Review
Copyright ©The Author(s) 2015.
World J Virology. Nov 12, 2015; 4(4): 323-342
Published online Nov 12, 2015. doi: 10.5501/wjv.v4.i4.323
Table 4 Main characteristics of hepatitis B virus and hepatitis C molecular techniques
HBV and HCV molecular diagnosisApplicationMethodAdvantagesDisadvantagesRef.
HBV DNA qualitative methodsDiagnose infectionPCRLow cost; high sensitivityIt only determines the presence or absence of HBV DNA[81,82,85]
HBV occult cases identification
Screening on blood donors
HBV DNA quantitative methodsEvaluate the prognosis and risk of progressionbDNAWide dynamic rangeLow sensitivity to detect low HBV DNA levels[81,82]
Define the beginning of antiviral treatment
Monitor antiviral treatment
Hybrid captureMore sensitive than bDNA; less labor-intensiveLow sensitivity to detect low HBV DNA levels; individual probes are required[83]
Real time PCRCapacity of detecting low viral loads; broad dynamic range; do not carry over contamination; can be fully automatedHigh cost[85,91]
HBV DNA genotyping methodsDetermination of HBV genotypeRFLPEasily done; low cost; simple, rapid and suitable for large number of samplesLow sensitivity for typing samples with low HBV DNA levels; poor accurate to determine some genotypes[85,104]
Genotype specific PCR assaysAutomated systems; high sensitivity; easy to perform; suitable for detecting mixed genotype infectionsHigh cost[107]
Sequence analysisIdentification of patients infected with recombinant genotypesTechnically demanded; time consuming[107]
HBV DNA aminoacid substitution identificationIdentify antiviral resistance to treatmentDirect DNA sequencingAccurateTechnically demanded; time consuming; necessity of cloning for identification of mixed population[107,110,111]
Commercial methodsSequencing of mixed population, relative quantification of individual mutations with extremely high coverageDifferences between the statistical and biological/clinical relevance of HBV mutation maximal sequence read length and PCR amplification bias[114]
HCV RNA qualitative methodsTo confirm chronic hepatitis C in patients with positive HCV antibodiesRT-PCRHigh sensitivityIt only determines the presence or absence of HCV RNA[121,168]
To identify virological response during, at the end or after antiviral therapyEqual sensitivity for all genotypes
To screen blood donations for evidence of infection with HCV
Transcription-mediated amplificationHigh sensitivity; amplifies viral RNA; more sensitivity for detection of genotype 1It only determines the presence or absence of HCV RNA[121,168]
HCV RNA quantitative methodsTo guide treatment decisions; To evaluate the prognosis; To monitor the antiviral efficacy of treatmentbDNAWide range of detection of HCV independent of HCV genotype (615 IU/mL to 8 million IU/mL)Low sensitivity to detect samples presenting low HCV RNA levels[172]
qRT-PCRCapacity of detecting low viral loads; broad dynamic range; not carry over contamination; can be fully automatedHigh cost[170-173]
HCV RNA genotyping methodsHCV genotyping is mandatory for double antiviral treatment (interferon and ribavirin), since patients infected with genotypes 1 or 4 are treated for longer times than patients infected by genotypes 2 and 3RFLPEasily done; low cost; simple, rapid and suitable for large number of samplesLow sensitivity for typing samples with low HCV RNA levels; Poor accurate to determine some genotypes[186,187]
ProbesEasily done; low cost; useful to detect HCV genotypes and subtypes based on region 5’UTR and core and has a low limit of detectionIdentify only subtypes 1a and 1b; discrepant results among subtypes when compared to sequence analysis of NS5B region[180-184]
qPCRCan be fully automated avoiding contamination; determines the viral genotype and subtypes 1a, 1b, 2a, 2b, 3, 4, 5 and 6High cost[186,187]
Direct sequencingGold standard; identification of patients infected with recombinant genotypesTechnically demanded; time consuming[120,182]
HCV RNA aminoacid substitution identificationIdentify antiviral resistance to treatmentDirect SequencingIdentification of antiviral resistance in majority populationTechnically demanded; time consuming; necessity of cloning for identification of quasispecies[188,189,193]
Deep SequencingIdentification on resistant variants predominate in the HCV population; powerful tool for obtaining more profound insight into the dynamics of variants in the HCV quasispeciesNeed for in-depth knowledge to analyze the results[112,113,195,196]