• Hepatitis C
• cirrhosis
• direct-acting antivirals
• drug-to-drug interactions
• liver disease
• personalized therapy

• Humans
• Antiviral Agents/adverse effects
• HIV Infections/drug therapy
• Hepatitis C, Chronic/drug therapy
• Hepatitis C/drug therapy
• Hepacivirus/genetics

• health care
• medical research
• risk factors

• Humans
• Hepacivirus
• Areca/adverse effects
• Taiwan/epidemiology
• Alcohol Drinking/adverse effects
• Alcohol Drinking/epidemiology
• Hepatitis C/epidemiology
• Liver Neoplasms
• Tobacco Products

• KMU-TC111A01 the Higher Education Sprout Project by the Ministry of Education (MOE) in Taiwan and by Kaohsiung Medical University Research Center
• KMUTC111IFSP01 the Higher Education Sprout Project by the Ministry of Education (MOE) in Taiwan and by Kaohsiung Medical University Research Center

• drug discovery and development
• viral infection

• Animals
• COVID-19
• Antiviral Agents/pharmacology
• Antiviral Agents/therapeutic use
• SARS-CoV-2
• Drug Discovery
• Pandemics

• Wellcome Trust
• U19 AI171399 NIAID NIH HHS

• Humans
• Antiviral Agents/pharmacology
• Hepacivirus/genetics
• Acute-On-Chronic Liver Failure/diagnosis
• Acute-On-Chronic Liver Failure/drug therapy
• Phylogeny
• Hepatitis C, Chronic/drug therapy
• Hepatitis C/complications
• Hepatitis C/drug therapy
• Genotype
• Hepatitis A/drug therapy
• Genomics
• Drug Resistance, Viral/genetics
• Viral Nonstructural Proteins/genetics
• Treatment Failure

• Hepatitis
• cancer
• diagnosis
• hepatocellular carcinoma.
• liver disease
• therapy

• Humans
• Hepacivirus
• Antiviral Agents/pharmacology
• Antiviral Agents/therapeutic use
• Hepatitis C, Chronic/drug therapy
• Viral Nonstructural Proteins
• Hepatitis C/drug therapy

The viral genome quasispecies composition of hepatitis C virus (HCV) could have important implications to viral pathogenesis and resistance to anti-viral treatment. The purpose of the present study was to profile the HCV RNA quasispecies. We developed a strategy to determine the full-length HCV genome sequences co-existing within a single patient serum by using next-generation sequencing technologies. The isolated viral clones were divided into the groups that can be distinguished by core amino acid 70 substitution. Subsequently, we determined HCV full-length genome sequences of three independent dominant species co-existing in the sequential serum with a 7-year interval. From phylogenetic analysis, these dominant species evolved independently. Our study demonstrated that multiple dominant species co-existed in patient sera and evolved independently.

•

HCV RNA genome forms quasispecies which may contribute viral pathogenesis.

• Influenza
• low-frequency variants
• next-generation sequencing
• patient data
• surveillance

Introduction of effective drugs in the treatment of hepatitis C virus (HCV) infection has prompted the World Health Organization to declare a global eradication target by 2030. Propositions have been made to screen the general population and treat all HCV carriers irrespective of the disease status. A year ago the new severe acute respiratory syndrome coronavirus 2 virus appeared causing a worldwide pandemic of coronavirus disease 2019 disease. Huge financial resources were redirected, and the pandemic became the first priority in every country. In this review, we examined the feasibility of the World Health Organization elimination program and the actual natural course of HCV infection. We also identified and analyzed certain comorbidity factors that may aggravate the progress of HCV and some marginalized subpopulations with characteristics favoring HCV dissemination. Alcohol consumption, HIV coinfection and the presence of components of metabolic syndrome including obesity, hyperuricemia and overt diabetes were comorbidities mostly responsible for increased liver-related morbidity and mortality of HCV. We also examined the significance of special subpopulations like people who inject drugs and males having sex with males. Finally, we proposed a different micro-elimination screening and treatment program that can be implemented in all countries irrespective of income. We suggest that screening and treatment of HCV carriers should be limited only in these particular groups.

• Hepatitis C, Comorbidities, Screening and treatment policy
• Hepatitis C virus
• Review

The hepatitis C virus (HCV) is a common cause of chronic liver disease and liver cancer worldwide. Despite advances in curative therapies for HCV, the incidence of new infections is not decreasing at the expected rate to hit the World Health Organization (WHO) target for the elimination of HCV by 2030. In fact, there are still more new cases of infection in the United States and worldwide than are being cured. The reasons for the rise in new cases include poor access to care and the opioid epidemic. The clinical burden of HCV requires a multimodal approach to eradicating the infection. Vaccination would be an excellent tool to prevent incidence of new infections; however, the genetic diversity of HCV and its ability to generate quasispecies within an infected host make creating a broadly reactive vaccine difficult. Multiple vaccine candidates have been identified, but to date, there has not been a target that has led to a broadly reactive vaccine, though several of the candidates are promising. Additionally, the virus is very difficult to culture and testing candidates in humans or chimpanzees is ethically challenging. Despite the multiple barriers to creating a vaccine, vaccination still represents an important tool in the fight against HCV.

• chronic liver disease
• direct acting anti-virals (DAAs)
• hepatitis C virus
• vaccination

• COVID-19
• Challenges
• Hepatitis C virus
• Nucleic acid-based vaccines
• Recombinant vector-based vaccines
• Vaccine candidates

• hepatitis c virus
• broadly neutralizing antibody
• antibody
• escape
• resistance

• Antibodies, Neutralizing
• Broadly Neutralizing Antibodies
• Hepacivirus
• Hepatitis C
• Hepatitis C Antibodies
• Humans
• Viral Envelope Proteins

• U19 AI159822 NIAID NIH HHS
• R01 AI127469 NIAID NIH HHS
• R21 AI151353 NIAID NIH HHS
• K99 AI153465 NIAID NIH HHS
• U19 AI088791 NIAID NIH HHS

• Grouper
• Nodavirus
• Quasispecies
• Tumor necrosis factor

• Animals
• Bass
• Capsid Proteins/genetics
• Capsid Proteins/immunology
• Fish Diseases/immunology
• Fish Proteins/genetics
• Fish Proteins/immunology
• Nodaviridae/genetics
• Quasispecies/physiology
• RNA Virus Infections/veterinary
• RNA Virus Infections/virology
• Tumor Necrosis Factor-alpha/genetics
• Tumor Necrosis Factor-alpha/immunology

Despite the recent development of safe and highly effective direct-acting antivirals, hepatitis C virus (HCV) infection remains a significant health problem. In 2016, the World Health Organization set out to reduce the rate of new HCV infections by 90% by 2030. Still, global control of the virus does not seem to be achievable in the absence of an effective vaccine. Current approaches to the development of a vaccine against HCV include the production of recombinant proteins, synthetic peptides, DNA vaccines, virus-like particles, and viral vectors expressing various antigens. In this review, we focus on the development of vaccines targeting the humoral immune response against HCV based on the cumulative evidence supporting the important role of neutralizing antibodies in protection against HCV infection. The main targets of HCV-specific neutralizing antibodies are the glycoproteins E1 and E2. Recent advances in the knowledge of HCV glycoprotein structure and their epitopes, as well as the possibility of getting detailed information on the human antibody repertoire generated by the infection, will allow rational structure-based antigen design to target specific germline antibodies. Although obtaining a vaccine capable of inducing sterilizing immunity will be a difficult task, a vaccine that prevents chronic hepatitis C infections, a more realistic goal in the short term, would have a considerable health impact.

• Antibody
• Glycoprotein E1
• Glycoprotein E2
• HCV
• Humoral immune response
• Vaccine
• Virus neutralization

Despite successful treatments, hepatitis C virus (HCV) infections continue to be a significant world health problem. High treatment costs, the high number of undiagnosed individuals, and the difficulty to access to treatment, particularly in marginalized susceptible populations, make it improbable to achieve the global control of the virus in the absence of an effective preventive vaccine. Current vaccine development is mostly focused on weakly immunogenic subunits, such as surface glycoproteins or non-structural proteins, in the case of HCV. Adjuvants are critical components of vaccine formulations that increase immunogenic performance. As we learn more information about how adjuvants work, it is becoming clear that proper stimulation of innate immunity is crucial to achieving a successful immunization. Several hepatic cell types participate in the early innate immune response and the subsequent inflammation and activation of the adaptive response, principally hepatocytes, and antigen-presenting cells (Kupffer cells, and dendritic cells). Innate pattern recognition receptors on these cells, mainly toll-like receptors, are targets for new promising adjuvants. Moreover, complex adjuvants that stimulate different components of the innate immunity are showing encouraging results and are being incorporated in current vaccines. Recent studies on HCV-vaccine adjuvants have shown that the induction of a strong T- and B-cell immune response might be enhanced by choosing the right adjuvant.

• HCV
• PRRs
• adjuvant
• innate immune response
• vaccine

The hepatitis C virus (HCV) causes both acute and chronic infection and continues to be a global problem despite advances in antiviral therapeutics. Current treatments fail to prevent reinfection and remain expensive, limiting their use to developed countries, and the asymptomatic nature of acute infection can result in individuals not receiving treatment and unknowingly spreading HCV. A prophylactic vaccine is therefore needed to control this virus. Thirty years since the discovery of HCV, there have been major gains in understanding the molecular biology and elucidating the immunological mechanisms that underpin spontaneous viral clearance, aiding rational vaccine design. This review discusses the challenges facing HCV vaccine design and the most recent and promising candidates being investigated.

• animal models
• hepatitis C virus
• immune responses
• neutralising antibodies
• vaccines

• G0801169 Medical Research Council
• MR/R010307/1 Medical Research Council

• genome viability
• genome-wide mutagenesis
• hepatitis C virus
• quasispecies
• reverse genetic analysis
• tolerance

• Science and Technology Commission of Shanghai Municipality

The rate of HIV infection in Bulgaria is low. However, the rate of HCV-HIV-coinfection and HCV infection is high, especially among high-risk communities. The molecular epidemiology of those infections has not been studied before.

Consensus Sanger sequences of HVR1 and NS5B from 125 cases of HIV/HCV coinfections, collected during 2010–2014 in 15 different Bulgarian cities, were used for preliminary phylogenetic evaluation. Next-generation sequencing (NGS) data of the hypervariable region 1 (HVR1) analyzed via the Global Hepatitis Outbreak and Surveillance Technology (GHOST) were used to evaluate genetic heterogeneity and possible transmission linkages. Links between pairs that were below and above the established genetic distance threshold, indicative of transmission, were further examined by generating k-step networks.

Preliminary genetic analyses showed predominance of HCV genotype 1a (54%), followed by 1b (20.8%), 2a (1.4%), 3a (22.3%) and 4a (1.4%), indicating ongoing transmission of many HCV strains of different genotypes. NGS of HVR1 from 72 cases showed significant genetic heterogeneity of intra-host HCV populations, with 5 cases being infected with 2 different genotypes or subtypes and 6 cases being infected with 2 strains of same subtype. GHOST revealed 8 transmission clusters involving 30 cases (41.7%), indicating a high rate of transmission.

Four transmission clusters were found in Sofia, three in Plovdiv, and one in Peshtera. The main risk factor for the clusters was injection drug use. Close genetic proximity among HCV strains from the 3 Sofia clusters, and between HCV strains from Peshtera and one of the two Plovdiv clusters confirms a long and extensive transmission history of these strains in Bulgaria.

Identification of several HCV genotypes and many HCV strains suggests a frequent introduction of HCV to the studied high-risk communities. GHOST detected a broad transmission network, which sustains circulation of several HCV strains since their early introduction in the 3 cities. This is the first report on the molecular epidemiology of HIV/HCV coinfections in Bulgaria.

• Adult
• Bulgaria/epidemiology
• Coinfection/virology
• Disease Outbreaks
• Female
• Genotype
• HIV Infections/epidemiology
• Hepacivirus/genetics
• Hepacivirus/pathogenicity
• Hepatitis C/epidemiology
• Hepatitis C/transmission
• High-Throughput Nucleotide Sequencing
• Humans
• Male
• Molecular Epidemiology
• Phylogeny
• Risk Factors
• Sex Workers
• Sexual and Gender Minorities
• Substance Abuse, Intravenous/epidemiology

• HCV
• Prophylactic Vaccination
• Vaccines
• Viral Hepatitis

• HCV genotypes
• next-generation sequencing
• treatment

• Adult
• Antiviral Agents/therapeutic use
• Female
• Genetic Variation
• Genetics, Population
• Genotype
• Hepacivirus/classification
• Hepacivirus/genetics
• Hepacivirus/isolation & purification
• Hepatitis C, Chronic/drug therapy
• Hepatitis C, Chronic/virology
• High-Throughput Nucleotide Sequencing
• Humans
• Interferons/therapeutic use
• Longitudinal Studies
• Male
• Middle Aged
• Quasispecies
• Ribavirin/therapeutic use
• Selection, Genetic
• Young Adult

Extraordinary genetic diversity is a hallmark of hepatitis C virus (HCV). Therefore, accurate measurement of the breadth of antibody neutralizing activity across diverse HCV isolates is key to defining correlates of immune protection against the virus, and essential to guide vaccine development. Panels of HCV pseudoparticle (HCVpp) or replication-competent cell culture viruses (HCVcc) can be used to measure neutralizing breadth of antibodies. These in vitro assays have been used to define neutralizing breadth of antibodies in serum, to characterize broadly neutralizing monoclonal antibodies, and to identify mechanisms of HCV resistance to antibody neutralization. Recently, larger and more diverse panels of both HCVpp and HCVcc have been described that better represent the diversity of circulating HCV strains, but further work is needed to expand and standardize these neutralization panels.

• Flaviviridae
• antibodies
• hepatitis C virus
• neutralizing breadth
• viral diversity

• Cell culture
• Chimeric HCV
• DAA
• Direct acting antivirals
• Full-length HCV
• HCV
• HCV treatment in vitro
• HCV vaccine
• HCVcc
• Hepatitis C virus
• In vitro
• Neutralizing antibodies
• Pseudo-particle
• Replicon

Virally induced liver cancer usually evolves over long periods of time in the context of a strongly oxidative microenvironment, characterized by chronic liver inflammation and regeneration processes. They ultimately lead to oncogenic mutations in many cellular signaling cascades that drive cell growth and proliferation. Oxidative stress, induced by hepatitis viruses, therefore is one of the factors that drives the neoplastic transformation process in the liver. This review summarizes current knowledge on oxidative stress and oxidative stress responses induced by human hepatitis B and C viruses. It focuses on the molecular mechanisms by which these viruses activate cellular enzymes/systems that generate or scavenge reactive oxygen species (ROS) and control cellular redox homeostasis. The impact of an altered cellular redox homeostasis on the initiation and establishment of chronic viral infection, as well as on the course and outcome of liver fibrosis and hepatocarcinogenesis will be discussed The review neither discusses reactive nitrogen species, although their metabolism is interferes with that of ROS, nor antioxidants as potential therapeutic remedies against viral infections, both subjects meriting an independent review.

• carcinogenesis
• hepatitis B virus
• hepatitis C virus
• pathogenesis
• reactive oxygen species

• error-prone PCR
• genetic diversity
• hepatitis E virus
• mutant cloud
• phenotype
• quasispecies
• random mutagenesis
• reverse genetics

• HCV
• Hepatitis
• Host-pathogen interaction
• Immune evasion
• Immunogenetics
• Infection

Hepatitis C virus (HCV) is a major cause of end-stage liver diseases. With 3–4 million new HCV infections yearly, a vaccine is urgently needed. A better understanding of virus escape from neutralizing antibodies and their corresponding epitopes are important for this effort. However, for viral isolates with high antibody resistance, or antibodies with moderate potency, it remains challenging to induce escape mutations in vitro. Here, as proof-of-concept, we used antibody-sensitive HVR1-deleted (ΔHVR1) viruses to generate escape mutants for a human monoclonal antibody, AR5A, targeting a rare cross-genotype conserved epitope. By analyzing the genotype 1a envelope proteins (E1/E2) of recovered Core-NS2 recombinant H77/JFH1_ΔHVR1 and performing reverse genetic studies we found that resistance to AR5A was caused by substitution L665W, also conferring resistance to the parental H77/JFH1. The mutation did not induce viral fitness loss, but abrogated AR5A binding to HCV particles and intracellular E1/E2 complexes. Culturing J6/JFH1_ΔHVR1 (genotype 2a), for which fitness was decreased by L665W, with AR5A generated AR5A-resistant viruses with the substitutions I345V, L665S, and S680T, which we introduced into J6/JFH1 and J6/JFH1_ΔHVR1. I345V increased fitness but had no effect on AR5A resistance. L665S impaired fitness and decreased AR5A sensitivity, while S680T combined with L665S compensated for fitness loss and decreased AR5A sensitivity even further. Interestingly, S680T alone had no fitness effect but sensitized the virus to AR5A. Of note, H77/JFH1_L665S was non-viable. The resistance mutations did not affect cell-to-cell spread or E1/E2 interactions. Finally, introducing L665W, identified in genotype 1, into genotypes 2–6 parental and HVR1-deleted variants (not available for genotype 4a) we observed diverse effects on viral fitness and a universally pronounced reduction in AR5A sensitivity. Thus, we were able to take advantage of the neutralization-sensitive HVR1-deleted viruses to rapidly generate escape viruses aiding our understanding of the divergent escape pathways used by HCV to evade AR5A.

Worldwide hepatitis C virus (HCV) is one of the leading causes of chronic liver diseases, including cirrhosis and cancer. Treatment accessibility is limited and development of a preventive vaccine has proven difficult, partly due to the high mutation rate of the virus. Recent studies of HCV antibody neutralization resistance have revealed important information about escape pathways and barriers to escape for several clinically promising human monoclonal antibodies. However, due to the varying levels of antibody shielding between HCV isolates these studies have been mostly limited to a few neutralization-sensitive HCV isolates. Here, we took advantage of the fact that deletion of the hypervariable region 1 (HVR1) increased antibody sensitivity of HCV isolates by increasing the exposure of important epitopes, thus facilitating studies of antibody escape for neutralization resistant isolates. We identified escape mutations in the envelope glycoprotein E2, at amino acid position L665, which conferred antibody resistance in parental HCV viruses from genotypes 1–6. We found that antibody escape was associated with loss of binding to HCV particles and intracellular envelope protein complexes. We also identified escape substitutions at L665 that were isolate-specific. Thus, our data sheds new light on antibody resistance mechanisms across diverse HCV isolates.

• Antibodies, Monoclonal/immunology
• Antibodies, Neutralizing/immunology
• Antibodies, Viral/immunology
• Cell Line
• Epitopes, B-Lymphocyte/immunology
• Hepacivirus/immunology
• Hepatitis C/immunology
• Humans
• Immune Evasion/immunology
• Immunoprecipitation
• Mutation
• Viral Envelope Proteins/genetics
• Viral Envelope Proteins/immunology
• Viral Proteins/genetics
• Viral Proteins/immunology

• R01 AI079031 NIAID NIH HHS
• R01 AI106005 NIAID NIH HHS
• R01 AI123365 NIAID NIH HHS
• U19 AI123861 NIAID NIH HHS
• Faculty of Health and Medical Sciences, University of Copenhagen
• Danish Council of Independent Research
• Lundbeckfonden
• Novo Nordisk Foundation
• Danish Council for Independent Research
• National Institutes of Health

• Animal models
• Antivirals
• Evolution
• Genotypes
• HCV
• MicroRNA
• Neutralizing antibodies
• Phylogeny
• Receptors
• Vaccine

• HCV GT1a and GT1b
• RNA structure
• T- and B-cell epitopes
• conservation
• protein secondary structure
• selective pressure

• antigens
• envelope proteins
• hepatitis C
• peptides
• vaccine
• virus

• Adolescent
• Adult
• Aged
• Antiviral Agents/therapeutic use
• Drug Resistance, Viral
• Female
• Hepatitis B virus/classification
• Hepatitis B virus/drug effects
• Hepatitis B virus/genetics
• Hepatitis B virus/isolation & purification
• Hepatitis B, Chronic/drug therapy
• Hepatitis B, Chronic/virology
• Humans
• Male
• Middle Aged
• Mutation/drug effects
• Telbivudine
• Thymidine/analogs & derivatives
• Thymidine/therapeutic use
• Young Adult

Bovine viral diarrhea virus (BVDV), a Pestivirus in the family Flaviviridae, is an economically important pathogen of cattle worldwide. The primary propagators of the virus are immunotolerant persistently infected (PI) cattle, which shed large quantities of virus throughout life. Despite the absence of an acquired immunity against BVDV in these PI cattle there are strong indications of viral variability that are of clinical and epidemiological importance. In this study the variability of E2 and NS5B sequences in multiple body compartments of PI cattle were characterized using clonal sequencing. Phylogenetic analyses revealed that BVDV exists as a quasispecies within PI cattle. Viral variants were clustered by tissue compartment significantly more often than expected by chance alone with the central nervous system appearing to be a particularly important viral reservoir. We also found strong indications for a genetic bottleneck during vertical transmission from PI animals to their offspring. These quasispecies analyses within PI cattle exemplify the role of the PI host in viral propagation and highlight the complex dynamics of BVDV pathogenesis, transmission and evolution.

Genetic and cellular studies have shown that the host’s innate and adaptive immune responses are an important correlate of viral infection outcome. The features of the host’s immune response (host resistance) reflect the coevolution between hosts and pathogens that has occurred over millennia, and that has also resulted in a number of strategies developed by viruses to improve fitness and survival within the host (viral adaptation). In this review, we discuss viral adaptation to host immune pressure via protein–protein interactions and sequence-specific mutations. Specifically, we will present the “state of play” on viral escape mutations to host T-cell responses in the context of the hepatitis C virus, and their influence on infection outcome.

• adaptive immune response
• hepatitis C virus
• immune escape
• viral adaptation

• 454 sequencing
• Clonal sequencing
• Hepatitis C virus
• NS3
• Next-generation sequencing
• Telaprevir
• Viral variants

The population of influenza virus consists of a huge variety of variants, called quasispecies, due to error-prone replication. Previously, we reported that progeny virions of influenza virus become infected to adjacent cells via cell-to-cell transmission pathway in the presence of oseltamivir. During cell-to-cell transmission, viruses become infected to adjacent cells at high multiplicity since progeny virions are enriched on plasma membrane between infected cells and their adjacent cells. Co-infection with viral variants may rescue recessive mutations with each other. Thus, it is assumed that the cell-to-cell transmission causes expansion of virus quasispecies. Here, we have demonstrated that temperature-sensitive mutations remain in progeny viruses even at non-permissive temperature by co-infection in the presence of oseltamivir. This is possibly due to a multiplex infection through the cell-to-cell transmission by the addition of oseltamivir. Further, by the addition of oseltamivir, the number of missense mutation introduced by error-prone replication in segment 8 encoding NS1 was increased in a passage-dependent manner. The number of missense mutation in segment 5 encoding NP was not changed significantly, whereas silent mutation was increased. Taken together, we propose that oseltamivir expands influenza virus quasispecies via cell-to-cell transmission, and may facilitate the viral evolution and adaptation.

• Antiviral Agents/isolation & purification
• Antiviral Agents/pharmacology
• Garcinia mangostana/chemistry
• Hepacivirus/drug effects
• Hepacivirus/physiology
• Humans
• Microbial Sensitivity Tests
• Plant Extracts/isolation & purification
• Plant Extracts/pharmacology
• Virus Replication/drug effects
• Xanthones/isolation & purification
• Xanthones/pharmacology

• Hepatitis C virus
• Molecular evolution
• Vertical transmission

The disproportionate imbalance between the systemic manifestation of reactive oxygen species and body’s ability to detoxify the reactive intermediates is referred to as oxidative stress. Several biological processes as well as infectious agents, physiological or environmental stress, and perturbed antioxidant response can promote oxidative stress. Oxidative stress usually happens when cells are exposed to more electrically charged reactive oxygen species (ROS) such as H2O2 or O2-. The cells’ ability to handle such pro-oxidant species is impeded by viral infections particularly within liver that plays an important role in metabolism and detoxification of harmful substances. During liver diseases (such as hepatocellular or cholestatic problems), the produced ROS are involved in transcriptional activation of a large number of cytokines and growth factors, and continued production of ROS and Reactive Nitrogen Species (RNS) feed into the vicious cycle. Many human viruses like HCV are evolved to manipulate this delicate pro- and antioxidant balance; thus generating the sustainable oxidative stress that not only causes hepatic damage but also stimulates the processes to reduce treatment of damage. In this review article, the oxidant and antioxidant pathways that are perturbed by HCV genes are discussed. In the first line of risk, the pathways of lipid metabolism present a clear danger in accumulation of viral induced ROS. Viral infection leads to decrease in cellular concentrations of glutathione (GSH) resulting in oxidation of important components of cells such as proteins, DNA and lipids as well as double strand breakage of DNA. These disorders have the tendency to lead the cells toward cirrhosis and hepatocellular carcinoma in adults due to constant insult. We have highlighted the importance of such pathways and revealed differences in the extent of oxidative stress caused by HCV infection.

About sixty thousand new cases of Hepatitis C virus (HCV) infection are recorded in Brazil each year. These cases are currently treated with pegylated interferon (PEG-IFN) and ribavirin (RBV) with an overall success rate of 50%. New compounds for anti-HCV therapy targeted to the HCV NS3 protease are being developed and some already form the components of licensed therapies. Mapping NS3 protease resistance mutations to protease inhibitors or anti-viral drug candidates is important to direct anti-HCV drug treatment.

Sequence analysis of the HCV NS3 protease was conducted in a group of 68 chronically infected patients harboring the HCV genotype 1. The patients were sampled before, during and after a course of PEG-IFN-RBV treatment.

Resistance mutations to the protease inhibitors, Boceprevir and Telaprevir were identified in HCV isolated from three patients (4.4%); the viral sequences contained at least one of the following mutations: V36L, T54S and V55A. In one sustained virological responder, the T54S mutation appeared during the course of PEG-IFN and RBV therapy. In contrast, V36L and V55A mutations were identified in virus isolated from one relapsing patient before, during, and after treatment, whereas the T54S mutation was identified in virus isolated from one non-responding patient, before and during the treatment course.

The incidence and persistence of protease resistance mutations occurring in HCV from chronically infected patients in Brazil should be considered when using protease inhibitors to treat HCV disease. In addition, patients treated with the current therapy (PEG-IFN and RBV) that are relapsing or are non-responders should be considered candidates for protease inhibitor therapy.

The quasispecies composition of Hepatitis C virus (HCV) could have important implications with regard to viral persistence and response to interferon-based therapy. The complete NS5A was analyzed to evaluate whether the composition of NS5A quasispecies of HCV 1a/1b is related to responsiveness to combined interferon pegylated (PEG-IFN) and ribavirin therapy.

Viral RNA was isolated from serum samples collected before, during and after treatment from virological sustained responder (SVR), non-responder (NR) and the end-of-treatment responder patients (ETR). NS5A region was amplified, cloned and sequenced. Six hundred and ninety full-length NS5A sequences were analyzed.

This study provides evidence that lower nucleotide diversity of the NS5A region pre-therapy is associated with viral clearance. Analysis of samples of NRs and the ETRs time points showed that genetic diversity of populations tend to decrease over time. Post-therapy population of ETRs presented higher genetic distance from baseline probably due to the bottleneck phenomenon observed for those patients in the end of treatment. The viral effective population of those patients also showed a strong decrease after therapy. Otherwise, NRs demonstrated a continuous variation or stability of effective populations and genetic diversity over time that did not seem to be related to therapy. Phylogenetic relationships concerning complete NS5A sequences obtained from patients did not demonstrate clustering associated with specific response patterns. However, distinctive clustering of pre/post-therapy sequences was observed. In addition, the evolution of quasispecies over time was subjected to purifying or relaxed purifying selection. Codons 157 (P03), 182 and 440 (P42), 62 and 404 (P44) were found to be under positive selective pressure but it failed to be related to the therapy.

These results confirm the hypothesis that a relationship exists between NS5A heterogeneity and response to therapy in patients infected with chronic hepatitis C.

• Adult
• Aged
• Evolution, Molecular
• Female
• Genetic Variation
• Genome, Viral
• Genotype
• Hepacivirus/classification
• Hepacivirus/genetics
• Hepatitis C, Chronic/genetics
• Hepatitis C, Chronic/virology
• Humans
• Interferons
• Interleukins/genetics
• Male
• Middle Aged
• Phylogeny
• Risk Factors
• Sequence Analysis, DNA

• hepatitis c
• oxidative stress
• nrf2/are pathway
• antioxidant defense
• iron homeostasis
• regulation.

• Animals
• Antioxidants/metabolism
• Hepacivirus/physiology
• Hepatitis C, Chronic/complications
• Hepatitis C, Chronic/genetics
• Hepatitis C, Chronic/metabolism
• Hepatitis C, Chronic/virology
• Humans
• Iron Overload/metabolism
• Liver/metabolism
• Liver/virology
• NF-E2-Related Factor 2/metabolism
• Oxidative Stress
• Reactive Oxygen Species/metabolism
• Response Elements
• Signal Transduction

Hepatitis C virus (HCV) is present in the host with multiple variants generated by its error prone RNA-dependent RNA polymerase. Little is known about the initial viral diversification and the viral life cycle processes that influence diversity. We studied the diversification of HCV during acute infection in 17 plasma donors, with frequent sampling early in infection. To analyze these data, we developed a new stochastic model of the HCV life cycle. We found that the accumulation of mutations is surprisingly slow: at 30 days, the viral population on average is still 46% identical to its transmitted viral genome. Fitting the model to the sequence data, we estimate the median in vivo viral mutation rate is 2.5×10⁻⁵ mutations per nucleotide per genome replication (range 1.6–6.2×10⁻⁵), about 5-fold lower than previous estimates. To confirm these results we analyzed the frequency of stop codons (N = 10) among all possible non-sense mutation targets (M = 898,335), and found a mutation rate of 2.8–3.2×10⁻⁵, consistent with the estimate from the dynamical model. The slow accumulation of mutations is consistent with slow turnover of infected cells and replication complexes within infected cells. This slow turnover is also inferred from the viral load kinetics. Our estimated mutation rate, which is similar to that of other RNA viruses (e.g., HIV and influenza), is also compatible with the accumulation of substitutions seen in HCV at the population level. Our model identifies the relevant processes (long-lived cells and slow turnover of replication complexes) and parameters involved in determining the rate of HCV diversification.

Hepatitis C virus (HCV) is a RNA virus that infects over 170 million people across the world. It leads to a chronic infection in the majority of people who are infected (>70%). Most people only discover that they are infected long after initial infection. Thus, it is difficult to study the very early events in infection. Here we study 17 individuals during the earliest possible stages of infection, from before the virus is detectable in the plasma to around 35 days post-infection. We focus on understanding the viral kinetics and the diversification of HCV during this acute phase of infection. During chronic infection HCV is present in the host as a swarm of multiple variants generated by its error prone copying. We studied the early diversification of HCV during acute infection using a new mathematical model of HCV replication. We found that after a phase of fast increase in viral load, accompanied by viral diversification, there is a stabilization of viral load and diversity levels. Using our model, we were able to estimate for the first time the HCV mutation rate during acute infection. We estimated the median in vivo viral mutation rate is 2.5×10⁻⁵ mutations per nucleotide per genome replication (range 1.6–6.2×10⁻⁵), about 5-fold lower than previous estimates. We also used a different approach, based on results of classical genetics, to calculate HCV's mutation rate and obtained consistent results (2.8–3.2×10⁻⁵).

• Acute Disease
• Female
• Hepacivirus/genetics
• Hepatitis C/genetics
• Humans
• Male
• Models, Biological
• Mutation Rate
• Time Factors
• Viral Load/genetics

• U01 AI067854 NIAID NIH HHS
• AI45008 NIAID NIH HHS
• AI028433 NIAID NIH HHS
• R37 AI028433 NIAID NIH HHS
• R01 AI028433 NIAID NIH HHS
• P20 RR018754 NCRR NIH HHS
• U19 AI067854 NIAID NIH HHS
• 8R01-OD011095-21 NIH HHS
• P20-RR018754 NCRR NIH HHS
• AI27767 NIAID NIH HHS
• P30 AI027767 NIAID NIH HHS
• R01 OD011095 NIH HHS
• P30 AI045008 NIAID NIH HHS
• AI67854 NIAID NIH HHS

• hepatitis C virus
• immune evasion
• neutralizing antibodies

• Animals
• Antibodies, Neutralizing/immunology
• Hepacivirus/genetics
• Hepacivirus/immunology
• Hepatitis C/immunology
• Hepatitis C/virology
• Hepatitis C Antibodies/immunology
• Humans
• Immune Evasion

• MC_U130184144 Medical Research Council

Porcine reproductive and respiratory syndrome virus (PRRSV) crosses the placenta during late gestation and productively infects the fetus. Virus replication and cytokine responses were measured in tissues of fetuses recovered at 109–112 days of gestation, just prior to parturition. At the time of recovery, gross anatomical abnormalities were evident in both infected and non-infected fetuses from the infected dams. Virus isolation and immunohistochemistry identified the thymus as the primary site of virus replication. Steady state RT-PCR amplification of inflammatory, Th1 and Th2 cytokines, showed elevated IFN-γ and TNF-α mRNAs in tissues from infected fetuses, which corresponded to elevated cytokine proteins in serum but not amniotic fluid. Further evidence for induction of immunity was found in the hyperplastic response of lymph nodes, which included the development of germinal centers occupied CDw75+ B cells. Collectively, these data support the notion that the immunocompetent fetus is capable of initiating an antiviral response, which is compartmentalized within the infected fetus. Furthermore, fetal pathology may not be a direct result of virus replication in the fetus.