcohol intake is an important cofactor in this progression. Most specialists prefer to include an examination of liver histology in the management of patients with chronic HCV infection to aid prognostic and treatment decisions. The current standard of pharmacologic treatment of chronic HCV is weekly subcutaneous peginterferon in combination with daily oral ribovarin, which results in sustained virologic response in approximately 55% of chronically infected patients. Side effects of interferon therapy include myalgias, fever, nausea, irritability, and depression. The cost-effectiveness of interferon therapy is similar to that of many commonly accepted medical interventions. The primary care physician serves a vital role in identifying patients with chronic HCV infection, educating patients about risk factors for transmission, advising patients about the avoidance of alcohol, and aiding patients in making treatment decisions.

Lauer GM, Walker BD. Hepatitis C virus infection. N Engl J Med. 2001;(345):41-52.

Hepatitis C virus (HCV) infects an estimated 170 million persons worldwide and this represents a viral pandemic, one that is five times as widespread as infection with the human immunodeficiency virus type 1 (HIV-1). The institution of blood-screening measures in developed countries has decreased the risk of transfusion-associated hepatitis to a negligible level, but new cases continue to occur mainly as a result of injection-drug use and, to a lesser degree, through other means of percutaneous or mucous-membrane exposure. Progression to chronic disease occurs in the majority of HCV-infected persons, and infection with the virus has become the main indication for liver transplantation. HCV infection also increases the number of complications in persons who are coinfected with HIV-1. Although research advances have been impeded by the inability to grow HCV easily in culture, there have been new insights into pathogenesis of the infection and improvements in treatment options.

Lemon SM, Brown EA. Hepatitis C Virus. In Mandell GL, Bennett JE, Dolin, eds. Mandell, Douglas and Bennett's Principles and Practice of Infectious Diseases, 4th edition, Vol. 2. New York, NY: Churchill Livingstone; 1995:1474-1486.

This chapter appears in the 2nd volume of the definite textbook on infectious diseases. Chapter 132 discusses the history and molecular structure of hepatitis infections in general, and the pathogenesis, clinical manifestations, diagnosis, epidemiology treatment and prevention of HCV, in particular.

Pirsch JD, Becker BN, Becker YT, Odorico JS, Chin LT et al. HCV-related kidney disease after kidney transplantation. American Transplant Congress; 2003. Abstract 576.

This study examined the outcomes of HCV positive recipients receiving an HCV positive or negative kidney, HCV negative recipients receiving an HCV positive kidney and patients without HCV infection receiving a cadaver transplant at this center since 1990. It would appear from the data generated that there is a significant risk for graft loss in the late transplant period due to CAN. There is also a significant mortality risk in patients with HCV infection or in recipients who are HCV negative and receive a HCV positive kidney transplant.

Rodriguez-Luna H, Vargas H, De Petris G, et al. Hepatitis C virus recurrence in living donor transplantation vs. cadaveric liver transplantation. American Transplant Congress; 2003. Abstract 34.

It has been suggested that HCV has a more severe recurrence in living donor liver transplantation due to regeneration and inflammation. Most of the regeneration occurs in the first 2 months post-transplantation. In our study, using protocol biopsies, there was no statistically significant histologic difference in recurrence at four months post-transplantation in living donor liver transplantation vs. cadaveric liver transplantation.

Zigneno AL, Gragnani L, Di Pietro E, et al. HCV infection, malignancy, and liver transplantation. Transplantation Proc. 2003;(35):1031-1033.

End-stage liver disease due to hepatitis C virus (HCV) infection is one of the most common indications for liver transplantation (LT) in Western countries. Although the pathogenesis is unknown, epidemiological studies have demonstrated an association between HCV infection and the development of hepatocellular carcinoma (HCC), complications of which account for 33% of deaths among patients with HCV cirrhosis. HCV infection may lead not only to liver damage but also to B-cell lymphoproliferative disorders (LPDs), ranging from benign, prelymphomatous conditions, such as mixed cryoglobulinemia (MC), to frank malignancies. The possible evolution of both hepatic and extrahepatic HCV-related damage into malignant conditions makes HCV one of the most interesting oncogenic agents in humans. Both aspects must be taken into account in the medical approach to HCV-positive patients undergoing liver transplantation.