Introduction of an automated system for the diagnosis and quantification of hepatitis B and hepatitis C viruses.

Hepatitis B virus (HBV) and Hepatitis C virus (HCV) infections pose major public health problems because of their prevalence worldwide. Consequently, screening for these infections is an important part of routine laboratory activity. Serological and molecular markers are key elements in diagnosis, prognosis and treatment monitoring for HBV and HCV infections. Today, automated chemiluminescence immunoassay (CLIA) analyzers are widely used for virological diagnosis, particularly in high-volume clinical laboratories. Molecular biology techniques are routinely used to detect and quantify viral genomes as well as to analyze their sequence; in order to determine their genotype and detect resistance to antiviral drugs. Real-time PCR, which provides high sensitivity and a broad dynamic range, has gradually replaced other signal and target ampli���cation technologies for the quantification and detection of nucleic acid. The next-generation DNA sequencing techniques are still restricted to research laboratories.The serological and molecular marker methods available for HBV and HCV are discussed in this article, along with their utility and limitations for use in Chronic Hepatitis B (CHB) diagnosis and monitoring.

Management of hepatitis B virus infection after liver transplantation.

Chronic hepatitis B virus (HBV) infection is responsible for up to 30% of cases of liver cirrhosis and up to 53% of cases of hepatocellular carcinoma. Liver transplantation (LT) is the best therapeutic option for patients with end-stage liver failure caused by HBV. The success of transplantation, though, depends on receiving prophylactic treatment against post-transplant viral reactivation. In the absence of prophylaxis, liver transplantation due to chronic hepatitis B (CHB) is associated with high rates of viral recurrence and poor survival. The introduction of treatment with hepatitis B immunoglobulins (HBIG) during the 1990s and later the incorporation of oral antiviral drugs have improved the prognosis of these patients. Thus, LT for CHB is now a universally accepted option, with an estimated 5 years survival of around 85% vs the 45% survival seen prior to the introduction of HBIG. The combination of lamivudine plus HBIG has for many years been the most widely used prophylactic regimen. However, with the appearance of new more potent oral antiviral agents associated with less resistance (e.g., entecavir and tenofovir) for the treatment of CHB, new prophylactic strategies are being designed, either in combination with HBIG or alone as a monotherapy. These advances have allowed for more personalized prophylaxis based on the individual risk profile of a given patient. In addition, the small pool of donors has required the use of anti-HBc-positive donors (with the resulting possibility of transmitting HBV from these organs), which has been made possible by suitable prophylactic regimens.

Association of Patients' Geographic Origins with Viral Hepatitis Co-infection

To determine if hepatitis C virus seropositivity and active hepatitis B virus infection in HIV-positive patients vary with patients' geographic origins, we studied co-infections in HIV-seropositive adults. Active hepatitis B infection was more prevalent in persons from Africa, and hepatitis C seropositivity was more common in persons from eastern Europe.

Incidence of liver damage of uncertain origin in HIV patients not co-infected with HCV/HBV.

BACKGROUND AND AIMS Several studies have reported that a significant number of HIV patients not co-infected with HCV/HBV develop liver damage of uncertain origin (LDUO). The objective of our study was to evaluate the incidence of and risk factors for the development of LDUO in HIV infected patients not co-infected with HCV/HBV. METHODS Prospective longitudinal study that included HIV-infected patients free of previous liver damage and viral hepatitis B or C co-infections. Patients were followed up at 6-monthly intervals. Liver stiffness was measured at each visit. Abnormal liver stiffness (ALS) was defined as a liver stiffness value greater than 7.2 kPa at two consecutive measurements. For patients who developed ALS, a protocol was followed to diagnose the cause of liver damage. Those patients who could not be diagnosed with any specific cause of liver disease were diagnosed as LDUO and liver biopsy was proposed. RESULTS 210 patients matched the inclusion criteria and were included. 198 patients completed the study. After a median (Q1-Q3) follow-up of 18 (IQR 12-26) months, 21 patients (10.6%) developed ALS. Of these, fifteen patients were diagnosed as LDUO. The incidence of LDUO was 7.64 cases/100 patient-years. Histological studies were performed on ten (66.6%) patients and all showed liver steatosis. A higher HOMA-IR value and body mass index were independently associated with the development of LDUO. CONCLUSION We found a high incidence of LDUO in HIV-infected patients associated with metabolic risk factors. The leading cause of LDUO in our study was non-alcoholic fatty liver disease.

Assisted reproductive technology and obstetric outcome in couples when the male partner has a chronic viral disease.

BACKGROUND Assisted reproductive technology (ART) with washed semen can achieve pregnancy with minimal risk of horizontal and vertical transmission of chronic viral diseases (CVD) such as human immunodeficiency virus (HIV), hepati- tis C virus (HCV) and hepatitis B virus (HBV) among serodiscordant couples. How- ever, few studies have been made of the use made by these couples of ARTs or of the obstetric results achieved. MATERIALS AND METHODS In this retrospective study, 93 men who were seropositive for HIV, HCV or HBV and who underwent assisted reproduction treatment at our centre (Hospital Universitario Virgen de las Nieves, Granada, Spain) were included. Washed semen was tested to detect viral particles. Non-infected women were tested before and after each treatment, as were the neonates at birth and after three months. RESULTS A total of 62 sperm samples were washed, and none were positive for the detec- tion of viral molecules. Semen samples from 34 HBV positive males were not washed since the female partner had immunity to hepatitis B. In total, 38 clinical pregnancies were achieved (22% per cycle and 40.9% per couple) out of 173 cycles initiated, and 28 births were achieved (16.2% per cycle and 30.1% per couple), producing 34 live births. The rate of multiple pregnancies was 21.4%. Obstetric and neonatal results were similar in the groups of couples studied. At follow-up, no seroconversion was detected in the women or neonates. CONCLUSION Sperm washing and intracytoplasmic sperm injection are shown to be a safe and effective option for reducing the risk of transmission or super infection in serodiscordant or concordant couples who wish to have a child. Pregnancies ob- tained by ART in couples when the male is CVD infected achieve good obstetric and neonatal results.

Comparison of oxidative stress markers in HIV-infected patients on efavirenz or atazanavir/ritonavir-based therapy.

INTRODUCTION Chronic low-grade inflammation and immune activation may persist in HIV patients despite effective antiretroviral therapy (ART). These abnormalities are associated with increased oxidative stress (OS). Bilirubin (BR) may have a beneficial role in counteracting OS. Atazanavir (ATV) inhibits UGT1A1, thus increasing unconjugated BR levels, a distinctive feature of this drug. We compared changes in OS markers in HIV patients on ATV/r versus efavirenz (EFV)-based first-line therapies. MATERIALS AND METHODS Cohort of the Spanish Research Network (CoRIS) is a multicentre, open, prospective cohort of HIV-infected patients na��ve to ART at entry and linked to a biobank. We identified hepatitis C virus/hepatitis B virus (HCV/HBV) negative patients who started first-line ART with either ATV/r or EFV, had a baseline biobank sample and a follow-up sample after at least nine months of ART while maintaining initial regimen and being virologically suppressed. Lipoprotein-associated Phospholipase A2 (Lp-PLA2), Myeloperoxidase (MPO) and Oxidized LDL (OxLDL) were measured in paired samples. Marker values at one year were interpolated from available data. Multiple imputations using chained equations were used to deal with missing values. Change in the OS markers was modelled using multiple linear regressions adjusting for baseline marker values and baseline confounders. Correlations between continuous variables were explored using Pearson's correlation tests. RESULTS 145 patients (97 EFV; 48 ATV/r) were studied. Mean (SD) baseline values for OS markers in EFV and ATV/r groups were: Lp-PLA2 [142.2 (72.8) and 150.1 (92.8) ng/mL], MPO [74.3 (48.2) and 93.9 (64.3) ��g/L] and OxLDL [76.3 (52.3) and 82.2 (54.4) ��g/L]. After adjustment for baseline variables patients on ATV/r had a significant decrease in Lp-PLA2 (estimated difference -16.3 [CI 95%: -31.4, -1.25; p=0.03]) and a significantly lower increase in OxLDL (estimated difference -21.8 [-38.0, -5.6; p<0.01] relative to those on EFV, whereas no differences in MPO were found. Adjusted changes in BR were significantly higher for the ATV/r group (estimated difference 1.33 [1.03, 1.52; p<0.01]). Changes in BR and changes in OS markers were significantly correlated. CONCLUSIONS In virologically suppressed patients on stable ART, OS was lower in ATV/r-based regimens compared to EFV. We hypothesize these changes could be in part attributable to increased BR plasma levels.

Liver safety assessment: required data elements and best practices for data collection and standardization in clinical trials.

A workshop was convened to discuss best practices for the assessment of drug-induced liver injury (DILI) in clinical trials. In a breakout session, workshop attendees discussed necessary data elements and standards for the accurate measurement of DILI risk associated with new therapeutic agents in clinical trials. There was agreement that in order to achieve this goal the systematic acquisition of protocol-specified clinical measures and lab specimens from all study subjects is crucial. In addition, standard DILI terms that address the diverse clinical and pathologic signatures of DILI were considered essential. There was a strong consensus that clinical and lab analyses necessary for the evaluation of cases of acute liver injury should be consistent with the US Food and Drug Administration (FDA) guidance on pre-marketing risk assessment of DILI in clinical trials issued in 2009. A recommendation that liver injury case review and management be guided by clinicians with hepatologic expertise was made. Of note, there was agreement that emerging DILI signals should prompt the systematic collection of candidate pharmacogenomic, proteomic and/or metabonomic biomarkers from all study subjects. The use of emerging standardized clinical terminology, CRFs and graphic tools for data review to enable harmonization across clinical trials was strongly encouraged. Many of the recommendations made in the breakout session are in alignment with those made in the other parallel sessions on methodology to assess clinical liver safety data, causality assessment for suspected DILI, and liver safety assessment in special populations (hepatitis B, C, and oncology trials). Nonetheless, a few outstanding issues remain for future consideration.

A Change in the Epidemiology of Infections Due to Extended-Spectrum b-Lactamase���Producing Organisms

Extended-spectrum ��-lactamases (ESBLs) form a heterogeneous group that share the property of hydrolytic activity against the oxyimino-��-lactams while remaining susceptible to inhibition by ��-lactamase inhibitors, such as clavulanic acid. From a clinical point of view, they are important because they confer resistance to penicillins, aztreonam, and cephalosporins, and ESBL-producing organisms are typically also resistant to aminoglycosides, trimethoprim-sulfamethoxazole, and quinolones [1]. Until recently, the main problem posed by ESBLs was related to nosocomial outbreaks caused by ESBL-producing Klebsiella species. These outbreaks are usually clonal, the strains are mainly spread through cross-transmission, and the risk factors are similar to those found for other multidrug-resistant nosocomial pathogens [2]. In Europe and the United States, most ESBL-producing Klebsiella isolates harbored enzymes belonging to the TEM and SHV families [3]. Detection of colonized patients by performing surveillance cultures within affected units, isolation precautions for colonized patients, and restriction of oxyimino-��-lactam use are frequently useful for the control of these outbreaks [1]. There is no evidence that hospital-acquired ESBL-producing klebsiellae are decreasing in importance���in fact, data from the Centers for Disease Control and Prevention show that 20.6% of Klebsiella pneumoniae isolates from United States intensive care units in 2003 were probable producers of ESBL [4]. This represented a 47% increase, compared with the preceding 5 years. However, during the last few years, an impressive increase in the number of ESBL-producing Escherichia coli (and, less frequently, other Enterobacteriaceae) is being described in several parts of the world [5���8]. This emergent phenomenon shows some differences from the problem posed by Klebsiella species; many of these ESBL-producing E. coli are isolated ���

The Presence of Precursors of Benign Pre-B Lymphoblasts (Hematogones) in the Bone Marrow of a Paediatric Patient with Cytomegalovirus Infection

Hematogones are normal B-lymphoid precursors that multiply in the bone marrow of small children and of adults with ferropenic anaemia, neuroblastoma or idiopathic thrombocytopenic purpura. They are not normally found in peripheral blood, and the immunophenotype is virtually indistinguishable from that of B lymphoblasts. We discuss the case of a 3-month infant with an active cytomegalovirus infection, with hepatitis and pancytopenia associated with 13% hematogones in the bone marrow

Bone Mineral Density and Serum Levels of Soluble Tumor Necrosis Factors, Estradiol, and Osteoprotegerin in Postmenopausal Women with Cirrhosis after Viral Hepatitis

CONTEXT: Cirrhosis after viral hepatitis has been identified as a risk factor for osteoporosis in men. However, in postmenopausal women, most studies have evaluated the effect of primary biliary cirrhosis, but little is known about the effect of viral cirrhosis on bone mass [bone mineral density (BMD)] and bone metabolism. OBJECTIVE: Our objective was to assess the effect of viral cirrhosis on BMD and bone metabolism in postmenopausal women. DESIGN: We conducted a cross-sectional descriptive study. SETTING AND PATIENTS: We studied 84 postmenopausal female outpatients with viral cirrhosis and 96 healthy postmenopausal women from the general community. BMD was measured by dual-energy x-ray absorptiometry at lumbar spine (LS) and femoral neck (FN). RESULTS: The percentage with osteoporosis did not significantly differ between patients (LS, 43.1%; FN, 32.2%) and controls (LS, 41.2%; FN, 29.4%), and there was no difference in BMD (z-score) between groups. Serum concentrations of soluble TNF receptors, estradiol, and osteoprotegerin (OPG) were significantly higher in patients vs. controls (P < 0.001, P < 0.05, and P < 0.05, respectively). No significant difference was observed in urinary deoxypyridinoline. Serum OPG levels were positively correlated with soluble TNF receptors (r = 0.35; P < 0.02) and deoxypyridinoline (r = 0.37; P < 0.05). CONCLUSIONS: This study shows that bone mass and bone resorption rates do not differ between postmenopausal women with viral cirrhosis and healthy postmenopausal controls and suggests that viral cirrhosis does not appear to increase the risk of osteoporosis in these women. High serum estradiol and OPG concentrations may contribute to preventing the bone loss associated with viral cirrhosis in postmenopausal women.

Influence of IL28B polymorphisms on response to a lower-than-standard dose peg-IFN-�� 2a for genotype 3 chronic hepatitis C in HIV-coinfected patients.

BACKGROUND Data on which to base definitive recommendations on the doses and duration of therapy for genotype 3 HCV/HIV-coinfected patients are scarce. We evaluated the efficacy of a lower peginterferon-�� 2a dose and a shorter duration of therapy than the current standard of care in genotype 3 HCV/HIV-coinfected patients. METHODS AND FINDINGS Pilot, open-label, single arm clinical trial which involved 58 Caucasian HCV/HIV-coinfected patients who received weekly 135 ��g peginterferon-�� 2a plus ribavirin 400 mg twice daily during 20 weeks after attaining undetectable viremia. The relationships between baseline patient-related variables, including IL28B genotype, plasma HCV-RNA, ribavirin dose/kg, peginterferon-�� 2a and ribavirin levels with virological responses were analyzed. Only 4 patients showed lack of response and 5 patients dropped out due to adverse events related to the study medication. Overall, sustained virologic response (SVR) rates were 58.3% by intention-to-treat and 71.4% by per protocol analysis, respectively. Among patients with rapid virologic response (RVR), SVR and relapses rates were 92.6% and 7.4%, respectively. No relationships were observed between viral responses and ribavirin dose/kg, peginterferon-�� 2a concentrations, ribavirin levels or rs129679860 genotype. CONCLUSIONS Weekly 135 ��g pegIFN-�� 2a could be as effective as the standard 180 ��g dose, with a very low incidence of severe adverse events. A 24-week treatment duration appears to be appropriate in patients achieving RVR, but extending treatment up to just 20 weeks beyond negativization of viremia is associated with a high relapse rate in those patients not achieving RVR. There was no influence of IL28B genotype on the virological responses.

Variation of transaminases, HCV-RNA levels and Th1/Th2 cytokine production during the post-partum period in pregnant women with chronic hepatitis C.

This study analyses the evolution of liver disease in women with chronic hepatitis C during the third trimester of pregnancy and the post-partum period, as a natural model of immune modulation and reconstitution. Of the 122 mothers recruited to this study, 89 were HCV-RNA+ve/HIV-ve and 33 were HCV-RNA-ve/HIV-ve/HCVantibody+ve and all were tested during the third trimester of pregnancy, at delivery and post-delivery. The HCV-RNA+ve mothers were categorized as either Type-A (66%), with an increase in ALT levels in the post-partum period (>40 U/L; P<0.001) or as Type-B (34%), with no variation in ALT values. The Type-A mothers also presented a significant decrease in serum HCV-RNA levels in the post-delivery period (P<0.001) and this event was concomitant with an increase in Th1 cytokine levels (INF��, P���=���0.04; IL12, P���=���0.01 and IL2, P���=���0.01). On the other hand, the Type-B mothers and the HCV-RNA-ve women presented no variations in either of these parameters. However, they did present higher Th1 cytokine levels in the partum period (INF�� and IL2, P<0.05) than both the Type-A and the HCV-RNA-ve women. Cytokine levels at the moment of delivery do not constitute a risk factor associated with HCV vertical transmission. It is concluded that differences in the ALT and HCV-RNA values observed in HCV-RNA+ve women in the postpartum period might be due to different ratios of Th1 cytokine production. In the Type-B women, the high partum levels of Th1 cytokines and the absence of post-partum variation in ALT and HCV-RNA levels may be related to permanent Th1 cytokine stimulation.