Prevalence and risk of hepatitis E virus infection in the HIV population of Nepal

Background

Infection with the hepatitis E virus (HEV) can cause acute hepatitis in endemic areas in immune-competent hosts, as well as chronic infection in immune-compromised subjects in non-endemic areas. Most studies assessing HEV infection in HIV-infected populations have been performed in developed countries that are usually affected by HEV genotype 3. The objective of this study is to measure the prevalence and risk of acquiring HEV among HIV-infected individuals in Nepal.

Methods

We prospectively evaluated 459 Human Immunodeficiency Virus (HIV)-positive individuals from Nepal, an endemic country for HEV, for seroprevalence of HEV and assessed risk factors associated with HEV infection. All individuals were on antiretroviral therapy and healthy blood donors were used as controls.

Results

We found a high prevalence of HEV IgG (39.4%) and HEV IgM (15.3%) in HIV-positive subjects when compared to healthy HIV-negative controls: 9.5% and 4.4%, respectively (OR: 6.17, 95% CI 4.42–8.61, p?<?0.001 and OR: 3.7, 95% CI 2.35–5.92, p?<?0.001, respectively). Individuals residing in the Kathmandu area showed a significantly higher HEV IgG seroprevalance compared to individuals residing outside of Kathmandu (76.8% vs 11.1%, OR: 30.33, 95% CI 18.02–51.04, p?=?0.001). Mean CD4 counts, HIV viral load and presence of hepatitis B surface antigen correlated with higher HEV IgM rate, while presence of hepatitis C antibody correlated with higher rate of HEV IgG in serum. Overall, individuals with HEV IgM positivity had higher levels of alanine aminotransferase (ALT) than IgM negative subjects, suggesting active acute infection. However, no specific symptoms for hepatitis were identified.

Conclusions

HIV-positive subjects living in Kathmandu are at higher risk of acquiring HEV infection as compared to the general population and to HIV-positive subjects living outside Kathmandu.

     

戊型肝炎病毒疫苗的研发和评价

由戊型肝炎病毒(Hepatitis E virus,HEV)导致的疾病日渐引起人们的关注。中国目前已经上市的HEV疫苗为万泰沧海生物技术公司(INNOVAX)研发的大肠杆菌表达的I型中国分离株ORF2 p239重组疫苗HEV 239(Hecolin),它能形成病毒样颗粒从而产生更好的免疫原性。该疫苗在HEV高危人群和高流行地区的使用将对戊型肝炎的流行起到积极的防控作用。回顾了HEV疫苗研究的历史过程,并探讨了目前HEV疫苗评价方法面临的问题及研究思路。     

重组戊型肝炎疫苗质量控制和评价研究

2012年全球首个戊型肝炎(简称戊肝)疫苗获准在中国上市,为防控戊型肝炎的流行提供了有效办法。提升戊肝疫苗的质量控制和评价标准不但对保证疫苗的质量,推动戊肝疫苗走向世界起着至关重要的作用;而且对今后同类重组疫苗的研究开发具有指导意义。本文就重组戊肝疫苗的质量控制和评价研究作一综述。     

戊型肝炎疫苗上市后质量控制

目的对中国独创的基因重组戊型肝炎(hepatitis E,HE)疫苗Hecolin~?上市后质量的一致性进行评价。方法比较了Hecolin~?批签发疫苗与III期临床试验疫苗的效力试验检测结果,进而通过趋势分析评价Hecolin~?疫苗上市5年来的批间质量一致性。结果批签发疫苗与Ⅲ期临床试验疫苗的体内效力试验结果一致。批签发疫苗体内效力、铝含量、硫柳汞含量、p H等关键指标,中国食品药品检定研究院与企业的趋势分析结果均显示具有较好的一致性。结论趋势分析结果表明,上市后Hecolin~?疫苗批间质量一致性较好,质量稳定。     

Clinical trial with inactivated hepatitis A vaccine and recommendations for its use.

OBJECTIVE--To compare the reactogenicity and immunogenicity of an inactivated hepatitis A vaccine in two different immunisation schedules. DESIGN--Randomised trial. SETTING--One London teaching hospital. SUBJECTS--104 healthy adult volunteers (71 men, 33 women aged 19-60). INTERVENTIONS--Hepatitis A vaccine to group 1 (54 volunteers) at 0, 1, and 2 months and to group 2 (50) at 0, 1, and 6 months. MAIN OUTCOME MEASURES--Symptoms at and after each dose; liver function, hepatitis A virus specific serum immune response; and responses in saliva and parotid fluid in immunised volunteers and subjects with natural immunity. RESULTS--The vaccine was well tolerated; 97% (96/99) and 100% of those immunised developed serum antibody after one and two doses of vaccine respectively. Geometric mean titres increased progressively after each dose and were significantly higher in men but not women in group 2 after the third dose (ratio between geometric mean titres 0.265, 95% confidence interval 0.18 to 0.39; p less than 0.001). At one year this group-sex interaction was absent; geometric mean titres for both sexes were significantly higher in group 2 (ratio 0.330, 0.227 to 0.478; p less than 0.0001). Antibody responses were not significantly different between the groups at two years. Compared with naturally infected subjects immunised volunteers developed poor or undetectable virus specific IgG and IgA responses in saliva and parotid fluid. CONCLUSIONS--The vaccine was safe and highly immunogenic, and the differences in the immune responses in saliva and parotid fluid are unlikely to affect its efficacy.     

Understanding and enhancing compliance with the second dose of hepatitis B vaccine: a cohort analysis and a randomized controlled trial

OBJECTIVE: To determine the predictors and extent of noncompliance with a second dose of hepatitis B vaccine and the effectiveness of a compliance enhancement strategy. DESIGN: Cohort analysis and randomized clinical trial. SUBJECTS: A total of 256 consecutive adults attending a sexually transmitted diseases clinic from October 1992 to July 1993 who were seronegative for hepatitis B virus and agreed to receive hepatitis B vaccination. SETTING: Hamilton, Ont. INTERVENTION: Subjects were followed up for 4 months. Those who did not return for the second dose of vaccine by 6 weeks after the first (2 weeks overdue) were randomly assigned to the enhanced intervention group (telephone and mail reminders) or the regular intervention group (mail reminder only). Subjects were considered noncompliant if they did not return for the second dose by 4 months after the first. RESULTS: The risk of not returning for the second dose of vaccine within 4 months after the first was strongly and linearly associated with level of education (p = 0.004). The noncompliance rate among those with less than a grade 10 education was 50%, grade 10-13 education 34%, some college education 15% and some university education 9%. In the randomized controlled trial the enhanced intervention group had twice the compliance rate of the regular intervention group (48% v. 25%; p = 0.008). Subjects with no postsecondary education were highly responsive to the enhanced intervention (relative risk 2.1; p = 0.02) compared with those with a higher level of education (relative risk 1.0; p = 1.0). CONCLUSION: Hepatitis B vaccine recipients with lower educational levels are at increased risk of noncompliance with the second dose of vaccine but are highly responsive to telephone reminders.     

Immunogenicity of hepatitis B vaccine in guinea-pigs, mice and man

The immunogenicity of the hepatitis B vaccine Hevac B Pasteur was assayed in guinea-pigs. The reproducibility of the test was determined by comparing the ImD50 of two samples of vaccine in 12 and eight groups of 40 animals each. The variation coefficient between groups was approximately 30%, similar to that found for other vaccines assayed in biological tests in vivo. The sensitivity of the test was determined by comparing the dose response curves in mice, guinea-pigs and humans. The results showed that one ImD50 in man (after one injection) was approximately equivalent to 3 ImD50 in guinea pigs and to 100 ImD50 in Balb/C mice. Stability of Hevac B at 37 degrees C was assayed by determining in guinea-pigs the ImD50 of a series of samples of vaccine that had been incubated at 37 degrees C for varying amounts of time. The immunogenicity of the vaccine was found to decay with a half-life of ten to 15 days at 37 degrees C.     

Purification and characterization of a recombinant hepatitis E protein vaccine candidate by liquid chromatography-mass spectrometry

A protein with a molecular mass of approximately 62·10³, derived from open reading frame 2 (ORF-2) of the hepatitis E virus (HEV; Burma strain), was expressed in a baculovirus expression vector and purified to homogeneity. The recombinant 62 kDa protein appeared to be a doublet, as determined by sodium dodecyl sulfate polycrylamide gel electrophoresis (SDS-PAGE). Tryptic digestion in conjunction with laser desorption mass spectrometry (LD-MS) and sequence analysis of the tryptic peptides indicated that the amino terminus was blocked, although no proteolytic degradation occurred. The determined internal sequences of peptides were in agreement with the predicted ORF-2 protein. Reversed-phase liquid chromatography coupled to electrospray mass spectrometry (LC-MS) resolved the doublet proteins into two major components with molecular masses of 56 548.5 and 58 161.4. Confirmation of the amino terminus of the molecule by LD-MS post-ion decay enabled us to tentatively assign the carboxyl terminus of each species at residues 540 and 525. Sequencing of the intact protein by automated carboxyl terminal sequencing confirmed that the carboxyl terminus was truncated and that the sequence assignment predicted by LC-MS was correct.     

IRIV-adjuvanted hepatitis A vaccine: in vivo absorption and biophysical characterization

Immunopotentiating reconstituted influenza virosomes (IRIV) are 150-nm proteoliposomes composed of influenza surface glycoproteins and a mixture of natural and synthetic phospholipids. Due to size, structure and composition of the IRIVs, they serve as an antigen carrier system for efficacious vaccination, as was demonstrated for hepatitis A and influenza. This paper reviews the unique properties of IRIVs and describes the in vivo biodistribution of model antigens using 14C-labeled IRIVs and 125I-labeled streptavidin. IRIV formulated streptavidin induced a strong depot effect after intra muscular (i.m.) vaccination of mice, whereas soluble streptavidin was soon eliminated via the kidney of the animals. A mixture of antigen and IRIVs yielded higher antibody titers after i.m. inoculation than streptavidin alone. The highest immunostimulation was achieved by the binding of the antigen to the investigated adjuvant. The potential penetration of inactivated hepatitis A virions into lipid membranes was assessed by measuring the area increase of a lipid monolayer kept at a constant surface pressure corresponding to that of lipid bilayer vesicles. The monolayers were composed of phosphatidylcholine (POPC) and phosphatidylethanolamine (POPE) (75/25 mol/mol), thus resembling the lipid composition of the IRIV. The results suggested that the hepatitis A antigen may spontaneously bind to the reconstituted IRIV membranes.     

戊型肝炎病毒遗传多样性和进化机制的研究进展

戊型肝炎病毒(hepatitis E virus, HEV)是一种引发全球急性病毒性肝炎的人兽共患病病原。HEV具有丰富的遗传多样性,不同基因型或基因亚型的流行与地理位置、宿主物种以及防控策略等密切相关。欧洲和美洲HEV流行株为HEV-3,包括3a-i亚型,而亚洲流行株含HEV-3和HEV-4;我国的流行毒株已从HEV-1进化到HEV-4。近年来,研究发现HEV进化的影响机制,包括同义密码子使用模式、氨基酸突变和基因重组等,其中氨基酸突变是病毒持续流行的主要驱动力。因此,本文就HEV的分类、全球流行特征、进化机制等进行综述,以期为戊型肝炎的有效防控以及疫苗开发提供参考。     

A randomized controlled phase IIb trial of antigen-antibody immunogenic complex therapeutic vaccine in chronic hepatitis B patients

Background

The safety of the immune complexes composed of yeast-derived hepatitis B surface antigen (HBsAg) and antibodies (abbreviated as YIC) among healthy adults and chronic hepatitis B patients has been proved in phase I and phase IIa trial. A larger number of patients for study of dosage and efficacy are therefore needed.

Methods and Principal Findings

Two hundred forty two HBeAg-positive chronic hepatitis B patients were immunized with six injections of either 30 µg YIC, 60 µg of YIC or alum adjuvant as placebo at four-week intervals under code. HBV markers and HBV DNA were monitored during immunization and 24 weeks after the completion of immunization. The primary endpoint was defined as loss of HBeAg, or presence of anti-HBe antibody or suppression of HBV DNA, while the secondary endpoint was both HBeAg seroconversion and suppression of HBV DNA. Statistical significance was not reached in primary endpoints four weeks after the end of treatment among three groups, however, at the end of follow-up, HBeAg sero-conversion rate was 21.8%(17/78) and 9% (7/78) in the 60 µg YIC and placebo groups respectively (p = 0.03), with 95% confidence intervals at 1.5% to 24.1%. Using generalized estimating equations (GEEs) model, a significant difference of group effects was found between 60 µg YIC and the placebo groups in terms of the primary endpoint. Eleven serious adverse events occurred, which were 5.1%, 3.6%, and 5.0% in the placebo, 30 µg YIC and 60 µg YIC groups respectively (p>0.05).

Conclusions

Though statistical differences in the preset primary and secondary endpoints among the three groups were not reached, a late and promising HBeAg seroconversion effect was shown in the 60 µg YIC immunized regimen. By increasing the number of patients and injections, the therapeutic efficacy of YIC in chronic hepatitis B patients will be further evaluated.

Trial Registration

ChiCTR.org ChiCTR-TRC-00000022     

Recent topics on hepatitis E virus: emerging, zoonotic, and animal-to-human transmission in Japan

Hepatitis E is undoubtedly a zoonosis. Recent observations suggest that the zoonotic food-borne mode of transmission has played an important role in the spread of hepatitis E virus (HEV) among Japanese people (who in general likes eating everything uncooked or undercooked: Sushi, Sashimi, Tataki, Namagimo, Shabu-shabu, etc). Moreover, the situation seems to be worsening. Wild boar (and deer also) has recently been increasing in its number, becoming a more potent HEV reservoir to humans than before. Pork, replacing beef in people's recent fear of BSE, is being consumed increasingly, particularly in Hokkaido. It may be Japanese people that an effective HEV vaccine is most longed for by.     

Engaging the community in research: lessons learned from the malaria vaccine trial

When staff of the Papua New Guinea Institute of Medical Research first went into Wosera in 1979, to try and interest the community into participating in malaria research, the traditional warning sign of crossed bamboo sticks constantly blocked their path as they tried to enter the villages. In 2003, the site is a thriving research centre, with many projects under its belt, a successfully executed malaria vaccine trial to its credit, and approximately 13000 people from 29 villages currently participating in demographic surveillance and various research projects. It has been a long road from community suspicion to sustainable community participation, and the field workers have learned many important lessons along the way that can be applicable to research programs in other disease-endemic areas.     

Studies in chimpanzees of live, attenuated hepatitis A vaccine candidates

Human hepatitis A virus was attenuated in virulence for chimpanzees by passage in FRhK6 and human diploid lung fibroblast cell cultures. A number of variants were developed by passage in cell cultures which showed different levels of virulence/attenuation for chimpanzees. These results were compared to those obtained with marmosets and reported previously. In general, most variants behaved similarly in the two animal types. Two chimpanzees which gave vaccine-like responses following inoculation with HAV cell culture variants were challenged with virulent HAV. Both animals were immune to HAV infection. These findings provide further evidence for the feasibility of developing live, attenuated vaccines against human hepatitis A.     

Immune response to a hepatitis B DNA vaccine in Aotus monkeys: a comparison of vaccine formulation, route, and method of administration.

BACKGROUND: Attempts to optimize DNA vaccines in mice include using different routes of administration and different formulations. It may be more relevant to human use to carry such studies out in nonhuman primates. Here we compare different approaches to delivery of a DNA vaccine against the hepatitis B virus (HBV) in Aotus monkeys. MATERIALS AND METHODS: Thirty-two adult Aotus l. lemurinus monkeys divided into 8 groups of four were immunized with 400 microg of a DNA vaccine which encoded hepatitis B surface antigen (HBsAg). DNA in saline was administered by intradermal (ID) or intramuscular (IM) injection with needle and syringe, IM injection with the Biojector needleless injection system or combined ID (needle) and IM (Biojector). DNA formulated with cationic liposomes (CellFECTIN) was injected IM with needle or Biojector. DNA with added E. coli DNA (100 microg) was injected IM with the Biojector or ID. A ninth group of 4 monkeys was injected IM (needle) with Engerix-B, a commercial vaccine containing recombinant HBsAg (10 microg) adsorbed onto alum. Monkeys were boosted in an identical fashion to their prime at 8 weeks, but all received the protein vaccine (Engerix-B) at 16 weeks. Sera was assessed for antibodies against HBsAg (anti-HBs) by enzyme-linked imunosorbent assay (ELISA). RESULTS: The primary humoral response induced by IM delivery of the DNA vaccine was very poor. In most cases there was no detectable anti-HBs even after 2 DNA doses but the kinetics of the response to subsequent protein indicated that a memory B cell response had been induced. In contrast, following IM-administration of DNA using the Biojector, detectable anti-HBs were observed in 3 of 8 animals and evidence for immunological priming was apparent in an additional 4 of the 8 monkeys. ID injection of DNA vaccine in saline induced a potent antibody response which was augmented 6-fold by the addition of E. coli DNA. Combining ID and IM administration did not improve humoral immunity over ID injection alone. CONCLUSIONS: For immunization of primates with DNA vaccines, ID may be a preferable route to IM, although it is not clear whether the Aotus monkey is a relevant model for humans in this respect. Nevertheless, the use of the Biojector needleless injection system may improve responses with IM delivery of DNA vaccines. As well, the immunostimulatory action of E. coli DNA may be used to augment the humoral response induced by a DNA vaccine.     

The PhyloCode, types, ranks and monophyly: a response to Pickett

A report from the First International Phylogenetic Nomenclature Meeting recently published in Cladistics conveys several misconceptions about the PhyloCode and presents an erroneous interpretation of discussions that took place at that meeting. Contrary to Pickett's assertions, the PhyloCode is designed to name clades, not paraphyletic groups; the rejection of ranks has never been a fundamental principle of phylogenetic nomenclature; and specifiers under the PhyloCode differ in several ways from types under rank‐based nomenclature. © The Willi Hennig Society 2005.     

Hemoglobin E: report of the first 2 cases in French subjects

Hemoglobin E has been discovered casually in the blood of two French donors: one coming from the Alsace region, the other one coming from the Champagne region. In the two cases, the hemoglobin E is in an heterozygote state and takes the place of 33 per cent of the total haemoglobin in the first and 24 per cent in the second. We investigated in their families and found that other members of these families had hemoglobin E. Each time, it was associated with a microcytosis and polycythaemia without anemy or iron deficiency. The red cells morphology shows many microspherocytes and target-cells. There is no relationship between these two families and the research of an asiatic antecedent proved negative. These two observations give a supplementary proof that the geographic repartition of the hemoglobin E is larger than what we read in the first publications and shows the interest to study the hemoglobin of unexplained polycythaemia with microcytes in the blood of blood donors.