Immunogenicity of oral poliovirus vaccine administered in mass campaigns versus routine immunization programmes.

Reported are the results of a study to investigate the immunogenicity of oral poliovirus vaccine (OPV) when administered in mass campaigns compared with that following routine immunization programmes. For this purpose, paired sera were collected from a cohort of children before and after a mass vaccination with OPV in Morocco in 1987. Serum samples and information on vaccination status and other confounding factors that could influence antibody responses to OPV were collected. Neutralizing antibody titres to poliovirus types 1, 2 and 3 were determined using a standardized assay. OPV doses administered exclusively during the mass campaign were consistently associated with higher type-specific seroprevalence rates than the same number of doses administered in the routine programme. These findings could not be attributed to differences in confounding factors. Enhanced secondary spread of vaccine virus may have occurred but could not be demonstrated because of limitations in the study design. Mass campaigns appear to be highly effective in raising the dose-related poliovirus type-specific immunity of the population above that achieved by the routine immunization programme. Our findings support the continued use of mass campaigns as an adjunct to routine programmes in order to both enhance and catalyse current efforts to achieve the global eradication of poliomyelitis by the year 2000.

PIP: Reported are the results of a study to investigate the immunogenicity of oral poliovirus vaccine (OPV) when administered in mass campaigns compared with that following routine immunization programs. For this purpose, paired sera were collected from a cohort of children before and after a mass vaccination with OPV in Morocco in 1987. Serum samples and information on vaccination status and other confounding factors that could influence antibody responses to OPV were collected. Neutralizing antibody titers to poliovirus types 1, 2, and 3 were determined using a standardized assay. OPV doses administered exclusively during the mass campaign were consistently associated with higher type-specific seroprevalence rates than the same number of doses administered in the routine program. These findings could not be attributed to differences in confounding factors. Enhanced secondary spread of vaccine virus may have occurred but could not be demonstrated because of limitations in the study design. Mass campaigns appear to be highly effective in raising the dose-related poliovirus type-specific immunity of the population above that achieved by the routine immunization program. These findings support the continued use of mass campaigns as an adjunct to routine programs in order to both enhance and catalyze current efforts to achieve the global eradication of poliomyelitis by the year 2000. (author's)

Availability of vaccines of an assured quality.

In establishing standards of quality for vaccines used in vaccination programmes, it is necessary to protect the recipients without demanding unnecessarily high specifications and standards of purity. Such standards reduce profitability, jeopardizing further vaccine supplies and future research, and also diminish the probability of vaccine manufacture outside the industrialized countries. Funds will be needed to encourage research and local manufacture, and internationally accepted procedures of quality assurance must be established.

Durability of immunity to diphtheria, tetanus and poliomyelitis after a three dose immunization schedule completed in the first eight months of life.

Blood samples were obtained from school entrants whose primary immunization schedule had consisted of three doses of DT or DTP vaccine and three doses of OPV all given before the age of 8 months. The sera were separated and assayed for diphtheria antitoxin, tetanus antitoxin and antibodies to the three serotypes of poliovirus. The results of the assays showed that the abbreviated three dose schedule induced satisfactory immunity to all five infections until school entry and that a reinforcing dose at 18 months was unnecessary.

The temperature sensitivity of the Sabin type 3 vaccine strain of poliovirus: molecular and structural effects of a mutation in the capsid protein VP3.

The growth of the Sabin strain of type 3 poliovirus is reduced at high temperatures compared to that of its virulent precursor strain Leon. Recombinant viruses have been generated from infectious cDNA clones and demonstrate that the temperature-sensitive (ts) phenotype is mainly attributable to a difference in residue 91 of the virion protein VP3. Examination of non-ts mutants derived in vitro or in vivo reveals the existence of second site mutations some of which are clearly able to suppress the ts phenotype. The location of residue 91 of VP3, and of a number of candidate suppressor mutations, in the atomic structure of the virion suggests that the ts phenotype may result in destabilization of the particle and that the suppressors may function by stabilizing specific interfaces. It is not yet clear whether the ts phenotype is expressed at the level of the particle or in the form of defects in assembly or uncoating of the virion, or all three.

Genetic basis of attenuation of the Sabin type 3 oral poliovirus vaccine.

The poliovirus type 3 Sabin oral poliovirus vaccine strain P3/Leon/12a1b differs in nucleotide sequence from its neurovirulent progenitor P3/Leon/37 by just 10 point mutations. The contribution of each mutation to the attenuation phenotype of the vaccine strain was determined by the construction of a series of recombinant viruses from infectious cDNA clones. The neurovirulence testing of recombinant viruses indicated that the attenuation phenotype is determined by just two point mutations: a C to U in the noncoding region at position 472 and a C to U at nucleotide 2034 which results in a serine-to-phenylalanine amino acid substitution in the structural protein VP3.

Antibody state to poliovirus in first year university students, 1984.

Circulating antibodies to poliovirus were estimated in a group of 300 British and 84 foreign first year students who registered at the health centre of Nottingham University in 1984. Detectable antibodies to all three poliovirus serotypes were found in 212 (71%) of the British students but in only 47 (56%) of those from abroad. Most of the British students (280; 93%) had been born in 1965 or 1966, when uptake of poliomyelitis vaccine was declining. Immunisation histories showed that 10 British and 29 foreign students (3% and 35%) had no record of any immunisation; only five British and two foreign students, however, were negative for all three poliovirus serotypes. These findings provide evidence that a high proportion of British born people aged 18-29 have adequate circulating poliovirus antibodies despite incomplete immunisation schedules. Though this is reassuring, the absence of antibodies in some students and the lack of previous immunisation against poliomyelitis in 39 suggest that reinforcing doses of vaccine at the time of leaving school or beginning further education are still warranted, particularly for students from other countries. The findings also emphasise the need for accurate immunisation records.

Conservation in vivo of protease cleavage sites in antigenic sites of poliovirus.

Polioviruses possess three major antigenic sites which have been located chemically and structurally on the particle. One of these sites, designated site 1, is strongly immunodominant for serotype 3, but highly immunorecessive for type 1. We report that monoclonal antibodies directed against site 1 of type 1 poliovirus may be isolated by an altered route of immunization of the donor mice. Site 1 is shown to be highly variable for type 1, but highly conserved for type 3 poliovirus, although the converse would be predicted from their immunodominance. The evidence presented suggests that the antigenic conservation is associated with a strong selective pressure for a proteolytic cleavage site within site 1 of type 3. As proteolytic cleavage results in the loss of the antigenicity of site 1 the presence of the cleavage site in a virus replicating in the gut in the presence of proteases would protect the virus from neutralizing antibodies directed against uncleaved site 1. The conservation of the site in type 3 is thus consistent with the view that site 1 is a significant target of a human as well as a murine immune response against type 3.

Antigenic and molecular properties of type 3 poliovirus responsible for an outbreak of poliomyelitis in a vaccinated population.

Virus isolated from an outbreak of poliomyelitis in Finland has been examined serologically and at the molecular level. The causative agent was an antigenically unusual strain of type 3 poliovirus, which was unrelated to the strains used to manufacture either live or killed poliovaccines. It is likely that the antigenic properties of the virus played a part in establishing a limited outbreak of poliomyelitis in a vaccinated population.

WHO collaborative study on the use of monoclonal antibodies for the intratypic differentiation of poliovirus strains.

An international collaborative study was carried out to identify monoclonal antibodies that could reliably discriminate between wild polioviruses and strains derived from Sabin vaccine viruses. For poliovirus types 2 and 3, monoclonal antibodies were identified that reacted specifically with type 2 or type 3 strains which gave T1-oligonucleotide maps similar to or indistinguishable from that of Sabin vaccine virus, thus indicating their vaccine origin. These monoclonal antibodies failed to react with strains which gave T1 maps unrelated to that of Sabin vaccine virus. However for type 1, five of the six antibodies examined in the study reacted only with strains with a T1 map indistinguishable from that of type 1 Sabin vaccine virus. In contrast, other monoclonal antibodies against poliovirus types 1, 2 and 3 reacted broadly within a serotype.

Induction by synthetic peptides of broadly reactive, type-specific neutralizing antibody to poliovirus type 3.

A region of virus capsid protein VP1 located 89-100 amino acids from the N-terminus has been proposed to comprise a major antigenic site involved in the neutralization of poliovirus type 3. Synthetic peptides 10-18 amino acids in length, containing all or part of this sequence, were tested for their ability to induce antiviral antibodies. Rabbits, but not guinea pigs or mice, immunized with the most active peptide, developed hightitered, type-specific, neutralizing antibodies for a wide range of poliovirus type 3 strains. Consistent with the broad type specificity of the antibody response was the observation that amino acids 89-100 of VP1 are highly conserved among different poliovirus type 3 strains. This sequence thus appears to provide, at least in part, a molecular basis for serotype antigenic specificity. Individual amino acids from 93 to 98 within this sequence were shown to be important for the neutralization of virus by antipeptide sera by examination of the ability of the sera to neutralize laboratory-derived poliovirus type 3 mutants with known single amino acid substitutions in the proposed antigenic site.

The standardization of infectivity titrations of poliovaccines--a WHO collaborative study.

The reproducibility of a method for infectivity titrations of live poliovaccines using microtitre plates was investigated in a WHO collaborative study involving eight laboratories. The large variation (up to 100-fold) in estimates of infectivity observed between laboratories using their local methods of assay was reduced to no more than fourfold when a common method was used. However, expressing infectivities relative to those of the monovalent reference viruses improved the agreement between the laboratories irrespective of the titration method employed. On the basis of these results, WHO adopted the common method used in the study as its recommended method for poliovirus titrations and established the preparations studied as international reference materials for the infectivity titrations of live poliovaccines.

Antigenic characterization of poliovirus type 3 using monoclonal antibodies.

Hybridoma cell lines secreting monoclonal antibodies to type 3 poliovirus were prepared, and their reactivity with infectious virus (D antigen), empty particles (C antigen), and isolated virion capsid proteins ( VPs ) were examined. Eight antibodies reacted with epitopes common to D and C antigens, and all of these possessed high titers of neutralizing activity. However, only 12 of 19 antibodies that reacted exclusively with D antigen neutralized virus infectivity, and some of these reacted only with strains of virus with T1-oligonucleotide maps identical or similar to that of Sabin vaccine polio virus. These antibodies will be of value in identifying strains of virus derived from Sabin vaccine. None of the 20 monoclonal antibodies that neutralized type 3 poliovirus strains reacted in immunoblot experiments with isolated virion capsid proteins. However, six of the 24 antibodies that reacted only with noninfectious C antigen bound to VP1 and VP3, and three of these antibodies also reacted with proteins of poliovirus types 1 or 2. The lack of reactivity of neutralizing monoclonal antibodies with isolated viral proteins suggests that the antigenic properties of proteins are determined by their arrangement in the virus and not simply by amino acid sequence.

Antigenic analysis of poliovirus type 3 using monoclonal antibodies.

Hybridoma cell lines secreting monoclonal antibodies to type 3 poliovirus have been prepared and their reactivity with infectious virus ('D' antigen), empty particles ('C' antigen) and isolated virus capsid proteins examined. Eight antibodies reacted with epitopes common to 'D' and 'C' antigen and all of these possessed high titres of neutralizing activity. However only 12 out of 19 antibodies which reacted exclusively with 'D' antigen neutralized virus infectivity and some of these reacted only with strains of virus with T1 oligonucleotide maps identical or similar to that of Sabin vaccine virus. These antibodies will be of value in identifying strains of virus derived from Sabin vaccine. None of the 20 monoclonal antibodies which neutralized type 3 poliovirus strains reacted in immunoblot experiments with isolated virus capsid proteins. However, 6 of the 24 antibodies which reacted only with non-infectious 'C' antigen bound to VP1 and VP3 and 3 of these antibodies also reacted with proteins of types 1 or 2 poliovirus. The lack of reactivity of neutralizing monoclonal antibodies with isolated viral proteins suggests that the antigenic properties of proteins are determined by their arrangement in the virus.

Neutralization epitopes on poliovirus type 3 particles: an analysis using monoclonal antibodies.

Monoclonal antibodies to poliovirus type 3 secreted by 51 hybridoma cell clones have been characterized in terms of (i) virus-neutralizing properties, (ii) reactivity in antigen-blocking tests with infectious, 155S ('D' antigen) and empty 80S ('C' antigen) poliovirus particles and (iii) reactivity in immunoblot tests with the isolated protein components of the poliovirus capsid. The antibodies could be separated into three groups on the basis of their reactivities with 'D' and 'C' antigens. All antibodies that reacted with both 'D' and 'C' antigen had potent neutralizing activity. Only a proportion of antibodies that reacted uniquely with 'D' antigen possessed neutralizing activity. Unexpectedly, one of 24 'C' antigen-specific antibodies inhibited virus growth. None of the antibodies that possessed virus-neutralizing activity reacted with isolated poliovirus capsid proteins, although the majority of these have been shown in previous studies to be specific for VP1 on intact virus particles. These findings suggest that antigenic determinants involved in virus neutralization do not survive the denaturing conditions required for the isolation of poliovirus capsid proteins and consequently are likely to be specified by the structural conformation of VP1 rather than by amino acid sequence alone. However, several of the antibodies which bound uniquely to 'C' antigen reacted in immunoblot tests, five with VP1 and one with VP3. Some of these antibodies also possessed heterotypic reactivity with the corresponding capsid proteins separated from other poliovirus types.

A W.H.O. collaborative study of in vitro and in vivo methods for the assay of yellow fever vaccines.

An international collaborative study was carried out to compare the plaque assay in monolayer cell cultures with the mouse assay for the potency testing of yellow fever vaccines. Twelve laboratories assayed four preparations of yellow fever virus by both methods. Differences were found between estimates of infectivities obtained by different laboratories using the plaque assay. There was also large variation between the estimates of mouse LD50 obtained by different laboratories. However, expressing potencies of the preparations relative to a common preparation greatly reduced the variation between laboratories for both the plaque and the mouse tests; it also resulted in remarkably close agreement between the methods.