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1.
Gene ; 820: 146235, 2022 Apr 30.
Article in English | MEDLINE | ID: covidwho-1778131

ABSTRACT

The relationship of single nucleotide polymorphisms (SNPs) in patatin-like phospholipase domain containing 3 (PNPLA3) rs738409, transmembrane 6 superfamily member 2 (TM6SF2) rs58542926, and membrane bound O-acyltransferase domain containing 7 (MBOAT7) rs641738 with outcomes in patients with hepatitis C infection (HCV) is unclear. This study aimed to evaluate the association of PNPLA3, TM6SF2, and MBOAT7 with the baseline fibrosis stage and progression of liver fibrosis after HCV eradication with direct antiviral agents (DAAs). A total of 171 patients who received the DAAs at the Peking University First Hospital between June 2015 and June 2020 were included in the retrospective cohort. Transient elastography was used to determine liver stiffness measurements (LSMs) at the baseline, the end of treatment (EOT), 24 weeks after treatment (W24), and the last follow-up (LFU) visit. We used the QIAamp Blood Mini Kit (Qiagen) for whole blood genomic DNA extraction and polymerase chain reaction for PNPLA3, TM6SF2, and MBOAT7 amplification of the target gene. The PNPLA3 rs738409 SNP was associated with the baseline fibrosis stage in multivariate logistic regression analysis adjusted for other factors, and the adjusted odds ratio (OR) for advanced fibrosis (≥F3) at baseline was 2.52 (95% confidence interval[CI] = 1.096-5.794, p = 0.03). The G and GG alleles were predictive of advanced fibrosis (OR = 1.98, 95% CI = 1.021-4.196, p = 0.015; OR = 3.12, 95% CI = 1.572-6.536, p = 0.005). Similarly, the OR of TM6SF2 rs58542926 at baseline was 2.608 (95% CI = 1.081-6.29, p = 0.033). T and TT alleles were predictive of advanced fibrosis (OR = 2.3, 95% CI = 1.005-5.98, p = 0.007; OR = 3.05, 95% CI = 1.32-6.87, p = 0.001). After adjustment, the MBOAT7 rs641738 T plus TT alleles were not independently associated with the baseline fibrosis stage (95% CI = 0.707-2.959, p = 0.312). At the EOT, there were 35 patients and 136 patients in the fibrosis improvement and fibrosis non-improvement group, respectively. Logistic regression analysis showed that the G allele in PNPLA3 rs738409 was associated with fibrosis progression (OR = 2.47, 95% CI = 1.125-5.89, p = 0.003). The GG alleles were predictive of fibrosis progression (OR = 2.95, 95% CI = 1.35-6.35, p = 0.005). Similarly, the ORs of the T and TT alleles in TM6SF2 rs58542926 for fibrosis progression were 1.82 and 2.21, respectively (95% CI = 1.006-5.373, p = 0.045; 95% CI = 1.18-5.75, p = 0.01). At the W24 visit, we found that there was an association between the G allele in PNPLA3 rs738409 and fibrosis progression (OR = 2.218, 95% CI = 1.095-5.631, p = 0.015). Moreover, GG alleles were also predictive for fibrosis progression (OR = 2.558, 95% CI = 1.252-5.15, p = 0.008). Similarly, the OR of T allele and TT alleles in TM6SF2 rs58542926 for fibrosis progression was 2.056 and 2.652 (95% CI = 1.013-5.592, p = 0.038; 95% CI = 1.25-5.956, p = 0.015). For additional affirmation, we surveyed fibrosis progression utilizing the Cox proportional hazards model. G and GG alleles in PNPLA3 rs738409 were associated with an increased risk of progression to advanced fibrosis in multivariate model (hazard ratio [HR]1.566, 95% CI = 1.02-2.575, p = 0.017; and HR2.109, 95% CI = 1.36-3.271, p = 0.001, respectively). Besides, T and TT alleles in TM6SF2 rs58542926 were associated with an increased risk of progression to advanced fibrosis in multivariate model (HR = 1.322, 95% CI = 1.003-1.857, p = 0.045; and HR = 1.855, 95% CI = 1.35-2.765, p = 0.006, respectively). In contrast, rs641738 in MBOAT7 did not show a significant trend in the univariate and multivariate models. The PNPLA3 CG/GG SNP at rs738409 and TM6SF2 CT/TT SNP at rs58542926 were associated with the baseline fibrosis stage and fibrosis progression after HCV eradication with DAAs.


Subject(s)
Acyltransferases/economics , Acyltransferases/genetics , Liver Cirrhosis/genetics , Membrane Proteins/economics , Membrane Proteins/genetics , Phospholipases A2, Calcium-Independent/genetics , Polymorphism, Single Nucleotide , Adult , Aged , Alleles , Disease Progression , Female , Genetic Predisposition to Disease , Hepacivirus , Hepatitis C/complications , Hepatitis C/virology , Humans , Male , Middle Aged , Non-alcoholic Fatty Liver Disease/genetics , Prognosis , Retrospective Studies
2.
Viruses ; 14(2)2022 02 02.
Article in English | MEDLINE | ID: covidwho-1715769

ABSTRACT

A hepatitis C virus (HCV) screening and treatment program was conducted in Hungarian prisons on a voluntary basis. After HCV-RNA testing and genotyping for anti-HCV positives, treatments with direct-acting antiviral agents were commenced by hepatologists who visited the institutions monthly. Patients were supervised by the prisons' medical staff. Data were retrospectively collected from the Hungarian Hepatitis Treatment Registry, from the Health Registry of Prisons, and from participating hepatologists. Eighty-four percent of Hungarian prisons participated, meaning a total of 5779 individuals (28% of the inmate population) underwent screening. HCV-RNA positivity was confirmed in 317/5779 cases (5.49%); 261/317 (82.3%) started treatment. Ninety-nine percent of them admitted previous intravenous drug use. So far, 220 patients received full treatment and 41 patients are still on treatment. Based on the available end of treatment (EOT) + 24 weeks timepoint data, per protocol sustained virologic response rate was 96.8%. In conclusion, the Hungarian prison screening and treatment program, with the active participation of hepatologists and the prisons' medical staff, is a well-functioning model. Through the Hungarian experience, we emphasize that the "test-and-treat" principle is feasible and effective at micro-eliminating HCV in prisons, where infection rate, as well as history of intravenous drug usage, are high.


Subject(s)
Antiviral Agents/administration & dosage , Hepacivirus/isolation & purification , Hepatitis C/drug therapy , Adolescent , Adult , Female , Hepacivirus/drug effects , Hepacivirus/genetics , Hepacivirus/immunology , Hepatitis C/blood , Hepatitis C/diagnosis , Hepatitis C/virology , Humans , Hungary , Male , Mass Screening , Middle Aged , Prisons/statistics & numerical data , Retrospective Studies , Sustained Virologic Response , Young Adult
3.
Sci Rep ; 12(1): 330, 2022 01 10.
Article in English | MEDLINE | ID: covidwho-1617001

ABSTRACT

We conducted a systematic review and meta-analysis of studies assessing HCV infection rates in haemodialysis patients in Brazil (Prospero CRD #42021275068). We included studies on patients under haemodialysis, comprising both convenience samples and exhaustive information from selected services. Patients underwent HCV serological testing with or without confirmation by HCV RNA PCR. Exclusion criteria were the following: absence of primary empirical information and studies without information on their respective settings, study year, accurate infection rates, or full specification of diagnostic tests. Studies with samples ≤ 30 and serial assessments with repeated information were also excluded. Reference databases included PubMed, LILACS, Scopus, and Web of Science for the period 1989-2019. A systematic review was carried out, followed by two independent meta-analyses: (i) studies with data on HCV prevalence and (ii) studies with a confirmatory PCR (i.e., active infection), respectively. A comprehensive set of different methods and procedures were used: forest plots and respective statistics, polynomial regression, meta-regression, subgroup influence, quality assessment, and trim-and-fill analysis. 29 studies and 11,290 individuals were assessed. The average time patients were in haemodialysis varied from 23.5 to 56.3 months. Prevalence of HCV infection was highly heterogeneous, with a pronounced decrease from 1992 to 2001, followed by a plateau and a slight decrease in recent years. The summary measure for HCV prevalence was 34% (95% CI 26-43%) for studies implemented before 2001. For studies implemented after 2001, the corresponding summary measure was 11% (95% CI 8-15%). Estimates for prevalence of active HCV infection were also highly heterogeneous. There was a marked decline from 1996 to 2001, followed by a plateau and a slight increase after 2010. The summary measure for active HCV infection was 19% (95% CI 15-25%) in studies carried out before 2001. For studies implemented after 2001, the corresponding summary measure was 9% (95% CI 6-13%). Heterogeneity was pervasive, but different analyses helped to identify its underlying sources. Besides the year each study was conducted, the findings differed markedly between geographic regions and were heavily influenced by the size of the studies and publication biases. Our systematic review and meta-analysis documented a substantial decline in HCV prevalence among Brazilian haemodialysis patients from 1992 to 2015. CKD should be targeted with specific interventions to prevent HCV infection, and if prevention fails, prompt diagnosis and treatment. Although the goal of HCV elimination by 2030 in Brazil remains elusive, it is necessary to adopt measures to achieve micro-elimination and to launch initiatives towards targeted interventions to curb the spread of HCV in people with CKD, among other high-risk groups. This is of particular concern in the context of a protracted COVID-19 pandemic and a major economic and political crisis.


Subject(s)
COVID-19/diagnosis , Hepacivirus/genetics , Hepatitis C/diagnosis , Renal Dialysis/statistics & numerical data , SARS-CoV-2/genetics , Brazil/epidemiology , COVID-19/epidemiology , COVID-19/virology , Hepacivirus/physiology , Hepatitis C/epidemiology , Hepatitis C/virology , Humans , Nucleic Acid Amplification Techniques/methods , Pandemics , Prevalence , RNA, Viral/genetics , Renal Dialysis/methods , SARS-CoV-2/physiology
4.
Med Sci Monit ; 27: e935075, 2021 Dec 31.
Article in English | MEDLINE | ID: covidwho-1592562

ABSTRACT

BACKGROUND Thyroiditis is an important extrahepatic association in chronic hepatitis C virus (HCV) infection. There have been reports of an association between SARS-CoV-2 infection and the onset or re-activation of autoimmune hypothyroidism. Therefore, we performed this prospective observational study of 42 patients with COVID-19 infection and a history of hepatitis C virus infection and thyroid disease with follow-up thyroid function and autoantibody testing. MATERIAL AND METHODS From April 2020 to October 2020, we performed a prospective observational study of patients with cured hepatitis C virus (HCV) infection and documented thyroid disease who became infected with SARS-CoV-2 (confirmed by SARS-CoV-2 RNA detection via reverse-transcription polymerase chain reaction [RT-PCT] from the upper respiratory tract, both nasal and pharyngeal swabs). Evaluation at 1 and 3 months after SARS-CoV-2 infection included serum determination of antithyroid antibodies (anti-thyroglobulin [anti-Tg] and antithyroid peroxidase [ATPO]), thyroid-stimulating hormone (TSH), free thyroxine (fT4), free triiodothyronine (fT3), and evaluation of thyroid medication, with dose adjustment if required. RESULTS One-month follow-up showed that both patients with autoimmune thyroiditis as well as patients without antibodies had increased ATPO levels. Also, levels of TSH, fT3, and fT4 were significantly decreased. At 3-month follow-up, levels of ATPO were decreased in all patient groups and the levels of thyroid hormones increased to normal values. CONCLUSIONS This study supports previous reports of an association between SARS-CoV-2 infection and thyroid dysfunction associated with thyroid autoantibodies. Thyroid function tests may be considered as part of the laboratory work-up in patients with COVID-19.


Subject(s)
COVID-19/complications , Hepatitis C/complications , Hypothyroidism/etiology , Adult , Aged , COVID-19/virology , Female , Follow-Up Studies , Hepacivirus/pathogenicity , Hepatitis C/virology , Humans , Hypothyroidism/physiopathology , Hypothyroidism/virology , Male , Middle Aged , Prospective Studies , RNA, Viral , Romania/epidemiology , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Thyroid Diseases/physiopathology , Thyroid Function Tests , Thyroid Gland/physiology , Thyroiditis, Autoimmune/blood , Thyroiditis, Autoimmune/immunology , Thyrotropin/blood , Thyroxine/blood , Triiodothyronine/blood
6.
Viruses ; 13(4)2021 04 03.
Article in English | MEDLINE | ID: covidwho-1389536

ABSTRACT

Replication of RNA viruses is characterized by exploration of sequence space which facilitates their adaptation to changing environments. It is generally accepted that such exploration takes place mainly in response to positive selection, and that further diversification is boosted by modifications of virus population size, particularly bottleneck events. Our recent results with hepatitis C virus (HCV) have shown that the expansion in sequence space of a viral clone continues despite prolonged replication in a stable cell culture environment. Diagnosis of the expansion was based on the quantification of diversity indices, the occurrence of intra-population mutational waves (variations in mutant frequencies), and greater individual residue variations in mutant spectra than those anticipated from sequence alignments in data banks. In the present report, we review our previous results, and show additionally that mutational waves in amplicons from the NS5A-NS5B-coding region are equally prominent during HCV passage in the absence or presence of the mutagenic nucleotide analogues favipiravir or ribavirin. In addition, by extending our previous analysis to amplicons of the NS3- and NS5A-coding region, we provide further evidence of the incongruence between amino acid conservation scores in mutant spectra from infected patients and in the Los Alamos National Laboratory HCV data banks. We hypothesize that these observations have as a common origin a permanent state of HCV population disequilibrium even upon extensive viral replication in the absence of external selective constraints or changes in population size. Such a persistent disequilibrium-revealed by the changing composition of the mutant spectrum-may facilitate finding alternative mutational pathways for HCV antiviral resistance. The possible significance of our model for other genetically variable viruses is discussed.


Subject(s)
Hepacivirus/genetics , Hepacivirus/physiology , Hepatitis C/virology , Antiviral Agents/pharmacology , COVID-19 , Cell Line , Drug Resistance, Viral/drug effects , Hepacivirus/drug effects , Humans , Mutation , RNA, Viral , Ribavirin/pharmacology , Sequence Analysis , Viral Nonstructural Proteins/genetics , Virus Replication/drug effects
7.
Viruses ; 13(8)2021 08 18.
Article in English | MEDLINE | ID: covidwho-1376994

ABSTRACT

Viral infection is a global public health threat causing millions of deaths. A suitable small animal model is essential for viral pathogenesis and host response studies that could be used in antiviral and vaccine development. The tree shrew (Tupaia belangeri or Tupaia belangeri chinenesis), a squirrel-like non-primate small mammal in the Tupaiidae family, has been reported to be susceptible to important human viral pathogens, including hepatitis viruses (e.g., HBV, HCV), respiratory viruses (influenza viruses, SARS-CoV-2, human adenovirus B), arboviruses (Zika virus and dengue virus), and other viruses (e.g., herpes simplex virus, etc.). The pathogenesis of these viruses is not fully understood due to the lack of an economically feasible suitable small animal model mimicking natural infection of human diseases. The tree shrew model significantly contributes towards a better understanding of the infection and pathogenesis of these important human pathogens, highlighting its potential to be used as a viable viral infection model of human viruses. Therefore, in this review, we summarize updates regarding human viral infection in the tree shrew model, which highlights the potential of the tree shrew to be utilized for human viral infection and pathogenesis studies.


Subject(s)
Disease Models, Animal , Tupaia , Virus Diseases , Adenoviridae Infections/immunology , Adenoviridae Infections/virology , Animals , COVID-19/virology , Dengue/immunology , Dengue/pathology , Dengue/virology , HIV Infections/virology , Hepatitis B/immunology , Hepatitis B/virology , Hepatitis C/immunology , Hepatitis C/pathology , Hepatitis C/virology , Herpes Simplex/pathology , Herpes Simplex/virology , Humans , Influenza, Human/immunology , Influenza, Human/virology , Orthomyxoviridae Infections/immunology , Orthomyxoviridae Infections/virology , Zika Virus Infection/immunology , Zika Virus Infection/pathology , Zika Virus Infection/virology
8.
Microbiol Spectr ; 9(1): e0043921, 2021 09 03.
Article in English | MEDLINE | ID: covidwho-1329042

ABSTRACT

Hepatitis C virus (HCV) can cause acute and chronic infection that is associated with considerable liver-related morbidity and mortality. In recent years, there has been a shift in the treatment paradigm with the discovery and approval of agents that target specific proteins vital for viral replication. We employed a cell culture-adapted strain of HCV and human hepatoma-derived cells lines to test the effects of our novel small-molecule compound (AO13) on HCV. Virus inhibition was tested by analyzing RNA replication, protein expression, and virus production in virus-infected cells treated with AO13. Treatment with AO13 inhibited virus spread in cell culture and showed a 100-fold reduction in the levels of infectious virus production. AO13 significantly reduced the level of viral RNA contained within cell culture fluids and reduced the cellular levels of HCV core protein, suggesting that the compound might act on a late step in the viral life cycle. Finally, we observed that AO13 did not affect the release of infectious virus from infected cells. Docking studies and molecular dynamics analyses suggested that AO13 might target the NS5B RNA polymerase, however, real-time RT-PCR analyses of cellular levels of HCV RNA showed only an ∼2-fold reduction in viral RNA levels in the presence of AO13. Taken together, this study revealed that AO13 showed consistent, but low-level antiviral effect against HCV, although the mechanism of action remains unclear. IMPORTANCE The discovery of curative antiviral drugs for a chronic disease such as HCV infection has encouraged drug discovery in the context of other viruses for which no curative drugs currently exist. Since we currently face a novel virus that has caused a pandemic, the need for new antiviral agents is more apparent than ever. We describe here a novel compound that shows a modest antiviral effect against HCV that could serve as a lead compound for future drug development against other important viruses such as SARS-CoV-2.


Subject(s)
Antiviral Agents/pharmacology , Cell Culture Techniques , Hepacivirus/drug effects , Virus Replication/drug effects , Antiviral Agents/therapeutic use , Carcinoma, Hepatocellular , Cell Line , Hepacivirus/genetics , Hepacivirus/physiology , Hepatitis C/drug therapy , Hepatitis C/virology , Humans , Life Cycle Stages , Liver , Liver Neoplasms , Molecular Docking Simulation , RNA, Viral , SARS-CoV-2 , Viral Nonstructural Proteins , Virus Release/drug effects
9.
Viruses ; 13(6)2021 05 29.
Article in English | MEDLINE | ID: covidwho-1282636

ABSTRACT

An effective vaccine for the hepatitis C virus (HCV) is a major unmet medical and public health need, and it requires an antigen that elicits immune responses to multiple key conserved epitopes. Decades of research have generated a number of vaccine candidates; based on these data and research through clinical development, a vaccine antigen based on the E1E2 glycoprotein complex appears to be the best choice. One bottleneck in the development of an E1E2-based vaccine is that the antigen is challenging to produce in large quantities and at high levels of purity and antigenic/functional integrity. This review describes the production and characterization of E1E2-based vaccine antigens, both membrane-associated and a novel secreted form of E1E2, with a particular emphasis on the major challenges facing the field and how those challenges can be addressed.


Subject(s)
Hepacivirus/chemistry , Hepatitis C/prevention & control , Viral Envelope Proteins/chemistry , Viral Hepatitis Vaccines/chemistry , Animals , Epitopes/immunology , HEK293 Cells , Hepacivirus/genetics , Hepacivirus/immunology , Hepatitis C/virology , Humans , Mice , Models, Molecular , Protein Conformation , Protein Multimerization , Viral Envelope Proteins/immunology , Viral Envelope Proteins/metabolism
10.
Am J Trop Med Hyg ; 105(1): 117-124, 2021 05 17.
Article in English | MEDLINE | ID: covidwho-1231559

ABSTRACT

Hepatitis C virus (HCV) infection large-scale diagnosis and treatment are hampered by lack of a simple, rapid, and reliable point-of-care (POC) test, which poses a challenge for the elimination of hepatitis C as a public health problem. This study aimed to evaluate Cepheid Xpert® HCV Viral Load performance in comparison with the Roche Cobas® TaqMan® HCV Test using serum samples of HCV-infected patients in Indonesia. Viral load quantification was performed on 243 anti-HCV positive patients' samples using both Xpert HCV VL and Roche HCV tests, followed by HCV genotyping by reverse hybridization. Strength of the relationship between the assays was measured by Pearson correlation coefficient, while level of agreement was analyzed by Deming regression and Bland-Altman plot analysis using log10-transformed viral load values. Quantifiable viral load was detected in 180/243 (74.1%), with Xpert HCV VL sensitivity of 100% (95% CI 0.98, 1.00) and specificity of 98.4% (95% CI 0.91, 0.99) based on the Roche HCV test, while HCV genotypes were determined in 172/180 (95.6%) samples. There was a good correlation between both assays (r = 0.97, P < 0.001), overall and per genotype, with good concordance by Deming regression and a mean difference of -0.25 log10 IU/mL (95% CI -0.33, -0.18) by Bland-Altman plot analysis. Xpert HCV VL test was demonstrated as a POC platform with good performance for HCV diagnosis and treatment decision that would be beneficial for decentralized services in resource-limited areas. HCV testing sites, alongside additional GeneXpert modular systems distributed toward the fight against COVID-19, could ensure some continuity, once this pandemic is controlled.


Subject(s)
Hepacivirus/genetics , Hepatitis C/diagnosis , Hepatitis C/virology , Point-of-Care Testing , Viral Load , Adolescent , Adult , Aged , Aged, 80 and over , Child , Female , Genotype , Hepacivirus/isolation & purification , Hepatitis C/epidemiology , Humans , Indonesia/epidemiology , Male , Middle Aged , RNA, Viral/genetics , Sensitivity and Specificity , Young Adult
11.
PLoS One ; 16(5): e0250833, 2021.
Article in English | MEDLINE | ID: covidwho-1223798

ABSTRACT

Despite the availability of highly effective and well-tolerated direct-acting antivirals, not all patients with chronic hepatitis C virus infection receive treatment. This retrospective, multi-centre, noninterventional, case-control study identified patients with chronic hepatitis C virus infection initiating (control) or not initiating (case) treatment at 43 sites in Germany from September 2017 to June 2018. It aimed to compare characteristics of the two patient populations and to identify factors involved in patient/physician decision to initiate/not initiate chronic hepatitis C virus treatment, with a particular focus on historical barriers. Overall, 793 patients were identified: 573 (72%) who received treatment and 220 (28%) who did not. In 42% of patients, the reason for not initiating treatment was patient wish, particularly due to fear of treatment (17%) or adverse events (13%). Other frequently observed reasons for not initiating treatment were in accordance with known historical barriers for physicians to initiate therapy, including perceived or expected lack of compliance (14.5%), high patient age (10.9%), comorbidities (15.0%), alcohol abuse (9.1%), hard drug use (7.7%), and opioid substitution therapy (4.5%). Patient wish against therapy was also a frequently reported reason for not initiating treatment in the postponed (35.2%) and not planned (47.0%) subgroups; of note, known historical factors were also common reasons for postponing treatment. Real-world and clinical trial evidence is accumulating, which suggests that such historical barriers do not negatively impact treatment effectiveness. Improved education is key to facilitate progress towards the World Health Organization target of eliminating viral hepatitis as a major public health threat by 2030.


Subject(s)
Antiviral Agents/therapeutic use , Hepacivirus/isolation & purification , Hepatitis C/drug therapy , Hepatitis C/psychology , Patient Compliance/psychology , Adolescent , Adult , Aged , Aged, 80 and over , Case-Control Studies , Drug Therapy, Combination , Female , Germany/epidemiology , Hepatitis C/epidemiology , Hepatitis C/virology , Humans , Male , Middle Aged , Retrospective Studies , Sustained Virologic Response , Young Adult
12.
J Prim Care Community Health ; 11: 2150132720969554, 2020.
Article in English | MEDLINE | ID: covidwho-939996

ABSTRACT

INTRODUCTION: Coronavirus disease 2019 (COVID-19) has led to unprecedented modifications to healthcare delivery in the U.S. To preserve resources in preparation for a COVID-19 surge, Boston Medical Center (BMC) implemented workflows to decrease ambulatory in-person visits effective March 16th, 2020. Telemedicine was incorporated into clinical workflows and much preventive care, including Hepatitis C (HCV) testing, was not routinely performed. OBJECTIVE: To explore the impact that the COVID-19 rapid restructuring response has had on HCV testing and identification hospital-wide and in ambulatory settings. METHODS: BMC utilizes reflex confirmatory testing for HCV. When a sample is HCV Ab positive, it is automatically reflexed for confirmatory RNA and genotype testing. HCV test results for patients were collected daily. We compared unique patient tests for 3.5 month periods before and after March 16th, 2020. Descriptive statistics showed total tests and total new HCV RNA+ before versus after, both hospital-wide and in ambulatory clinics alone. Mean daily tests completed were compared. RESULTS: Hospital-wide, total HCV testing decreased by 49.6%, and new HCV+ patient identification decreased by 42.1%. In ambulatory clinics, testing decreased by 71.9%, and new HCV+ identification decreased by 63.3%. Hospital-wide, mean daily tests decreased by 22.9 tests per day (95% CI: 17.9-28.0, P < .001), and mean daily new HCV+ identification decreased by 0.36 (95% CI: 0.20-0.53, P < .001). In ambulatory clinics, mean daily tests decreased by 22.1 tests per day (95% CI: 17.5-26.7, P < .001) and mean daily HCV+ decreased by 1.40 (95% CI: 1.03-1.76, P < .001). CONCLUSION: The COVID-19 systematic emergency response led to decreased HCV testing and identification, and in this regard telemedicine acts as a barrier to HCV care. Other public health initiatives must be monitored in the context of telemedicine workflows. Continued monitoring of HCV screening trends is vital, and adaptive approaches to work toward the goal of HCV elimination are needed.


Subject(s)
Ambulatory Care Facilities , COVID-19 , Delivery of Health Care , Hepatitis C/diagnosis , Mass Screening , Pandemics , Telemedicine , Adolescent , Adult , Aged , Antibodies, Viral/blood , Boston , COVID-19/prevention & control , Coronavirus , Delivery of Health Care/methods , Delivery of Health Care/organization & administration , Emergencies , Female , Health Services Accessibility , Hepacivirus/genetics , Hepacivirus/immunology , Hepatitis C/virology , Hospitals , Humans , Male , Mass Screening/methods , Mass Screening/statistics & numerical data , Middle Aged , RNA, Viral , Young Adult
13.
Theranostics ; 10(26): 12223-12240, 2020.
Article in English | MEDLINE | ID: covidwho-934619

ABSTRACT

Rationale: Many viral infections are known to activate the p38 mitogen-activated protein kinase (MAPK) signaling pathway. However, the role of p38 activation in viral infection and the underlying mechanism remain unclear. The role of virus-hijacked p38 MAPK activation in viral infection was investigated in this study. Methods: The correlation of hepatitis C virus (HCV) infection and p38 activation was studied in patient tissues and primary human hepatocytes (PHHs) by immunohistochemistry and western blotting. Coimmunoprecipitation, GST pulldown and confocal microscopy were used to investigate the interaction of p38α and the HCV core protein. In vitro kinase assays and mass spectrometry were used to analyze the phosphorylation of the HCV core protein. Plaque assays, quantitative real time PCR (qRT-PCR), western blotting, siRNA and CRISPR/Cas9 were used to determine the effect of p38 activation on viral replication. Results: HCV infection was associated with p38 activation in clinical samples. HCV infection increased p38 phosphorylation by triggering the interaction of p38α and TGF-ß activated kinase 1 (MAP3K7) binding protein 1 (TAB1). TAB1-mediated p38α activation facilitated HCV replication, and pharmaceutical inhibition of p38α activation by SB203580 suppressed HCV infection at the viral assembly step. Activated p38α interacted with the N-terminal region of the HCV core protein and subsequently phosphorylated the HCV core protein, which promoted HCV core protein oligomerization, an essential step for viral assembly. As expected, SB203580 or the HCV core protein N-terminal peptide (CN-peptide) disrupted the p38α-HCV core protein interaction, efficiently impaired HCV assembly and impeded normal HCV replication in both cultured cells and primary human hepatocytes. Similarly, severe fever with thrombocytopenia syndrome virus (SFTSV), herpes simplex virus type 1 (HSV-1) or severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection also activated p38 MAPK. Most importantly, pharmacological blockage of p38 activation by SB203580 effectively inhibited SFTSV, HSV-1 and SARS-CoV-2. Conclusion: Our study shows that virus-hijacked p38 activation is a key event for viral replication and that pharmacological blockage of p38 activation is an antiviral strategy.


Subject(s)
COVID-19/metabolism , Hepacivirus/metabolism , Hepatitis C/metabolism , Mitogen-Activated Protein Kinase 14/metabolism , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/metabolism , A549 Cells , Adaptor Proteins, Signal Transducing/metabolism , Animals , COVID-19/virology , Chlorocebus aethiops , Enzyme Activation , HEK293 Cells , Hepatitis C/pathology , Hepatitis C/virology , Hepatocytes/metabolism , Humans , Imidazoles/pharmacology , MAP Kinase Kinase Kinases/metabolism , MAP Kinase Signaling System/drug effects , Mitogen-Activated Protein Kinase 14/antagonists & inhibitors , Phosphorylation , Pyridines/pharmacology , Vero Cells , Viral Core Proteins/metabolism , Virus Replication/drug effects
14.
Antiviral Res ; 185: 104974, 2021 01.
Article in English | MEDLINE | ID: covidwho-927811

ABSTRACT

Vaccines and antiviral agents are in urgent need to stop the COVID-19 pandemic. To facilitate antiviral screening against SARS-CoV-2 without requirement for high biosafety level facility, we developed a bacterial artificial chromosome (BAC)-vectored replicon of SARS-CoV-2, nCoV-SH01 strain, in which secreted Gaussia luciferase (sGluc) was encoded in viral subgenomic mRNA as a reporter gene. The replicon was devoid of structural genes spike (S), membrane (M), and envelope (E). Upon transfection, the replicon RNA replicated in various cell lines, and was sensitive to interferon alpha (IFN-α), remdesivir, but was resistant to hepatitis C virus inhibitors daclatasvir and sofosbuvir. Replication of the replicon was also sensitive overexpression to zinc-finger antiviral protein (ZAP). We also constructed a four-plasmid in-vitro ligation system that is compatible with the BAC system, which makes it easy to introduce desired mutations into the assembly plasmids for in-vitro ligation. This replicon system would be helpful for performing antiviral screening and dissecting virus-host interactions.


Subject(s)
Antiviral Agents/pharmacology , COVID-19/virology , Chromosomes, Artificial, Bacterial , Replicon/drug effects , SARS-CoV-2/drug effects , SARS-CoV-2/genetics , Adenosine Monophosphate/analogs & derivatives , Adenosine Monophosphate/pharmacology , Alanine/analogs & derivatives , Alanine/pharmacology , Animals , COVID-19/drug therapy , Cell Line , Chlorocebus aethiops , HEK293 Cells , Hepacivirus/drug effects , Hepatitis C/drug therapy , Hepatitis C/virology , Humans , Interferon-alpha/pharmacology , RNA-Binding Proteins/biosynthesis , RNA-Binding Proteins/genetics , Sofosbuvir/pharmacology , Vero Cells , Virus Replication/drug effects
16.
J Gen Virol ; 102(1)2021 01.
Article in English | MEDLINE | ID: covidwho-910292

ABSTRACT

Great strides have been made in understanding and treating hepatitis C virus (HCV) thanks to the development of various experimental systems including cell-culture-proficient HCV, the HCV pseudoparticle system and soluble envelope glycoproteins. The HCV pseudoparticle (HCVpp) system is a platform used extensively in studies of cell entry, screening of novel entry inhibitors, assessing the phenotypes of clinically observed E1 and E2 glycoproteins and, most pertinently, in characterizing neutralizing antibody breadth induced upon vaccination and natural infection in patients. Nonetheless, some patient-derived clones produce pseudoparticles that are either non-infectious or exhibit infectivity too low for meaningful phenotyping. The mechanisms governing whether any particular clone produces infectious pseudoparticles are poorly understood. Here we show that endogenous expression of CD81, an HCV receptor and a cognate-binding partner of E2, in producer HEK 293T cells is detrimental to the infectivity of recovered HCVpp for most strains. Many HCVpp clones exhibited increased infectivity or had their infectivity rescued when they were produced in 293T cells CRISPR/Cas9 engineered to ablate CD81 expression (293TCD81KO). Clones made in 293TCD81KO cells were antigenically very similar to their matched counterparts made parental cells and appear to honour the accepted HCV entry pathway. Deletion of CD81 did not appreciably increase the recovered titres of soluble E2 (sE2). However, we did, unexpectedly, find that monomeric sE2 made in 293T cells and Freestyle 293-F (293-F) cells exhibit important differences. We found that 293-F-produced sE2 harbours mostly complex-type glycans whilst 293T-produced sE2 displays a heterogeneous mixture of both complex-type glycans and high-mannose or hybrid-type glycans. Moreover, sE2 produced in 293T cells is antigenically superior; exhibiting increased binding to conformational antibodies and the large extracellular loop of CD81. In summary, this work describes an optimal cell line for the production of HCVpp and reveals that sE2 made in 293T and 293-F cells are not antigenic equals. Our findings have implications for functional studies of E1E2 and the production of candidate immunogens.


Subject(s)
Hepacivirus/physiology , Viral Envelope Proteins/metabolism , Antibody Affinity , Gene Knockdown Techniques , HEK293 Cells , Hepacivirus/immunology , Hepatitis C/virology , Hepatitis C Antibodies/immunology , Hepatitis C Antigens/immunology , Hepatitis C Antigens/metabolism , Humans , Mannose/chemistry , Polysaccharides/chemistry , Protein Binding , Receptors, Virus/genetics , Receptors, Virus/metabolism , Tetraspanin 28/genetics , Tetraspanin 28/metabolism , Viral Envelope Proteins/chemistry , Viral Envelope Proteins/immunology
17.
Channels (Austin) ; 14(1): 403-412, 2020 12.
Article in English | MEDLINE | ID: covidwho-889445

ABSTRACT

The COVID-19 pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has prompted an urgent need to identify effective medicines for the prevention and treatment of the disease. A comparative analysis between SARS-CoV-2 and Hepatitis C Virus (HCV) can expand the available knowledge regarding the virology and potential drug targets against these viruses. Interestingly, comparing HCV with SARS-CoV-2 reveals major similarities between them, ranging from the ion channels that are utilized, to the symptoms that are exhibited by patients. Via this comparative analysis, and from what is known about HCV, the most promising treatments for COVID-19 can focus on the reduction of viral load, treatment of pulmonary system damages, and reduction of inflammation. In particular, the drugs that show most potential in this regard include ritonavir, a combination of peg-IFN, and lumacaftor-ivacaftor. This review anaylses SARS-CoV-2 from the perspective of the role of ion homeostasis and channels in viral pathomechanism. We also highlight other novel treatment approaches that can be used for both treatment and prevention of COVID-19. The relevance of this review is to offer high-quality evidence that can be used as the basis for the identification of potential solutions to the COVID-19 pandemic.


Subject(s)
Betacoronavirus/metabolism , Coronavirus Infections/metabolism , Hepacivirus/metabolism , Ion Channels/metabolism , Pneumonia, Viral/metabolism , Animals , Betacoronavirus/pathogenicity , COVID-19 , Coronavirus Infections/pathology , Coronavirus Infections/virology , Hepacivirus/pathogenicity , Hepatitis C/metabolism , Hepatitis C/virology , Humans , Pandemics , Pneumonia, Viral/pathology , Pneumonia, Viral/virology , SARS-CoV-2
18.
Sci Rep ; 10(1): 16577, 2020 10 06.
Article in English | MEDLINE | ID: covidwho-834912

ABSTRACT

SARS-CoV-2 is responsible for COVID-19, resulting in the largest pandemic in over a hundred years. After examining the molecular structures and activities of hepatitis C viral inhibitors and comparing hepatitis C virus and coronavirus replication, we previously postulated that the FDA-approved hepatitis C drug EPCLUSA (Sofosbuvir/Velpatasvir) might inhibit SARS-CoV-2. We subsequently demonstrated that Sofosbuvir triphosphate is incorporated by the relatively low fidelity SARS-CoV and SARS-CoV-2 RNA-dependent RNA polymerases (RdRps), serving as an immediate polymerase reaction terminator, but not by a host-like high fidelity DNA polymerase. Other investigators have since demonstrated the ability of Sofosbuvir to inhibit SARS-CoV-2 replication in lung and brain cells; additionally, COVID-19 clinical trials with EPCLUSA and with Sofosbuvir plus Daclatasvir have been initiated in several countries. SARS-CoV-2 has an exonuclease-based proofreader to maintain the viral genome integrity. Any effective antiviral targeting the SARS-CoV-2 RdRp must display a certain level of resistance to this proofreading activity. We report here that Sofosbuvir terminated RNA resists removal by the exonuclease to a substantially higher extent than RNA terminated by Remdesivir, another drug being used as a COVID-19 therapeutic. These results offer a molecular basis supporting the current use of Sofosbuvir in combination with other drugs in COVID-19 clinical trials.


Subject(s)
Adenosine Monophosphate/analogs & derivatives , Alanine/analogs & derivatives , Antiviral Agents/pharmacology , Betacoronavirus/drug effects , Coronavirus Infections/drug therapy , Exonucleases/metabolism , Pneumonia, Viral/drug therapy , Prodrugs/pharmacology , RNA, Viral/drug effects , Sofosbuvir/pharmacology , Adenosine Monophosphate/chemistry , Adenosine Monophosphate/pharmacology , Adenosine Monophosphate/therapeutic use , Alanine/chemistry , Alanine/pharmacology , Alanine/therapeutic use , Antiviral Agents/chemistry , Antiviral Agents/therapeutic use , Betacoronavirus/enzymology , COVID-19 , Coronavirus Infections/virology , Coronavirus RNA-Dependent RNA Polymerase , Drug Discovery/methods , Drug Repositioning/methods , Hepacivirus/drug effects , Hepacivirus/enzymology , Hepatitis C/drug therapy , Hepatitis C/virology , Humans , Pandemics , Pneumonia, Viral/virology , Prodrugs/therapeutic use , RNA, Viral/chemistry , RNA, Viral/metabolism , RNA-Dependent RNA Polymerase/antagonists & inhibitors , RNA-Dependent RNA Polymerase/metabolism , SARS-CoV-2 , Sofosbuvir/chemistry , Sofosbuvir/therapeutic use , Viral Nonstructural Proteins/antagonists & inhibitors , Viral Nonstructural Proteins/metabolism , Virus Replication/drug effects
19.
Trends Microbiol ; 28(12): 1022-1033, 2020 12.
Article in English | MEDLINE | ID: covidwho-593617

ABSTRACT

Viruses, as obligate intracellular parasites, exploit cellular pathways and resources in a variety of fascinating ways. A striking example of this is the remodelling of intracellular membranes into specialized structures that support the replication of positive-sense ssRNA (+RNA) viruses infecting eukaryotes. These distinct forms of virus-induced structures include double-membrane vesicles (DMVs), found during viral infections as diverse and notorious as those of coronaviruses, enteroviruses, noroviruses, or hepatitis C virus. Our understanding of these DMVs has evolved over the past 15 years thanks to advances in imaging techniques and modern molecular biology tools. In this article, we review contemporary understanding of the biogenesis, structure, and function of virus-induced DMVs as well as the open questions posed by these intriguing structures.


Subject(s)
Intracellular Membranes/virology , Virus Replication/physiology , Animals , Coronavirus/physiology , Enterovirus/physiology , Hepacivirus/physiology , Hepatitis C/virology , Host Microbial Interactions/physiology , Humans , Norovirus/physiology , Organelle Biogenesis , RNA, Viral , Viral Proteins
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