Distribution of Hepatitis C Virus (HCV) RNA in Whole Blood and Blood Cell Fractions: Plasma HCV RNA Analysis Underestimates Circulating Virus Load

ABSTRACT In this study, we established a fully automated molecular assay for qualitative detection of hepatitis C virus (HCV) in serum and whole-blood samples and compared it with conventional molecular assays, including manual HCV RNA extraction protocols. Whole-blood samples were collected from patients with and without chronic HCV infection in EDTA tubes and nucleic acid stabilization tubes (NASTs). Prior to HCV RNA extraction, the HCV Internal Control (IC), derived from the COBAS AMPLICOR HCV test, version 2.0 (Roche Molecular Diagnostics), was added. The new assay was based on an automated extraction protocol on the MagNA Pure LC instrument (Roche Applied Science), followed by automated reverse transcription, amplification, hybridization, and detection on the Cobas Amplicor analyzer (Roche Molecular Diagnostics). The detection limit of the new assay was found to be similar to those of conventional molecular assays. In clinical samples, 100% agreement between the new assay and conventional methods was observed. The introduced amount of IC was detected in 45 of 45 serum samples, 41 of 45 EDTA tube whole-blood samples, and 43 of 45 NAST whole-blood samples. Retesting led to more frequent IC detection. The fully automated molecular assay was found to be suitable for detection of HCV RNA in different kinds of sample materials. It may be recommended for use in the high-throughput routine molecular diagnostic laboratory.

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Vertical transmission of hepatitis C virus (HCV) detected by HCV-RNA analysis.

Gut ◽

10.1136/gut.34.2_suppl.s52 ◽

1993 ◽

Vol 34 (2 Suppl) ◽

pp. S52-S53 ◽

Cited By ~ 14

Author(s):

T Kuroki ◽

S Nishiguchi ◽

K Fukuda ◽

N Ikeoka ◽

R Murata ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Vertical Transmission ◽

Hcv Rna ◽

Rna Analysis

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Assessing Molecular Point-of-Care Testing and Dried Blood Spot for Hepatitis C Virus Screening in People Who Inject Drugs

Open Forum Infectious Diseases ◽

10.1093/ofid/ofaa196 ◽

2020 ◽

Vol 7 (6) ◽

Cited By ~ 2

Author(s):

Stéphane Chevaliez ◽

Mélanie Wlassow ◽

Johann Volant ◽

Françoise Roudot-Thoraval ◽

Antoine Bachelard ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Whole Blood ◽

People Who Inject Drugs ◽

Point Of Care ◽

Dried Blood Spot ◽

World Health ◽

Hcv Rna ◽

Rna Detection ◽

Blood Spot

Abstract Background Injecting drug use is a major driver of hepatitis C virus (HCV) spread worldwide, and the World Health Organization (WHO) has identified people who inject drugs (PWID) as a key population to target for HCV screening and care. Point-of-care (POC) hepatitis C tests and dried blood spot (DBS) sampling offer benefits for the management of patients with HCV infection by increasing HCV testing and linkage to care in different nonclinical settings. The aims of this prospective study were to evaluate the feasibility and the acceptability of use HCV ribonucleic acid (RNA) POC and fingerstick DBS testing in social-medical risk-reduction centers and to describe the cascade of care among PWID in France. Methods Between June 2018 and February 2019, 89 consecutive HCV-seropositive PWID attending 2 drug treatment services and 1 supervised consumption room in inner Paris were invited to participate in further evaluation, undergoing a clinical review with a liver assessment and blood tests including fingerstick capillary whole blood POC HCV RNA testing and fingerstick DBS sampling. Results Of the 89 participants enrolled, HCV RNA was detected in 34 (38.6%) participants. Fingerstick whole blood POC RNA testing and HCV RNA detection from DBS sample were feasible and acceptable among PWID with no major difference in terms of HCV RNA detection rate. Overall, 16 participants received pan-genotypic antiviral treatment. The proportion of PWID with sustained virologic response at 12 weeks was 81.2%, with data for 3 patients still pending. Conclusions One-step screening strategy based on the detection of HCV RNA would engage people in care for treatment scale-up and HCV elimination.

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Genetic Clustering of Hepatitis C Virus Strains and Severity of Recurrent Hepatitis after Liver Transplantation

Journal of Virology ◽

10.1128/jvi.75.23.11292-11297.2001 ◽

2001 ◽

Vol 75 (23) ◽

pp. 11292-11297 ◽

Cited By ~ 24

Author(s):

Michèle Gigou ◽

Anne Marie Roque-Afonso ◽

Bruno Falissard ◽

François Penin ◽

Elisabeth Dussaix ◽

...

Keyword(s):

Liver Transplantation ◽

Hepatitis C Virus ◽

Hepatitis C ◽

Genetic Distance ◽

Correlation Method ◽

Genetic Distances ◽

Hcv Rna ◽

Genotype 1 ◽

Virus Load ◽

Recurrent Hepatitis

ABSTRACT The influence of viral factors on the severity of hepatitis C virus (HCV)-related liver disease is controversial. We studied 68 liver transplant patients with recurrent hepatitis C, of whom 53 were infected by genotype 1 strains. Relationships between core sequences, serum HCV RNA levels, and fibrosis scores for each patient were analyzed in pairwise fashion 5 years after transplantation. We used Mantel's test, a matrix correlation method, to evaluate the correspondence between measured genetic distances and observed phenotypic differences. No clear relationship was found when all 68 patients were analyzed. In contrast, when the 53 patients infected by genotype 1 strains were analyzed, a strong positive relationship was found between genetic distance and differences in 5-year fibrosis scores (P = 0.001) and differences in virus load (P = 0.009). In other words, the smaller the genetic distance between two patients' viral core sequences, the smaller the difference between the two patients' fibrosis scores and viral replication levels. No relationship was found between genetic distance and differences in age, sex, or immunosuppression. In multivariate analysis, the degree of fibrosis was negatively related to the virus load (r = −0.68; P = 0.003). In the particular setting of liver transplantation, and among strains with closely related phylogenetic backgrounds (genotype 1), this study points to a correlation between the HCV genetic sequence and the variability of disease expression.

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Hepatitis C virus (HCV) RNA quantitation in small-volume whole blood samples

Zeitschrift für Gastroenterologie ◽

10.1055/s-0029-1191927 ◽

2009 ◽

Vol 47 (01) ◽

Author(s):

T Bruns ◽

K Steinmetzer ◽

E Ermantraut ◽

A Stallmach

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Whole Blood ◽

Small Volume ◽

Hcv Rna ◽

Blood Samples ◽

Whole Blood Samples

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Evaluation of Nitric Oxide (NO) Levels in Hepatitis C Virus (HCV) Infection: Relationship to Schistosomiasis and Liver Cirrhosis among Egyptian Patients

Disease Markers ◽

10.1155/2002/647961 ◽

2002 ◽

Vol 18 (3) ◽

pp. 137-142 ◽

Cited By ~ 8

Author(s):

Mahmoud Ismail Hassan ◽

Samar Kamal Kassim ◽

Hebatalla Said Ali ◽

El-Dieb Abd ElSattar Sayed ◽

Ali Khalifa

Keyword(s):

Nitric Oxide ◽

Polymerase Chain Reaction ◽

Liver Cirrhosis ◽

Hepatitis C Virus ◽

Hepatitis C ◽

Hcv Rna ◽

Chain Reaction ◽

Virus Load ◽

Egyptian Patients ◽

Polymerase Chain

Nitric oxide (NO), a recently discovered free radical, is overproduced in liver cirrhosis. Hepatitis C virus (HCV) might increase NO levels via increased inducible NO synthase (iNOS). This work was carried out to study the effect of HCV-induced liver cirrhosis on NO levels among Egyptian patients. The study included 46 patients with liver cirrhosis, and 30 healthy individuals of matched age and sex. NO levels determined as the stable endproduct nitrate, showed a statistically significant increase among patients compared to the control group (P< 0.001). Furthermore, NO levels increased proportionally with the severity of liver cirrhosis as assessed by Child’s classification (P< 0.05). Moreover, schistosomial infection enhanced NO levels in cirrhotic patients with HCV infection compared to non-bilharzial patients (P< 0.001). Polymerase chain reaction (PCR) and branched DNA assays were used for detection of HCV RNA positivity, and measurement of the virus load, respectively. Both showed a positive correlation with the NO levels (P< 0.001). At a nitrate cutoff value of 70μmol/L, the sensitivity and specificity were 83.0% and 37.0% respectively. Chi square analysis showed a significant correlation between ALT levels and both HCV RNA positivity by polymerase chain reaction (PCR) (P< 0.02), and virus load (P< 0.05). Interestingly enough, there was a significant positive correlation between HCV RNA and schistosomal antibody titer as measured by hemaglutination inhibition assay (HAI) (P< 0.05). The data presented in this report indicated an association between NO levels and the development and progression of liver cirrhosis. Furthermore, the findings obtained from this study demonstrated that schistomiasis is an important risk factor involved in enhancement of NO levels and virus replication. The latter may aggravate liver cell injury and hence the development of cirrhosis.

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Use of Whole Blood Specimens for Routine Clinical Quantitation of Hepatitis C Virus RNA Does Not Increase Assay Sensitivity

Journal of Clinical Microbiology ◽

10.1128/jcm.38.12.4326-4331.2000 ◽

2000 ◽

Vol 38 (12) ◽

pp. 4326-4331 ◽

Cited By ~ 9

Author(s):

Linda Cook ◽

Angela M. Ross ◽

Glenn B. Knight ◽

Vincent Agnello

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Whole Blood ◽

Extraction Method ◽

White Blood Cells ◽

Clinical Samples ◽

Hcv Rna ◽

Clinical Diagnostic Laboratory ◽

Diagnostic Laboratory ◽

Assay Sensitivity

The measurement of hepatitis C virus (HCV) RNA levels in the blood has, in the last few years, become a critical component in the therapy of patients with HCV infections. Initially, extraction methods for serum and plasma were used, but a newer method that uses Catrimox-14 as the extraction agent for whole blood has been reported. Because the whole blood extraction method may yield higher virus levels if significant levels of virus are present in the white blood cells (WBC), the method was evaluated for use in our clinical diagnostic laboratory despite its higher reagent costs and more time-consuming methodology. RNA was simultaneously extracted from 39 clinical samples by four different methods: Catrimox-14–Trizol extraction from whole blood, Trizol extraction from whole blood, Trizol extraction from serum, and a commercial serum extraction method, the EZNA total RNA kit. In addition, in an effort to quantitate the amount of HCV RNA virus in the WBC, Trizol extraction from isolated WBC was also performed. Quantitative results for samples from which RNA was extracted by all four methods were essentially the same; the Catrimox-14–Trizol method did not yield increased virus levels. Insignificant levels of virus were found in the WBC. The results did not demonstrate a clinical usefulness for the Catrimox-14–Trizol method.

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