Diagnostic performance of hepatitis C core antigen assay to identify active infections: A systematic review and meta-analysis.

Hepatitis C virus (HCV) core antigen (HCVcAg) assay is an alternative for diagnosing HCV infection in a single step. This meta-analysis aimed to evaluate the Abbott ARCHITECT HCV Ag assay's diagnostic performance (validity and utility) for diagnosing active hepatitis C. PubMed, EMBASE, Scopus, Web of Science, and Cochrane Library were searched until 10 January 2023. The protocol was registered at the prospective international register of systematic reviews (PROSPERO: CRD42022337191). Abbott ARCHITECT HCV Ag assay was the test for evaluation, and nucleic acid amplification tests with a cut-off ≤50 IU/mL were the gold standard. Statistical analysis was performed using STATA with the MIDAS module and random-effects models. The bivariate analysis was conducted on 46 studies (18,116 samples). The pooled sensitivity was 0.96 (95% CI = 0.94-0.97), specificity 0.99 (95% CI = 0.99-1.00), positive likelihood ratio 141.81 (95% CI = 72.39-277.79), and negative likelihood ratio 0.04 (95% CI = 0.03-0.06). The area under the summary receiver operating characteristic curve was 1.00 (95% CI = 0.34-1.00). For active hepatitis C prevalence values of 0.1%-15%, the probability that a positive test was a true positive was 12%-96%, respectively, indicating that a confirmatory test should be necessary, particularly with a prevalence ≤5%. However, the probability that a negative test was a false negative was close to zero, indicating the absence of HCV infection. The validity (accuracy) of the Abbott ARCHITECT HCV Ag assay for screening active HCV infection in serum/plasma samples was excellent. Although the HCVcAg assay showed limited diagnostic utility in low prevalence settings (≤1%), it might help diagnose hepatitis C in high prevalence scenarios (≥5%).

The Transgene Expression of the Immature Form of the HCV Core Protein (C191) and the LncRNA MEG3 Increases Apoptosis in HepG2 Cells.

Long non-coding RNAs (lncRNAs) are regulated in cancer cells, including lncRNA MEG3, which is downregulated in Hepatocellular Carcinoma (HCC). In addition, hepatitis C virus (HCV) core proteins are known to dysregulate important cellular pathways that are linked to HCC development. In this study, we were interested in evaluating the overexpression of lncRNA MEG3, either alone or in combination with two forms of HCV core protein (C173 and C191) in HepG2 cells. Cell viability was assessed by MTT assay. Transcripts' levels of key genes known to be regulated in HCC, such as p53, DNMT1, miRNA152, TGF-b, and BCL-2, were measured by qRT-PCR. Protein expression levels of caspase-3 and MKI67 were determined by immunocytochemistry and apoptosis assays. The co-expression of lncRNA MEG3 and C191 resulted in a marked increase and accumulation of dead cells and a reduction in cell viability. In addition, a marked increase in the expression of tumor suppressor genes (p53 and miRNA152), as well as a marked decrease in the expression of oncogenes (DNMT1, BCL2, and TGF-b), were detected. Moreover, apoptosis assay results revealed a significant increase in total apoptosis (early and late). Finally, immunocytochemistry results detected a significant increase in apoptotic marker caspase-3 and a decrease in tumor marker MKI67. In this study, transgene expression of C191 and lncRNA MEG3 showed induction in apoptosis in HepG2 cells greater than the expression of each one alone. These results suggest potential anticancer characteristics.

Critical role of triglycerides for adiponectin levels in hepatitis C: a joint study of human and HCV core transgenic mice.

BACKGROUND: Both hepatitis C virus (HCV) infection and adiponectin are critically involved in metabolism. The reversal and associations of altering adiponectin levels after sustained virological responses (SVRs) following direct-acting antivirals (DAA) in HCV-infected patients remained elusive. METHODS: A joint study was conducted in a prospective cohort of 427 HCV-infected patients and a line of HCV core transgenic mice. RESULTS: Of 427, 358 had completed a course of DAA therapy and 353 had SVRs. At baseline, male sex (95% CI ß: - 1.44 to - 0.417), estimated glomerular filtration rate (eGFR) (- 0.025 to - 0.008), triglycerides (- 0.015 to - 0.005), and fibrosis-4 levels (0.08-0.297) were associated with adiponectin levels; BMI (0.029-0.327) and triglycerides levels (0.01-0.03) were associated with homeostatic model assessment for insulin resistance (HOMA-IR) in HCV-infected patients. At 24-week post-therapy, in SVR patients, male sex (- 1.89 to - 0.5) and eGFR (- 0.02 to - 0.001) levels were associated with adiponectin levels, levels of BMI (0.094-0.335) and alanine transaminase (0.018-0.078) were associated with HOMA-IR; compared with baseline levels, adiponectin levels decreased (6.53 ± 2.77 vs. 5.45 ± 2.56 µg/mL, p < 0.001). In 12-month-old HCV core transgenic mice with hepatic steatosis, triglyceride levels (0.021-0.111) were associated with adiponectin levels, and hepatic adipopnectin expression was comparable with that of control mice. CONCLUSIONS: Triglycerides and hepatic fibrosis are associated with HCV-specific alteration of adiponectin levels, and adiponectin may affect insulin sensitivity through triglycerides during HCV infection. In DAA-treated patients, after SVR, adiponectin levels decreased and the linking function of triglycerides between adiponectin and insulin sensitivity vanished. Moreover, HCV core with hepatic steatosis might affect extrahepatic adiponectin expression through triglycerides.

Hepatitis C virus core antigen as alternative diagnostic algorithm for active hepatitis C virus infection among haemodialysis population: Cost implications.

AIMS: In Malaysia, majority anti-HCV positive haemodialysis patients do not undergo hepatitis C confirmation due to the high cost of HCV RNA. HCV Core Antigen might be a cost-effective diagnostic test to identify HD patients who have active HCV infection eligible for Direct Acting Anti-viral therapy. METHODS: A cross-sectional study was conducted to assess the correlation between HCV Ag and HCV RNA and the cost implications of different diagnostic algorithms to diagnose active HCV infection using Anti-HCV, HCV Ag, and HCV RNA. Pre-dialysis blood was tested for both HCV Ag and HCV RNA. HCV Ag was tested with Abbott ARCHITECT HCV Ag test. RESULTS: Two-hundred twenty-seven haemodialysis patients were recruited from 20 centres with mean age of 57.68 ± 12.48 years, and male constitutes 56.8% (129) of the study population. HCV Ag correlated well with HCV RNA (Spearman test coefficient 0.943, p < .001) with sensitivity of 93.9%, specificity 99.3%, and the accuracy was 97.36%. Cost analysis indicated that a sequential test involving Anti-HCV antibody as initial screening, followed by HCV Ag on Anti-HCV positive and HCV RNA on HCV Ag negative cases translated to a modest cost-saving algorithm compared to standard diagnostic algorithm. CONCLUSION: HCV Ag correlated well with HCV RNA and can potentially be fused in an alternative diagnostic algorithm to generate cost savings methods to diagnose active HCV infection among haemodialysis patients. This alternative algorithm is especially relevant in low to middle-income countries such as Malaysia to optimize the use of the healthcare resource and gains in clinical outcomes.

Ultrasensitive electrochemical detection of hepatitis C virus core antigen using terminal deoxynucleotidyl transferase amplification coupled with DNA nanowires.

Early diagnosis of hepatitis C virus (HCV) infection is essential to prevent disease from spreading and progression. Herein, a novel electrochemical biosensor was developed for ultrasensitive detection of HCV core antigen (HCVcAg) based on terminal deoxynucleotidyl transferase (TdT) amplification and DNA nanowires (DNW). After sandwich-type antibody-antigen recognition, the antibody-conjugated DNA was pulled to the electrode surface and further extended into a long DNA sequence by robust TdT reaction. Then, large numbers of methylene blue-loaded DNW (MB@DNW) as signal labels are linked to the extended DNA sequence. This results in an amplified electrochemical signal for HCVcAg determination, typically measured at around -0.25 V (Ag/AgCl). Under the optimum conditions, the proposed biosensor achieved a wide linear range for HCVcAg from 0.1 to 312.5 pg/mL with a low limit of detection of 32 fg/mL. The good practicality of the biosensor was demonstrated by recovery experiment (recoveries from 98 to 104% with RSD of 2.5-4.4%) and comparison with enzyme-linked immunosorbent assay (ELISA). Given the highlighted performance, the biosensor is expected to act as a reliable sensing tool for HCVcAg determination in clinics. Schematic representation of the ultrasensitive electrochemical biosensor based on terminal deoxynucleotidyl transferase (TdT) amplification linked with methylene blue-loaded DNA nanowires (MB@DNW), which can be applied to the determination of hepatitis C virus core antigen (HCVcAg) in clinical samples. dTTPs, 2'-deoxythymidine 5'-triphosphate.

Hepatitis C Virus RNA Levels Following Virologic Failure With Direct-acting Antivirals: Implications for Lower Sensitivity Diagnostic Assays.

We analyzed post-treatment hepatitis C virus (HCV) RNA levels from 330 subjects who experienced virologic failure in clinical trials of direct-acting antivirals. We demonstrated that 97% had post-treatment Week 12 HCV RNA >10 000 IU/mL, above reported sensitivity limits of novel diagnostic assays being considered for simplified HCV treatment monitoring.

Hepatitis C core antigen testing to diagnose active hepatitis C infection among haemodialysis patients.

BACKGROUND: Hepatitis C virus (HCV) infects more than 71 million people worldwide and chronic HCV infection increases the risk of liver cirrhosis and failure. Haemodialysis (HD) is one of the renal replacement therapies with risk of HCV transmission. Anti-HCV antibodies are the serological screening test for HCV infection that does not detect active phase of infection. Majority HCV infected HD patients in Malaysia do not have further HCV RNA performed due to high cost and thus HCV treatment is less frequently offered. HCV Core Antigen (HCV Ag) can potentially be used to diagnose active HCV infection in HD population in comparison to HCV RNA, at lower cost. METHODS: We conducted a cross-sectional study to assess the correlation between HCV Ag and HCV RNA and to identify the prevalence of active HCV infection among HCV seropositive HD patients from dialysis centres across West Malaysia from July 2019 to May 2020. Pre-dialysis blood was taken and tested for both HCV Ag and HCV RNA tests. HCV Ag was tested with Abbott ARCHITECT HCV Ag test. RESULTS: We recruited 112 seropositive HD patients from 17 centres with mean age of 54.04 ± 11.62 years, HD vintage of 14.1 ± 9.7 years, and male constitute 59.8% (67) of the study population. HCV Ag correlates well with HCV RNA (Spearman test coefficient 0.833, p < 0.001). The sensitivity was 90.7%, specificity 100%, positive predictive value (PPV) 100%, negative predictive value (NPV) 76.5%, and accuracy 92.9%. For HCV RNA level > 3000 IU/mL, HCV Ag had a higher sensitivity of 95.1% and greater correlation (Spearman test coefficient 0.897, p < 0.001). The prevalence of active HCV infection was 76.8% among HCV seropositive HD patients. CONCLUSIONS: Although HCV Ag is less sensitive, it shows an excellent correlation with HCV RNA and has 100% PPV. HCV Ag can be considered as an alternative diagnostic tool for chronic active HCV infection among HD cohort, who can then be considered for HCV treatment. For seropositive HD patient with negative HCV Ag, we recommend to follow-up with HCV RNA test.

N-terminal HCV core protein fragment decreases 20S proteasome activity in the presence of PA28γ.

The Hepatitis C virus (HCV) core protein plays a crucial role in the development of chronic liver diseases such as chronic hepatitis, cirrhosis, and hepatocellular carcinoma (HCC). Its involvement in these diseases is reportedly abolished by a knockout of the proteasome activator PA28γ gene in transgenic mice, suggesting an interaction between the core protein and the PA28γ-proteasome system. This study found a direct interaction between the N-terminal 1-71 fragment of HCV core protein (Core71) and PA28γ in vitro, and that this interaction was found to enhance PA28γ-20S proteasome complex formation. While 20S proteasome activity was increased by PA28γ, it was significantly reduced by Core71 attachment in a dose-dependent manner. These results suggest that the Core-PA28γ interaction has an important role in regulating 20S proteasome activity and furthers our understanding of the pathogenesis of HCV.

Evaluation of a hepatitis C virus core antigen assay from venepuncture and dried blood spot collected samples: A cohort study.

The global scale-up of hepatitis C virus (HCV) diagnosis requires simplified and affordable HCV diagnostic pathways. This study evaluated the sensitivity and specificity of the HCV Architect core antigen (HCVcAg) assay for detection of active HCV infection in plasma and capillary whole blood dried blood spots (DBS) compared with HCV RNA testing in plasma (Abbott RealTime HCV Viral Load). Samples were collected from participants in an observational cohort enrolled at three sites in Australia (two-drug treatment and alcohol clinics and one homelessness service). Of 205 participants, 200 had results across all samples and assay types and 186 were included in this analysis (14 participants receiving HCV therapy were excluded). HCV RNA was detected in 29% of participants ([95% CI: 22.6-36.1], 54 of 186). The sensitivity of HCVcAg for detection of active HCV infection in plasma was 98.1% (95% CI: 90-100) and 100% (95% CI: 93-100) when compared to HCV RNA thresholds of ≥12 and ≥1000 IU/mL, respectively. The sensitivity of the HCVcAg assay for detection of active HCV infection in DBS was 90.7% (95% CI: 80-97) and 92.5% (95% CI: 82-98) when compared to HCV RNA thresholds of ≥12 and ≥1000 IU/mL, respectively. The specificity of HCV core antigen for detection of active infection was 100% (95% CI: 97-100) for all samples and RNA thresholds. These data indicate that the detection of HCVcAg is a useful tool for determining active HCV infection; to facilitate enhanced testing, linkage to care and treatment particularly when testing plasma samples are collected by venepuncture.

Anti-viral activity of compounds from Agrimonia pilosa and Galla rhois extract mixture.

Hepatitis C virus (HCV) infection is a significant health problem, with a worldwide prevalence of about 170 million. Recently, the development of direct acting antiviral (DAA) as a therapeutic agent for HCV has been rapidly increasing. However, DAA has a side effect and is costly. Therefore, it is still necessary to develop a therapeutic agent to treat HCV infection using products. Agrimonia pilosa (AP) and Galla rhois (RG) are traditional medicines and are known to display therapeutic activity on various diseases. Notably, they have been reported to have an anti-viral effect on HBV and influenza virus infections. It is expected that anti-viral activity will increase when two extracts are mixed. To investigate their anti-viral activity, the expression level of HCV Core 1b and NS5A was measured. Remarkably, AP, RG, and their mixed compound (APRG64) strongly inhibited the expression of viral proteins, which led us to identify their metabolites. A total of 14 metabolites were identified using liquid chromatography mass spectrometry (LC-MS). These metabolites were evaluated for their anti-HCV activity to identify active ingredients. In conclusion, our results unveiled that anti-HCV activity of Agrimonia pilosa and Galla rhois extract mixture could lead to the development of a novel therapy for HCV infection.

The correlation of HCV RNA and HCV core antigen in different genotypes of HCV.

BACKGROUND: To analyze the correlation of HCV RNA and HCV core antigen (HCV cAg) in different genotypes of HCV. METHODS: One hundred and six patients who were diagnosed with HCV infection by HCV RNA test were included in the study. HCV genotypes were detected by PCR fluorescent probe. Detected HCV cAg's expression in serum quantitatively and qualitatively with chemiluminescent micro-particle immuno assay (CMIA) and enzyme-linked immunosorbent assay (ELISA), respectively, and compared positive rates. Analyzed the correlation of HCV RNA and HCV cAg in different genotypes. RESULTS: Distribution of HCV genotypes in 106 HCV infected patients were as follows: 1b genotype 46 (43.4%); 2a genotype 7 (6.6%); 3a genotype 18 (17.0%); 3b genotype 3 (2.8%); 6a genotype 9 (8.5%); 1b/3b mixed type 13 (12.3%); and unidentified type 10 (9.4%). Positive rates of HCV cAg detected by CMIA and ELISA were 100% and 56%, respectively, with statistical significance (χ2  = 60.38, P = 0.000). HCV cAg in 1b genotype group was higher than that in 3b and 1b/3b genotype groups, with statistical significance (U = 3.0, P = 0.006, U = 165, P = 0.014). HCV RNA and HCV cAg in genotype 1b demonstrated a positive correlation (r = 0.894, P = 0.04). CONCLUSION: Major genetic subtype of HCV genotype was 1b. Compared with ELISA, detection of HCV cAg by CMIA increased the positive rate and facilitated early diagnosis and treatment of HCV-infected patients. With the increase in HCV RNA load and the expression of HCV cAg, HCV cAg could be an early indicator for the diagnosis of HCV infection in 1b genotype.

Developments in the HCV Screening Technologies Based on the Detection of Antigens and Antibodies.

Hepatitis C virus (HCV) accounts for 15%-20% of cases of acute infection, and chronic HCV infection is developed in about 50%-80% of HCV patients. Unfortunately, due to the lack of proper medical care, difficulty in screening for HCV infection, and lack of awareness resulted in chronic HCV infection in 71 million people on a global scale, and about 399,000 deaths in 2016. It is crucial to recognize that the effective use of antiviral medicines can cure more than 95% of HCV infected people. The Global Health Sector Strategy (GHSS) aim is to reduce the new HCV infections and the HCV associated mortality by 90% and 65%, respectively. Therefore, the methods that are simple, yet powerful enough to detect HCV infections with high sensitivity, specificity, and a shorter window period are crucial to restrain the global burden of HCV healthcare. This article focuses on the technologies used for the detection of HCV in clinical specimens.

Confirmation of HCV viremia using HCV RNA and core antigen testing on dried blood spot in HIV infected peoples who inject drugs in Vietnam.

BACKGROUND: Nucleic acid tests performed on blood samples collected on Dried Blood Spot (DBS) and detection of HCV core antigen (HCVcAg) are two approaches that may facilitate access to HCV diagnosis in low and middle incomes countries. In this study we evaluate HCV RNA and HCV antigen testing on DBS in HIV/HCV co-infected peoples who inject drugs in Vietnam. METHOD: One hundred and four HIV/HCV seropositive patients managed in outpatient care at the Haiphong Viet Tiep hospital were included in this study from February to March, 2014 (ANRS 12262 study). RESULTS: Eighty-six subjects were tested positive for HCV RNA in serum, median (IQR): 6.9 log10 IU/ml (5.6-7.4 log10 IU/ml). Genotypes consisted of 57 G1 (69%), 3 G3 (4%), and 22 G6 (27%). HCV RNA was detected on DBS specimens in 79 out 86 subjects with chronic hepatitis C (sensitivity 92.5%; 95% CI: 85.1-96.9%). HCV RNA level on DBS and serum was moderately correlated (r = 0.24; p = 0.05) suggesting a degradation of HCV RNA due to transportation and storage conditions. HCVcAg was detected in 75/86 dB specimens (sensitivity: 87.2%; 95% CI: 78.3-93.4%), with a strong positive relationship between DBS HCVcAg and serum HCV RNA levels (r = 0.80; P < 0.0001). CONCLUSIONS: Quantification of HCVcAg on DBS appears to benefit from substantial stability under prolonged storage conditions but with a lower analytical sensitivity compared to DBS HCV RNA testing. Detection of HCV RNA on DBS is an interesting approach for confirming viral replication in HCV seropositive persons but the impact of pre-analytical conditions on the integrity of HCV RNA needs to be controlled.

HCV core antigen comes of age: a new opportunity for the diagnosis of hepatitis C virus infection.

The diagnosis of hepatitis C virus (HCV) infection has been traditionally based on the detection of the host antibody response. Although antibody assays are available in different formats and are fairly accurate, they cannot distinguish between an ongoing infection with HCV replicative activity and a past infection where HCV has been cleared, spontaneously or after a successful therapy. As a chronic infection is mostly asymptomatic until the late clinical stages, there is a compelling need to detect active HCV infection by simple and reproducible methods. On this purpose, the clinical guidelines have suggested to search for the HCV ribonucleic acid (HCV-RNA) after anti-HCV has been detected, but this second step carries several limitations especially for population screening. The availability of fast and automated serological assays for the hepatitis C core antigen (HCVAg) has prompted an update of the guidelines that now encompass the use of HCVAg as a practical alternative to HCV-RNA, both for screening and monitoring purposes. In this paper, we summarize the features, benefits and limitations of HCVAg testing and provide an updated compendium of the evidences on its clinical utility and on the indications for use.

Persistence of Circulating Hepatitis C Virus Antigens-Specific Immune Complexes in Patients with Resolved HCV Infection.

BACKGROUND: Our recent study indicated the possible presence of detectable hepatitis C virus antigens (HCV-Ags) after denaturation of sera with resolved HCV (R-HCV) infection. The present study determined and characterized persistent HCV-Ags-specific immune complexes (ICs) in these patients. METHODS: Sixty-eight sera with R-HCV and 34 with viremic HCV (V-HCV) infection were tested for free and IC-bound HCV-Ags using HCV-Ags enzyme immunoassay (EIA), the presence of HCV-Ags-specific ICs by immunoprecipitation and Western blot (IP-WB), HCV ICs containing HCV virions using IP and HCV RNA RT-PCR, and correlation of HCV ICs with clinical presentation in these patients. RESULTS: Using HCV-Ags EIA, we found 57.4% of sera with R-HCV infection were tested positive for bound, but not free HCV-Ags. Using pooled or individual anti-HCV E1/E2, cAg, NS3, NS4b, and/or NS5a to precipitate HCV-specific-Ags, we confirmed persistent HCV-Ags ICs specific to various HCV structural and non-structural proteins not only in V-HCV infection, but also in R-HCV infection. Using IP and HCV RNA PCR, we then confirmed the presence of HCV virions within circulating ICs in V-HCV, but not in R-HCV sera. Multivariable analysis indicated significant and independent associations of persistent circulating HCV-Ags-specific ICs with both age and the presence of cirrhosis in patients with R-HCV infection. CONCLUSIONS: Various HCV-Ag-specific ICs, but not virions, persist in 57.4% of patients who had spontaneous or treatment-induced HCV clearance for 6 months to 20 years. These findings enriched our knowledge on HCV pathogenesis and support further study on its long-term clinical relevance, such as extrahepatic manifestation, transfusion medicine, and hepatocarcinogenesis.

Role of Hepatitis C Virus (HCV) core antigen in improving blood transfusion safety in high prevalence, resource limited countries, a step forward.

OBJECTIVE: The aim of the present study was to evaluate the efficacy of hepatitis C virus [HCV] core Ag as an alternative affordable test in resource limited countries blood banks. BACKGROUND: Implementing nucleic acid testing in developing countries with low resources is still unaffordable. Egypt has the highest prevalence of hepatitis C in the world and still in need to efficient affordable transfusion program that reduces the window period for the virus before implementing the complex high cost NAT. STUDY DESIGN AND METHODS: HCV core Ag by ELISA in serum, in the presence or absence of anti-HCV antibodies was compared to HCV- RNA by PCR on total number of 1850 first time and repeat donations from Fayoum University Hospital and Badr University Hospital. RESULTS: Among 1850 healthy voluntary donors, 143 donors with anti-HCV antibody positivity, 105 were determined as positive, 38 were negative for HCV core Ag, and 107 were positive for HCV RNA. CONCLUSION: Hepatitis C virus core antigen-ELISA can be a useful alternative in the developing nations and Greater consideration should be given to its implementation as an additional serological test for blood donors in Egypt as the most cost-effective measure for further improvement of transfusion safety.

Ultra-sensitive aptasensor based on a GQD nanocomposite for detection of hepatitis C virus core antigen.

In the present study, by using the aptamer proximity binding assay strategy, a novel electrochemical aptasensor is described for ultrasensitive detection of hepatitis C virus (HCV) core antigen. The immobilization surface is prepared by the modification of a glassy carbon electrode (GCE) with a graphene quantum dots (GQD). GQD were introduced as a novel and suitable substrate for aptamers through π-π stacking interactions, the richness of hydrophilic edges as well as hydrophobic plane in GQD which enhances the aptamer absorption on the electrode surface. Electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) and differential pulse voltammetry (DPV) were performed at each stage of the chemical modification process to confirm the resulting surface changes. EIS technique was used as an efficient alternative detection system for HCV core antigen measurement with detection limit 3.3 pg mL-1 and two linear concentration range 10-70 pg mL-1 and 70-400 pg mL-1. Moreover, the fabricated aptasensor could accurately detect HCV core antigen concentration in human serum samples. Such an aptasensor opens a rapid, selective and sensitive route for HCV core antigen detection and provides a promising strategy for potential applications in clinical diagnostics.

Performance comparison of new generation HCV core antigen test versus HCV RNA test in management of hepatitis C virus infection.

The study has evaluated the performance of HCV core antigen (Cag) test by comparing HCV RNA PCR assay which is considered the gold standard for management of HCV infection. Totally, 132 samples sent for HCV RNA (real-time PCR) test were included in the study. Anti-HCV antibody test and HCV Cag test were performed by chemiluminescent enzyme immunoassay (CMEI). Anti-HCV test was positive in all samples. HCV RNA was detected in 112/132 (84.8%) samples, and HCV Cag in 105/132 (79.5%). The most common HCV genotype was genotype 1 (86%). Considering the HCV RNA test as gold standard; the sensitivity, specificity, positive predictive value, negative predictive value and accuracy of Cag test were found to be 93.75%, 100%, 100%, 74.07% and 94.69%, respectively, and paired test results were detected as highly concordant. A high level of correlation was seen between HCV RNA and Cag tests, however, the concordance between the two tests appeared to be disrupted at viral loads lower than 103IU/mL. On the contrary, the correlation reached significance for the values higher than 103IU/mL. Viral loads were in the 17-2500IU/mL range for the negative results for Cag test. Pearson's correlation coefficient revealed a considerably high correlation. The concordance between HCV RNA and Cag tests was disrupted under a viral load lower than 103IU/mL. Therefore, it would be appropriate to consider cost effectiveness, advantages and limitations of the HCV RNA and Cag tests during the decision on which method to use for patient management.

Comparison of Three Different Hepatitis C Virus Genotyping Methods: 5'NCR PCR-RFLP, Core Type-Specific PCR, and NS5b Sequencing in a Tertiary Care Hospital in South India.

BACKGROUND: Based on genetic heterogeneity, hepatitis C virus (HCV) is classified into seven major genotypes and 64 subtypes. In spite of the sequence heterogeneity, all genotypes share an identical complement of colinear genes within the large open reading frame. The genetic interrelationships between these genes are consistent among genotypes. Due to this property, complete sequencing of the HCV genome is not required. HCV genotypes along with subtypes are critical for planning antiviral therapy. Certain genotypes are also associated with higher progression to liver cirrhosis. METHODS: In this study, 100 blood samples were collected from individuals who came for routine HCV genotype identification. These samples were used for the comparison of two different genotyping methods (5'NCR PCR-RFLP and HCV core type-specific PCR) with NS5b sequencing. RESULTS: Of the 100 samples genotyped using 5'NCR PCR-RFLP and HCV core type-specific PCR, 90% (κ = 0.913, P < 0.00) and 96% (κ = 0.794, P < 0.00) correlated with NS5b sequencing, respectively. Sixty percent and 75% of discordant samples by 5'NCR PCR-RFLP and HCV core type-specific PCR, respectively, belonged to genotype 6. All the HCV genotype 1 subtypes were classified accurately by both the methods. CONCLUSION: This study shows that the 5'NCR-based PCR-RFLP and the HCV core type-specific PCR-based assays correctly identified HCV genotypes except genotype 6 from this region. Direct sequencing of the HCV core region was able to identify all the genotype 6 from this region and serves as an alternative to NS5b sequencing.

Molecular characterization of hepatitis C virus core region in moroccan intravenous drug users.

Intravenous drug users (IDUs) represent a highly-infected reservoir for Hepatitis C virus (HCV) worldwide, harboring some of the most elevated prevalences and majority of the epidemic in developed nations. Studies aimed at sequencing regions of the viral genome uncovered amino acid mutations, some of which have been implicated in resistance to standard of care pegylated interferon/Ribavirin double therapy. Using the nested PCR method on the Core region of HCV strains in Moroccan IDUs living in the Tangier region this study sought to identify genotype-specific amino acid mutations, followed by Phylogenetic methods in order to compare them with international strains so as to identify sequences of highest homology. Genotyping was confirmed and recombination events excluded by line-probe assay. Italy was found most homologous for genotypes 1a and 3a, Iran for genotype 1a and Egypt for genotype 4a. Amino Acid Mutation analysis revealed the following novel genotype 3a-specific mutations: N16I, L36V, T49A, P71S, T75S, and T110N. The outcome of this work describes the HCV genetic heterogeneity in high-risk intravenous drug users, and it gives clues to the global migratory flow of genotypes as they cross geographical boundaries between various IDU populations and identifies "signature" amino acid mutations traceable to HCV genotype 3a. Identification of key amino acid positions in the HCV Core region with higher rates of mutations paves the way for eventual clinical trials seeking to establish a link between these recurrent mutations and response to standard of care Interferon and Ribavirin antiviral therapy. J. Med. Virol. 88:1376-1383, 2016. © 2016 Wiley Periodicals, Inc.