Antigenic and biochemical characterization of poliovirus type 1 isolates of non-vaccine origin

During the past 15 years, five poliovirus type 1 strains with non-vaccine-like antigenicity have been isolated in Japan. Of these isolates, two were from paralytic poliomyelitis patients not associated with the use of Sabin vaccine, and three were apparently introduced from abroad. All the isolates could be readily distinguished from the corresponding Sabin type 1 vaccine strain by oligonucleotide mapping of the viral RNA and by polyacrylamide gel electrophoresis of the viral proteins. The oligonucleotide map of the virulent Mahoney strain which has non-vaccine-like antigenicity was very similar to the map of Sabin type 1 strain. These data indicate that none of the isolates were derived from Sabin type 1 vaccine or its parental Mahoney strain. In addition, some isolates had close antigenic relationship with one another. It is probable that all these strains were introduced from foreign lands where wild poliovirus strains are prevalent.     

New Strains Intended for the Production of Inactivated Polio Vaccine at Low-Containment After Eradication

Poliomyelitis has nearly been eradicated through the efforts of the World Health Organization’s Global Eradication Initiative raising questions on containment of the virus after it has been eliminated in the wild. Most manufacture of inactivated polio vaccines currently requires the growth of large amounts of highly virulent poliovirus, and release from a production facility after eradication could be disastrous; WHO have therefore recommended the use of the attenuated Sabin strains for production as a safer option although it is recognised that they can revert to a transmissible paralytic form. We have exploited the understanding of the molecular virology of the Sabin vaccine strains to design viruses that are extremely genetically stable and hyperattenuated. The viruses are based on the type 3 Sabin vaccine strain and have been genetically modified in domain V of the 5’ non-coding region by changing base pairs to produce a cassette into which capsid regions of other serotypes have been introduced. The viruses give satisfactory yields of antigenically and immunogenically correct viruses in culture, are without measurable neurovirulence and fail to infect non-human primates under conditions where the Sabin strains will do so.     

Sporadic Isolation of Sabin-Like Polioviruses and High-Level Detection of Non-Polio Enteroviruses during Sewage Surveillance in Seven Italian Cities,after Several Years of Inactivated Poliovirus Vaccination

Sewage surveillance in seven Italian cities between 2005 and 2008, after the introduction of inactivated poliovirus vaccination (IPV) in 2002, showed rare polioviruses, none that were wild-type or circulating vaccine-derived poliovirus (cVDPV), and many other enteroviruses among 1,392 samples analyzed. Two of five polioviruses (PV) detected were Sabin-like PV2 and three PV3, based on enzyme-linked immunosorbent assay (ELISA) and PCR results. Neurovirulence-related mutations were found in the 5′ noncoding region (5′NCR) of all strains and, for a PV2, also in VP1 region 143 (Ile > Thr). Intertypic recombination in the 3D region was detected in a second PV2 (Sabin 2/Sabin 1) and a PV3 (Sabin 3/Sabin 2). The low mutation rate in VP1 for all PVs suggests limited interhuman virus passages, consistent with efficient polio immunization in Italy. Nonetheless, these findings highlight the risk of wild or Sabin poliovirus reintroduction from abroad. Non-polio enteroviruses (NPEVs) were detected, 448 of which were coxsackievirus B (CVB) and 294 of which were echoviruses (Echo). Fifty-six NPEVs failing serological typing were characterized by sequencing the VP1 region (nucleotides [nt] 2628 to 2976). A total of 448 CVB and 294 Echo strains were identified; among those strains, CVB2, CVB5, and Echo 11 predominated. Environmental CVB5 and CVB2 strains from this study showed high sequence identity with GenBank global strains. The high similarity between environmental NPEVs and clinical strains from the same areas of Italy and the same periods indicates that environmental strains reflect the viruses circulating in the population and highlights the potential risk of inefficient wastewater treatments. This study confirmed that sewage surveillance can be more sensitive than acute flaccid paralysis (AFP) surveillance in monitoring silent poliovirus circulation in the population as well as the suitability of molecular approaches to enterovirus typing.     

Antigenic Analysis of Polioviruses Isolated from a Child with Agammaglobulinemia and Paralytic Poliomyelitis after Sabin Vaccine Administration

A 3-year-old boy with agammaglobulinemia developed paralytic poliomyelitis on day 553 after being fed poliovaccine. Non-vaccine-like type 2 polioviruses were isolated from 22 stools obtained within 684 days after the onset of illness. Antigenic variations were observed among these viruses. The non-vaccine-like virus isolated 1 week after the onset of paralysis differed in virulence from the Sabin type 2 vaccine strain in the neurovirulence test in monkeys, and did not have the same antigenic character as the wild virulent strains. Another virus isolated on day 348 before the onset of illness was also classified as non-vaccine-like. However, the Sabin type 2 strain was shown to be homologous with this strain by the McBride test. Some Sabin-like particles were found in this stock virus. We may conclude that the non-vaccine-like virus isolates were derived from Sabin vaccine by antigenic variation that occurred during long-term multiplication in the intestinal tract.     

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.     

Effects of arildone on the immunogenicity of formalin-inactivated polioviruses

Concentrated and purified Sabin and virulent strains of poliovirus types 1, 2 and 3 were inactivated with formalin at 37 C. By addition of 5.4 microM arildone, an antiviral agent, to the virus suspension, the stability of D antigen increased in both Sabin and virulent strains of all types, especially in virulent type 1 Mahoney strain. The drug had neither any inhibitory nor enhancing effect on the formalin inactivation. When antibody response was compared in guinea pigs, Sabin strains inactivated in the absence of arildone were less immunogenic against homotypic virulent strains than inactivated vaccine prepared from virulent strains. On the other hand, Sabin strains inactivated in the presence of arildone were equally immunogenic. These results indicate that it is possible to prepare from Sabin strains a potent and safe inactivated vaccine having an immunogenicity comparable to that prepared from virulent strains.     

Assessment of poliovirus eradication in Japan: genomic analysis of polioviruses isolated from river water and sewage in toyama prefecture

Seventy-eight poliovirus strains isolated from river water and sewage in Toyama Prefecture, Japan, during 1993 to 1995 were characterized by the PCR-restriction fragment length polymorphism (RFLP) method and by partially sequencing the VP3 and VP1 regions of the viral genome. Of these isolates, 36 were identified as Sabin vaccine strains, and 42 were identified as vaccine variant strains that had less than 1.4% nucleotide divergence from the Sabin strains, including 7 isolates with patterns different from those of Sabin strains as determined by PCR-RFLP analysis. These findings suggest that wild-type poliovirus was not circulating in Toyama Prefecture.     

Assessment of Poliovirus Eradication in Japan: Genomic Analysis of Polioviruses Isolated from River Water and Sewage in Toyama Prefecture

Seventy-eight poliovirus strains isolated from river water and sewage in Toyama Prefecture, Japan, during 1993 to 1995 were characterized by the PCR-restriction fragment length polymorphism (RFLP) method and by partially sequencing the VP3 and VP1 regions of the viral genome. Of these isolates, 36 were identified as Sabin vaccine strains, and 42 were identified as vaccine variant strains that had less than 1.4% nucleotide divergence from the Sabin strains, including 7 isolates with patterns different from those of Sabin strains as determined by PCR-RFLP analysis. These findings suggest that wild-type poliovirus was not circulating in Toyama Prefecture.     

2010年山东省环境污水中脊髓灰质炎病毒的监测及其基因特征分析

本研究在山东省开展了脊髓灰质炎病毒(Poliovirus,PV)的外环境监测,从济南、临沂两地采集污水标本,浓缩处理后进行病毒分离,对分离到的PV采用中和试验进行血清定型,并对其VP1及3D区进行序列测定,分析其基因突变和重组情况。2010年,共采集污水标本32份,PV阳性10份,阳性率31.3%;分离到18株PV(PV1型3株,PV2型9株,PV3型6株),均为疫苗相关株,VP1完整编码区核苷酸变异数在0~4个之间,在3株PV2型病毒和4株PV3型病毒的基因组中发现重组;对VP1区影响神经毒力的减毒位点分析发现,PV1型病毒中有1株在nt 2 749发生突变(A→G),PV2型病毒中有1株在nt2 908发生A→G突变,3株在nt2 909发生U→C突变,6株PV3型病毒全部在nt2 493发生C→U突变。环境污水中可以分离到PV,其基因重组率和主要减毒位点的回复突变率较高,未发现脊灰野毒株和疫苗衍生株脊灰病毒(Vaccine-derived poliovirus,VDPV)。     

Comparative sensitivities of Sabin and Mahoney poliovirus type 1 prototype strains and two recent isolates to low concentrations of glutaraldehyde.

Significant intratypic differences in the glutaraldehyde (GTA) sensitivity of echovirus isolates have been shown. While exploring ways to optimize the study of GTA sensitivity of enteroviruses, we also observed intratypic differences in poliovirus type 1 isolates collected in France. A suspension procedure was used for assessing the virucidal effect of GTA at low concentrations (< or = 0.10%) against purified viruses. Two recent isolates of poliovirus type 1 tested were first fully characterized by the PCR restriction fragment length polymorphism (RFLP) test. The RFLP pattern of clinical isolate 5617 was similar to that of poliovirus type 1 LS-c, 2ab (Sabin strain), confirming the vaccine origin of strain 5617. The RFLP pattern of strain 5915 recovered from sewage was different from that of the Mahoney strain, suggesting a genetic variation in this wild isolate. We then analyzed under the same controlled conditions the GTA sensitivities of both isolates and their respective prototype strains. The wild Mahoney and 5915 strains exhibited significantly lower sensitivities to GTA than did the vaccine Sabin and 5617 strains. The inactivation rates of clinical isolates 5617 and 5915 were very similar to those of their corresponding reference Sabin and Mahoney strains. Both the conformational structure of the capsid of each strain and the amino acid constitution of structural polypeptides could be involved in the variations observed. The relevance of our comparative sensitivity studies to standardization of virucidal tests is discussed.