Antonio Madejón

High-Resolution Hepatitis C Virus Subtyping Using NS5B Deep Sequencing and Phylogeny, an Alternative to Current Methods

ABSTRACT Hepatitis C virus (HCV) is classified into seven major genotypes and 67 subtypes. Recent studies have shown that in HCV genotype 1-infected patients, response rates to regimens containing direct-acting antivirals (DAAs) are subtype dependent. Currently available genotyping methods have limited subtyping accuracy. We have evaluated the performance of a deep-sequencing-based HCV subtyping assay, developed for the 454/GS-Junior platform, in comparison with those of two commercial assays (Versant HCV genotype 2.0 and Abbott Real-time HCV Genotype II) and using direct NS5B sequencing as a gold standard (direct sequencing), in 114 clinical specimens previously tested by first-generation hybridization assay (82 genotype 1 and 32 with uninterpretable results). Phylogenetic analysis of deep-sequencing reads matched subtype 1 calling by population Sanger sequencing (69% 1b, 31% 1a) in 81 specimens and identified a mixed-subtype infection (1b/3a/1a) in one sample. Similarly, among the 32 previously indeterminate specimens, identical genotype and subtype results were obtained by direct and deep sequencing in all but four samples with dual infection. In contrast, both Versant HCV Genotype 2.0 and Abbott Real-time HCV Genotype II failed subtype 1 calling in 13 (16%) samples each and were unable to identify the HCV genotype and/or subtype in more than half of the non-genotype 1 samples. We concluded that deep sequencing is more efficient for HCV subtyping than currently available methods and allows qualitative identification of mixed infections and may be more helpful with respect to informing treatment strategies with new DAA-containing regimens across all HCV subtypes.

Lack of hepatitis E virus infection in HIV patients with advanced immunodeficiency or idiopathic liver enzyme elevations.

Summary.u2002 Hepatitis E virus (HEV) is an enterically transmissible RNA agent that causes self‐limited acute hepatitis. Recent reports have highlighted that organ‐transplant recipients may develop chronic hepatitis E and progress to cirrhosis. Similar cases could occur in HIV patients. We have investigated 50 HIV‐infected individuals with CD4 counts <200u2003cells/mm3 and 43 with cryptogenic hepatitis. None of them showed HEV viremia. Thus, HEV infection does not seem to be prevalent in the HIV population and accordingly universal HEV vaccination is not warranted in these patients.

Emerging drugs for hepatitis C.

Background: Chronic hepatitis C virus (HCV) infection remains a global health threat with approximately 200 million carriers worldwide. Current treatment consists of the use of peginterferon (pegIFN)/ribavirin (RBV) for 24 or 48 weeks depending on HCV genotype. Serious side effects and the fact that less than half of patients infected with HCV genotypes 1 and 4 (which are the most common) accomplish sustained virological response with this medication warrant the need for novel anti-HCV therapies. Objective: Description of specifically targeted antiviral therapies for hepatitis C (STAT-C) designed to inhibit the serine protease and the RNA-dependent HCV-RNA polymerase. Methods: Review of available data reported in peer-reviewed journals and medical conferences. Results/conclusions: Early preclinical studies using these compounds produced encouraging results, but the initial enthusiasm has been hampered by toxicity issues and rapid selection of resistance. Therefore, combination therapy with a backbone of pegIFN/RBV, or perhaps in the future using several of these small molecules, preferably having distinct modes of action and resistance profiles, will be required.

Incidence of liver cirrhosis in HIV-infected patients with chronic hepatitis B or C in the era of highly active antiretroviral therapy.

BACKGROUNDnLongitudinal assessment of liver fibrosis with transient elastometry (TE) in patients with chronic viral hepatitis is becoming routine clinical practice in many clinics, as this procedure is non-invasive, easy to perform and relatively inexpensive, allowing early detection of cirrhosis. Herein, we examine the incidence of cirrhosis, using TE assessment, in HIV-infected individuals with chronic hepatitis B or C receiving highly active antiretroviral therapy (HAART).nnnMETHODSnA longitudinal study was performed on a cohort of HIV-infected patients with chronic hepatitis B or C who were followed since 2004 at Hospital Carlos III (Madrid, Spain) with periodic TE assessments. The primary outcome was the development of cirrhosis, defined as liver stiffness >12.5 KPa.nnnRESULTSnA total of 508 HIV-infected patients were examined, of whom 54 developed liver cirrhosis during a mean ±(SD) follow-up of 2.6 ±1.0 years (overall incidence was 41.13 cases per 1,000 person-years). The risk of developing cirrhosis was significantly higher in 297 HCV-RNA-positive patients (either untreated or non-responders to hepatitis C therapy) compared with 55 patients who had cleared HCV with therapy (odds ratio 3.73, 95% confidence interval 1.06-13.17; P=0.04). By contrast, the risk of developing cirrhosis was low and similar in 24 HIV-HBV-coinfected patients under long-term suppressive HBV therapy (mainly tenofovir disoproxil fumarate), 132 HIV-infected patients without chronic liver disease and those who had cleared HCV with therapy.nnnCONCLUSIONSnDevelopment of liver cirrhosis in HIV-infected individuals in the HAART era is mainly associated with active HCV coinfection. The risk of developing cirrhosis is negligible in patients who cleared HCV with therapy, as well as in HIV-HBV-coinfected patients on long-term suppressive tenofovir disoproxil fumarate therapy.

Hepatitis C virus (HCV) treatment uptake and changes in the prevalence of HCV genotypes in HIV/HCV-coinfected patients.

Summary.u2002 The efficacy of current hepatitis C therapy in HIV/HCV‐coinfected patients is largely dependent on HCV genotype. The annual prevalence of HCV genotypes/subtypes and their influence on HCV clearance with antiviral treatment were examined in a dynamic cohort of HIV/HCV‐coinfected patients followed up in Madrid since 2000. Patients entered the cohort at first visit and left the cohort when HCV clearance was achieved with HCV therapy or when follow‐up was interrupted for any reason, including death. A total of 672 HIV/HCV‐coinfected patients constituted the cohort. The mean follow‐up time was 5.5u2003years, corresponding to 4108 patient‐years. Mean age at entry was 37u2003years, and 73% were men and 86% were intravenous drug users. Overall distribution of HCV genotypes was as follows: 57.1% HCV‐1 (1a: 29.2%, 1b: 20.4%, unknown: 7.6%), 1.3% HCV‐2, 25.4% HCV‐3 and 15.9% HCV‐4. A total of 274 (40.8%) patients were treated with peginterferon–ribavirin, of whom 116 (42.3%) achieved HCV clearance following 1–3 courses of therapy. The proportion of HCV‐1/4 rose from 71.7% in 2000 to 76.8% in 2008, whereas the proportion of HCV‐2/3 fell from 28.1% in 2000 to 23.2% in 2008. The yearly prevalence increased for HCV‐1 (R2: 0.92, b: 0.59, Pu2003<u20030.001) and HCV‐4 (R2: 0.77, b: 0.33, Pu2003<u20030.005) and conversely diminished for HCV‐3 (R2: 0.94, b: −0.82, Pu2003<u20030.001). In summary, the prevalence of HCV‐1 and HCV‐4 has increased over the last decade in HIV/HCV‐coinfected patients, whereas conversely it has declined for HCV‐3, in association with the wider use of HCV therapy (41%) in this population.

Hepatitis B, C, and D and HIV infections among immigrants from Equatorial Guinea living in Spain.

A total of 1,220 subjects from Equatorial Guinea living in Spain (median age = 41 years; 453 male and 767 female) was examined for antibodies to human immunodeficiency virus (HIV) and Hepatitis B (HBV), C (HCV), and D (HDV) viruses. Extracted RNA and DNA from the positive samples were used to quantify viral load. The prevalence of HIV antibodies, HCV RNA, and HBV surface antigen (HBsAg) was 10.8% (N = 132), 11.6% (N = 141), and 7.9% (N = 96), respectively. The most prevalent HIV variant was CRF02_AG (38.5%; N = 40). HCV genotype 4 (60%; N = 36) and HBV genotype A3 (32%; N = 8) were the hepatitis variants most frequently found. Superinfection with HDV was seen in 20.9% (N = 24) of HBsAg carriers. A control group of 276 immigrants from other sub-Saharan countries showed similar rates of HIV and HBsAg, although no HCV cases were found. Immigrants constitute a major source of HIV and hepatitis viruses in Spain; therefore, it is important that control measures are intensified.

Increased soluble CD36 is linked to advanced steatosis in nonalcoholic fatty liver disease.

Soluble CD36 (sCD36) clusters with insulin resistance, but no evidence exists on its relationship with hepatic fat content. We determined sCD36 to assess its link to steatosis in nonalcoholic fatty liver disease (NAFLD) and chronic hepatitis C (CHC) patients.

Hepatitis B virus genotype A2 harbours an L217R polymorphism which may account for a lower response to adefovir

Sir, Hepatitis B virus (HBV) infection is a major health problem worldwide. The World Health Organization estimates that more than 400 million people are affected. Chronic hepatitis B frequently leads to end-stage liver disease and is the leading cause of liver cancer worldwide. Current HBV therapies are inadequate to eradicate chronic infection. However, anti-HBV drugs may ameliorate liver disease progression in the short term (with normalization of transaminases, reduction in serum HBV-DNA levels and/or HBeAg seroreversion) and in the long term (with prevention of liver cirrhosis and/or hepatocellular carcinoma). After many years of lamivudine being the only oral drug to treat chronic hepatitis B, marketing of adefovir was very welcome. Moreover, this nucleotide analogue was shown to be also effective in lamivudine-experienced patients, with a relatively low incidence of resistance, at least within the first 2 years of therapy but increasing to � 30% at 5 years. 1 Suboptimal response to adefovir, and subsequent selection of resistance mutations, has been associated with pharmacological issues, mainly derived from the low dose (10 mg daily) approved in an attempt to minimize renal toxicity. Adefovir resistance has been classically linked to a single mutation (rtN236T) within the D domain of the HBV polymerase gene. 2 A number of other mutations, such as V84M and S85A (A domain), A181T/V (B

Detection of hepatitis B virus genotype A3 and primary drug resistance mutations in African immigrants with chronic hepatitis B in Spain

BACKGROUNDnUniversal vaccination and antiviral therapy have reduced chronic hepatitis B virus (HBV) in natives in the Western world. However, immigration from high HBV endemic areas continues to maintain a relatively stable prevalence of chronic hepatitis B in most developed countries.nnnMETHODSnAll foreigners attending a referral infectious diseases department in Madrid, Spain, from January 2007 to December 2008, were evaluated for serum HBV surface antigen (HBsAg). Positive cases underwent further virological characterization.nnnRESULTSnA total of 1718 foreigners were examined, of whom 1322 (77%) were sub-Saharan Africans. Serum HBsAg was positive in 121 (7%), HIV in 135 (7.9%) and hepatitis C virus antibodies in 212 (12.3%). HBV subgenotype A3, which so far had only been reported in people originating from Cameroon, was found in nearly half (14/29) of the tested specimens with detectable serum HBV-DNA. Interestingly, the lamivudine resistance mutation rtM204V was found in two Africans (6.9%), one infected with HBV-A3 and the other with HBV-E. Lack of prior exposure to antiviral therapy in these two patients was confirmed retrospectively.nnnCONCLUSIONSnCirculation of uncommon HBV variants, including strains with primary drug resistance, may follow large immigrant flows from HBV endemic regions to Western countries. Close surveillance of this population is warranted, as early diagnosis and early antiviral therapy may reduce transmission and prevent clinical complications.

Short Communication: Transmission of Hepatitis B Viruses with Lamivudine Resistance Mutations in Newly Diagnosed HIV Individuals

From 1519 newly diagnosed HIV individuals seen in Madrid between the years 2000 and 2008, 65 (4.3%) were HBsAg(+). Two HIV/HBV-coinfected patients showed the lamivudine resistance mutation M204V in HBV while no drug resistance mutations were recognized in HIV. None of them admitted prior exposure to antiretroviral drugs. Thus, HIV/HBV-coinfected patients might benefit from baseline drug resistance testing for both HIV and HBV to optimize the selection of anti-HBV active antiviral therapy.