Sabin Oral Poliovirus Vaccine Research Articles

Safety of the novel oral poliovirus vaccine type 2 (nOPV2) in infants and young children aged 1 to

Novel oral poliovirus vaccine type 2 (nOPV2) has been engineered to improve the genetic stability of Sabin oral poliovirus vaccine (OPV) and reduce the emergence of circulating vaccine-derived polioviruses. This trial aimed to provide key safety and immunogenicity data required for nOPV2 licensure and WHO prequalification. This phase 3 trial recruited infants aged 18 to <52 weeks and young children aged 1 to <5 years in The Gambia. Infants randomly assigned to receive one or two doses of one of three lots of nOPV2 or one lot of bivalent OPV (bOPV). Young children were randomised to receive two doses of nOPV2 lot 1 or bOPV. The primary immunogenicity objective was to assess lot-to-lot equivalence of the three nOPV2 lots based on one-dose type 2 poliovirus neutralising antibody seroconversion rates in infants. Equivalence was declared if the 95% CI for the three pairwise rate differences was within the -10% to 10% equivalence margin. Tolerability and safety were assessed based on the rates of solicited adverse events to 7 days, unsolicited adverse events to 28 days, and serious adverse events to 3 months post-dose. Stool poliovirus excretion was examined. The trial was registered as PACTR202010705577776 and is completed. Between February and October, 2021, 2345 infants and 600 young children were vaccinated. 2272 (96·9%) were eligible for inclusion in the post-dose one per-protocol population. Seroconversion rates ranged from 48·9% to 49·2% across the three lots. The minimum lower bound of the 95% CIs for the pairwise differences in seroconversion rates between lots was -5·8%. The maximum upper bound was 5·4%. Equivalence was therefore shown. Of those seronegative at baseline, 143 (85·6%) of 167 (95% CI 79·4-90·6) infants and 54 (83·1%) of 65 (71·7-91·2) young children seroconverted over the two-dose nOPV2 schedule. The post-two-dose seroprotection rates, including participants who were both seronegative and seropositive at baseline, were 604 (92·9%) of 650 (95% CI 90·7-94·8) in infants and 276 (95·5%) of 289 (92·4-97·6) in young children. No safety concerns were identified. 7 days post-dose one, 78 (41·7%) of 187 (95% CI 34·6-49·1) infants were excreting the type 2 poliovirus. nOPV2 was immunogenic and safe in infants and young children in The Gambia. The data support the licensure and WHO prequalification of nOPV2. Bill & Melinda Gates Foundation.

Health Economic Analysis of Antiviral Drugs in the Global Polio Eradication Endgame.

Polio antiviral drugs (PAVDs) may provide a critical tool in the eradication endgame by stopping poliovirus infections in immunodeficient individuals who may not clear the virus without therapeutic intervention. Although prolonged/chronic poliovirus excreters are rare, they represent a source of poliovirus reintroduction into the general population. Prior studies that assumed the successful cessation of all oral poliovirus vaccine (OPV) use estimated the potential upper bound of the incremental net benefits (INBs) of resource investments in research and development of PAVDs. However, delays in polio eradication, OPV cessation, and the development of PAVDs necessitate an updated economic analysis to reevaluate the costs and benefits of further investments in PAVDs. Using a global integrated model of polio transmission, immunity, vaccine dynamics, risks, and economics, we explore the risks of reintroduction of polio transmission due to immunodeficiency-related vaccine-derived poliovirus (iVDPV) excreters and reevaluate the upper bound of the INBs of PAVDs. Under the current conditions, for which the use of OPV will likely continue for the foreseeable future, even with successful eradication of type 1 wild poliovirus by the end of 2023 and continued use of Sabin OPV for outbreak response, we estimate an upper bound INB of 60 million US$2019. With >100 million US$2019 already invested in PAVD development and with the introduction of novel OPVs that are less likely to revert to neurovirulence, our analysis suggests the expected INBs of PAVDs would not offset their costs. While PAVDs could play an important role in the polio endgame, their expected economic benefits drop with ongoing OPV use and poliovirus transmissions. However, stakeholders may pursue the development of PAVDs as a desired product regardless of their economic benefits.HighlightsWhile polio antiviral drugs could play an important role in the polio endgame, their expected economic benefits continue to drop with delays in polio eradication and the continued use of oral poliovirus vaccines.The incremental net benefits of investments in polio antiviral drug development and screening for immunodeficiency-related circulating polioviruses are small.Limited global resources are better spent on increasing global population immunity to polioviruses to stop and prevent poliovirus transmission.

Oral Vaccination Approaches for Anti-SHIV Immunity.

We modified a Sabin Oral Poliovirus Vaccine (OPV) vector to permit secretion of the antigens of interest with the goal of improving anti-HIV Env humoral responses in a SHIV mucosal immunization composed of DNA and recombinant OPVs. We evaluated stimulation of systemic and mucosal cell-mediated and humoral immunity in Rhesus macaques by two regimens, both involving a prime with a SHIVBG505 DNA construct producing non-infectious particles formulated in lipid nanoparticles, administered in the oral cavity, and two different viral vector boostings, administered in the oral cavity and intestinally. Group 1 was boosted with rMVA-SHIVBG505, expressing SIV Gag/Pol and HIVBG505 Env. Group 2 was boosted with a SHIVBG505-OPV vaccine including a non-secreting SIVmac239CA-p6-OPV, expressing Gag CA, NC and p6 proteins, and a HIVBG505C1-V2-OPV, secreting the C1-V2 fragment of HIV EnvBG505, recognized by the broadly neutralizing antibody PG16. A time course analysis of anti-SHIV Gag and Env CD4+ and CD8+ T-cell responses in PBMC and in lymph node, rectal, and vaginal MNC was carried out. Both regimens stimulated significant cell-mediated responses in all compartments, with SHIVBG505-OPV immunization stimulating more significant levels of responses than rMVA- SHIVBG505. Boolean analysis of these responses revealed predominantly monofunctional responses with multifunctional responses also present in all tissues. Stimulation of antibody responses was disappointing in both groups with negative anti-SHIV IgG in plasma, and IgA in salivary, rectal and vaginal secretions being restricted to a few animals. After repeated rectal challenge with SHIVBG505, two Group 1 animals remained uninfected at challenge termination. No significant differences were observed in post-infection viral loads between groups. After the acute phase decline, CD4+ T cell percentages returned to normal levels in vaccinated as well as control animals. However, when compared to controls, vaccinate groups had more significant preservation of PBMC and rectal MNC Th17/Treg ratios, considered the strongest surrogate marker of progression to AIDS. We conclude that the vaccine platforms used in this study are insufficient to stimulate significant humoral immunity at the tested doses and schedule but sufficient to stimulate significant mucosal and systemic cell-mediated immunity, impacting the preservation of key Th17 CD4+ T cells in blood and rectal mucosa.

Development of a new oral poliovirus vaccine for the eradication end game using codon deoptimization

Enormous progress has been made in global efforts to eradicate poliovirus, using live-attenuated Sabin oral poliovirus vaccine (OPV). However, as the incidence of disease due to wild poliovirus has declined, vaccine-derived poliovirus (VDPV) has emerged in areas of low-vaccine coverage. Coordinated global cessation of routine, type 2 Sabin OPV (OPV2) use has not resulted in fewer VDPV outbreaks, and continued OPV use in outbreak-response campaigns has seeded new emergences in low-coverage areas. The limitations of existing vaccines and current eradication challenges warranted development of more genetically stable OPV strains, most urgently for OPV2. Here, we report using codon deoptimization to further attenuate Sabin OPV2 by changing preferred codons across the capsid to non-preferred, synonymous codons. Additional modifications to the 5′ untranslated region stabilized known virulence determinants. Testing of this codon-deoptimized new OPV2 candidate (nOPV2-CD) in cell and animal models demonstrated that nOPV2-CD is highly attenuated, grows sufficiently for vaccine manufacture, is antigenically indistinguishable from Sabin OPV2, induces neutralizing antibodies as effectively as Sabin OPV2, and unlike Sabin OPV2 is genetically stable and maintains an attenuation phenotype. In-human clinical trials of nOPV2-CD are ongoing, with potential for nOPV strains to serve as critical vaccine tools for achieving and maintaining polio eradication.

A New Live-Attenuated Poliovirus Vaccine Constructed by Rational Design Improves Safety by Preventing Reversion to Virulence

The life-attenuated Sabin oral poliovirus vaccine (OPV), created over 60 years ago, replicate efficiently in the vaccinee’s gut and, therefore, elicits robust mucosal and systemic immunity, which limits poliovirus spread. Accordingly, OPV has been used in the context of the Global Polio Eradication Initiative, which aims to end the circulation of all polioviruses. However, the program has met a significant hurdle imposed by the ability of OPV to evolve to regain transmissibility and virulence. This represents a risk of widespread polio re-emergence should immunization cease. We exploited molecular features of poliovirus replication to develop a safer, genetically stable, strain by limiting its ability to increase fitness and regain neurovirulence. This novel attenuated poliovirus strain is immunogenic in preclinical and clinical tests and may enable completion of poliovirus eradication as well as pave the way for the rational design of effective and safe vaccines for polio-related pathogens.

Update on Vaccine-Derived Poliovirus Outbreaks - Democratic Republic of the Congo and Horn of Africa, 2017-2018.

Widespread use of live attenuated (Sabin) oral poliovirus vaccine (OPV) has resulted in marked progress toward global poliomyelitis eradication (1). However, in underimmunized populations, extensive person-to-person transmission of Sabin poliovirus can result in genetic reversion to neurovirulence and paralytic vaccine-derived poliovirus (VDPV) disease (1). This report updates (as of February 26, 2019) previous reports on circulating VDPV type 2 (cVDPV2) outbreaks during 2017-2018 in the Democratic Republic of the Congo (DRC) and in Somalia, which experienced a concurrent cVDPV type 3 (cVDPV3) outbreak* (2,3). In DRC, 42 cases have been reported in four cVDPV2 outbreaks; paralysis onset in the most recent case was October 7, 2018 (2). Challenges to interrupting transmission have included delays in outbreak-response supplementary immunization activities (SIAs) and difficulty reaching children in all areas. In Somalia, cVDPV2 and cVDPV3 were detected in sewage before the detection of paralytic cases (3). Twelve type 2 and type 3 cVDPV cases have been confirmed; the most recent paralysis onset dates were September 2 (cVDPV2) and September 7, 2018 (cVDPV3). The primary challenge to interrupting transmission is the residence of >300,000 children in areas that are inaccessible for vaccination activities. For both countries, longer periods of surveillance are needed before interruption of cVDPV transmission can be inferred.

Identification of vaccine-derived polioviruses using dual-stage real-time RT-PCR

Vaccine-derived polioviruses (VDPVs) are associated with polio outbreaks and prolonged infections in individuals with primary immunodeficiencies. VDPV-specific PCR assays for each of the three Sabin oral poliovirus vaccine (OPV) strains were developed, targeting sequences within the VP1 capsid region that are selected for during replication of OPV in the human intestine. Over 2400 Sabin-related isolates and identified 755 VDPVs were screened. Sensitivity of all assays was 100%, while specificity was 100% for serotypes 1 and 3, and 76% for serotype 2. The assays permit rapid, sensitive identification of OPV-related viruses and flag programmatically important isolates for further characterization by genomic sequencing.

Immunity status of adults and children against poliomyelitis virus type 1 strains CHAT and Sabin (LSc-2ab) in Germany.

BackgroundIn October 2007, the working group CEN/TC 216 of the European Committee for standardisation suggested that the Sabin oral poliovirus vaccine type 1 strain (LSc-2ab) presently used for virucidal tests should be replaced by another attenuated vaccine poliovirus type 1 strain, CHAT. Both strains were historically used as oral vaccines, but the Sabin type 1 strain was acknowledged to be more attenuated. In Germany, vaccination against poliomyelitis was introduced in 1962 using the oral polio vaccine (OPV) containing Sabin strain LSc-2ab. The vaccination schedule was changed from OPV to an inactivated polio vaccine (IPV) containing wild polio virus type 1 strain Mahoney in 1998. In the present study, we assessed potential differences in neutralising antibody titres to Sabin and CHAT in persons with a history of either OPV, IPV, or OPV with IPV booster.MethodsNeutralisation poliovirus antibodies against CHAT and Sabin 1 were measured in sera of 41 adults vaccinated with OPV. Additionally, sera from 28 children less than 10 years of age and immunised with IPV only were analysed. The neutralisation assay against poliovirus was performed according to WHO guidelines.ResultsThe neutralisation activity against CHAT in adults with OPV vaccination history was significantly lower than against Sabin poliovirus type 1 strains (Wilcoxon signed-rank test P < 0.025). In eight sera, the antibody titres measured against CHAT were less than 8, although the titre against Sabin 1 varied between 8 and 64. Following IPV booster, anti-CHAT antibodies increased rapidly in sera of CHAT-negative adults with OPV history. Sera from children with IPV history neutralised CHAT and Sabin 1 strains equally.ConclusionThe lack of neutralising antibodies against the CHAT strain in persons vaccinated with OPV might be associated with an increased risk of reinfection with the CHAT polio virus type 1, and this implies a putative risk of transmission of the virus to polio-free communities. We strongly suggest that laboratory workers who were immunised with OPV receive a booster vaccination with IPV before handling CHAT in the laboratory.

Interrupting wild poliovirus transmission using oral poliovirus vaccine: Environmental surveillance in high‐risks area of India

Global eradication of poliomyelitis has reached critical stage. Sabin Oral Poliovirus Vaccine (OPV) has been successful in three major regions of the world. In India eradication of poliomyelitis from states of Uttar Pradesh (UP) and Bihar has been difficult due to high population and low-socioeconomic standards of living. Acute flaccid paralysis (AFP) surveillance and intensive OPV rounds continues with the World Health Organization (WHO) operational strategies. Yet apparent lack of progress in reducing the number of wild cases has resulted in occasional impatience and frustration, even leading to questions about ultimate feasibility of global eradication using OPV. Lucknow in UP is in geographical area endemic for poliomyelitis and is surrounded by high-risk areas yet maintains a polio-free status since 2002. Environmental surveillance study was conducted (2004-2006) to authenticate the decline in the wild poliovirus (PV) cases in Lucknow. Sewage sample analyses were compared with stools of AFP patients and healthy children from same geographical area. Study reveals useful information on OPV circulation and proves important epidemiological tool to trust WHO's OPV immunization program. Genetic sequencing had detected silent wild PV-1 circulation of RCP1PGI (EU049849), RCP2PGI (EU049850), RCP3PGI (EU049851), and RCP4PGI (EU049852) in sewage waters. Properties of isolates from sewage reflected those of viruses excreted from human. This study provides valuable information and encouragement to AFP surveillance to maintain high levels of OPV immunization campaigns in the most difficult endemic region of India to interrupt the wild PV transmission.

Antigenic evolution of vaccine-derived polioviruses: changes in individual epitopes and relative stability of the overall immunological properties.

The Sabin oral poliovirus vaccine (OPV) readily undergoes changes in antigenic sites upon replication in humans. Here, a set of antigenically altered descendants of the three OPV serotypes (76 isolates) was characterized to determine the driving forces behind these changes and their biological implications. The amino acid residues of OPV derivatives that lie within or close to the known antigenic sites exhibited a marked tendency to be replaced by residues characteristic of homotypic wild polioviruses, and these changes may occur very early in OPV evolution. The specific amino acid alterations nicely correlated with serotype-specific changes in the reactivity of certain individual antigenic sites, as revealed by the recently devised monoclonal antibody-based enzyme-linked immunosorbent assay. In comparison to the original vaccine, small changes, if any, in the neutralizing capacity of human or rabbit sera were observed in highly diverged vaccine polioviruses of three serotypes, in spite of strong alterations of certain epitopes. We propose that the common antigenic alterations in evolving OPV strains largely reflect attempts to eliminate fitness-decreasing mutations acquired either during the original selection of the vaccine or already present in the parental strains. Variability of individual epitopes does not appear to be primarily caused by, or lead to, a significant immune evasion, enhancing only slightly, if at all, the capacity of OPV derivatives to overcome immunity in human populations. This study reveals some important patterns of poliovirus evolution and has obvious implications for the rational design of live viral vaccines.

Shedding of sabin poliovirus Type 3 containing the nucleotide 472 uracil-to-cytosine point mutation after administration of oral poliovirus vaccine.

A uracil-to-cytosine point mutation at nucleotide (nt) 472 of Sabin oral poliovirus vaccine (OPV) type 3 is found in conjunction with vaccine-associated paralytic poliomyelitis (VAPP). Direct RNA extraction and mutant analysis by polymerase chain reaction and restriction enzyme cleavage were used to identify this point mutation in clinical samples. A total of 238 stool samples were obtained from 28 healthy infants for 6 weeks after OPV vaccination. More than 25% of infants shed OPV3 in the week after vaccination, with a decrease on day 6. A second wave of OPV3 shedding occurred beginning the second week after vaccination and was maintained through the end of the study period. During the first week after vaccination, the proportion of nt 472 mutants in the shed OPV3 increased from undetectable to almost 100%. During the second shedding period, the proportion of nt 472 mutants remained close to 100%. These results suggest that selective mutation drives the VAPP-associated nt 472 point mutation for OPV3 in the human gastrointestinal tract.

Direct extraction of Sabin poliovirus genomes from human fecal samples using a guanidine thiocyanate extraction method

To permit rapid and efficient detection of Sabin poliovirus type 3 from human fecal samples, we developed a guanidine thiocyanate (GuSCN) extraction and reverse transcriptase polymerase chain reaction (RT-PCR) method. Using 10-fold serial dilutions from stock Sabin–Leon 12 a1b poliovirus type 3 at 10 7 TCID 50 per 0.1 ml, genome was detected to a dilution of 10 3 TCID 50 per 0.1 ml. A total of 40 archived fecal samples were examined using this GuSCN extraction method followed by RT-PCR. Fourteen of 20 poliovirus type 3 tissue culture-positive specimens (70%) and two of 20 tissue culture-negative specimens (10%) were detected by GuSCN extraction and RT-PCR. All positive and negative extraction and RT-PCR controls were identified accurately. This GuSCN extraction and RT-PCR technique is rapid, inexpensive, and can be readily adapted to identify genome sequences of other enterovirus types in large numbers of fecal samples. Moreover, the GuSCN technique extracts viral RNA directly from fecal samples, allowing observation of in vivo alterations of genome sequences. Further studies are underway to examine the development of revertant point mutations in the Sabin poliovirus type 3 genome following oral administration of trivalent Sabin Oral Poliovirus Vaccine to humans.

Sabin type 2 polioviruses with intertypic vaccine/vaccine recombinant genomes.

Attenuated strains of the Sabin oral poliovirus vaccine replicate in the human gut and, in rare cases, cause vaccine-associated paralytic poliomyelitis. In the present study, 15 vaccine-derived strains isolated from patients with vaccine-associated paralytic poliomyelitis and from healthy vaccinees were examined. Four distant sequences of the poliovirus genome (5' NCR, VP3/VP1, VP1/2A, and 3DPol/3' NCR) were targeted, and the reverse-transcribed segments were amplified by polymerase chain reaction followed by restriction fragment length polymorphism analysis with four restriction enzymes. Among the 15 isolates (11 Sabin type 2, 3 Sabin type 1, and 1 Sabin type 3), four Sabin type 2 isolates (36%) were found to be intertypic vaccine/vaccine recombinant in the 3DPol/3' NCR region of the viral genome. The recombinant genotypes identified were S2/S2/S1 for two isolates and S2/S2/S2/S3 and S2/S2/S1/S2 for each of the other two isolates, respectively. Recombinant viruses with unmodified segments in the 5' NCR and the VP3/VP1 regions of the viral genome, a modified segment in the VPI/2A region only for one strain, and an often recombinant segment in the 3DPol/3' NCR parts of the genome were so identified. These findings provide strong evidence that recombination is a frequent phenomenon in type 2 poliovirus vaccine strains and suggest that recombination may be an important mechanism of the natural evolution of polioviruses of Sabin type 2 origin, perhaps even one of the mechanisms of reversion of attenuated vaccine strains toward neurovirulence.

Evolution of the Sabin type 1 poliovirus in humans: characterization of strains isolated from patients with vaccine-associated paralytic poliomyelitis.

Attenuated strains of the Sabin oral poliovirus vaccine replicate in the human gut and in rare cases cause vaccine-associated paralytic poliomyelitis (VAPP). Reversion of vaccine strains toward a pathogenic phenotype is probably one of the main causes of VAPP, a disease most frequently associated with type 3 and type 2 strains and more rarely with the type 1 (Sabin 1) strain. To identify the determinants and mechanisms of safety versus pathogenicity of the Sabin 1 strain, we characterized the genetic and phenotypic changes in six Sabin 1-derived viruses isolated from immunocompetent patients with VAPP. The genomes of these strains carried either few or numerous mutations from the original Sabin 1 genome. As assessed in transgenic mice carrying the human poliovirus receptor (PVR-Tg mice), all but one strain had lost the attenuated phenotype. Four strains presented only a moderate neurovirulent phenotype, probably due at least in part to reversions to the wild-type genotype, which were detected in the 5' noncoding region of the genome. The reversions found in most strains at nucleotide position 480, are known to be associated with an increase in neurovirulence. The construction and characterization of Sabin 1 mutants implicated a reversion at position 189, found in one strain, in the phenotypic change. The presence of 71 mutations in one neurovirulent strain suggests that a vaccine-derived strain can survive for a long time in humans. Surprisingly, none of the strains analyzed were as neurovirulent to PVR-Tg mice as was the wild-type parent of Sabin 1 (Mahoney) or a previously identified neurovirulent Sabin 1 mutant selected at a high temperature in cultured cells. Thus, in the human gut, the Sabin 1 strain does not necessarily evolve toward the genetic characteristics and high neuropathogenicity of its wild-type parent.

Ending polio immunization.

The World Health (WHO) has implemented a plan to eradicate poliovirus that is widely viewed as having made enormous progress. If all goes as planned, polio will be eradicated on this planet by the year 2003. However, there is a debate, as highlighted in a pair of Policy Forums in this issue, over when vaccination against polio can be stopped. Dove and Racaniello believe that the reliance of the WHO on the live Sabin oral polio virus vaccine (OPV) means that there will be a continuing threat of release of potentially pathogenic virus into the environment. They are also concerned that the planned destruction of all wild-type polio stocks will not be possible. They recommend a switch to the inactivated polio vaccine. In response, Hull and Aylward set out the reasons for thinking that a switch from the OPV is not necessary and describe the studies being sponsored by the WHO to determine how and when immunization can safely be ended.

The Polio Eradication Effort: Should Vaccine Eradication Be Next?

The World Health Organization (WHO) has implemented a plan to eradicate poliovirus that is widely viewed as having made enormous progress. If all goes as planned, polio will be eradicated on this planet by the year 2003. However, there is a debate, as highlighted in a pair of Policy Forums in this issue, over when vaccination against polio can be stopped. Dove and Racaniello believe that the reliance of the WHO on the live Sabin oral poliovirus vaccine (OPV) means that there will be a continuing threat of release of potentially pathogenic virus into the environment. They are also concerned that the planned destruction of all wild-type polio stocks will not be possible. They therefore recommend a switch to the inactivated polio vaccine (IPV). In response, Hull and Aylward explain why a switch from OPV is not necessary and describe the studies being sponsored by the WHO to determine how and when immunization can safely be ended.

Elucidating Mechanisms of Thermostabilization of Poliovirus by D2O and MgCl2

To understand a significant reduction in the loss of poliovirus infectivity by D2O and a combination of D2O and MgCl2at 37–45°C, this paper attempts to elucidate the mechanisms underlying the thermostabilization of poliovirus. Three serotypes of Sabin oral poliovirus vaccine strains were investigated. Temperature-dependent fluorescence emission intensity studies showed that the effects of D2O and MgCl2on the stability and conformation of poliovirus are correlated with those of the infectivity of poliovirus. Fluorescence steady-state polarization revealed that the conformation of poliovirus capsid is sensitive to D2O medium and MgCl2salt, and that the rigidity of poliovirus conformation is increased in their presence. The exposure of poliovirus tryptophan residues to water is modified by D2O and MgCl2, as evidenced by changes in fluorescence emission intensity excited at 295 nm. The involvement of hydrogen bonding in the D2O effect was demonstrated by the greatly increased value of relative fluorescence intensity. Conformational alteration was also shown by changes in the positive band (193–230 nm) of circular dichroism spectra. D2O and MgCl2were also found to reduce the interaction of virus with water as examined by differential scanning microcalorimetry, leading to a decline in the extent of water penetration into the poliovirus capsid. All these observations were found to be more profound in a combination of D2O with MgCl2than D2O or MgCl2alone. By inducing a conformation favorable to maintaining the poliovirus assembly and by reducing virus–water interaction to decrease water penetration into the poliovirus capsid, D2O, MgCl2, or D2O–MgCl2is able to exert its thermostabilization effect. Thus, to maintain the virus assembly and conformation of the virus and to reduce the swelling of the virus capsid are key factors in increasing the thermostability of poliovirus. These two factors are mutually complementary. The latter can provide a favorable environment for the formers and the formers, in turn, lead to the latter.

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.

Transport modelling of biological tracers from septic systems

The potential of contamination of individual ground water supplies from septic systems was investigated. Chloride (Cl) and electrical conductivity ( C e ) as tracers of chemicals and indicator bacteria—total coliforms (TC), fecal coliforms (FC) and fecal streptococci (FS)—were determined for 7 months in septic tank effluent and ground water samples collected downgradient from 17 new or replacement septic systems. Indicator bacteria were also determined occasionally in ground water samples from wells near the systems for a continuing period of 2 years. Following 5 years of system operation a last round of analyses was conducted. Water table fluctuations in the vicinity of the septic systems were monitored for 2 years. As a model for viruses, poliovirus in feces from babies who had received the trivalent Sabin oral poliovirus vaccine was introduced into one septic system as a single dose by flushing stools down the toilet. Effluent and ground water were assayed for poliovirus over a period of approx. 6 months. The data were evaluated statistically and a biological contaminant transport model was tested to simulate ground water contamination by the tracers of pathogens. No tracer bacteria reached ground water from any of the 17 septic systems. Presumably, the bacteria were removed by the soil under the seepage bed; mean transport was < 1 count/100 ml for each bacterial type, in keeping with U.S. drinking water regulations. Poliovirus entered and spread in ground water from the one septic system tested, even though this system was functioning properly. Of 3.3 × 10 8 PFU of poliovirus inoculated into the septic tank, a mean of 70 PFU/100 ml escaped from the tank with the effluent and an overall mean of 62 PFU/100 ml were transported to ground water. Septic systems had little influence on water table fluctuations in their vicinity, compared to the seasonal climatic factors.

Antigenic and biochemical characterization of poliovirus type 1 isolates.

By the introduction of Sabin oral poliovirus vaccine, the circulation of wild type polioviruses has virtually disappeared in Japan. However, an outbreak of poliomyelitis associated with sporadic transmission of type 1 wild strain occurred in Nagano in 1980. Furthermore, we found that some type 1 wild strains were introduced into Japan from abroad in 1981. In recent surveys, the two poliovirus type 1 isolates which have non-vaccine-like antigenic character were detected in Aichi. Then, an investigation to trace the origin of these strains was performed, by using intratypic serodifferentiation and biochemical techniques. Electrophoretic migration patterns of their structural polypeptides were quite different from the vaccine virus. In the oligonucleotide mapping, however, one of them gave patterns very similar to those of the vaccine virus. We could conclude that one originated most probably from wild strains, and the other was an antigenic variant derived from the vaccine virus. It showed that oligonucleotide mapping was a very useful method for identification of antigenic modified Sabin type 1 derivatives.