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PubMed1999

Characterization of the nucleoside triphosphatase activity of poliovirus protein 2C reveals a mechanism by which guanidine inhibits poliovirus replication.

The highly conserved non-structural protein 2C of picornaviruses is involved in viral genome replication and encapsidation and in the rearrangement of intracellular structures. 2C binds RNA, has nucleoside triphosphatase activity, and shares three motifs with superfamily III helicases. Motifs "A" and "B" are involved in nucleotide triphosphate (NTP) binding and hydrolysis, whereas a function for motif "C" has not yet been demonstrated. Poliovirus RNA replication is inhibited by millimolar concentrations of guanidine hydrochloride (GdnHCl). Resistance and dependence to GdnHCl map to 2C. To characterize the nucleoside triphosphatase activity of 2C, we purified poliovirus recombinant 2C fused to glutathione S-transferase (GST-2C) from Escherichia coli. GST-2C hydrolyzed ATP with a Km of 0.7 mM. Other NTPs, including GTP, competed with ATP for binding to 2C but were poor substrates for hydrolysis. Mutation of conserved residues in motif A and B abolished ATPase activity, as did mutation of the conserved asparagine residue in motif C, an observation indicating the involvement of this motif in ATP hydrolysis. GdnHCl at millimolar concentrations inhibited ATP hydrolysis. Mutations in 2C that confer poliovirus resistant to or dependent on GdnHCl increased the tolerance to GdnHCl up to 100-fold.

Acid Anhydride Hydrolases↗

PubMedDate not supplied

Long-term immunity following vaccination with killed poliovirus vaccine in Sweden, a country with no circulating poliovirus.

The long-term immunity following vaccination with killed poliovirus vaccine was analyzed by making repeated antibody surveys. A comparison between the levels of antibody of vaccinated groups of children and young adults in 1968 and 1978 indicated that mean levels of antibody of the age groups that, during the period 1968-1978, had not received any extra booster differed by less than or equal to 0.3 log10. Children born in the period 1964-1967 and young adults born in the period 1960-1963 were compared in 1978. The two groups had received their last booster an average of six and 10 years ago, respectively. The mean level of antibody in the older group, which had received booster approximately 11 years previously, was 0.2 log10 lower against type 1 poliovirus and 0.1 log10 lower against type 3 virus than the corresponding titers of the younger children who had received booster approximately six years earlier. The levels of antibody to poliovirus type 2 were identical in both age groups.

Adolescent↗

PubMed1997

Inactivated poliovirus vaccine alone or sequential inactivated and oral poliovirus vaccine in two-, four- and six-month-old infants with combination Haemophilus influenzae type b/hepatitis B vaccine.

BACKGROUND: Advisory committees have recommended the increased use of inactivated poliovirus vaccine (IPV) for children. OBJECTIVES: The purpose of this study was to assess the safety and immunogenicity of three schedules using IPV administered with diphtheria and tetanus toxoids and whole cell pertussis vaccines in a dual-chambered syringe. Children also received a combination of Haemophilus influenzae type b (Hib) and hepatitis B vaccines (COMVAX). METHODS: All infants (n = 295) received IPV and COMVAX at 2 and 4 months of age and IPV, oral poliovirus vaccine (OPV) or both vaccines at 6 months and OPV at 15 months of age. RESULTS: After two doses of IPV 96 to 100% of infants had antibodies to poliomyelitis viruses types 1, 2 and 3, and after a third dose of vaccine (IPV or OPV) all but one child had antibodies to all three poliovirus types. After two doses of COMVAX 89 and 96% of children had protective levels of antibody to Hib and hepatitis B, respectively. CONCLUSIONS: IPV is highly immunogenic in a two-dose schedule. Administration of a third dose of IPV or a dose of OPV at 6 months of age is highly effective. Simultaneous administration of a combination H. influenzae type b/hepatitis B vaccine did not interfere with the response to IPV.

Bacterial Capsules↗

PubMed1987

Evolution of poliovirus during an outbreak: sequential type 3 poliovirus isolates from several persons show shifts of neutralization determinants.

An outbreak of poliomyelitis in Finland resulted in the widespread circulation of wild-type 3 poliovirus strains that had antigenic properties distinct from the strains used to produce the attenuated and inactivated vaccines. Considerable variation was observed in the ability of broadly reacting monoclonal antibodies directed against type 3 poliovirus to neutralize the 54 strains examined. Sequential isolates from several persons showed an antigenic drift with these monoclonal antibodies and selected human sera. In addition, some faecal specimens were found to contain more than one antigenic variant. Primer extension sequencing of genomic RNAs of three plaque-purified antigenic variants isolated from one patient showed base substitutions in the region coding for the major antigenic site of poliovirus type 3. The resulting difference in the amino acid sequence in the virion protein VP1 could explain the differences observed in the neutralization of these strains by the monoclonal antibodies. Whether the observed changes in the antigenic characteristics of the sequential isolates represent true antigenic drift under immunological pressure or whether the emergence of the new variants is based on other modes of selection during replication is not known.

Antibodies, Monoclonal↗

PubMed1991

Analysis of neutralization-escape mutants selected from a mouse virulent type 1/type 2 chimeric poliovirus: identification of a type 1 poliovirus with antigenic site 1 deleted.

A chimeric type 1/type 2 poliovirus (v510), in which the antigenic site 1 (Ag1) of poliovirus type 1 (PV-1) Mahoney was replaced by the corresponding site of poliovirus type 2 (PV-2) Lansing, is known to be neurovirulent for mice and neutralized by both type 1 and type 2 monoclonal antibodies. Neutralization-escape mutants to monoclonal antibodies specifically recognizing the PV-2 sequence were obtained from v510. The nucleotide sequence and the mouse neurovirulence of mutants were determined. Amino acid substitutions obtained inside the replaced sequence, at positions 95 and 99, and outside this site, at positions 93 or 104, rendered the virus attenuated for mice. One of the escape mutants harboured a deletion of the entire substituted nonapeptide sequence in v510. This particular virus, which is a PV-1 Mahoney lacking the natural Ag1 loop, does not react with PV-2-specific monoclonal antibodies, has a ts phenotype, is heat-labile and is devoid of neurovirulence for mice.

Amino Acid Sequence↗

PubMed2006

Risk of introduction of poliovirus into a Dutch Cape Verdian community during an outbreak of poliovirus in Cape Verde, 2000.

OBJECTIVE: To assess the risk of introduction of polio virus in a Cape Verdian community of Rotterdam, during the polio epidemic in Cape Verde in 2000. METHODS: All 225 insufficiently vaccinated 0-14-year-old Cape Verdian children (n=4188) and a random sample of 285 out of all 15-30-year-old Cape Verdians (n=5074) in Rotterdam were surveyed to assess travel behaviour and vaccination coverage. Faecal specimens were collected and sewage samples taken in neighbourhoods with a sizable Cape Verdian population for testing of polio virus. RESULTS: During the polio epidemic in Cape Verde, 10% of insufficiently vaccinated children aged 0-14 years and 17% of adults aged 15-30 years living in Rotterdam reported travelling to Cape Verde. 94.6% of Cape Verdians in Rotterdam aged 0-14 years were sufficiently vaccinated against polio, but 9 of 91 insufficiently vaccinated children had travelled to Cape Verde during the epidemic. Of those aged 15-30 years, 10% were not vaccinated against polio. In the faeces of 80 insufficiently vaccinated individuals aged 0-14 years and in 74 adults aged 15-30 years, no poliovirus was detected. Samples of sewage from six sites were negative for poliovirus. CONCLUSION: No evidence of poliovirus infection was found in the Cape Verde population in Rotterdam despite extensive travel to the Cape Verde during the outbreak.

Adolescent↗

PubMed1991

Specific antibodies to poliovirus type I in breastmilk of unvaccinated mothers before and seven years after start of community-wide vaccination of their infants with live, oral poliovirus vaccine.

Secretory IgA (SIgA) antibodies against poliovirus type 1 were determined using the ELISA method in breastmilk samples obtained each month from 100 young, healthy, unvaccinated mothers living in urban slum areas of Lahore, Pakistan. The study covered two different groups, one in 1980-1981 and the other in 1987, before and seven years after a nation-wide expanded programme of childhood immunization (EPI) had started. The SIgA titres did not change neither with duration of lactation nor with time after vaccination in the infants of the mothers studied. The seasonal breastmilk IgA antibody titres to poliovirus type 1 corresponded to the epidemiological conditions existing both before (1980-81) and after general vaccination coverage with live, oral poliovirus vaccine (OPV) had reached 80% of the infant population (1987). Neutralization titres did not seem to correlate well with ELISA titres although colostrum samples had high levels of neutralizing antibodies. The wide variation between high (greater than 10,000) and low (less than 500) individual breastmilk IgA antibody titres observed during various seasons could be of consequence for the breast-fed baby. Colostrum, which was also found to have significant neutralization capacity, might interfere with the OPV now often given on the day of birth.

Adult↗

PubMed1970

Characterization of some poliovirus temperature-sensitive mutants and poliovirus-related particle formation under nonpermissive conditions.

Investigation of 15 poliovirus temperature-sensitive (ts) mutants by using physiological tests [formation of virus-specific antigen and ribonucleic acid (RNA) under nonpermissive conditions] permitted us to divide them into three groups. From each group, one mutant was selected (ts 2, 5, 11), and a comparative study of poliovirus-related particle (5, 10, 73, and 150S) formation under permissive (36 C) and nonpermissive (40 C) conditions was carried out. The ts 2 and ts 11 are mutants with greatly reduced RNA synthesis which at 40 C produce particles with a sedimentation constant of 5S, and the ts 5 (RNA(+)) mutant produces both 5 and 10S particles. The relationship between different temperature-sensitive defects in the mutants is discussed. The results obtained indicate a possible role of 5S protein structures in morphogenesis of poliovirus.

Adsorption↗

PubMed1997

Poliovirus infection and expression of the poliovirus protein 2B provoke the disassembly of the Golgi complex, the organelle target for the antipoliovirus drug Ro-090179.

Infection of Vero cells with poliovirus results in complete disassembly of the Golgi complex. Milestones of the process of disassembly are the release to the cytosol of the beta-COP bound to Golgi membranes, the disruption of the cis-Golgi network into fragments scattered throughout the cytoplasm, and the disassembly of the stacked cisternae by a process mediated by long tubular structures. Transient expression of the viral protein 2B in COS-7 cells also causes the disassembly of the Golgi complex by a process preceded by the accumulation of the protein in the Golgi area. Vero cells infected for 3 h show no recognizable Golgi complexes at the ultrastructural level and display an enormously swollen endoplasmic reticulum (ER) with extensive areas of its surface heavily coated. Ro-090179 (Ro), a flavonoid isolated from the herb Agastache rugosa, provokes the specific swelling and disruption of the Golgi complex and strongly inhibits poliovirus infection. Ro provokes the swelling and the disruption of the stacked cisternae and trans-Golgi elements without affecting the cis-most Golgi cisternae much. Moreover, Ro inhibits the fusion of the Golgi complex with the ER in cells treated with brefeldin A and provokes the accumulation of the intermediate compartment membrane protein p58 into ERD2-positive Golgi elements but has no effect on the anterograde transport involved in protein secretion. Our results indicate that the secretory pathway and specifically the Golgi complex are preferential targets of poliovirus.

Animals↗

PubMed1976

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.

Animals↗

PubMed1994

Studies with poliovirus polymerase 3Dpol. Stimulation of poly(U) synthesis in vitro by purified poliovirus protein 3AB.

The synthesis in vitro of poly(U) on a poly(A) template with oligo(dT)15 primer by poliovirus RNA polymerase 3Dpol (280 ng/ml) is strongly stimulated (50-100 fold) by the addition of purified poliovirus polypeptide 3AB. The synthesis of product continues linearly with time for up to 90 min. The reaction with 3Dpol alone can be reactivated and similarly enhanced by the addition of 3AB at 30 min of incubation. Optimal stimulation is achieved under conditions where the concentration of 3Dpol and of template is low, when the molar ratio of 3AB to 3Dpol is about 100:1 and that of 3AB to poly(A) is about 25:1. In the presence of 3AB, the yield of product made by 3Dpol is much increased but its size is unchanged. From a number of basic proteins and peptides tested, a few were found which also exhibited limited enhancement of polymerase activity. The stimulatory effect of 3AB is probably related to its ability to bind both the template-primer, poly(A).oligo(dT)15, and 3Dpol (Molla, A., Harris, K. S., Paul, A. V., Shin, S. H., Mugavero, J., and Wimmer, E. J. (1994) J. Biol. Chem. 269, 27015-27020). RNA synthesis on purified poliovirus RNA with oligo(dT)15 primer is enhanced by 3AB about 5-10 fold, and this reaction is highly sensitive to detergent.

Amino Acid Sequence↗

PubMed1997

Combined immunization of infants with oral and inactivated poliovirus vaccines: results of a randomized trial in The Gambia, Oman, and Thailand. WHO Collaborative Study Group on Oral and Inactivated Poliovirus Vaccines.

To assess an immunization schedule combining oral (OPV) and inactivated poliovirus vaccines (IPV), a clinical trial was conducted in The Gambia, Oman, and Thailand. Children were randomized to receive OPV at birth and at 6, 10, and 14 weeks of age; OPV at birth followed by both OPV and IPV at 6, 10, and 14 weeks of age; or placebo at birth followed by IPV at 6, 10, and 14 weeks of age. Serum specimens were available at 24 weeks for 1291 (77%) of 1685 enrolled infants. In the combined-schedule group, the proportion of children seropositive at 24 weeks was 95%-99% for type 1, 99%- 100% for type 2, and 97%-100% for type 3. In The Gambia and Oman, the combined schedule performed significantly better than OPV for type 1 (95%-97% vs. 88%-90%) and type 3 (97%-99% vs. 72%-73%). Across the study sites, IPV given at 6, 10, and 14 weeks of age provided inadequate protection against poliovirus. The combined schedule provided the highest levels of serum antibody response, with mucosal immunity equivalent to that produced by OPV alone.

Antibodies, Viral↗

PubMed1999

Poliovirus vaccination: current understanding of poliovirus interactions in humans and implications for the eradication of poliomyelitis.

Poliomyelitis is a paralytic disease of the motor neurones of the central nervous system, which is caused by poliovirus. The virus is transmitted by the faecal-oral route, and if virus replication is confined to the gut, it is harmless. Poliomyelitis is an ancient human disease, but was rare until the beginning of the 20th century, when children began to be exposed to the virus at older ages and were, therefore, no longer protected by maternal antibody, which had already been lost. Inactivated polio vaccines are increasingly being used in those countries in which poliomyelitis has been brought under control; however, live vaccines are still the most widely used types and the World Health Organization (WHO) have set the goal of using such vaccines to eliminate the wild-type virus throughout the world by the year 2000. Substantial progress has been made to this end; however, the strains of poliovirus that are used as vaccines are able to adapt rapidly to the human gut, losing their attenuated (weakened) character within a few weeks. Currently, there is urgent debate about the best method of stopping vaccination against poliomyelitis once the wild-type poliovirus has been eliminated completely, so that the vaccine-strain virus will also be eliminated. Proposed strategies include the abrupt cessation of vaccination with the live virus worldwide, followed by the optional use of inactivated vaccines for an appropriate period. Further information about both the epidemiology and the pathogenesis of the disease is required before an informed choice can be made. The topics covered in this article include a brief history of studies of the disease, its pathogenesis and its control by vaccination, the molecular biology of the live vaccines, which have been extremely successful in controlling poliomyelitis so far, and the concerns that are raised as the eradication of the wild-type virus approaches.

Journal Article↗

PubMed1995

Microevolution of type 3 Sabin strain of poliovirus in cell cultures and its implications for oral poliovirus vaccine quality control.

Screening for sequence heterogeneities in Sabin Type 3 strains of attenuated poliovirus demonstrated mutations that consistently accumulate to significant levels following 10 passages in cultures of primary African green monkey kidney (AGMK) cells or continuous cultures of Vero cells. Fourteen newly identified mutations were quantified by mutant analysis by PCR and restriction enzyme cleavage in passages and in batches of commercial vaccines made in AGMK and Vero cells from the Sabin original (SO) seed virus and from a seed virus rederived by RNA plaque purification (RSO or "Pfizer" seed). Nine of the 14 mutations were reproducibly observed in more than one series of passages. Although 5 other mutations were observed in only one set of passages each, their content gradually increased to a high percentage, suggesting that all the mutations that we found accumulated consistently. SO-derived samples accumulated more mutations than did RSO-derived ones, and the number of mutations and the rates of their accumulation were higher in Vero than in AGMK cells. While the rates of accumulation of most mutations were higher when passaging was performed at 37 degrees, a U-->C transition at nucleotide 5832 occurred faster at 34 degrees, the temperature used for vaccine production. Analysis of Type 3 oral poliovirus vaccine (OPV) monopools made by six manufacturers found only 5 of these newly identified mutations in vaccine batches (nucleotides 3956, 4935, 5357, 5788, and 5832). Some of the mutations were found in trace amounts (less than 0.1%) while others were present at up to 1.8% levels. The pattern of these mutations was characteristic for the type of seed virus and the cell substrate but demonstrated no correlation with results of the monkey neurovirulence test. Therefore the only mutation occurring in Type 3 OPV which contributed to neurovirulence in monkeys was the previously described reversion at nucleotide 472. Quantitation of reversion at nucleotide 472 can be utilized for assessment of acceptability of vaccine lots, while other mutations can be used for monitoring the consistency of vaccine production.

Animals↗

PubMed1985

Natural variants of the Sabin type 1 vaccine strain of poliovirus and correlation with a poliovirus neutralization site.

Independent substitution mutations have been detected in capsid polypeptide VP1 of the type 1 oral poliovirus vaccine isolated from normal infant vaccine recipients. These mutations map at amino acid residues 142 and 147 of VP1, a region only minimally hydrophilic. A synthetic peptide, corresponding to residues 141 to 147 of VP1 was synthesized, conjugated to a carrier polypeptide of bovine serum albumin. The conjugate was found to elicit a weak poliovirus neutralizing antibody response. It was also capable of priming the immune system for the production of IgG-type antibodies able to neutralize greater than 99.999% of infectious type 1 virus. It is suggested that region 141 to 147 of VP1 may be involved in neutralization of the virus and that the mutants may have accumulated by antibody selection.

Antigen-Antibody Complex↗

PubMed1993

Long-term immunity to poliovirus in children immunized with live attenuated and enhanced-potency inactivated trivalent poliovirus vaccines.

Eighty-six children who completed immunization with the two trivalent poliovirus vaccines, live attenuated (OPV) and enhanced potency inactivated (EIPV), in one of four schedules (OPV-OPV-OPV, EIPV-EIPV-EIPV, EIPV-OPV-OPV, and EIPV-EIPV-OPV) at 1 year of age were monitored serologically over the subsequent 4 years and challenged with OPV at 5 years of age. Each of the immunization groups exhibited an initial 10- to 100-fold decline in neutralizing antibody to poliovirus types 1, 2, and 3 during the first 2 years of follow-up; thereafter antibody titers stabilized. The EIPV-EIPV-OPV group maintained the highest antibody levels throughout the observation period, including the response to OPV challenge at 5 years of age. These data suggest that immunization with OPV, EIPV, and combinations of the two vaccines confers long-term immunity. Optimal systemic immunity was associated with two or more doses of EIPV.

Antibodies, Viral↗

PubMed1995

Factors affecting the immunogenicity of oral poliovirus vaccine: a prospective evaluation in Brazil and the Gambia. World Health Organization Collaborative Study Group on Oral Poliovirus Vaccine.

To assess factors that may influence the immunogenicity of oral poliovirus vaccine (OPV), neutralizing antibody responses were measured in 1409 infants in Brazil and the Gambia who were randomized to receive one of four different formulations of OPV at approximately birth and at 6, 10, and 14 weeks. Overall seroconversion rates at the end of the trial were 85% for poliovirus type 1 (P1), 94% for type 2 (P2), and 68% for type 3 (P3). Factors associated with vaccine failure included high levels of maternal antibody (P1, P2, and P3), vaccination during the rainy season (P1, P2, and P3), diarrhea at the time of vaccination (P2 and P3), household exposure to other OPV recipients (P1), and breast-feeding (P3) (P < .05 for each factor, logistic regression analysis). OPV containing twice the standard potency of Sabin type 1 virus increased seroconversion rates to P1 by 8% in Brazil (P < .05) and 15% in the Gambia (P < .001). Suboptimal responses to OPV in developing countries are determined by a complex array of factors related to the vaccine, host, and environment.

Antibodies, Viral↗

PubMed1983

The nucleotide sequence of poliovirus type 3 leon 12 a1b: comparison with poliovirus type 1.

The complete nucleotide sequence of the genome of the Sabin vaccine strain of poliovirus type 3 (P3/Leon 12 a1 b) has been determined from cDNA cloned in E. coli. The genome comprises a 5' non-coding region of 742 nucleotides, a large open reading frame of 6618 nucleotides (89% of the sequence) and a 3' non-coding region of 72 nucleotides. There is 77.4% base-sequence homology and 89.6% predicted amino-acid homology between types 1 and 3. Conservation of all glutamine-glycine and tyrosine-glycine cleavage sites suggests a mechanism of polyprotein processing similar to that established for poliovirus type 1.

Amino Acid Sequence↗