Hepatitis C Virus - HCV

Diagnosis

Indications for Testing

  • New onset of jaundice, anorexia, or dark urine
  • Known exposure to hepatitis
  • Suspicion of chronic hepatitis (elevated liver enzymes)

Laboratory Testing

  • Testing recommendations for chronic hepatitis C (CDC)
  • Initial testing – rule out hepatitis A virus (HAV) or hepatitis B virus (HBV) in acute presentation
    • Perform testing for HAV antibody IgM, HBV core antibody IgM, HBV surface antigen, and HCV antibody
      • Positive HCV from hepatitis panel – perform quantitative HCV RNA PCR test
        • Documents baseline level of viremia for acute disease
        • Quantitative PCR test negative – infected but recovered, or false-positive screen
          • Inform patient that s/he does not have active infection 
        • Quantitative PCR test positive – currently infected (acute versus chronic designation depends on initial presentation)
      • Coinfection with HBV – predicts poorer prognosis
  • Consider further testing when HCV testing is positive
    • HIV testing – coinfection associated with poorer prognosis 
    • HCV genotyping – guides selection of most appropriate antiviral regimen
      • Subtyping for 1a, 1b, and 6 may be useful
    • Interleukin 28 B (IL28B) gene and/or inosine triphosphatase (ITPA) gene for patients with HCV genotype 1
      • IL28B genotype – predicts response to peginterferon (PEG-IFNα) and ribavirin (RBV) therapy for chronic genotype 1 HCV
      • ITPA genotype – predicts response to PEG-IFNα/RBV therapy for chronic HCV-1 infection, aids in dosage planning, and predicts risk of RBV treatment-related anemia
    • Liver biopsy
      • Patients with HCV genotype 1
      • More advanced disease is associated with lower response to therapy

Prognosis

  • Viral load (as measured by quantitative PCR) predicts likelihood of treatment response
    • Lower viral load at therapy initiation is associated with improved therapeutic response

Differential Diagnosis

Screening

  • CDC (2012), Infectious Diseases Society of America (IDSA) (2014), and U.S. Preventive Services Task Force (USPSTF) (2013) recommend screening at least once for all individuals born between 1945-1965
    • All others with risk factors for HCV infection (see Background section) – one-time testing should be performed
  • WHO (2013), American Gastroenterological Association, National Gastroenterology Society (2009), National Hepatology Society (2003), and European Association for the Study of the Liver (EASL) (2014) recommend screening for those at high risk (eg, IV drug users, immigrants from endemic areas)  
    • Initial screening for HCV antibodies by CIA, EIA, ELISA
    • Follow-up testing required for positive result
  • Pregnant females – routine HCV screening is not recommended
  • Injection drug users and HIV-seropositive men who have unprotected sex with men – annual HCV testing (IDSA, 2014)

Monitoring

  • HCV RNA PCR quantitative test – monitor effectiveness of treatment and perform when treatment is complete
    • Monthly until week 12 of treatment
    • Negative result confirms successful treatment

Clinical Background

Hepatitis C is a virally mediated disease of the liver with a propensity to cause chronic infection, leading to cirrhosis and an increased risk of hepatocellular carcinoma.

Epidemiology

  • Prevalence – 2% of U.S. population is infected  
    • >50% of new cases are caused by IV drug use
    • ~25,000 laboratory-confirmed cases of chronic hepatitis C (National Notifiable Diseases Surveillance System, 2010)
  • Age – peaks in 30s-40s
  • Sex – M:F, equal

Organism

  • Single-stranded RNA virus; member of Flaviviridae family (genus Hepacivirus)
  • Six major genotypes with multiple subtypes (1a, 1b, 1c, etc.)
    • Genotype is an important predictor of virological response to HCV treatment
      • Type 1 is predominant genotype in U.S. and more difficult to treat
      • Types 2 and 3 are less aggressive and easier to treat

Genetics

  • Interleukin 28 B (IL28B) genotype

    IL28B Genotype Interpretation

    Genotype

    Interpretation

    rs12979860 C/C

    Favorable

    • Two- to three-fold greater rate of sustained virological response (SVR) following PEG-IFNα/RBV therapy
    • Three-fold increase in natural clearance of HCV

    rs8099917 T/T

    Favorable                                                             

    • Higher rate of SVR following PEG-IFNα/RBV therapy
    • Increased natural clearance of HCV
    rs12979860 C/T
    rs12979860 T/T
    rs8099917 T/G
    rs8099917 G/G

    Unfavorable

    • Less likely to respond to treatment and achieve SVR

    One favorable and one unfavorable single-nucleotide polymorphism (SNP) identified

    Indeterminate

    • Likelihood of SVR following PEG-IFNα/RBV therapy not well-defined
    Inosine triphosphatase (ITPA) genotype

    ITPA Genotype Interpretation

    Genotype

    Interpretation

    rs1127354 A/A
    rs1127354 A/C
    rs7270101 C/C
    rs7270101 C/A

    Protective

    • Decreased ITPase activity
    • Protection against RBV treatment-related anemia in individuals with HCV
    rs1127354 C/C
    rs7270101 A/A

    Not Protective

    • Susceptible to RBV-induced hemolytic anemia

    One protective and one nonprotective single-nucleotide polymorphism (SNP) identified

    Indeterminate

    • Moderate decrease in ITPase activity
    • Decreased hemolytic side effects from RBV therapy

Risk Factors (from IDSA Practice Guidelines, 2014)

  • Risk behaviors
    • Injection drug use (current or ever, including those who injected once)
    • Intranasal illicit drug use
  • Risk exposures
    • Long-term hemodialysis (ever)
    • Getting a tattoo in an unregulated setting
    • Healthcare, emergency medical, and public safety workers after needle stick, sharps, or mucosal exposure to HCV-infected blood
    • Babies born to HCV-infected mothers
    • Prior recipients of transfusions or organ transplants, including individuals who
      • Were notified that they received blood from a donor who later tested positive for HCV infection
      • Received a transfusion of blood or blood components, or underwent an organ transplant before July 1992
      • Received clotting factor concentrates produced before 1987
      • Were ever incarcerated
  • Other medical conditions
    • HIV infection
    • Unexplained chronic liver disease and chronic hepatitis, including elevated alanine aminotransferase levels

Clinical Presentation

  • HCV is typically asymptomatic as acute infection
    • Infection may be initially identified when patient has positive anti-HCV in a blood donor screen or has high alanine aminotransferase (10-20 times the upper limit of normal) in blood chemistry testing for flu-like symptoms
  • Chronic asymptomatic hepatitis may manifest with other systemic symptoms
    • Mixed cryoglobulinemia – systemic vasculitis involving skin, kidneys, nervous system
    • Sjögren syndrome – anti-SSA and SSB antibodies are usually absent or are present in low levels
    • Lichen planus – violaceous papules on any skin site; oral most common
    • Porphyria cutanea tarda
    • Non-Hodgkin lymphoma – B-cell type most common
  • Chronic disease occurs in ~10-20% of patients
    • Cirrhosis (20%) and hepatocellular carcinoma (1-5%)
  • Pregnant females
    • Not transmitted to infant via breast-feeding 
    • Pregnancy not contraindicated

Treatment

  • Genotypes 2 and 3 have more favorable prognosis and treatment response

Indications for Laboratory Testing

  • Tests generally appear in the order most useful for common clinical situations
  • Click on number for test-specific information in the ARUP Laboratory Test Directory
Test Name and Number Recommended Use Limitations Follow Up
Hepatitis Panel, Acute with Reflex to HBsAg Confirmation 0020457
Method: Qualitative Chemiluminescent Immunoassay

Order to evaluate viral etiology in patients with acute hepatitis

Not recommended for screening asymptomatic patients

Panel includes HAV IgM, HBV core antibody IgM, HBV surface antigen with reflex to confirmation, HCV antibody

   
Hepatitis B Virus Surface Antigen with Reflex to Confirmation 0020089
Method: Qualitative Chemiluminescent Immunoassay 

Initial testing for suspected chronic HBV infection

   
Hepatitis C Virus Antibody by CIA with Reflex to HCV by Quantitative PCR 2010784
Method: Qualitative Chemiluminescent Immunoassay/Quantitative Polymerase Chain Reaction

Initial testing for individuals at risk for HCV infection

Reflex pattern – if CIA screening result is low positive or high positive, HCV by quantitative PCR will be added

   
Hepatitis C Virus (HCV) by Quantitative PCR with Reflex to HCV Genotype by Sequencing 2002685
Method: Quantitative Polymerase Chain Reaction/Sequencing

Preferred reflex test to confirm active HCV infection following positive HCV screen

Limit of quantification for HCV PCR – 1.2 log IU/mL (15 IU/mL)

Reflex pattern – if PCR result is ≥3.6 log IU/mL, then HCV genotype by sequencing will be added

If virus is not detected, result will be reported as <1.2 log IU/mL; if virus is detected but number of copies not accurately quantified, result will be reported as not quantified

 
Hepatitis C Virus by Quantitative PCR 0098268
Method: Quantitative Polymerase Chain Reaction

Confirm active HCV infection following positive HCV screen

Establish baseline viral load prior to initiation of therapy

Monitor therapy

Evaluate prognosis and disease progression

Assess transmission of HCV in newborns from HCV-positive mothers

Quantitative range – 1.2-8.0 log IU/mL

If virus is not detected, result will be reported as <1.2 log IU/mL; if virus is detected but number of copies not accurately quantified, result will be reported as not quantified  
Hepatitis C Virus Genotype by Sequencing 0055593
Method: Polymerase Chain Reaction/Sequencing

Order before initiating HCV therapy to aid in determining therapy of choice, likelihood of response, and probable therapeutic duration

Assay does not differentiate between type 1a and type 1b

Do not order prior to molecular confirmation of positive HCV screen

Assay does not subtype

Test may be unsuccessful if HCV RNA viral load is <log 3.6 or 4,000 IU/mL

 
Hepatitis C Virus (HCV) by Quantitative PCR with Reflex to HCV High-Resolution Genotype by Sequencing 2010793
Method: Quantitative Polymerase Chain Reaction/Sequencing

Confirm active HCV infection following positive screen when a higher level of subtype resolution is required

Limit of quantification for HCV PCR – 1.2 log IU/mL (15 IU/mL)

Reflex pattern – if PCR ≥3.6 log IU/mL, HCV high-resolution genotype by sequencing will be added

If virus is not detected, result will be reported as <1.2 log IU/mL; if virus is detected but number of copies not accurately quantified, result will be reported as not quantified

 
Hepatitis C Virus (HCV) Genotype with Reflex to HCV High-Resolution Genotype by Sequencing 2009255
Method: Polymerase Chain Reaction/Sequencing

Reflex panel to use for prognosis and treatment selection when a higher level of subtype resolution is required (eg, 1a vs.1b; 1a or 1b vs. type 6)

Do not order prior to molecular confirmation of positive HCV screen

Reflex pattern – if subtype 1a or 1b is detected, then genotyping will be added

Test may be unsuccessful if HCV RNA viral load is <log 3.6 or 4,000 IU/mL

 
Interleukin 28 B (IL28B)-Associated Variants, 2 SNPs 2004680
Method: Polymerase Chain Reaction/High Resolution Melt Analysis

Predict response to PEG-IFNα and RBV therapy for chronic HCV-1 infection

Clinical sensitivity/specificity – unknown

SNPs other than those targeted will not be detected

Usefulness of IL28B-associated SNPs for predicting therapy response for HCV genotypes other than HCV-1 is unknown; lack of favorable genetic factors should not be used to deny therapy

Mutations in genes and nongenetic factors that may affect response to HCV therapy are not detected

Diagnostic errors can occur due to rare sequence variations

 
Inosine Triphosphatase (ITPA) and Interleukin 28 B (IL28B)-Associated Variants, 4 SNPs 2006344
Method: Polymerase Chain Reaction/Single Nucleotide Extensions

Predict response to PEG-IFNα/RBV therapy for chronic HCV-1

Aid in dose planning for chronic HCV-1 infection

Predict risk of RBV treatment-related anemia

Clinical sensitivity/specificity – unknown

Usefulness of IL28B-associated SNPs for predicting therapy response for HCV genotypes other than HCV-1 is unknown; lack of favorable genetic factors should not be used to deny therapy

Mutations in genes and nongenetic factors that may affect response to HCV therapy are not detected

Diagnostic errors can occur due to rare sequence variations

 
Hepatitis C Virus RNA Quantitative bDNA 0051811
Method: Quantitative Branched Chain DNA

Not recommended to confirm active HCV infection; quantitative PCR generally preferred

If used, provide a baseline viral load for monitoring treatment efficacy

Negative result does not rule out the presence of PCR inhibitors in the patient sample or the presence of HCV RNA concentrations below the level of detection by the assay

Low-positive values may occasionally be seen in specimens from patients who are not infected

 
Additional Tests Available
 
Click the plus sign to expand the table of additional tests.
Test Name and NumberComments
Hepatitis C Virus Antibody by CIA 2002483
Method: Qualitative Chemiluminescent Immunoassay

Initial testing for individuals at risk for HCV infection

One-time screening for population born between 1945-1965

If positive, order quantitative HCV quantitative PCR

Hepatitis C Virus RNA Quantitative bDNA 0051811
Method: Quantitative Branched Chain DNA

Not recommended to confirm active HCV infection; quantitative PCR is generally preferred

If used, provide a baseline viral load for monitoring treatment efficacy

Hepatitis A Virus Antibody, IgM 0020093
Method: Qualitative Chemiluminescent Immunoassay

Rule out acute HAV

Hepatitis B Virus Core Antibody, IgM 0020092
Method: Qualitative Chemiluminescent Immunoassay

Rule out HBV

Hepatitis B Virus Surface Antibody 0020090
Method: Quantitative Chemiluminescent Immunoassay

Determine immunity to HBV

Hepatitis C Virus RNA Quantitative bDNA with Reflex to Hepatitis C Virus RNA Quantitative, Real-Time PCR 2002682
Method: Quantitative Branched Chain DNA/Polymerase Chain Reaction

Limited use; HCV quantitative PCR is preferred

Hepatitis C Virus RNA Quantitative bDNA with Reflex to Genotype 2002681
Method: Quantitative Branched Chain DNA/Sequencing

Preferred test is HCV quantitative PCR following a positive HCV screen

Liver Fibrosis, Chronic Viral Hepatitis (Echosens FibroMeter) 2005661
Method: Quantitative Nephelometry/Quantitative Enzymatic/Quantitative Spectrophotometry/Automated Cell Count/ Electromagnetic Mechanical Clot Detection

Not a substitute for liver biopsy – may be considered for the following

  • Assess liver status in individuals at increased risk of complications from liver biopsy
  • Determine liver status before beginning hepatitis C virus (HCV) treatment
  • Assess liver status after completion of HCV treatment

Surrogate marker of liver fibrosis, cirrhosis, and necroinflammatory activity

Proprietary algorithm calculates and compares results from 7 blood markers along with age and gender to provide a patient score and a correlated fibrosis stage and activity grade

Blood markers – alpha-2-macroglobulin, alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen (BUN), gamma glutamyl transferase (GGT), platelets, and prothrombin index

Hepatitis C Virus High-Resolution Genotype by Sequencing 2006898
Method: Polymerase Chain Reaction/Sequencing

Order before initiating HCV therapy to aid in determining therapy of choice, likelihood of response, and probable therapeutic duration

Use for prognosis and treatment selection when a higher level of subtype resolution is required (eg, non 6a/b vs. type 1 and type 1a vs. 1b)

Do not order prior to molecular confirmation of positive HCV screen

Test may be unsuccessful if HCV RNA viral load <log 3.6 or 4,000 IU/mL

Hepatitis C Virus (HCV) NS3/4A Protease Inhibitor Resistance, GenoSure 2010647
Method: Polymerase Chain Reaction/Sequencing

Recommended testing for HCV genotype 1 patients prior to initiating simeprevir therapy