35 research outputs found
Risks associated with the use of live-attenuated vaccine poliovirus strains and the strategies for control and eradication of paralytic poliomyelitis
The Global Polio Eradication Initiative was launched in 1988 with the aim to eliminate paralytic poliomyelitis. Two effective vaccines are available: inactivated polio vaccine (IPV) and oral polio vaccine (OPV). Since 1964, OPV has been used instead of IPV in most countries due to several economic and biological advantages. However, in rare cases, the live-attenuated Sabin strains of OPV revert to neurovirulence and cause vaccine-associated paralytic poliomyelitis in vaccinees or lead to emergence of vaccine-derived poliovirus strains. Attenuating mutations and recombination events have been associated with the reversion of vaccine strains to neurovirulence. The substitution of OPV with an improved new-generation IPV and the availability of new specific drugs against polioviruses are considered as future strategies for outbreak control and the eradication of paralytic poliomyelitis worldwide
Retrospective molecular and phenotypic analysis of poliovirus vaccine strains isolated in Greece
The live oral poliovirus vaccine (OPV) strains are genetically unstable, causing, in rare cases, vaccine-associated paralytic poliomyelitis. Reversions of the known attenuating mutations in OPV strains and intertypic recombination have been identified as the underlying causes of the increased neurovirulence of poliovirus isolates. In this study, three OPV isolates (one non-recombinant and two recombinants) were tested in order to correlate phenotypic traits such as temperature sensitivity (Rct test) and growth kinetics (one-step growth curve test) with mutations and recombination events of the viral genome. Moreover, the immunity level of the western Greek population aged 1-40 years was evaluated against OPV isolates and Sabin vaccine strains, with a microneutralization assay. Members of the 1-40-year age group (both pooled and individual sera) showed no significant differences in neutralization test (NT) titres against OPV isolates in comparison with the Sabin vaccine strains. However, all three OPV isolates showed reverted phenotypic traits in Rct or one-step growth curve assays. The results of our study revealed a significant decrease in immunity level from the 1-10-year age group to the 21-30-year age group (pooled sera) for both poliovirus types 1 and 3. For both poliovirus types, the highest NT titres were observed in the 1-10-year age group, and the lowest NT titre was observed in the 21-30-year age group, towards poliovirus type 3. Our study underlines the need for immunological studies in all age groups, in order to allow reconsideration of the current vaccination policies and to avoid epidemics caused by the circulation of highly evolved OPV derivatives
Use of mutational pattern in 5 '-NCR and VP1 regions of polioviruses for molecular diagnosis
Polioviruses are members of the enterovirus genus, belonging to the Picornaviridae family. They are the causative agents of poliomyelitis, a paralytic and sometimes fatal disease in humans. The number of poliomyelitis cases caused by wild poliovirus infections has been dramatically reduced by the extensive use of two available vaccines: the inactivated poliovirus vaccine (IPV) and the oral poliovirus vaccine (OPV). Despite the importance of OPV in the reduction of poliomyelitis cases, one of the disadvantages associated with this vaccine is the rare occurrence of vaccine-associated paralytic poliomyelitis (VAPP) in vaccinees or their healthy contacts through the accumulation of mutations and/or recombination in Sabin strains genome. Thirteen clinical isolates originating from healthy vaccinees and VAPP cases were investigated in order to identify genomic modifications in 5' non-coding region (5'-NCR) and VP1 genomic regions. The analysis of samples was conducted by RT-PCR, RFLP, sequencing and bioinformatics analysis. All clinical isolates were characterized as OPV-Iike viruses. Our results showed that analysis of 5'-NCR and VP1 regions of Poliovirus Sabin strains is important in order to identify mutations that increase the neurovirulence conducting to the eventuality of emergence of VAPP cases. (C) 2007 Elsevier Ltd. All rights reserved
Growth kinetic analysis of bi-recombinant poliovirus vaccine strains
Attenuated strains of Sabin poliovirus vaccine replicate in the human gut and in rare cases may cause vaccine-associated paralytic poliomyelitis (VAPP). Mutations at specific sites of the genome and recombination between Sabin strains may result in the loss of the attenuated phenotype of OPV (Oral Poliovirus Vaccine) strains and the acquisition of traits characteristic of wild polioviruses, such as increased neurovirulence and loss of temperature sensitivity. In this study, we determined the phenotypic traits such as temperature sensitivity and growth kinetics of eight OPV isolates (six bi-recombinant and two non-recombinant). The growth phenotype of each isolate as well as of Sabin vaccine strains in Hep2 cell line at two different temperatures (37 and 40A degrees C) was evaluated using two different assays, RCT test (Reproductive Capacity at different Temperatures) and one-step growth curve analysis. Moreover, the nucleotide and amino acid positions in the genomes of the isolates that have been identified as being involved in the attenuated and thermo sensitive phenotype of Sabin vaccine strains were investigated. Mutations that result in loss of the attenuated and thermo sensitive phenotype of Sabin vaccine strains were identified in the genomes of all isolates. Both mutations and recombination events correlated well with the reverted phenotypic traits of OPV-derivatives. In the post-eradication era of wild polioviruses, the identification and the characterization (genomic and phenotypic) of vaccine-derived polioviruses become increasingly important in order to prevent cases or even outbreaks of paralytic poliomyelitis caused by neurovirulent strains
Molecular and phylogenetic analysis of the HPV 16 E4 gene in cervical lesions from women in Greece
The HPV16 E1(a )E4 protein is thought to contribute to the release of newly formed viral particles from infected epithelia. In order to investigate amino acid mutations in the HPV16 E1(a )E4 protein, the complete E4 ORF was amplified by PCR in 27 HPV16-positive cervical samples, and the amplicons were cloned. Fifteen nucleic acid variations were identified in the E4 ORF, including seven silent nucleic acid mutations. In addition, nine amino acid mutations (A7V, A7P, L16I, D45E, L59I, L59T, Q66P, S72F, H75Q) were detected in the E1(a )E4 protein, and these were associated with the severity of cervical malignancy. A maximum-likelihood phylogenetic tree was constructed based on the E4 ORF, and nucleotide sequence analysis of the E4, E6 and E7 genes from the same samples was conducted in order to determine the phylogenetic origin of the cloned sequences from the amplified HPV16 E4. Based on the nucleotide sequence and phylogenetic analysis it was revealed that even though E4 ORF constitutes a small polymorphic portion of the viral genome (288 bp), it could provide valuable information about the origins of the HPV16 genome. In addition, molecular evolutionary analysis of the E4 coding region revealed that neutral selection is dominant in the overlapping region of the E4 and E2 ORFs
Molecular identification and full genome analysis of an echovirus 7 strain isolated from the environment in Greece
Two enteroviruses from river water and four from sewage treatment plant were isolated in Larissa, Greece, that all shared the same sequence. A full genome analysis was conducted in an attempt to reveal the evolutionary pathways of one of the isolated strains (LR11F7). VP1 nucleotide and phylogenetic analysis revealed that the isolated strain had 78% homology with the echovirus 7 prototype strain Wallace. Full genome analysis revealed that LR11F7 P1 region is related to echoviruses 7 and that P2 and P3 regions are originating from contemporary enteroviruses isolated in South Asia. Two recombination events were shown to be involved into the evolutionary history of LR11F7, the one event concerning 3A, 3B, and 2C, and the other concerning 3D genomic region, both with new types of HEV-B. The contribution of recombination to enterovirus evolution is substantial, giving rise to new genetic lineages with unknown properties
Structure, Optical and Electric Properties of Opal-Bismuth Silicate Nanocomposites
Synthetic opals composed of 300 nm silica spheres are impregnated with a Bi₁₂SiO₂₀ melt at 1190 K. Structure and properties of the as-prepared samples are studied by employing the scanning electron microscopy, X-ray diffraction, and optical spectroscopy and direct current conductivity techniques. The nanocomposites are found to be multi-phase systems composed of Bi₁₂SiO₂₀, Bi₄Si₃O₁₂ and SiO₂ crystallites with an average linear size not less than 20 nm. Formation of Bi₄Si₃O₁₂ crystallites becomes possible as a result of changing in the Bi₂O₃-SiO₂ molar ratio due to the melting of silica spheres. The Raman intensity redistribution observed by surface scanning may be caused by both composition inhomogeneity and concentration of the exciting radiation field at composite defects. The "red" shift of photoluminescence band is observed. Activation energy of direct current conductivity is estimated as 1.1 eV
Direct extraction and molecular characterization of enteroviruses genomes from human faecal samples
Routine diagnosis of acute flaccid paralysis (AFP) is still based on classical virological procedures. Several enteroviruses serotypes are not easily isolated in cell cultures system used and routinely more than one passage in cell culture is performed. A total of 54 archived faecal samples were examined. The heterogeneous nature of faecal samples may contribute to variations in the yields of viral nucleic acids with different extraction methods and specimen types. PCR inhibitors are frequently encountered in stool specimens. From the three methods initially compared for extraction of viral RNA, QIAamp Viral RNA Mini Kit was retained as it yielded the highest amount of viral RNA without the interference of RTPCR inhibitors. Evaluation of 54 archived stool specimens by RT-PCR and cell culture resulted in a higher frequency of detection by RT-PCR. With the use of RT-PCR we were able to detect two additional samples otherwise considered negative for enterovirus isolation if only the cell culture standard methodology was employed. RNA extraction with QIAamp Viral RNA Mini Kit coupled with RT-PCR in the 5'NCR (sub-grouping into distinct genetic clusters of all enteroviruses) and VP1 (reliable serotyping by sequencing) is a rapid and sensitive technique of direct poliovirus/non-polio enteroviruses recovery and molecular characterization from human faecal specimens without further passage in cell culture, which may select for genetic variants that may not accurately reflect the virus composition in the original specimen. (C) 2008 Elsevier Ltd. All rights reserved