Studies on the genetic basis of attenuation of the Sabin type 3 poliovirus vaccine.
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Cloning, Molecular↗
Biomedical subjects
D M Evans
Publications and source records attributed to D M Evans.
Botulinum neurotoxin type B. Its purification, radioiodination and interaction with rat-brain synaptosomal membranes.
Neurotoxin from Clostridium botulinum type B was purified to homogeneity by by affinity and ion-exchange chromatography; specific neurotoxicity of this protein (Mr of approximately equal to 155 000) following trypsinisation attained a level of 2 X 10(8) mouse LD50 units/mg protein. 125I-iodination of the toxin to high specific radioactivities (19-63 TBq/mmol) yielded typically greater than 65% of its original toxicity; dodecyl sulphate gel electrophoresis under reducing conditions, after trypsinisation, showed that the larger polypeptide (Mr of approximately equal to 101 000) was labelled preferentially. Saturable binding of the 125I-labelled neurotoxin to rat cerebrocortical synaptosomes was observed and Scatchard analysis showed a low content of acceptors with high affinity (Kd = 0.3-0.5 nM;Bmax approximately equal to 30-60 fmol/mg protein, together with a much larger population of weak-affinity sites. No significant differences in binding affinity were seen in competition experiments using native or fully activated (trypsinized) neurotoxin, indicating that chain cleavage is not essential for acceptor-toxin interaction. Type A botulinum neurotoxin showed a limited capacity to inhibit the synaptosomal binding of labelled type B toxin, even at high concentrations (1 muM), and other neurotoxins were without effect, emphasising the acceptor selectivity. Near-complete loss of specific toxin binding was produced by preincubation of synaptosomes with neuraminidase whereas inhibition of the low-affinity sites with wheat-germ agglutinin was less pronounced; such inactivation was prevented by inclusion of selective inhibitors (2,3-dehydro-2-deoxy-N-acetylneuraminic acid and N-acetylglucosamine, respectively). These observations implicate N-acetylneuraminic acid and, possibly, other sugar moieties as constituents of the toxin acceptors. Trypsinisation of synaptosomes gave incomplete inhibition of binding when assayed with 1 nM or 10 nM 125I-iodinated toxin. Detailed analysis of the actions of neuraminidase, trypsin and heat treatment on the concentration dependence of toxin binding suggest the existence of at least two distinguishable populations of sites that contain N-acetylneuraminic acid, with a protein component being associated with the acceptors of lower affinity. These findings are discussed in relation to those previously reported for type A neurotoxin and to the possible physiological significance of such membrane acceptors.
Animals↗
Soft-tissue anatomy of mirror hand.
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Child, Preschool↗
Mirror hand treated by true pollicization.
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Child↗
Aspiration of ganglia.
Thirty four patients with a simple ganglion have been treated by aspiration alone, using a short-bevel wide-bore needle. In 41% there was no recurrence after more than one year, and most of the remaining 59% recurred with three months.
Foot Diseases↗
The nucleotide sequence of a type 3 poliovirus isolated during a recent outbreak of poliomyelitis in Finland.
We have cloned and sequenced the complete genome of a strain of poliovirus type 3 (23127) isolated during an outbreak of poliomyelitis in Finland. The genome is 7435 nucleotides long excluding the 3' poly(A) stretch and is 95.5% homologous at the amino acid level to the previously sequenced type 3 strain, P3/Leon/37. The most striking feature of the presented sequence is the extent of amino acid substitutions relative to P3/Leon/37 and other type 3 strains in areas of known antigenic importance. The major antigenic determinant for virus neutralization (site 1), located at residues 89 to 100 of VP1, has three amino acid substitutions and there are six substitutions in site 3, a composite site made up of sequences from VP1 and VP3. The variation in these regions probably accounts for the observed unusual antigenicity and may explain why the virus was able to spread in a well-vaccinated community. Sequence comparisons imply that the virus is not derived from the currently used live attenuated vaccine.
Amino Acid Sequence↗
Antigenic and molecular properties of type 3 poliovirus responsible for an outbreak of poliomyelitis in a vaccinated population.
Virus isolated from an outbreak of poliomyelitis in Finland has been examined serologically and at the molecular level. The causative agent was an antigenically unusual strain of type 3 poliovirus, which was unrelated to the strains used to manufacture either live or killed poliovaccines. It is likely that the antigenic properties of the virus played a part in establishing a limited outbreak of poliomyelitis in a vaccinated population.
Amino Acid Sequence↗
Antigenic structure of polioviruses of serotypes 1, 2 and 3.
The antigenic sites recognized by monoclonal antibodies with neutralizing activity for the Sabin vaccine strains of poliovirus of serotypes 1, 2 and 3 have been studied by the isolation and characterization of mutants resistant to neutralization by antibody. Three distinct sites have been identified which are designated site 1, site 2 and site 3. Site 1 includes a region of 12 amino acids of VP1, from residues 89 to 100, and a corresponding region of VP1 has been identified as an antigenic site for poliovirus 2. This site was strongly immunodominant in type 2 and type 3 but was not detected for poliovirus 1. Site 2 is a complex site including residues 220 to 222 from VP1 (site 2a) with residues including 169 and 170 and others of VP2 (site 2b). Both site 2a and site 2b have been detected in type 1 poliovirus, while as yet only site 2b has been detected in type 3 poliovirus. Site 3 is a complex site including residues 286 to 290 from VP1 (site 3a) with residues including 58 and 59 and others of VP3 (site 3b). Both sites 3a and 3b have been detected in type 3 poliovirus, while as yet only site 3b has been detected in type 1 poliovirus.
Amino Acid Sequence↗
Construction of poliovirus intertypic recombinants by use of cDNA.
We investigated the use of infectious cDNA for the production of poliovirus type 1-type 3 recombinants. One such recombinant virus was produced, but a second construct involving the transfer of part of the capsid protein region was not infectious. Our results suggest that the approach may prove valuable but that not all cDNA constructs will give rise to viable viruses.
Base Sequence↗
Terminology in gynaecological cytopathology: report of the Working Party of the British Society for Clinical Cytology.
The report defines and recommends terms for use in cervical cytology.
Cell Nucleus↗
Correction of prominent ears using Mustardé's technique: an out-patient procedure under local anaesthetic in children and adults.
Experience in correction of prominent ears as an out-patient procedure under local anaesthetic in 52 patients is described. Adults and children aged 9 and over are suitable for this form of treatment. A review of 44 patients has revealed 100% patient satisfaction and a low complication rate. Infection has occurred in two ears. An objective appraisal of the results is given.
Adolescent↗
Induction by synthetic peptides of broadly reactive, type-specific neutralizing antibody to poliovirus type 3.
A region of virus capsid protein VP1 located 89-100 amino acids from the N-terminus has been proposed to comprise a major antigenic site involved in the neutralization of poliovirus type 3. Synthetic peptides 10-18 amino acids in length, containing all or part of this sequence, were tested for their ability to induce antiviral antibodies. Rabbits, but not guinea pigs or mice, immunized with the most active peptide, developed hightitered, type-specific, neutralizing antibodies for a wide range of poliovirus type 3 strains. Consistent with the broad type specificity of the antibody response was the observation that amino acids 89-100 of VP1 are highly conserved among different poliovirus type 3 strains. This sequence thus appears to provide, at least in part, a molecular basis for serotype antigenic specificity. Individual amino acids from 93 to 98 within this sequence were shown to be important for the neutralization of virus by antipeptide sera by examination of the ability of the sera to neutralize laboratory-derived poliovirus type 3 mutants with known single amino acid substitutions in the proposed antigenic site.
Amino Acid Sequence↗
Identification and initial characterisation of a pyrimidine dimer UV endonuclease (UV endonuclease beta) from Deinococcus radiodurans; a DNA-repair enzyme that requires manganese ions.
An endonuclease that incises lightly ultraviolet-irradiated supercoiled plasmid DNA was identified in cell-free extracts of Deinococcus radiodurans R1 wild-type. The endonuclease was absent from strains mutant in the uvsC, uvsD or uvsE genes identifying it as 'UV endonuclease beta' responsible for the initial incision step of one excision-repair pathway for the removal of pyrimidine dimers from D. radiodurans DNA in vivo. The enzyme was purified free from contaminating nuclease activities and was partially characterised. The enzyme has an apparent molecular weight of 36 000, is ATP-independent, caffeine-insensitive and is inactivated by N-ethylmaleimide. It also has a novel requirement for manganese ions distinguishing it from all other known DNA-repair enzymes.
Chromatography, Gel↗
Principal and subsidiary antigenic sites of VP1 involved in the neutralization of poliovirus type 3.
The characterization of over 300 mutants, derived from two strains of poliovirus type 3 and selected for resistance to neutralization by monoclonal antibodies, has led to the further definition of the major antigenic site involved in neutralization. The site encompasses amino acids 89 to 100 of VP1. A subsidiary antigenic site near the C-terminus of VP1 has been identified for the Sabin vaccine strain of poliovirus type 3. Of 59 monoclonal antibodies to poliovirus type 3 examined, 27 had virus-neutralizing activity and 25 of these were identified as directed against the major site on VP1 (designated site 1), indicating the immunodominant role of this site. One of the six monoclonal antibodies that recognized the subsidiary antigenic site on VP1 (designated site 2) possessed virus-neutralizing activity. The identification of the principal antigenic site of the virus provides a rational basis for attempts at the development of synthetic oligopeptide vaccines against poliovirus type 3.
Amino Acid Sequence↗
Reversion to neurovirulence of the live-attenuated Sabin type 3 oral poliovirus vaccine.
The complete nucleotide sequence has been determined of a strain of poliovirus type 3, P3/119, isolated from the central nervous system of a victim of fatal vaccine-associated poliomyelitis. Comparison of this sequence with those obtained previously for the Sabin type 3 vaccine, P3/Leon 12a1b and its neurovirulent progenitor, P3/Leon/37, reveals that these three strains are on a direct geneaological lineage and therefore that P3/119 is a bona fide revertant of the vaccine. P3/119 differs in sequence from its attenuated vaccine parent at just seven positions. Only one of these differences, a mutation from U to C at position 472 in the presumed noncoding region of the genome, is a back mutation to the wild type sequence. Of the six other differences, three give rise to coding changes in virus structural proteins, two are silent changes in the major open reading frame of the genome and one affects the 3'-terminus just prior to the poly A tract. These differences indicate that there are three possible types of molecular change which could, singly or collectively, result in attenuation and reversion to neurovirulence of the Sabin type 3 vaccine.
Amino Acid Sequence↗
New poliovirus vaccines: a molecular approach.
This article summarizes recent work on the determinants of antigenicity in poliovirus type 3 and reports on experiments in progress aimed at understanding the molecular basis of attenuation in Sabin's type 3 vaccines. Ways in which this new information might be used to produce alternative, safe, inexpensive, multivalent vaccines against polio and other enteroviruses are discussed.
Amino Acid Sequence↗
Assessment of hand function.
Hand assessment involves analysis of developing deformities and the pathological changes causing them, and a study of how these changes affect the various components of hand function both individually and combined into activities required by the patient. A detailed recording of this information helps in the planning of treatment, and is essential for reliable evaluation of the effects of treatment.
Activities of Daily Living↗
The PIP joint.
The anatomy of the PIP joint has been reviewed, regarding it as a functional complex including extensor and flexor mechanisms, rather than as a simple hinge joint. The importance of stability and mobility has been stressed. The pathological changes affecting this joint, resulting from primary joint disease, primary extensor tendon disease and primary flexor tendon disease have been described. The management of the resulting deformities has been discussed under the headings of general management, splintage, steroid injection, and surgery, including synovectomy, restoration of flexion, restoration of extension, arthroplasty and arthrodesis.
Arthritis, Rheumatoid↗