Arie J. Zuckerman

Vaccine-induced escape mutant of hepatitis B virus

In southern Italy, 44 contacts of hepatitis B virus carriers, including infants of carrier mothers, became HBsAg positive despite passive and active immunisation according to standard protocols. In 32 of these vaccinees infection was confirmed by the presence of additional markers of viral replication. In 1 infant, serious disease occurred. The virus from this patient is an escape mutant with a different sequence from that of the isolate from the mother. A point mutation from guanosine to adenosine at nucleotide position 587 resulted in an aminoacid substitution from glycine to arginine in the highly antigenic a determinant of HBsAg. This mutation is stable: it is present in an isolate from the child 5 years later. In some of these patients, including this child, the a determinant, to which a large part of the vaccine-induced immunity is directed, has been partly lost. Binding to HBsAg of a monoclonal antibody, previously mapped to the region of the mutation, was reduced in the child relative to that of the mother.

Molecular epidemiology of hepatitis B virus vaccine variants in Singapore

Perinatal infection with variants of hepatitis B virus occurs despite combined immunoprophylaxis with hepatitis B immunoglobulin and currently licensed plasma-derived and recombinant yeast hepatitis B vaccines. Several variants have been detected during a large study of infants born to carrier mothers in Singapore. The most frequent variant was a virus in which a single amino acid substitution Gly to Arg occurred at amino acid position 145 of the outer protein coat of the virus. Similar mutations have been described in Italy, Japan, the USA and a number of other countries. The emergence of antibody escape mutants is a cause for concern for the detection of virus and possibly for future immunization programmes.

The role of circulating hepatitis B antigen/antibody immune complexes in the pathogenesis of vascular and hepatic manifestations in polyarteritis nodosa.

To investigate further the role of hepatitis B antigen (HBs Ag) and specific immune complexes in polyarteritis, sera from 55 histologically confirmed cases were tested for the presence of hepatitis B antigen-associated particles and hepatitis B-antibody (anti HBs) by solid phase radio-immunoassay, electron microscopy, and passive haemagglutination. Results of these findings have been correlated with the clinical course of the disease. HBs Ag was detected in 30 patients (54·5%) and anti HBs in 13/45 (28%). Subtyping in 20 patients revealed that 11 were Y and 9 D. Thirty-seven cases (69%) demonstrated either HBs Ag or anti HBs and 5/45 (11%) had both. Electron microscopic examination showed 20 nm spherical and tubular particles in sera of 20/27 patients with 42 nm particles in 11 cases and clumped particles in 12 (60%). No correlation was found between detection of immune complexes and liver disease whereas the presence of coexisting hepatitis B antigen and antibody or aggregated particles was restricted to cases of active vasculitis. Seroconversion or the presence of hepatitis B antibody alone was associated with improved prognosis. Circulating hepatitis B antigen antibody complexes may be responsible for vasculitis or polyarteritis but do not appear to be pathogenic for the liver.

AFFINITY OF ANTIBODY RESPONSES IN MAN TO HEPATITIS B VACCINE DETERMINED WITH SYNTHETIC PEPTIDES

The affinity and level of antibody to hepatitis B surface antigen (anti-HBs) in recipients of a plasma-derived hepatitis B vaccine were determined with three different antigens. The first two antigens were prepared by chemical synthesis, to represent linear or cyclical forms of aminoacid sequences 139 to 147 of the major hepatitis B surface antigen (HBsAg) polypeptide. The binding of antibodies to these synthetic peptides was compared with that to a third antigen, prepared by solubilisation of the naturally occurring HBsAg, the basic component of the currently licensed hepatitis B vaccine in the United Kingdom. Antibody levels, expressed as total antibody combining sites (Abt) in fixed volumes of immune sera, increased throughout the course of immunisation and correlated with the development of antibody as measured by a commercially available radioimmunoassay. Abt values were similar for both forms of the synthetic peptide, although higher affinity values were found with the cyclical structure, which illustrates the importance of protein conformation in antibody responses to HBsAg. Antibody affinity for the three antigens increased progressively throughout the immunisation schedule but the pattern of affinity maturation varied according to the peptide used as an antigen probe and between subjects. Most subjects showed a significant rise in antibody affinity after the third (booster) dose of vaccine given at six months. The use of synthetic peptides allowed a quantitative and qualitative assessment of antibody responses to hepatitis B vaccine and confirmed that selected peptides corresponding to relevant HBsAg epitopes may be useful as alternative hepatitis B vaccines.

Independent emergence of a vaccine-induced escape mutant of hepatitis B virus.

We investigated the case of a child who was vaccinated at birth against hepatitis B and was also given hepatitis B immune globulin, but who nevertheless later became infected with the virus. Hepatitis B virus-specific deoxyribonucleic acid covering the region of the genome encoding the predominant a epitope of hepatitis B surface antigen was amplified using the polymerase chain reaction and the nucleotide sequence determined. We present evidence for the independent emergence of an escape mutant, similar to that previously identified in Italy, where a substitution of arginine for glycine has occurred in the second loop of the a epitope.

Immune response to a new hepatitis B vaccine in healthcare workers who had not responded to standard vaccine: randomised double blind dose-response study

Abstract Objective: To evaluate the immunogenicity and reactogenicity of a new triple S recombinant hepatitis B vaccine in a cohort of healthy people in whom currently licensed hepatitis B vaccines had persistently not induced an immune response. Design: Single centre, randomised, double blind, dose-response study. Setting: Research vaccine evaluation centre at a teaching hospital. Subjects: 100 healthcare workers aged 18-70 years with a history of failure to seroconvert after at least four doses of a licensed hepatitis B vaccine containing the S component. Intervention: Each subject was randomly allocated two doses of 5, 10, 20, or 40 µg of a new hepatitis B vaccine two months apart. Main outcome measures: Immunogenicity of the four doses. Seroconversion and seroprotection were defined as an antibody titre >10 IU/l and >100 IU/l respectively against an international antibody standard. Results: 69 subjects seroconverted after a single dose of the vaccine. After the booster vaccination one other subject seroconverted, bringing the overall seroconversion rate to 70%. Fifteen subjects given 5 µg of vaccine, 19 given 10 µg, 16 given 20 µg, and 20 given 40 µg seroconverted. Seroconversion rates in the four antigen dose groups were 60% (15/25), 76% (19/25), 64% (16/25), and 80% (20/25). After the booster dose there was no significant dose-response effect on the overall seroconversion rate, although the small sample size meant that a clinically important dose-response could not be ruled out. Conclusion: A single dose of 20 µg of the vaccine was as effective as two doses of either 40 µg or 20 µg of this vaccine formulation in terms of seroconversion, seroprotection, and geometric mean titres. Key messages Up to 15% of healthy people do not respond to currently licensed hepatitis B vaccines Incorporation of the pre-S1 and pre-S2 components with the S antigen overcame this non-response in 69% of healthcare workers with a history of persistent non-response to conventional hepatitis B vaccines Significantly higher geometric mean titre levels were obtained with increased dosage of vaccine A single dose of 20 µg of the new vaccine seems to be effective in terms of seroconversion, seroprotection, and geometric mean titres

Taxonomic classification of human hepatitis B virus

Sufficient data have accumulated to permit the ICTV Study Group on the Nomenclature of Hepatitis Viruses to recognize human hepatitis B virus as a member of a unique group of viruses and to classify it, together with a number of related animal viruses, into a new family called the Hepadnaviridae. Over the past decade, the International Committee on Taxonomy of Viruses (ICTV) has been active in the development of a classification system for viruses. The majority of viruses infecting vertebrate hosts have been classified into families and genera on the recommendations of the Vertebrate Virus Subcommittee (VVSC). In June 1980, the VVSC authorized the formation of an ad hoc Study Group on the Nomenclature of Hepatitis Viruses under the Chairmanship of Dr. Ian D. Gust. This paper represents the first report of the Study Group on the Taxonomic Classification of Human Hepatitis B Virus.

NON-A NON-B HEPATITIS ASSOCIATED WITH CHRONIC LIVER DISEASE IN A HÆMODIALYSIS UNIT

Abstract To clarify the aetiology of an outbreak of HB s Ag-negative acute hepatitis in the renal unit at Fulham Hospital in 1968-70, serological tests for antibody to hepatitis-A virus (anti-H.A.V.) were done retrospectively on serum samples obtained at the time of the outbreak. 7 patients had had two previous episodes of clinical HB s Ag-negative hepatitis. Serum samples were available from 24 of the 29 infected patients, and these were paired in 12 instances. There was a slight increase in the titre of anti-H.A.V. in 1 patient, and a further 2 patients who subsequently developed chronic hepatitis showed a decrease in titre, but no changes in titre were detected in the remaining 21 cases. These findings do not provide evidence for the involvement of hepatitis-A virus in the outbreak of hepatitis and effectively exclude a role for this virus in the chronic liver disease which developed subsequently in 8 (28%) of the patients. This outbreak is therefore probably non-A non-B hepatitis, which has not been reported previously in Great Britain in a haemodialysis unit. The results confirm that this form of hepatitis may be related to a high frequency of persistent hepatic dysfunction.

Hepatitis B virus DNA in the hepatocyte: a series of 160 biopsies

DNA-DNA hybridisation was used to examine 160 liver biopsies for the presence of the hepatitis B virus genome. HBeAg-positive HBsAg carriers were found to have replicating viral DNA in the hepatocytes and, very occasionally, HBV DNA was also integrated into the chromosomes. A high proportion of the anti-HBe-positive HBsAg carriers also have replicating HBV DNA and in the remainder integrated sequences are often, but not always, seen. No evidence was found, however, to implicate HBV in HBsAg-negative patients with alcoholic liver disease, nor in patients with a variety of other liver diseases including non-A, non-B hepatitis.

EPIDEMIC HEPATITIS B CAUSED BY COMMERCIAL HUMAN IMMUNOGLOBULIN

An epidemic of acute hepatitis B followed the administration of human immunoglobulin to members of the staff of a mission hospital in India and their families. Jaundice developed in 123 (38%) of 325 persons inoculated. Hepatitis-B surface antigen was detected in three of the batches of immunoglobulin which were available for testing.