Detection of poliovirus antigens and antibodies: microindirect haemagglutination and haemagglutination inhibition tests for poliovirus types I, II, and III.

Microtitre indirect haemagglutination (IHA) and IHA-inhibition (IHAI) procedures were adapted to determine the reactivities of type I, II, and III poliovirus antibodies and antigens. Glutaraldehyde-fixed sheep erythrocytes were sensitized for these tests with concentrated, partially purified preparations of type I, II, and III poliovirus. Antibody titres by IHA were generally 10 to 100 times greater than serum microneutralization (SN) titres. The SN and IHA reactivities of three kinds of sera were compared. Of these sera, virus type specific antibodies, in monospecific guinea pig sera one week after immunization and in sera from hyperimmunized horses, could be readily differentiated and measured; antibodies in human diagnostic specimens, however, showed some intertypic cross reactivity. Monovalent one-week immune guinea pig sera reacted specifically in the IHAI test to differentiate viruses, and could be used for virus typing and differentiating strains of poliovirus type III.     

The trend of acquired immunity with live poliovirus vaccine and the effect of revaccination: follow-up of vaccinees for ten years

The persistence of neutralizing antibody (NA) against three types of poliovirus acquired after two doses of trivalent live attenuated poliovirus vaccine (LPV) has been followed up for ten years in individual vaccinees. Sixty-seven children were bled once a year over a five year period following the primary vaccination. More than 80% of them retained NA against all three types of poliovirus. Thirty-two individuals whose NA titres were 1:16 or over for types 1 and 2 and 1:4 or over for type 3 at the fifth year were further followed up for a further five years and it was shown that during this period some of them had a naturally-acquired antibody rise, mostly against type 3 virus. At the sixth to eighth year after the primary vaccination, one further dose of the trivalent vaccine was administered to the children whose NA titres were down to 1:8 or less and the effect of booster vaccination on NA was followed. Other subjects were revaccinated with LPV and their fecal excretion of the vaccine virus was investigated. The results showed that a decrease in serum antibody level could be a good indicator of the local resistance of the alimentary tract and that reinfection could occur if serum NA had decreased to 1:8 or less, which allowed a virus excretion in the stools.     

Polio antibody outfit of newborns and infants born after 20 years of mass vaccination with live poliovaccine

Antibodies against type 1 and 2 poliovirus were tested in 250 infants and against type 3 poliovirus in 341 infants aged 0-13 months to compare the polio antibody outfit of newborns and infants born to three groups of mothers: (a) nonvaccinated, above 35 years of age; (b) vaccinated with attenuated poliovirus but having had chance of contact with wild poliovirus during childhood, age 22-35 years, and (c) vaccinated but not having had contact with wild poliovirus, below 22 years of age. Over 90% of newborns had specific neutralizing antibodies against all three poliovirus types. With increasing infant age the percentage of antibody-positive infants decreased: by age 9-11 months only 46%, 27% and 14% of infants displayed antibodies to polio-virus types 1, 2 and 3, respectively. Geometric mean titres decreased accordingly. Differences between infants born to mothers of the above three groups were nonsignificant: the results obtained suggest that future newborns will also possess satisfactory levels of antibodies acquired from mothers who have only had contact with attenuated vaccine poliovirus.     

Excretion of attenuated polioviruses in children vaccinated with live oral poliovirus vaccine

The excretion of attenuated polioviruses was studied in a group of nursery children vaccinated with 105TCD50 of each type of virus. The primovaccinated children were found to excrete type 1 poliovirus for 8 weeks, type 2 for 11 weeks after the vaccination with the type 1 + 2 bivaccine. Poliovirus type 1 as eliminated by 78% and type 2 by 98% of the vaccinees. The separately administered type 3 was detectable for 6 weeks and was isolated from 100% of the vaccinees. The highest per cent of children with type 1 excretion positivity was recorded at week 5, with type 2 positivity at week 1 and with type 3 positivity at week 2. The poliovirus excretion peaked early after the vaccination, the titres of the poliovirus type 2 were the highest. The children revaccinated next year with the type 1 + 2 bivaccine eliminated the respective types of virus 1 - 2 weeks; type 3 poliovirus was detectable for 6 weeks after revaccination and was excreted by the highest per cent of vaccines. The contact infections caused by the attenuated polioviruses developed in 9 from 22 children vaccinated previously. The excretion of polioviruses did not last longer than 1 week. The contact infections were most frequently caused by the poliovirus type 2. The examined children, particularly those vaccinated previously, turned out to excrete also other enteroviruses identified as Coxsakieviruses B 4 and B 5 and Echovirus 21. In the primovaccinated these viruses were isolated only from those with the negative excretion of polioviruses.     

Distribution of antibodies to poliovirus in the children of a large industrial city

The results of prolonged dynamic observations on the state of herd immunity against poliomyelitis virus in an industrial city are given. The survey covered 1304 children. The data thus obtained, when synchronized according to years, seasons, the age of the surveyed children and the methods used in the survey, indicated that in every age group 20-30% of children had no antibodies to group I poliomyelitis virus and 30% of children had no antibodies to group III poliomyelitis virus. The geometrical mean of antibody titers to different types of the virus fluctuated from 1.8 to 4.6 log2, the lowest value being obtained for the titer of antibodies to type III poliomyelitis virus. During the whole period of immunological control (1974-1978) no mass circulation of poliomyelitis virus and no outbreaks of poliomyelitis were registered despite the fact that a considerable proportion of children having no antibodies to one or several types of the virus was constantly present among the most susceptible part of children.     

Anti-kappa elastin antibody titer in the sera of patients with hyperlipoproteinemia type II, III and IV]

Titre of antibodies against elastin degradation product (kappa-elastin) was measured in patients with atherogenic types of hyperlipoproteinemia. The hemagglutination technique was used. A significant decrease in titres of the tested antibodies was found. It was the most prominent in IIa, III and IIb types of hyperlipoproteinemia and rather mild in IV type. The authors attempted to explain causes of antibodies titres decrease in tested patients and relate their results with those of other authors.     

Improved distribution of antigenic site specificity of poliovirus-neutralizing antibodies induced by a protease-cleaved immunogen in mice.

Previous studies showed that the distribution of antigenic site specificity of neutralizing antibodies to type 3 poliovirus obtained with the inactivated poliovirus vaccine can be deficient as compared with that obtained following poliovirus infection. This observation was shown by the relatively low capacity of sera from inactivated-poliovirus-vaccine-immunized persons to neutralize poliovirus cleaved at antigenic site 1. We investigated possibilities for improving the situation in a mouse model. Balb/c mice were immunized with intact or trypsin-cleaved type 3 poliovirus (Saukett strain). Sera from mice immunized with the intact virus readily neutralized the intact virus but neutralized the cleaved virus only rarely. In contrast, cleaved-virus-immunized mice produced antibodies that were able to neutralize the cleaved virus as well as the intact one. Mice immunized with a 100-fold-higher dose of the intact virus produced significant levels of antibodies to the cleaved virus, too. Somewhat surprisingly, mice immunized with high doses of the cleaved virus produced antibodies specific for the intact loop between beta sheets B and C of VP1 (virion protein 1), which should be cleaved in the immunogen. This was shown by a higher titer of antibodies to intact Saukett virus than to the corresponding cleaved virus, as well as to a type 1/type 3 hybrid poliovirus in which only the BC loop amino acids were derived from type 3 poliovirus. The cleavage-induced enhanced availability of antigenic determinants residing outside the BC loop was also shown by increased neutralization titers of monoclonal antibodies specific for some of these other determinants. These results indicate that by using a trypsin-cleaved type 3 poliovirus as a parenteral immunogen, it is possible to change the distribution of antigenic site specificities of neutralizing antibodies to resemble that following poliovirus infection.     

Intestinal trypsin can significantly modify antigenic properties of polioviruses: implications for the use of inactivated poliovirus vaccine.

It was recently reported that the intestinal protease trypsin cleaves in vitro the VP1 protein of type 3 poliovirus at antigenic site 1 (J. P. Icenogle, P. D. Minor, M. Ferguson, and J. M. Hogle, J. Virol. 60:297-301, 1986). We found that incubation of purified or crude type 3 poliovirus preparations with specimens of human intestinal fluid brings about a similar change in the virion structure. Sera from children immunized solely with the regular inactivated poliovirus vaccine (IPV) neutralized trypsin-cleaved Sabin 3 virus poorly, if at all, despite moderate levels of antibodies to the corresponding intact virus. Sera containing very high titers of the intact virus also neutralized the trypsin-cleaved virus but at a relatively weaker capacity. Most sera from older persons who may have been exposed to a natural poliovirus infection before the introduction of the poliovirus vaccines as well as sera from children infected with type 3 poliovirus during the recent outbreak in Finland were able to neutralize the trypsin-cleaved type 3 polioviruses. Serum specimens collected 1 month after a single dose of live poliovirus vaccine from children previously immunized with IPV were able to neutralize the trypsin-cleaved virus as well. During natural infection and after live poliovirus vaccine administration polioviruses are exposed to proteolytic enzymes in the gut. Our results may offer an alternative explanation for the relatively weak mucosal immunity obtained with IPV. Improvement of IPV preparations by incorporation of trypsin-treated type 3 polioviruses in the vaccine should be studied.     

Excretion of live attenuated polioviruses in the faeces of orally vaccinated children. Comparison of two immunization schedules

The excretion of live, attenuated poliovirus vaccine strains was determined in the feces of Prague Infants home children given 10(5) PFU of type 1 and 2 and 2.10(5) PFU of type 3 vaccine in a routine annual mass campaign. The first two faeces specimens examined in each vaccinee prior to immunization were negative for the virus. A total of 476 stool specimens were collected from 37 children at weekly intervals for a period of 18 weeks. The presence of type 1 poliovirus in the faeces of children given monovalent type 1 vaccine was detectable for 9 weeks, with a maximum in first week, and the virus was isolated in 74.2% of vaccinees. The timing of bivalent type 2 and type 3 vaccine was 9 weeks after monovalent type 1 immunization. The excretion of these two types of poliovirus was found to persist for at least 6 weeks. Type 2 poliovirus was isolated in all vaccinees, type 3 in 70.4% of children. The highest percentage of children excreting type 2 poliovirus was recorded in the first week, the excretion of type 3 peaked three weeks after bivaccine administration. The excretion peaks were reached relatively early postvaccination, with type poliovirus reaching the highest titre per 1 g of faeces. After revaccination (one year later) with monovalent type 1 vaccine, the vaccine strain of type 1 poliovirus could be detected for 6 weeks and was present in the highest percentage of positive stool samples.(ABSTRACT TRUNCATED AT 250 WORDS)     

A human-human hybridoma which produces antibodies to polioviruses

A human-human hybridoma which produced antibodies to Sabin strain poliovirus types 3 and 2 was established. This hybridoma was constructed by fusing a HAT-sensitive variant of a human lymphoblastoid subline R4-4-6 with peripheral blood lymphocytes of a donor who had been previously immunized with poliovirus Sabin vaccines and had shown high titer of neutralizing antibodies against these viruses. This hybridoma had been cultured for more than one year and the neutralizing activities of the culture supernatants against poliovirus type 3 could still be detected. Cross neutralization with poliovirus type 2 was also observed. Karyotypic analyses of this hybrid showed that the chromosome numbers were distributed mainly between 80 and 100. Dot immunobinding assays also confirmed the reactivities of this hybridoma supernatants with poliovirus type 3 and, slightly, with type 2.     

Colocalization of cytokeratin 18 and villin in type III alveolar cells (brush cells) of the rat lung

Alveoli of the rat lung are lined by three different cell types, the flat type I cells and the cuboidal type II and type III cells. Type III cells differ from type II cells by the presence of an apical tuft of microvilli and the absence of lamellar type secretory granules. In the present study we show by double immunolabelling that type III cells of the rat lung can be identified at the light-and electron microscope level by antibodies against both cytokeratin 18 and the actin-crosslinking protein villin. At the ultrastructural level, microvilli and their rootlets in the apical cytoplasm were labelled by the anti-villin antibodies, whereas a monoclonal antibody against cytokeratin 18 (Ks18.04) labelled bundles of intermediate filaments. In conclusion, antibodies against villin and certain monoclonal antibodies specific for cytokeratin 18 can be used as tools for selective visualization of type III cells in the rat lung.     

Transplacental antibodies. Part III: Maternal antibodies against polioviruses, M. parotitidis and M. parainfluenzae

In 1981 we examined 247 sera for the presence of antibodies against all three types of poliovirus and 253 sera for antibodies against M. parotitidis and three types of M. parainfluenzae viruses. The sera were obtained from the cord blood of mothers between 15 and 34 years of age. All mothers were divided into four age groups, each with primipara and multipara subgroups. The rate of seropositivity for type 1 and type 2 poliovirus-specific antibody was in all age groups higher than 90%, the overall seropositivity rate for type 3 poliovirus antibody was 83.6%, with 73.1% as the lowest rate for age group of youngest mothers. Significant seropositivity variations between the primipara and multipara subgroups were recorded only for type 3 antibody in the two age groups of oldest mothers (25-29 and 30-34 years). This is consistent with the assumed booster effect of Sabin vaccine strains on mothers of families with more than one child. Antibodies specific to mumps virus were present in the cord blood of 78.6% of all mothers and the rates of seropositivity were found to rise with the increasing age. Seropositivity for M. parainfluenzae type 1-specific antibody was demonstrated in 95.8%, for type 2-specific antibody in 98.9% and for type 3-specific antibody in 100% of mothers, which is suggestive of high herd immunity levels in the population.     

Neurovirulence of type 1 polioviruses isolated from sewage in Japan.

Sixteen type 1 poliovirus strains were isolated from a sewage disposal plant located downstream of the Oyabe River in Japan between October 1993 and September 1995. The isolates were intratypically differentiated as vaccine-derived strains. Neutralizing antigenicity analysis with monoclonal antibodies and estimation of neurovirulence by mutant analysis by PCR and restriction enzyme cleavage (MAPREC) were performed for 13 type 1 strains of these isolates. The isolates were classified into three groups. Group I (five strains) had a variant type of antigenicity and neurovirulent phenotype. Group II (four strains) had the vaccine type of antigenicity and neurovirulent phenotype. Group III (four strains) had the vaccine type of antigenicity and an attenuated phenotype. Furthermore, it was demonstrated that the virulent isolates were neutralized by human sera obtained after oral poliomyelitis vaccine (OPV) administration, and the sera of rats immunized with inactivated poliovirus vaccine. Although vaccination was effective against virulent polioviruses, virulent viruses will continue to exist in the environment as long as OPV is in use.     

Antibodies to poliomyelitis virus in cord blood of neonates born to from mothers in contact or not in contact with the wild virus

Neutralizing antibodies to polioviruses in cord blood of neonates born from 64 mothers under age 20 and in 53 mothers aged 30 years and over were investigated in order to know and compare the transfer to newborns of antibodies to polioviruses produced by live oral vaccine mainly and those antibodies induced by natural contact with wild poliovirus strains. Total immunity for the two groups was higher than 80% for the three types of polioviruses, with only virus 3 showing an immunity below 80% (77.4%) in mothers aged 30 years and over. Average geometric titers though relatively low may be considered satisfactory. However, there is a statistically significant difference in titers to poliovirus type 2 (24.9) in mothers over age 30 years as compared to those found in mothers below age 20 years (10.8), for which we have found no explanation. It is not deemed necessary for the time being to take special prophylactic measures with these children given the occurrent epidemiologic status quo.     

Strain-specific antibodies as an indicator of the circulation of wild poliomyelitis viruses and their role in the formation of collective immunity in the population

Essential differences in the intensity of collective immunity to poliomyelitis in the donors of Moscow and Kaluga were established. To find out the nature of high characteristics of collective immunity to poliovirus, types 1 and 2, in the donors of Kaluga, strain-specific antibodies to wild and vaccine polioviruses were studied. In a considerable number of donors strain-specific antibodies to poliovirus, types 1 and 2, were detected. This made it possible to presume a sufficiently wide circulation of these viruses among the population of the city in the middle of the 20th century and, as a consequence, high level of collective immunity appeared. Strain-specific antibodies to poliovirus of type 3 were rarely detected. This made it possible to suggest that the circulation of viruses of this type among the population was limited. Immunity to viruses of this type was due only to immunization. For this reason the characteristics of collective immunity in the donors of Moscow and Kaluga coincided. The detection of strain-specific antibodies to poliomyelitis virus allowed to retrospectively form the opinion of the spread and time of the circulation of wild poliomyelitis viruses in the population.     

Changes in collagen metabolism--another look at osteoarthrosis

Collagen types I and III were isolated from osteoarthrotic cartilage. Immunofluorescence study has shown that these two collagen types are present in the deep layers of cartilage. Additional staining for fibronectin revealed the presence of the cell-adhesive protein in osteoarthrotic cartilage. Neither collagen type I and type III nor fibronectin were found in control cartilage. A passive haemagglutination assay determined anticollagen antibodies (against types I, II and III) in sera of some osteoarthrotic patients.     

Evaluation of poliovirus antibody titers in orally vaccinated semi‐captive chimpanzees in Uganda

Background To understand immunological responses in chimpanzees vaccinated with live‐attenuated vaccine (oral polio vaccine; OPV), serum neutralizing antibodies against poliovirus types 1, 2, and 3 were investigated over time. Methods The neutralizing antibody titers against poliovirus types 1, 2, and 3 were determined by microneutralization test using 100 ID₅₀ of poliovirus types 1, 2, and 3 (Sabin strains). Results Neutralizing antibodies against poliovirus types 1, 2, and 3 were detected in 85.7%, 71.4%, and 65% of the serum from 42 chimpanzees tested 9 years post‐vaccination. The neutralizing antibody titers in chimpanzees were similar to the documented levels in human studies as an indicator of vaccine efficacy. Conclusions This study reveals persistence of neutralizing antibodies in chimpanzees for at least 9 years after vaccination with OPV. This first study in chimpanzees provides useful information for the evaluation of the success of vaccination with OPV in other captive apes.     

Cleavage of VP1 and modification of antigenic site 1 of type 2 polioviruses by intestinal trypsin.

We have exposed 22 independent type 2 poliovirus isolates to human intestinal fluid and purified trypsin. In all cases the virus retained its infectivity, while polyacrylamide gel electrophoresis of viral proteins showed disappearance of the VP1 bands. Concomitantly, the viruses became resistant to antigenic site 1-specific monoclonal antibodies, indicating that the cleavage took place at the antigenic site 1. Sera from persons immunized solely with the inactivated poliovirus vaccine (IPV) neutralized intact type 2 polioviruses more readily than the corresponding trypsin-cleaved virus preparations. The ratio between the neutralization indices for the intact and trypsin-cleaved type 2 polioviruses was not significantly changed by a dose of trivalent oral poliovirus vaccine given to children previously immunized with IPV. These results indicate that while the antigenic site 1 of type 2 poliovirus is immunogenic in humans when IPV is used, the relative role of this antigenic site in human immunity appears to be less critical than that in the case of type 3 polioviruses. Before we obtained these results, only antigenic site 1 had been shown to be immunogenic in type 2 polioviruses.     

Protective antibodies in human sera against encapsulated strains of Staphylococcus epidermidis

Passive protective antibodies in 100 samples of normal human sera against challenge with three representative capsular type strains of Staphylococcus epidermidis in mice were examined. Six of them passively protected mice against capsular type I; 17 protected against capsular type II; one against capsular type III; and one against both capsular types I and II. The activities were sensitive to 2-mercaptoethanol and were absorbed out either with rabbit anti-human IgG serum, rabbit anti-human IgA serum or rabbit anti-human IgM serum. Also, the sera activities absorbed out with cell surface polysaccharide extracted from three representative capsular type strains. These results indicate that the protective activities were specifically related to three major immunoglobulins against the above cell surface polysaccharides.