Hepatitis C Virus Diagnosis Research Articles

Prokaryotic Expression of Hepatitis C Virus-NS3 Protein and Preparation of a Monoclonal Antibody.

Hepatitis C virus (HCV) is a significant health threat that has been extensively investigated worldwide. Improving the sensitivity and specificity of laboratory tests for screening and early diagnosis of HCV in a relevant population is an effective measure to control the spread of HCV. To build a more reliable diagnostic method for HCV, we expressed gene fragments of HCV-NS3 linked to a carrier, pET28a, and then transformed this vector into Escherichia. coli. The produced recombinant NS3 protein with a molecular weight of 38 kDa, which was purified through Ni-chelating affinity chromatography, was used to immunize BALB/C mice, which generated a serum antibody titer of 1:160,000 against the immunogen. Three positive monoclonal isolates (2A5, 2A6, and 5B12) were screened and established. Western blot and enzyme-linked immunosorbent assay (ELISA) results of these monoclonal cells show that each could specifically recognize the recombinant protein. Antibodies 2A5 and 2A6 were developed into an ELISA sandwich antibody pair for the recombinant protein. The detection sensitivity of our developed ELISA was 1.6 ng/mL, with a linear range of 2.5-80 ng/mL (R2 = 0.998). Serum NS3 ELISA results show that the average value in the healthy group, liver disease group, and hepatitis C group was 3.71, 7.28, and 13.11 ng/mL, respectively. The positive rates of HCV-NS3 protein in the liver disease group and hepatitis C group was 17.2% and 41.7%, respectively. Detection of HCV-NS3 antigen can be used as an auxiliary test for anti-HCV antibody detection, thus reducing leakage detection and providing a reliable basis for clinical practice.

Global elimination of hepatitis C virus infection: Progresses and the remaining challenges.

Today, with the introduction of interferon-free direct-acting antivirals and outstanding progresses in the prevention, diagnosis and treatment of hepatitis C virus (HCV) infection, the elimination of HCV infection seems more achievable. A further challenge is continued transmission of HCV infection in high-risk population specially injecting drug users (IDUs) as the major reservoir of HCV infection. Considering the fact that most of these infections remain undiagnosed, unidentified HCV-infected IDUs are potential sources for the rapid spread of HCV in the community. The continuous increase in the number of IDUs along with the rising prevalence of HCV infection among young IDUs is harbinger of a forthcoming public health dilemma, presenting a serious challenge to control transmission of HCV infection. Even the changes in HCV genotype distribution attributed to injecting drug use confirm this issue. These circumstances create a strong demand for timely diagnosis and proper treatment of HCV-infected patients through risk-based screening to mitigate the risk of HCV transmission in the IDUs community and, consequently, in the society. Meanwhile, raising general awareness of HCV infection, diagnosis and treatment through public education should be the core activity of any harm reduction intervention, as the root cause of failure in control of HCV infection has been lack of awareness among young drug takers. In addition, effective prevention, comprehensive screening programs with a specific focus on high-risk population, accessibility to the new anti-HCV treatment regimens and public education should be considered as the top priorities of any health policy decision to eliminate HCV infection.

Management of hepatitis C in decentralised versus centralised drug substitution programmes and minimally invasive point-of-care tests to close gaps in the HCV cascade.

In Switzerland, intravenous drug use accounts for the majority of hepatitis C virus (HCV) infections. Early HCV treatment prevents further transmissions and reduces morbidity and mortality due to decompensated liver cirrhosis and hepatocellular carcinoma. Nevertheless, patients in drug substitution programmes are often insufficiently screened and treated. The aim was to compare the current state of HCV management in centralised and decentralised drug substitution programmes of the canton Aargau. Objectives were human immunodeficiency virus (HIV) and HCV prevalence, compliance with guidelines and gaps in the HCV cascade, as well as feasibility/acceptance/validity of HIV/HCV rapid tests on finger-prick blood and noninvasive liver fibrosis assessment with Fibroscan®. For the cross-sectional study, in June 2013, questionnaires and free rapid tests for HIV (Determine®) and HCV (OraQuick®) that used capillary blood (finger-stick) were sent to 161 physicians providing drug substitution treatment for 631 patients. Free liver fibrosis assessment with Fibroscan® by a member of the study team was offered to all patients. Additionally, patients were directly recruited by the study team in the heroin substitution programme and several addiction clinics visited every 4-6 months, as well as in the Infectious Diseases Outpatient Clinic (questionnaire, rapid tests and Fibroscan® in the same session). Between July 2013 and July 2015, 205 (32.5%) of the 631 patients receiving opioid substitution in the canton Aargau were enrolled, 192 (93.7%) with HIV/HCV rapid tests and 167 (81.5%) with Fibroscan®. Acceptance of Fibroscan® was higher when offered in the same session (94.1 vs 69.2%). Overall, 77.8% had ever used intravenous drugs. HCV seroprevalence was 53.7% (109/203), HCV RNA prevalence 27.8%. Overall, 7.4% (15/202) were HIV infected, all of whom were HCV co-infected and under antiretroviral treatment. Of the 205 patients included, 104 (50.7%) were recruited in a decentralised setting (family practice / pharmacy) and 101 (49.3%) in a centralised setting (heroin programme, addiction clinic, Infectious Diseases Outpatient Clinic). Compliance with guidelines (regular HIV/HCV screening, workup of HCV-positive patients, availability of HAV/HBV serology) was consistently lower in the decentralised setting, characterised by a higher proportion of females, longer median time in the programme, lower percentage of daily attendance, ever-use of intravenous drugs and HIV and HCV infections. We identified several gaps in the HCV cascade: 23.9% (49/205) had never been HCV screened; 18.9% (18/95) of the HCV positive patients had no HCV RNA test. Of the 61 patients developing chronic HCV infection, 19.7% (12) were not HCV genotyped, 52.5% (32) had no liver fibrosis assessment (liver biopsy) and 54.1% (33) never received treatment; 25.0% (7/28) did not achieve a sustained virological response with interferon-based treatment. The 192 HCV rapid tests showed a sensitivity of 90.4% (94/104; 95% confidence interval 84.7-96.1%) and a specificity of 100% (88/88), and provided 14 new HCV diagnoses. Eight of ten patients with a false-negative HCV rapid test were HCV RNA negative (2 unknown). Among the 88.6% (39/44) currently HCV RNA-positive individuals with valid Fibroscan® results, 24 (61.5%) had a liver stiffness <7.5 kPa. Both HIV co-infection and alcohol overconsumption doubled the risk of severe fibrosis/cirrhosis in HCV positive patients. In contrast to HIV, HCV transmission among intravenous drug users is still ongoing. The management of hepatitis C in drug substitution patients needs improvement, especially in family practices. Minimally invasive "point-of-care" diagnostics such as the HCV antibody rapid test using capillary blood and mobile Fibroscan® can close some of the gaps in the HCV cascade. HCV RNA determination in capillary blood is still an unmet need. A "one-stop strategy" might improve linkage to care. Restricting the new, highly efficient (90-100% sustained virological response for all genotypes) direct-acting antivirals to patients with at least stage F2 fibrosis withholds treatment from two thirds of the chronically infected and prevents us from reaching the WHO goal of 80% treatment uptake necessary to eliminate hepatitis C by 2030.

Increasing Prevalence of Hepatitis C among Hospitalized Children Is Associated with an Increase in Substance Abuse

To evaluate the impact of substance abuse on pediatric hepatitis C virus (HCV) prevalence, we examined geographic and demographic data on inpatient hospitalizations in children with HCV. We examined hospitalizations in children using the Kids' Inpatient Database, a part of the Healthcare Cost and Utilization Project. We identified cases using the International Classification of Diseases, 9th edition, codes for HCV infection during 2006, 2009, and 2012. Nonparametric tests for trend were used to calculate trend statistics. From 2006 to 2012 nationally, the number of hospitalizations of children with HCV increased 37% (2.69 to 3.69 per 10 000 admissions; P < .001). The mean age of children hospitalized was 17.6 years (95% CI, 17.4-17.8). HCV cases among those 19-20 years of age represented 68% of the total HCV diagnoses, with a 54% increase over the years sampled (P < .001 for trend). The burden of HCV in children was highest in whites, those in the lowest income quartile, and in the Northeast and Southern regions of the US (all P < .0001). The prevalence of substance use among children with HCV increased from 25% in 2006 to 41% in 2012 (P < .001). The increases of HCV in hospitalized children are largely in teenagers, highly associated with substance abuse, and concentrated in Northeast and Southern states. These results strongly suggest that public health efforts to prevent and treat HCV will also need to include adolescents.

Values, preferences and current hepatitis B and C testing practices in low- and middle-income countries: results of a survey of end users and implementers

BackgroundAccess to hepatitis B virus (HBV) and hepatitis C virus (HCV) diagnostics remains a key bottleneck in scale-up of access to HBV and HCV treatment, particularly in low- and middle-income countries (LMICs) that lack laboratory resources and skilled personnel. To inform the development of World Health Organization (WHO) testing guidelines on who to test and how to test, we performed a “values and preferences” survey of end users and implementers of hepatitis testing in LMICs on current hepatitis B and C testing practices and acceptability of diagnostic approaches, as well as preferences for the future.MethodsThe survey consisted of a four-part, 28 question online survey tool using SurveyMonkey software. The invitation to participate was sent via email to a network of contacts in hepatitis clinical care, research, advocacy and industry.ResultsThe survey collected responses on current testing practices from 48 respondents in 23 LMICs. Only a small proportion of hepatitis testing is currently funded through government-supported programmes. Most limit their testing programmes to blood donor screening and although testing is recommended in several populations, this is not well implemented. Also, there is still very limited access to virological testing.ConclusionsThe survey showed that HBV and HCV testing programmes in LMICs are inadequate and/or scarce. Lack of affordable diagnostic tests; lack of funding, public education and awareness; absence of national policies and guidelines; and a dearth of skilled health professionals are the most important barriers to scaling up HBV and HCV diagnosis and treatment.

The negative impact of HBV/HCV coinfection on cirrhosis and its consequences.

Hepatitis B virus (HBV)/hepatitis C virus (HCV) confection has been rarely studied in nonasian series. To compare the characteristics of HBV/HCV coinfected patients to those of HBV- or HCV-monoinfected patients in the ANRS CO22 HEPATHER cohort study. Of the 20 936 included patients, 95 had HBV/HCV coinfection (hepatitis B surface antigen, anti-HCV antibody and HCV RNA positive) and were matched with 375 HBV- and 380 HCV-monoinfected patients on age, gender and time since HBV or HCV diagnosis. F3-F4 fibrosis was more frequent in coinfected patients (58%) than in HBV- (32%, P < .0001), but similar in HCV-monoinfected patients (52%, P = .3142). Decompensated cirrhosis was more frequent in coinfected patients (11%) than in HBV- (2%, P = .0002) or HCV- (4%, P = .0275) monoinfected patients. Past excessive alcohol use was more frequent in coinfected patients (26%) than in HBV (12%, P = .0011), but similar in HCV monoinfected patients (32%, P = .2868). Coinfected patients had a higher proportion with arterial hypertension (42%) than HBV- (26%) or HCV-monoinfected patients (25%) (P < .003). Multivariable analysis confirmed the association between F3-F4 fibrosis and HCV infection in HBV-infected patients (OR = 3.84, 95% CI 1.99-7.43) and the association between decompensated cirrhosis and coinfection in HBV infected (OR = 5.58, 95% CI 1.42-22.0) or HCV infected patients (OR = 3.02, 95% CI 1.22-7.44). HCV coinfection harmfully affects liver fibrosis in HBV patients, while decompensated cirrhosis is increased in coinfected patients compared with HBV- or HCV-monoinfected patients. HCV treatment is as safe and effective in coinfected as monoinfected patients and should be considered following the same rules as HCV monoinfected patients.

Rising Rates of Hepatocellular Carcinoma Leading to Liver Transplantation in Baby Boomer Generation with Chronic Hepatitis C, Alcohol Liver Disease, and Nonalcoholic Steatohepatitis-Related Liver Disease.

We aim to study the impact of the baby boomer (BB) generation, a birth-specific cohort (born 1945–1965) on hepatocellular carcinoma (HCC)-related liver transplantation (LT) in patients with chronic hepatitis C virus (HCV), alcoholic liver disease (ALD), and non-alcoholic steatohepatitis (NASH). We performed a retrospective analysis using the United Network for Organ Sharing (UNOS)/Organ Procurement Transplant Network (OPTN) database from 2003 to 2014 to compare HCC-related liver transplant surgery trends between two cohorts—the BB and non-BB—with a secondary diagnosis of HCV, ALD, or NASH. From 2003–2014, there were a total of 8313 liver transplant recipients for the indication of HCC secondary to HCV, ALD, or NASH. Of the total, 6658 (80.1%) HCC-related liver transplant recipients were BB. The number of liver transplant surgeries for the indication of HCC increased significantly in NASH (+1327%), HCV (+382%), and ALD (+286%) during the study period. The proportion of BB who underwent LT for HCC was the highest in HCV (84.7%), followed by NASH (70.3%) and ALD (64.7%). The recommendations for birth-cohort specific HCV screening stemmed from a greater understanding of the high prevalence of chronic HCV and HCV-related HCC within BB. The rising number of HCC-related LT among BB with ALD and NASH suggests the need for increased awareness and improved preventative screening/surveillance measures within NASH and ALD cohorts as well.

Impact of an electronic health record alert in primary care on increasing hepatitis c screening and curative treatment for baby boomers.

Implementation of an electronic health record-based prompt increased HCV screening rates among baby boomers in primary care by 5-fold due to efficiency in determining needs for HCV screening and workflow design. Streamlined access to specialty care enabled patients with previously undiagnosed advanced disease to be cured. This intervention can be easily integrated into electronic health record systems to increase HCV diagnosis and linkage to care. (Hepatology 2017;66:1805-1813).

Universal opt-out screening for hepatitis C virus (HCV) within correctional facilities is an effective intervention to improve public health

PurposeWorldwide efforts to identify individuals infected with the hepatitis C virus (HCV) focus almost exclusively on community healthcare systems, thereby failing to reach high-risk populations and those with poor access to primary care. In the USA, community-based HCV testing policies and guidelines overlook correctional facilities, where HCV rates are believed to be as high as 40 percent. This is a missed opportunity: more than ten million Americans move through correctional facilities each year. Herein, the purpose of this paper is to examine HCV testing practices in the US correctional system, California and describe how universal opt-out HCV testing could expand early HCV detection, improve public health in correctional facilities and communities, and prove cost-effective over time.Design/methodology/approachA commentary on the value of standardizing screening programs across facilities by mandating all facilities (universal) to implement opt-out testing policies for all prisoners upon entry to the correctional facilities.FindingsCurrent variability in facility-level testing programs results in inconsistent testing levels across correctional facilities, and therefore makes estimating the actual number of HCV-infected adults in the USA difficult. The authors argue that universal opt-out testing policies ensure earlier diagnosis of HCV among a population most affected by the disease and is more cost-effective than selective testing policies.Originality/valueThe commentary explores the current limitations of selective testing policies in correctional systems and provides recommendations and implications for public health and correctional organizations.

Increased risk of concurrent hepatitis C among Male patients with schizophrenia

Prior studies attempted to explore the association between schizophrenia and hepatitis C virus (HCV). However, their conclusions were inconsistent. This study aimed to examine the association of schizophrenia with HCV using a population-based dataset in Taiwan. There were 6097 patients with schizophrenia and 6097 sex- and age-matched comparison patients without schizophrenia included in this study. We defined the dependent variable of interest as whether or not a patient had received a diagnosis of HCV. We found that of the sampled patients, 2.1% of patients with schizophrenia and 1.4% of comparison patients had concurrent HCV. We further found that schizophrenia was not significantly associated with concurrent HCV after adjusting for sex, age, urbanization level, geographic region, monthly income, and drug abuse. However, of the sampled male patients, the adjusted odds of concurrent hepatitis C for patients with schizophrenia were 1.72-times higher than the odds of concurrent HCV among comparison patients. We failed to observe this association among female sampled patients. We concluded that schizophrenia was not significantly associated with concurrent HCV. However, of the sampled male patients, the risk of concurrent HCV among patients with schizophrenia was higher than comparison patients.

Severe liver disease related to chronic hepatitis C virus infection in treatment-naive patients: epidemiological characteristics and associated factors at first expert centre visit, France, 2000 to 2007 and 2010 to 2014.

Given recent profound improvements in the effectiveness of antiviral treatment for chronic Hepatitis C virus (HCV) infection, we aimed to describe the characteristics of patients referred to hepatology expert centres in France from 2000 to 2007 and from 2010 to 2014, and to identify factors associated with severe liver disease at their first visit for evaluation. We analysed data from two sources covering all of France: the former hepatitis C surveillance network, which included patients between 2000 and 2007, and the ANRS CO22 HEPATHER multi-centre cohort, which included patients between 2012 and 2014. Severe liver disease (SLD) was defined as the presence of either cirrhosis (histological, biochemical or clinical) or hepatocellular carcinoma. Multivariable Poisson regression models were used to identify the factors associated with SLD in complete-case analysis and after multiple imputation. Overall, 16,851 patients were included in the analysis and SLD was diagnosed in 11.6%. SLD at first visit was significantly associated with known risk factors (male sex, history of excessive alcohol intake, HCV genotype 3), late referral to hepatologists after diagnosis and HCV diagnosis at an older age. Providing earlier specialised care and treatment may be an important target for public health action.

Late hepatitis B and C diagnosis in relation to disease decompensation and hepatocellular carcinoma development

We measured the timing of hepatitis B virus (HBV) and hepatitis C virus (HCV) diagnoses relative to the detection of decompensated cirrhosis (DC) and hepatocellular carcinoma (HCC) as an indicator of late hepatitis diagnosis. HBV and HCV diagnoses were defined relative to the diagnosis of DC or HCC such that HBV/HCV diagnoses within two years prior, at the time of or after HCC or DC diagnosis were considered late. We performed multivariable logistic regression to assess factors associated with late HBV/HCV diagnoses among those with DC or HCC. From 1990 to 2012, 778/32,664 HBV cases (2.4%) and 3,925/57,866 HCV cases (6.8%) developed DC while 628/32,644 HBV cases (1.9%) and 902/57,866 HCV cases (1.6%) developed HCC. Among HBV and HCV cases with DC, 49% and 40% respectively were late diagnoses, as were 46% and 31% of HBV and HCV cases with HCC, respectively. HBV late diagnosis declined from 100% in 1992 to 11% and 26% in 2011, while HCV late diagnosis declined from 100% in 1992 to 16% and 14% in 2011 for DC and HCC respectively. In multivariable modelling, late HBV diagnosis was associated with mental illness and a fewer number of physician visits in the five years prior to HBV diagnosis. Late HCV diagnosis was also associated with fewer physician visits, while those with illicit drug use were less likely to be diagnosed late. The proportion of late diagnoses has declined over time. People with better engagement with the healthcare system and with risk activities were diagnosed earlier. Lay summary: Late diagnosis of HBV and HCV represents a missed opportunity to reduce the risk of serious liver disease. Our results identify successes in earlier diagnosis over time using risk-based testing as well as groups that are being missed for screening such as those who do not see a physician regularly and those with serious mental illness.

The impact of hepatitis C virus infection on 90-day outcomes following major orthopaedic surgery: a propensity-matched analysis.

The impact of hepatitis C virus (HCV) infection on outcomes following major orthopaedic interventions, such as joint arthroplasty or spine surgery, has not been effectively studied in the past. Most prior studies are impaired by small samples, limited surveillance for adverse events, or the potential for selection bias to confound results. In this context, we sought to evaluate the impact of HCV infection on 90-day outcomes following joint arthroplasty or spine surgery using propensity-matched techniques. This study utilized 2006-2014 claims from TRICARE insurance. Adults who received spine surgical procedures, total knee and hip arthroplasty were identified. Covariates included demographic factors, a diagnosis of HCV and medical co-morbidities defined by International Classification of Disease-9th revision (ICD-9) code. Outcomes consisted of 30- and 90-day mortality, complications and readmission. A propensity score was used to balance the cohorts with logistic regression techniques employed to determine the influence of HCV infection on post-operative outcomes. The propensity-matched cohort consisted of 2262 patients (1131 with and without HCV). Following logistic regression, patients with HCV were found to have increased odds of 30-day complications (OR 1.87; 95% CI 1.33, 2.64; p<0.001), 90-day complications (OR 1.55; 95% CI 1.16, 2.08; p=0.003) and 30-day readmission (OR 1.46; 95% CI 1.04, 2.05; p=0.03). HCV infection was found to increase the risk of complication and readmission following spine surgery and total joint arthroplasty. Patients should be counseled on their increased risk prior to surgery. Health systems that treat a higher percentage of patients with HCV need to consider the increased risk of complications and readmission when negotiating with insurance carriers.

Access to treatment for Hepatitis C among injection drug users: results from the cross-sectional HOPE IV study

BackgroundEvidence suggests that Greece is among the European countries with increased trend in HCV prevalence among injecting drug users (IDUs) from 2008 to 2014. Nonetheless, the access of IDUs to treatment for Hepatitis C Virus (HCV) is very limited while the risk of co-infection and transmission remains high. In an effort to better understand the inhibitors to HCV treatment, the present study aimed to investigate the main barriers to access in a sample of IDUs.MethodsThe cross-sectional study was carried out between July and September 2015 using a 23-items questionnaire. Participants were recruited from urban primary services, mobile health vans, community health services, day-care centers as well as during street work, located in Athens, Greece. Inclusion criteria were age above 18 years, understanding and speaking Greek sufficiently, HCV diagnosis, intravenous drug use. Data collection was carried out by health professionals of Praksis, a non-governmental organization. For the comparisons of proportions chi-square and Fisher’s exact tests were used.ResultsThe study sample consisted of 101 HCV patients, 68% male. More than 80% of study participants experienced barriers in accessing their doctor and medication during the past 12 months. The most common obstacles in accessing a doctor were “delay in making the appointment and “difficulties in going to the doctor due to health condition or lack of means of transport”. Access to physician or medication was not differed according to gender, but significant differences were found according to economic status and health insurance coverage. 56.1% of participants reported loss or treatment delay due to barriers to treatment. The majority of participants had deteriorated financial status, health status, access to health services and medication, higher financial burden for health services, worse mental health and lower adherence to medical instructions in 2015 compared to 2009.ConclusionsThe findings from the present study revealed that the vast majority of IDUs experience significant barriers in seeking HCV care in Greece, thus highlighting the need for immediate action in this particular area due to the high risk of co-infection and transmission.

P109 Routine hepatitis C antibody testing in MSM – are we overtesting?

IntroductionThe hepatitis C virus (HCV) is rarely transmitted sexually. MSM with HIV are at increased transmission risk. Debate exists regarding sexual transmissibility of HCV in those without HIV or additional risk factors beyond receptive anal intercourse. Following outbreaks of HCV in Europe and London in MSM, Oxfordshire Sexual Health Services introduced annual unselected HCV antibody testing as a screening minimum for all MSM. Evidence now suggests this may not be necessary. We set out to audit our HCV testing to assess this and identify potential policy modification.MethodsWe reviewed all HCV antibody tests undertaken in a 12 month period. We identified all HCV positive patients to determine risk factors for infection in order to establish whether these patients were identified through annual screening or would have been identified using a selective basic risk analysis.ResultsWe found 13 positive results out of 1351 tests. 6 had previously known HCV, 4 were co-infected with HIV. 2 were heterosexual men with additional risk factors, one was an MSM with additional risk factors. No HIV negative MSM with HCV infection were identified through annual screening alone. Approximately 3.5% of tests undertaken were based on recognised risk factors for HCV, 96.5% were undertaken as part of annual screening. This equated to £1486 per new diagnosis, excluding service costs.DiscussionRoutine annual screening of HIV negative MSM in this study did not pick up any new HCV diagnoses. Cost per diagnosis may be reduced with targeted testing. The annual screening policy needs modification.

HIV-positive men who have sex with men are at high risk of development of significant liver fibrosis after an episode of acute hepatitis C.

Acute hepatitis C virus infection remains a major health concern in human immunodeficiency virus(HIV)-infected men who have sex with men (MSM). New direct-acting antiviral agent (DAA) combination therapy has not yet been approved for the treatment for acute hepatitis C virus(HCV), thereby potentially causing deferral of HCV treatment. Therefore, we aimed to study the course of liver disease after an episode of acute HCV. This study is a retrospective single-centre cohort of HIV-positive MSM with acute HCV infection. Liver fibrosis was estimated by Fibroscan® and Fibrotest® . Liver-related and non-liver-related outcomes were documented. Overall 213 episodes of acute HCV infection in 178 men were documented. Median follow-up for all included patients was 38.7months. Spontaneous HCV clearance was found in 10.8% of patients, which was significantly associated with older age, lower HCV RNA levels, and higher ALT levels upon initial acute HCV diagnosis. Treatment with interferon-based therapy was initiated in 86.3% of cases, resulting in a sustained virological response(SVR) rate of 70.7%. After 3years' follow-up, significant liver fibrosis of METAVIR F2 stage or higher was found in 39.4% of patients after first acute HCV diagnosis. Higher age, physician-declared alcoholism, and nonresponse to acute HCV therapy were independently associated with higher fibrosis stages. Ten patients died during the observation period (IR 1.4/100 patient-years) and four during interferon treatment. Significant liver fibrosis is a common finding in HIV-positive MSM following acute HCV infection despite high treatment uptake and cure rates, suggesting the need for close liver disease monitoring particularly if HCV treatment is deferred.

Internal control for real-time polymerase chain reaction based on MS2 bacteriophage for RNA viruses diagnostics.

BACKGROUND Real-time reverse transcription polymerase chain reaction (RT-PCR) is routinely used to detect viral infections. In Brazil, it is mandatory the use of nucleic acid tests to detect hepatitis C virus (HCV), hepatitis B virus and human immunodeficiency virus in blood banks because of the immunological window. The use of an internal control (IC) is necessary to differentiate the true negative results from those consequent from a failure in some step of the nucleic acid test.OBJECTIVES The aim of this study was the construction of virus-modified particles, based on MS2 bacteriophage, to be used as IC for the diagnosis of RNA viruses.METHODS The MS2 genome was cloned into the pET47b(+) plasmid, generating pET47b(+)-MS2. MS2-like particles were produced through the synthesis of MS2 RNA genome by T7 RNA polymerase. These particles were used as non-competitive IC in assays for RNA virus diagnostics. In addition, a competitive control for HCV diagnosis was developed by cloning a mutated HCV sequence into the MS2 replicase gene of pET47b(+)-MS2, which produces a non-propagating MS2 particle. The utility of MS2-like particles as IC was evaluated in a one-step format multiplex real-time RT-PCR for HCV detection.FINDINGS We demonstrated that both competitive and non-competitive IC could be successfully used to monitor the HCV amplification performance, including the extraction, reverse transcription, amplification and detection steps, without compromising the detection of samples with low target concentrations. In conclusion, MS2-like particles generated by this strategy proved to be useful IC for RNA virus diagnosis, with advantage that they are produced by a low cost protocol. An attractive feature of this system is that it allows the construction of a multicontrol by the insertion of sequences from more than one pathogen, increasing its applicability for diagnosing different RNA viruses.

Incidence of Hepatitis C Virus Infection in the Human Immunodeficiency Virus Outpatient Study Cohort, 2000-2013.

There are few recent studies of incident hepatitis C virus (HCV) infection among human immunodeficiency virus (HIV)-infected patients in the United States. We studied HIV Outpatient Study (HOPS) participants seen in 9 HIV-specialty clinics who had ≥1 clinical encounter during 2000-2013 and ≥2 HCV-related tests, the first of which was a negative HCV antibody test (Ab). Hepatitis C virus incident cases were identified by first positive HCV Ab, viral load, or genotype. We assessed rates of incident HCV overall, by calendar intervals, and by demographic and HIV risk strata, and we explored risk factors for incident HCV using Cox proportional hazards models. The 1941 eligible patients (median age 40 years, 23% female, 61% men who had sex with men [MSM], and 3% persons who injected drugs [PWID]) experienced 102 (5.3%) incident HCV infections for an overall incidence of 1.07 (95% confidence interval [CI], 0.87-1.30) per 100 person-years (py). Hepatitis C virus incidence decreased from 1.83 in 2000-2003 to 0.88 in 2011-2013 (P = .024), with decreases observed (P < .05) among PWID and heterosexuals, but not among MSM. Overall, MSM comprised 59% of incident cases, and PWID were at most risk for incident HCV infection (adjusted hazard ratio [aHR] for PWID = 4.62 and 95% CI = 2.11-10.13; for MSM, aHR = 1.48 and 95% CI = 0.86-2.55 compared with heterosexuals). Among HIV-infected patients in care during 2000-2013, incidence of HCV infection exceeded 1 case per 100 py. Our findings support recommendations for annual HCV screenings for HIV-infected persons, including persons with only MSM risk, to enable HCV diagnosis and treatment for coinfected individuals.

Comparing Child-Pugh, MELD, and FIB-4 to Predict Clinical Outcomes in Hepatitis C Virus-Infected Persons: Results From ERCHIVES.

Identifying hepatitis C virus (HCV)-positive persons at high risk of early complications can help prioritize treatment decisions. We conducted this study to compare Child-Turcotte-Pugh (CP), MELD, and FIB-4 scores for predicting clinical outcomes and to identify those at low risk of complications. Within electronically retrieved cohort of HCV-infected veterans, we identified HCV-positive persons and excluded those with human immunodeficiency virus (HIV), hepatitis B surface antigen (HBsAg), prevalent hepatic decompensation (HD), hepatocellular carcinoma (HCC), and those treated for HCV. We calculated incidence rates for HD, HCC, and all-cause mortality at 1, 3, and 5 years after HCV diagnosis. Using receiver operating characteristic (ROC) curves, we determined the optimal cut-off values for each score for these outcomes. Among 21 116 persons evaluated, 89.7% were CP Class-A, 79.9% had MELD<9, and 43.4% had FIB-4<1.45. AUROC for HD at 1, 3, and 5 years was higher for FIB-4 (0.84-0.86) compared with MELD (0.70-0.76) (P < .001). AUROC for HCC at 1, 3, and 5 years was 0.81-0.82 for FIB-4 but 0.61-0.68 for CP and MELD scores. (P < .001) AUROC for all-cause mortality at 3 and 5 years was 0.65-0.68. The optimal cut-off scores to identify persons at low risk of complications were as follows: CP <5; MELD <8; FIB-4 <3 for HD and HCC, and <2 for all-cause mortality, below which <1.5% developed HD and HCC and ≤2.5% died at 3 years. FIB-4 score is a better predictor of HD and HCC in HCV-positive persons. A score of <3 is associated with a low risk of HD and HCC 1 and 3 years after HCV diagnosis.

Evaluation of A Reverse Transcriptase (Rt) Loop Mediated Isothermal Amplification Assay for Detection of Hepatitis C Genotypes 1-4 Viruses Under Limited Logistical Conditions

The loop-amplification mediated isothermal amplification (LAMP) presents characteristics that overcome the limitations associated with the Current limitations of nucleic acid based technologies (NATs) hinder their use in the low resource settings hence overreliance on serological assays which miss hepatitis C virus (HCV) during the long seroconversion period. The LAMP assay would be ideal for early detection of HCV in routine diagnostics and blood transfusion setups in such settings drastically reducing transmission related transfusion. This study validated and tested a reverse transcriptase-LAMP for HCV detection under limited logistical conditions in a low resource setting. Under stringent laboratory conditions, analytical sensitivity and reproducibility were performed using a panel of HCV positive plasma of genotypes 1a, 1b, mixed 1a/1b, 2b, 3a and 4. Cell culture supernatants of HIV-1 B and plasma samples for Hepatitis B virus were used for specificity testing. Upto 227 plasma including 70 (40 RNA positive and 30 negative) from German patients and 157 (43 RNA positive and 114 negative) from Kenyan patients were tested. Kenyan samples were obtained from 121 sero-positive plasma screened from 1121 participants of various cohorts in Kenya. Although LAMP detected upto 102 IU/mL for genotypes 1a, 1b and 2b, a lower detection threshold was established at 103 IU/mL. Overall sensitivity was 94% (PPV 98%) and specificity was 98% (NPV 96%) for RT-LAMP. Sub optimal detection was noted for genotypes 2b, 3a and 4. Sequence analysis revealed mismatches affecting stringency of primer binding at the F1, B1 and the LB primer targets. RT-LAMP shows potential for early HCV diagnosis and screening in low resource settings. Its robustness is however genotype dependent and can be enhanced by designing primers targeting circulating and suspected genotypes. More studies should be done on the possibility of designing multiplex RT-LAMP primers to capture a wide variety of genotypes. The assay remains simple, rapid, and cost effective for nucleic acid detection and is ideal for use in the limited resource settings.