Frédéric Le Gal

Eighth major clade for hepatitis delta virus.

Hepatitis delta virus is the only representative of the Deltavirus genus, which consists of 7 differentiated major clades. In this study, an eighth clade was identified from 3 distinct strains. Deltavirus genetic variability should be considered for diagnostic purposes. Clinical consequences of the diversity have yet to be evaluated.

Quantification of Hepatitis Delta Virus RNA in Serum by Consensus Real-Time PCR Indicates Different Patterns of Virological Response to Interferon Therapy in Chronically Infected Patients

ABSTRACT Hepatitis delta virus (HDV), in association with hepatitis B virus, is responsible for severe acute and chronic hepatitis. Treatment of the infection relies on the long-term administration of high doses of alpha interferon (IFN), and the treatment efficiency is monitored by the detection of anti-HDV immunoglobulin M and HDV genome in serum. Like the case for other chronic viral infections, HDV genome quantification in serum should be useful for the follow-up of infected patients. The aims of this study were to develop a quantitative assay for the detection of any type of HDV in serum and to evaluate the benefit of HDV RNA quantification for the follow-up of chronically infected patients receiving IFN. A real-time reverse transcription-PCR assay was developed to quantify the HDV RNA load in serum. Its efficacy was evaluated with 160 serum samples, 76 of which were collected from 11 chronically infected patients who were treated with pegylated IFN. The assay was sensitive (100 copies/ml of serum) and efficient for all HDV types, including type 3 and the recently described types 5, 6, and 7. The viral load determinations for treated patients allowed us to identify different profiles of virological responses to IFN therapy with more accuracy than that attainable with the qualitative approach. In conclusion, we have developed a quantitative HDV RNA assay for serum which is adapted to the follow-up of antiviral treatment for patients infected with any HDV type. The assay will help us to understand the natural history of HDV infection and to define guidelines for the management of chronic delta hepatitis.

Phylogenetic Analyses Indicate an Atypical Nurse-to-Patient Transmission of Human Immunodeficiency Virus Type 1

ABSTRACT A human immunodeficiency virus (HIV)-negative patient with no risk factor experienced HIV type 1 (HIV-1) primary infection 4 weeks after being hospitalized for surgery. Among the medical staff, only two night shift nurses were identified as HIV-1 seropositive. No exposure to blood was evidenced. To test the hypothesis of a possible nurse-to-patient transmission, phylogenetic analyses were conducted using two HIV-1 genomic regions (pol reverse transcriptase [RT] and env C2C4), each compared with reference strains and large local control sets (57 RT and 41 C2C4 local controls). Extensive analyses using multiple methodologies allowed us to test the robustness of phylogeny inference and to assess transmission hypotheses. Results allow us to unambiguously exclude one HIV-positive nurse and strongly suggest the other HIV-positive nurse as the source of infection of the patient.

Resolution of chronic hepatitis Delta after 1 year of combined therapy with pegylated interferon, tenofovir and emtricitabine.

BACKGROUND Hepatitis Delta virus (HDV) Infection has a worldwide distribution, with approximately 20 millions infected persons. Interferon (IFN) is the only approved drug for the treatment of HDV infection which is still a difficult to treat disease. OBJECTIVES To report a successful treatment of a patient with a chronic severe hepatitis Delta using combination therapy with Pegylated interferon (PegIFN), Tenofovir disoproxil fumarate (TDF) and Emtricitabine (FTC). STUDY DESIGN The patient, a 47 years -old male patient, originating from Dagestan (East Asia), suffered of chronic hepatitis Delta infection. The patient was HBsAg, HBeAg, and anti-Delta Ab (IgG) positive. Serum HBV-DNA level was elevated (more than 9 log UI/mL). Serum HDV-RNA level was up to 5.6 log (copies/ml). Genotypes HBV/D and HDV-1 were demonstrated. The liver histology revealed chronic active hepatitis (Metavir score: A2F2). The treatment was started with PegIFN (180 microg/week) for two months and then TDF (300 mg/day) (combined later with FTC) was added. RESULTS Sustained response was obtained after 10 months of treatment and was accompanied by the clearance of serum hepatitis B virus surface antigen with seroconversion to anti-HBs. CONCLUSION This case report suggests that Delta infection may co-exist with high replicative HBV infection and that combination therapy with PegIFN and nucleoside/tide analogues seems to be more effective than IFN alone. Given that only a single case is reported, further studies including more patients are warranted.

First International External Quality Assessment for Hepatitis Delta Virus RNA Quantification in Plasma

Infection by the hepatitis delta virus (HDV), a satellite of the hepatitis B virus (HBV), increases viral liver disease severity. Its diagnosis is thus vital for HBV‐infected patients. HDV‐RNA load (HDVL) should be assessed and monitored in plasma using real‐time reverse‐transcriptase polymerase chain reaction assays. Taking advantage of the recently‐developed World Health Organization (WHO) HDV international standard (WHO‐HDV‐IS), the first international external quality control for HDVL quantification was performed. Two panels of samples were sent to 28 laboratories in 17 countries worldwide. Panel A comprised 20 clinical samples of various genotypes (1, 2, and 5‐8) and viral loads, including two negative controls. Panel B, composed of dilutions of the WHO‐HDV‐IS, allowed the conversion of results from copies/mL into IU/mL for HDVL standardization and interlaboratory comparisons. Comprehensive analysis revealed a very high heterogeneity of assay characteristics, including their technical steps and technologies. Thirteen labs (46.3%) properly quantified all 18 positive samples; 16 (57.1%) failed to detect one to up to 10 samples, and several others underestimated (>3 log IU/mL) HDVL of African genotype strains (1 and 5‐8). Discrepancies were mainly attributed to either primers or probe mismatches related to the high genetic variability of HDV and, possibly, to the complex secondary structure of the target genomic RNA. The labs were grouped in four clusters by the statistical analysis of their performances. The best clusters comprised the 17 labs that obtained the expected HDVL values, including five that otherwise failed to quantify one or two samples. Conclusion: The results of this international quality‐control study underline the urgent need to improve methods used to monitor HDV viremia and will be instrumental in achieving that goal. (Hepatology 2016;64:1483‐1494)

Absence of Hepatitis Delta Infection in a Large Rural HIV Cohort in Tanzania.

OBJECTIVES The epidemiological and clinical determinants of hepatitis delta virus (HDV) infection in Sub-Saharan Africa are ill-defined. The prevalence of HDV infection was determined in HIV/hepatitis B virus (HBV) co-infected individuals in rural Tanzania. METHODS All HBV-infected adults under active follow-up in the Kilombero and Ulanga Antiretroviral Cohort (KIULARCO) were screened for anti-HDV antibodies. For positive samples, a second serological test and nucleic acid amplification were performed. Demographic and clinical characteristics at initiation of antiretroviral therapy (ART) were compared between anti-HDV-negative and -positive patients. RESULTS Among 222 HIV/HBV co-infected patients on ART, 219 (98.6%) had a stored serum sample available and were included in the study. Median age was 37 years, 55% were female, 46% had World Health Organization stage III/IV HIV disease, and the median CD4 count was 179 cells/μl. The prevalence of anti-HDV positivity was 5.0% (95% confidence interval 2.8-8.9%). There was no significant predictor of anti-HDV positivity. HDV could not be amplified in any of the anti-HDV-positive patients and the second serological test was negative in all of them. CONCLUSIONS No confirmed case of HDV infection was found among over 200 HIV/HBV co-infected patients in Tanzania. As false-positive serology results are common, screening results should be confirmed with a second test.

Increasing prevalence of HDV/HBV infection over 15 years in France.

BACKGROUND In France, there are no consistent data estimating hepatitis delta virus (HDV) prevalence in the general population. OBJECTIVES To better characterize HDV/HBV infection and its trends over a 15-years period from 1997 to 2011, we used data retrieved from the National Epidemiological Donors database including viral and demographic characteristics of all French HBV infected blood donors. STUDY DESIGN Of the 39,911,011 donations collected over the 15 year-study-period, 6214 (1.56 in 10(4) donations) were confirmed positive for HBV from which 72.3% were tested for HDV antibodies (Ab). HDV viral load was performed using a real-time PCR assay on positive HDV Ab samples and HDV genotype determined for each positive viremic sample. RESULTS Among the 4492 HBV donations, 89 (1.98%) were HDV Ab positive. After being stable around 1.1% from 1997 to 2005, this rate has continuously increased to reach 6.5% in 2010, before declining to 0.85% in 2011. Of the 61 investigated HDV Ab positive individuals, 22.9% were viremic with a viral load ranging from 10(4) to 9.8 × 10(7) copies mL(-1). Genotyping revealed 12 HDV-1, 1 HDV-6 and 1 HDV-7 in accordance with the geographical origin of individuals. CONCLUSION Such a study gives unexpected features of HBV-HDV infection in the population of blood donors which is a priori, a healthy population. The increase of HDV prevalence mainly linked to migration of population from endemic countries, demonstrates that there is still no complete control of HBV infection and must encourage HBV vaccination campaigns and systematic screening for HDV in HBV-infected.

High Prevalence and Diversity of Hepatitis Viruses in Suspected Cases of Yellow Fever in the Democratic Republic of Congo

ABSTRACT The majority of patients with acute febrile jaundice (>95%) identified through a yellow fever surveillance program in the Democratic Republic of Congo (DRC) test negative for antibodies against yellow fever virus. However, no etiological investigation has ever been carried out on these patients. Here, we tested for hepatitis A (HAV), hepatitis B (HBV), hepatitis C (HCV), hepatitis D (HDV), and hepatitis E (HEV) viruses, all of which can cause acute febrile jaundice, in patients included in the yellow fever surveillance program in the DRC. On a total of 498 serum samples collected from suspected cases of yellow fever from January 2003 to January 2012, enzyme-linked immunosorbent assay (ELISA) techniques were used to screen for antibodies against HAV (IgM) and HEV (IgM) and for antigens and antibodies against HBV (HBsAg and anti-hepatitis B core protein [HBc] IgM, respectively), HCV, and HDV. Viral loads and genotypes were determined for HBV and HVD. Viral hepatitis serological markers were diagnosed in 218 (43.7%) patients. The seroprevalences were 16.7% for HAV, 24.6% for HBV, 2.3% for HCV, and 10.4% for HEV, and 26.1% of HBV-positive patients were also infected with HDV. Median viral loads were 4.19 × 105 IU/ml for HBV (range, 769 to 9.82 × 109 IU/ml) and 1.4 × 106 IU/ml for HDV (range, 3.1 × 102 to 2.9 × 108 IU/ml). Genotypes A, E, and D of HBV and genotype 1 of HDV were detected. These high hepatitis prevalence rates highlight the necessity to include screening for hepatitis viruses in the yellow fever surveillance program in the DRC.

Serological and Molecular Diagnosis of Hepatitis Delta Virus Infection: Results of a French National Quality Control Study

ABSTRACT A French national quality control study for the serological and molecular diagnosis of hepatitis delta virus (HDV) was organized. Total HDV antibodies were properly detected by all laboratories; 8/14 laboratories failed to detect low titers of IgM, and 6/11 failed to quantify and/or underestimated the RNA viral load in several samples. These discrepancies are likely related to the molecular diversity of HDV.

Performance Characteristics of a New Consensus Commercial Kit for Hepatitis D Virus RNA Viral Load Quantification

ABSTRACT Hepatitis D virus (HDV) is responsible for fulminant hepatitis and liver failure and accelerates evolution toward cirrhosis and hepatocellular carcinoma in hepatitis B virus (HBV)-infected patients. To date, treatment relies upon long-term administration of pegylated alpha-interferon with a sustained virological response in 30% of the patients. Very recently, new, promising anti-HDV therapies have been developed and are already being used in clinical trials. HDV RNA viral load (HDVL) monitoring must be an integral part of the management of the infected patients. However, HDV genus is characterized by a high genetic variability into eight genotypes (HDV-1 to -8), and most available in-house or commercial assays are useful for only a limited subset of genotypes. Results of a comparison of the performance of a new kit for HDVL quantification with the consensus in-house assay of the French National Reference Laboratory for HDV developed in 2005 are reported here. A total of 611 clinical samples of all HDV genotypes with various HDVL values, including several consecutive samples over several years from 36 patients, were studied. A specificity, sensitivity, and reproducibility evaluation was conducted using HDV-positive clinical samples, hepatitis A, B, C and E (HAV, HBV, HCV, and HEV, respectively) and HIV mono-infected samples, and the WHO HDV RNA international standard. Overall results were strictly comparable between the two assays (median difference, 0.07 log IU/ml), with high diagnosis precision and capacity. In summary, this new kit showed high performance in detection/quantification of HDVL, regardless of the genotype of the infecting strain used, and seems to be a suitable tool for patient management.