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      Evaluation of hepatitis C viral RNA persistence in HIV-infected patients with long-term sustained virological response by droplet digital PCR

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          Abstract

          Several studies have reported the persistence of HCV RNA in liver and/or peripheral blood mononuclear cells (PBMCs) in spite of undetectable viremia in patients who have achieved sustained virological response (SVR). This event, defined as occult HCV infection, remains controversial and low titers of persistent virus may be underestimated because it has not yet been analyzed by a highly sensitive test such as droplet digital PCR (ddPCR). This method provides an alternate ultra-sensitive detection technique for very low numbers of copies of viral RNA or DNA. The aim of this study was to evaluate the persistence of HCV in HIV-coinfected patients with long-term SVR using ddPCR. For each patient, the presence of HCV RNA in serum and PBMCs at baseline was determined by nested RT-ddPCR. Patients with HCV RNA in PBMCs at baseline were followed until the end of the study. One hundred and twenty-three patients were analyzed for persistence of HCV RNA in serum and PBMCs. Persistence of HCV was not found in serum in any patient. HCV RNA was detected in PBMCs in one patient (0.81%; 95% CI: 0.04–3.94) and resolved spontaneously during follow-up. Persistence of HCV RNA in PBMCs is not a common event in HIV/HCV co-infected patients with long-term SVR evaluated by RT-ddPCR.

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          Eradication of hepatitis C virus in patients successfully treated for chronic hepatitis C.

          It is unclear whether hepatitis C virus (HCV) is eradicated in patients with chronic hepatitis C who achieved a sustained virologic response (SVR). In this long-term follow-up study, including chronic hepatitis C patients who achieved SVR after interferon-based therapy, the presence of residual HCV RNA in serum, liver, and peripheral blood mononuclear cells (PBMCs) was assessed, using transcription-mediated amplification (sensitivity, <9.6 IU/mL). The benefit of SVR on liver fibrosis was evaluated using the METAVIR score. A total of 344 patients were followed up for a median duration of 3.27 years (range, 0.50-18 y; interquartile range [IQR], 1.68-5.35 y). A total of 114 patients had a posttreatment liver tissue (median, 0.50 y; range, 0-14 y; IQR, 0-3.5 y) and a total of 156 had one PBMC (median, 3.0 y; range, 0.50-18 y; IQR, 1.25-5.50 y). Serum HCV RNA remained undetectable (1300 samples), indicating that none of the patients had a relapse. HCV RNA was detectable in 2 of 114 (1.7%) liver specimens, and in none of 156 PBMC specimens. Histologic analysis of 126 paired pretreatment and posttreatment liver biopsy specimens (median, 0.50 y; range, 0-14 y; IQR, 0-3.5 y) showed that fibrosis stage was improved in 56%, stable in 32%, deteriorated in 12%. Regression of cirrhosis was observed in 9 of 14 (64%) (95% confidence interval, 39-89) patients. No cirrhosis decompensation was observed, and 3 patients developed hepatocellular carcinoma. In this large cohort of chronic hepatitis C patients, SVR was durable up to 18 years after treatment cessation, in addition to fibrosis stability/improvement (88%) and cirrhosis regression (64%). The presence of residual HCV RNA was observed only in liver tissue (1.7%). This result strongly suggests that SVR may be considered to show eradication of HCV infection.
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            The use of transient elastometry for assessing liver fibrosis in patients with HIV and hepatitis C virus coinfection.

            Transient elastometry (TE) is accurate for detecting significant liver fibrosis and cirrhosis in hepatitis C virus (HCV)-monoinfected patients. However, this procedure has been insufficiently validated in patients with human immunodeficiency virus (HIV) and HCV coinfection. The purpose of this study was to validate reported cutoff values of TE that discriminate significant liver fibrosis and cirrhosis in HIV-HCV-coinfected subjects. Liver stiffness measurements were obtained for 169 HIV-HCV-coinfected adult patients who had undergone a liver biopsy or who had received a nonhistologic diagnosis of cirrhosis within 12 months before or after a liver stiffness measurement. Patients had received no prior therapy for HCV infection. TE measurements ranged from 3.6 kPa to 75 kPa. The area under the receiver operating characteristic curve was 0.87 (95% confidence interval, 0.84-0.93) for significant liver fibrosis and 0.95 (95% confidence interval, 0.92-0.99) for cirrhosis. To diagnose significant liver fibrosis, a cutoff value of 7.2 kPa was associated with a positive predictive value of 88% and a negative predictive value of 75%. Thirty-four patients (20%) were misclassified when this cutoff value was used. Thirteen (24%) of 54 patients with liver stiffness values <7.2 kPa had significant liver fibrosis detected by liver biopsy. To diagnose cirrhosis, a cutoff value of 14.6 kPa was associated with a positive predictive value of 86% and a negative predictive value of 94%. Thus, 13 patients (10%) had disease that was misclassified using this cutoff value. We found that the diagnostic accuracy of TE was high for detecting cirrhosis and good for diagnosis of significant liver fibrosis. However, the performance of TE was low for discriminating mild fibrosis from significant liver fibrosis, which might limit the applicability of this technique in clinical practice.
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              Persistence of hepatitis C virus in patients successfully treated for chronic hepatitis C.

              It is unclear whether the current antiviral treatment for chronic hepatitis C virus (HCV) infection results in complete elimination of the virus, or whether small quantities of virus persist. Our study group comprised 17 patients with chronic HCV who had sustained virological response (SVR) after interferon/ribavirin treatment. Serum and peripheral blood mononudear cells were collected 2 to 3 times at 3- to 6-month intervals starting 40 to 109 months (mean, 64.2 +/- 18.5 months) after the end of therapy. In addition, lymphocyte and macrophage cultures were established at each point. In 11 patients, frozen liver tissue samples were available from follow-up biopsies performed 41 to 98 months (mean, 63.6 +/- 16.7 months) after therapy. Presence of HCV RNA was determined by sensitive reverse-transcriptase polymerase chain reaction, and concentration of positive and negative strands was determined by a novel quantitative real-time reverse transcriptase polymerase chain reaction. Only 2 of 17 patients remained consistently HCV RNA negative in all analyzed compartments. HCV RNA was detected in macrophages from 11 patients (65%) and in lymphocytes from 7 patients (41%). Viral sequences were also detected in 3 of 11 livers and in sera from 4 patients. Viral replicative forms were found in lymphocytes from 2 and in macrophages from 4 patients. In conclusion, our results suggest that in patients with SVR after therapy, small quantities of HCV RNA may persist in liver or macrophages and lymphocytes for up to 9 years. This continuous viral presence could result in persistence of humoral and cellular immunity for many years after therapy and could present a potential risk for infection reactivation.
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                Author and article information

                Contributors
                arjvet@gmail.com
                Journal
                Sci Rep
                Sci Rep
                Scientific Reports
                Nature Publishing Group UK (London )
                2045-2322
                29 August 2019
                29 August 2019
                2019
                : 9
                : 12507
                Affiliations
                [1 ]ISNI 0000 0001 2183 9102, GRID grid.411901.c, Unidad de Enfermedades Infecciosas. Instituto Maimónides de Investigación Biomédica de Córdoba (IMIBIC), Hospital Universitario Reina Sofía de Córdoba, Universidad de Córdoba, ; Córdoba, Spain
                [2 ]Unidad Gestión Clínica Enfermedades Infecciosas, Hospital La Línea, AGS Campo de Gibraltar, Cádiz, Spain
                [3 ]Unidad de Enfermedades Infecciosas. Hospital Universitario Virgen de la Victoria, Complejo Hospitalario Provincial de Málaga, Málaga, Spain
                [4 ]Unidad de Genómica. Instituto Maimónides de Investigación Biomédica de Córdoba (IMIBIC), Hospital Universitario Reina Sofía, Universidad de Córdoba, Córdoba, Spain
                [5 ]ISNI 0000 0004 1768 1690, GRID grid.412800.f, Unidad de Enfermedades Infecciosas y Microbiología, Hospital Universitario de Valme, Instituto de Biomedicina de Sevilla (iBiS), ; Sevilla, Spain
                [6 ]Unidad de Enfermedades Infecciosas, Hospitales Juan Ramón Jiménez e Infanta Elena de Huelva, Huelva, Spain
                [7 ]ISNI 0000 0004 1771 208X, GRID grid.418878.a, Unidad de Enfermedades Infecciosas, Complejo Hospitalario de Jaén, ; Jaén, Spain
                [8 ]ISNI 0000 0004 1768 1455, GRID grid.452455.7, Unidad de Enfermedades Infecciosas, Hospital de Poniente, ; El Ejido, Spain
                Article
                48966
                10.1038/s41598-019-48966-9
                6715682
                31467339
                806a1dbe-7807-492a-b8fd-675779f2033b
                © The Author(s) 2019

                Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

                History
                : 24 January 2019
                : 13 August 2019
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                © The Author(s) 2019

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                molecular biology,medical research
                Uncategorized
                molecular biology, medical research

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