Hepatitis B virus (HBV) infections continue to occur in adult hemodialysis units. A possible contributing factor is the presence of occult HBV (serum hepatitis B surface antigen [HBsAg] negative but HBV DNA positive). Two hundred forty-one adult hemodialysis patients were screened for occult HBV. HBV DNA testing was performed by real-time polymerase chain reaction (PCR) with 2 independent primer sets (core promoter and surface). Two (0.8%) of the 241 patients were HBsAg positive. Of the remaining 239 HBsAgnegative patients, 9 (3.8%) were HBV DNA positive. Viral loads in these individuals were low (10 2 -10 4 viral copies/mL). Seven of the 9 (78%) were nt 587 mutation (sG145R mutant) positive. Demographic, biochemical, and HBV serological testing did not help to identify those with occult HBV. In conclusion, the prevalence of occult HBV in adult hemodialysis patients in this North American urban center is approximately 4 to 5 times higher than standard HBsAg testing would suggest. The majority of these infections are associated with low viral loads and a high prevalence of the sG145R mutant. Finally, the demographic, biochemical, and/or serological features of HBV DNA-positive subjects do not distinguish these individuals from the remainder of the dialysis patient population. (HEPATOLOGY 2004; 40:1072-1077.) D espite the development of an effective hepatitis B virus (HBV) vaccine and extensive infection control guidelines, HBV infections continue to occur in dialysis units throughout North America and Europe. 1-3 Based on the results of hepatitis B surface antigen (HBsAg) testing, the incidence of such infections is thought to be 0.05%-1% per year. 4 However, this likely represents an underestimate of the true incidence, as the use of more sensitive monoclonal-based assays for HBsAg detection increase positive findings in dialysis patients by 120%. 5 The relatively low acceptance and response rates to the HBV vaccine among dialysis patients likely contributes to ongoing transmission, as does the need for vaccine boosts to maintain antibody to HBsAg (anti-HBs) at protective levels. 4,6 -8 Additional contributing factors include "breakdowns" in the application of universal and/or dialysis-specific infection control measures. 4,7,9 With the development of specific and sensitive polymerase chain reaction (PCR)-based testing for HBV DNA, the presence of occult HBV infection (HBV DNA positivity in the setting of negative serum HBsAg) represents yet another possible explanation for ongoing transmission.To date, few studies have documented the prevalence of occult HBV infection in renal dialysis patients. In 3 small studies of 33, 5, and 67 HBsAg-negative dialysis patients, 50%, 40%, and 0%, respectively, were HBV DNA positive. 10 -12 Of note, the majority of HBV DNApositive individuals in these studies had serological evidence of previous HBV infection (HBV seropositive), but as many as 39% were HBV seronegative.The present study documents the prevalence of HBV DNA positivity in a large, North American renal dialys...

In March 2003, a novel coronavirus was isolated from patients exhibiting atypical pneumonia and subsequently proven to be the causative agent of the disease now referred to as severe acute respiratory syndrome (SARS). The complete genome of the SARS coronavirus (SARS-CoV) has since been sequenced. The SARS-CoV nucleocapsid (SARS-CoV N) shares little homology with other members of the coronavirus family. To determine if the N protein is involved in the regulation of cellular signal transduction, an ELISA-based assay on transcription factors was used. We found that the amount of transcription factors binding to promoter sequences of c-Fos, ATF2, CREB-1, and FosB was increased by the expression of SARS-CoV N. Since these factors are related to AP-1 signal transduction pathway, we investigated whether the AP-1 pathway was activated by SARS-CoV N protein using the PathDetect system. The results demonstrated that the expression of N protein, not the membrane protein (M), activated AP-1 pathway. We also found that SARS-CoV N protein does not activate NF-kappaB pathway, demonstrating that activation of important cellular pathways by SAS-CoV N protein is selective. Thus our data for the first time indicate that SARS-CoV has encoded a strategy to regulate cellular signaling process.

Determining the longitudinal molecular evolution of hepatitis B virus (HBV) is difficult due to HBV's genomic complexity and the need to study paired samples collected over long periods of time. In this study, serial samples were collected from eight hepatitis B virus e antigen-negative asymptomatic carriers of HBV genotype B in 1979 and 2004, thus providing a 25-year period to document the long-term molecular evolution of HBV. The rate, nature, and distribution of mutations that emerged over 25 years were determined by phylogenetic and linear regression analysis of full-length HBV genome sequences. Nucleotide hypervariability was observed within the polymerase and pre-S/S overlap region and within the core gene. The calculated mean number of nucleotide substitutions/site/year (7.9 ؋ 10 ؊5 ) was slightly higher than published estimates (1.5 ؋ 10 ؊5 to 5 ؋ 10 ؊5 ). Nucleotide changes in the quasispecies population did not significantly alter the molecular evolutionary rate, based on linear regression analysis of evolutionary distances among serial clone pre-S region sequences. Therefore, the directly amplified or dominant sequence was sufficient to estimate the putative molecular evolutionary rate for these long-term serial samples. On average, the ratio of synonymous (d S ) to nonsynonymous (d N ) substitutions was highest for the polymerase-coding region and lowest for the core-coding region. The low d S /d N ratios observed within the core suggest that selection occurs within this gene region, possibly as an immune evasion strategy. The results of this study suggest that HBV sequence divergence may occur more rapidly than previously estimated, in a host immune phase-dependent manner.Determination of the molecular evolution of hepatitis B virus (HBV) involves an understanding of the accumulated sequence changes to the viral genome and the observed mutation rate over a long period. Determining the rate of sequence change is difficult due to the complex organization of the HBV genome, which involves multiple coding and regulatory functions within overlapping open reading frames (16). Two-thirds of the viral genome codes for multiple proteins, and thus a synonymous change in one open reading frame results in a nonsynonymous change in the overlapping open reading frame. In this way, it is believed that HBV genome evolution is "constrained" in order to maintain essential protein functions (22).Since HBV replication involves an error-prone reverse transcription step, the rate of nucleotide change during replication is higher than that found for other DNA viruses and is more similar to the rate observed for the slower-evolving RNA viruses (21, 24). The rate of HBV evolution in hepatitis B virus e antigen (HBeAg)-positive individuals has been estimated to be 1.5 ϫ 10 Ϫ5 to 5 ϫ 10 Ϫ5 nucleotide substitutions per site per year (1,13,24,29). However, the mutation rate or total accumulated number of mutations appears to be higher in HBeAgnegative patients (11, 33), suggesting that the host immune response plays an importan...

Hepatitis B virus (HBV) infection is an important public health problem in Canada. In keeping with evolving evidence and understanding of HBV pathogenesis, the Canadian Association for the Study of Liver Disease periodically publishes HBV management guidelines. The goals of the 2018 guidelines are to (1) highlight the public health impact of HBV infection in Canada and the need to improve diagnosis and linkage to care, (2) recommend current best-practice guidelines for treatment of HBV, (3) summarize the key HBV laboratory diagnostic tests, and (4) review evidence on HBV management in special patient populations and include more detail on management of HBV in pediatric populations. An overview of novel HBV tests and therapies for HBV in development is provided to highlight the recent advances in HBV clinical research. The aim and scope of these guidelines are to serve as an up-to-date, comprehensive resource for Canadian health care providers in the management of HBV infection.

Although endothelial cells have been speculated to be a target in the pathogenesis of dengue hemorrhagic fever (DHF), there has been little evidence linking dengue virus infection to any alteration in endothelial cell function. In this study, we show that human umbilical vein endothelial cells become activated when exposed to culture fluids from dengue virus-infected peripheral blood monocytes. Maximum activation was achieved with culture fluids from monocytes in which virus infection was enhanced by the addition of dengue virus-immune serum, thus correlating with epidemiological evidence that prior immunity to dengue virus is a major risk factor for DHF. Activation was strongest for endothelial cell expression of VCAM-1 and ICAM-1. In contrast, activation of endothelial cell E-selectin expression appeared to be more transient, as indicated by its detection at 3 h, but not at 16 h, of treatment. Treatment of monocyte culture fluids with anti-tumor necrosis factor alpha (TNF-␣) antibody largely abolished the activation effect (as measured by endothelial cell expression of ICAM-1), whereas treatment with IL-1␤ receptor antagonist had a much smaller inhibitory effect on activation. Endothelial cells inoculated directly with dengue virus or with virus-antibody combinations were poorly infectable (compared to Vero cells or peripheral blood monocytes), and virus-inoculated endothelial cells showed no increased expression of VCAM-1, ICAM-1, or E-selectin. Taken together, the results strongly indicate that dengue virus can modulate endothelial cell function by an indirect route, in which a key intermediary is TNF-␣ released from virus-infected monocytes.

Hepatitis B virus genotype B (HBV/B) has been classified into 5 subgenotypes. Except for Bj/B1 in Japan, the subgenotypes (Ba/B2-B5) have undergone recombination with HBV/C in the core promoter/precore/core genomic region. Phylogenetic analyses of complete sequences show that the Arctic strains belong to a novel subgenotype (HBV/B6) without the recombination, analogous to what is seen with Bj/B1. Comparison of 50 HBV/B6 carriers from the Arctic versus 50 Bj and 50 Ba age- and sex-matched carriers from Asia revealed that clinical characteristics of HBV/B6 carriers were similar to those of Bj/B1 carriers in Japan. The results suggest that HBV/B may be classified into nonrecombinant (Bj/B1 and B6) and recombinant (Ba/B2-B5) types.

Mother-to-child transmission of hepatitis B virus (HBV) continues to occur despite immunoprophylaxis. We examined maternal factors contributing to transmission in infants receiving adequate immunoprophylaxis in Alberta, Canada. Prenatal specimens from HBsAg-positive women whose babies developed HBV infection despite immunoprophylaxis (cases) and HBsAg-positive mothers whose babies did not (controls) were tested for HBsAg, HBeAg and HBV DNA. Specimens with detectable DNA underwent HBV genotyping. Routinely collected surveillance data and laboratory test results were compared between cases and controls. Twelve cases and 52 controls were selected from a provincial registry from 2000 to 2005. At the time of prenatal screening, median maternal age was 31 years [interquartile range (IQR): 27.5-34.5], and median gestational age was 12 weeks (IQR 10.0-15.5). Cases were more likely than controls to test positive for HBeAg (77.8% vs. 23.1%; P < 0.05). Of all mothers with detectable viral load (n = 51), cases had a significantly higher median viral load than did controls (5.6 × 10(8)  IU/mL vs. 1750 IU/mL, P < 0.0001). Of the two cases who were HBeAg negative, one had an undetectable viral load 8 months prior to delivery and a sP120T mutation. The viral load in the other case was 14,000 IU/mL. The majority of isolates were genotype B (31.3%) and C (31.3%) with no significant differences in genotype between cases or controls. In this case-control study, transmission of HBV to infants was more likely to occur in mothers positive for HBeAg and with high HBV DNA.

Hepatitis B virus (HBV) genotype G (HBV/G) is an unusual variant, and little is known about its epidemiology and natural history, particularly the requirement for a co-infecting HBV genotype and their relationship during infection. This study investigated the quasispecies nature of coinfecting genotypes in 39 samples collected over a 6 year period from 13 HBV/G-infected patients. HBV/G infections were found to occur predominantly in males (92 %) and were primarily associated with male homosexual sex (67 %). All patients were infected with HBV/G and HBV/A, or a recombinant HBV/A/G strain. Co-infecting genotypic prevalence was often observed to fluctuate over time, with periods of HBV/G monoinfection in some patients. The average sequence divergence among Canadian HBV/G strains was 1.57±0.62 %. Thus, all HBV/G infections in Canada occur in the context of co-infection or recombination with HBV/A, and strains display increased sequence divergence compared with all known HBV/G sequences described to date.