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Biomedical subjects

P J Watt

Publications and source records attributed to P J Watt.

At least 19 recordsLinked to original sources
PubMed1992

Genotyping of Chlamydia trachomatis from a trachoma-endemic village in the Gambia by a nested polymerase chain reaction: identification of strain variants.

Direct amplification of the major outer membrane protein (MOMP) gene by polymerase chain reaction (PCR) was used to identify Chlamydia trachomatis in eye swabs from clinically active cases of endemic trachoma in a Gambian village. Chlamydial DNA was detected in 51% of 96 subjects with clinically active disease and in 5% of 37 clinically negative individuals. The PCR detection was combined with typing, using nested primers to variable sequences (VS) 1, 2, and 4 of the MOMP genes to distinguish between trachoma genotypes A, B, and C, respectively. Genotypes A and B were detected in the village, with some individuals harboring both genotypes within the same eye. DNA sequencing revealed strain variants of both genotypes. Typing of genotype and strain variants is now in progress to study trachoma transmission within the village.

Bacterial Outer Membrane Proteins↗
PubMed1990

Determinants of susceptibility to challenge and the antibody response of adult volunteers given experimental respiratory syncytial virus vaccines.

The virulence and immunogenicity of a wild-type respiratory syncytial (RS) virus together with four temperature sensitive (ts) mutants derived from this isolate were tested by intranasal inoculation into adult volunteers. Resistance to challenge correlated with neutralizing antibody titres in nasal secretions and to a lesser extent in serum. All ts mutants were reduced in virulence. Mutant ts1B caused mild or asymptomatic infections yet induced antibody responses comparable with wild-type RS virus. Ts1B might be developed further to produce a live-virus vaccine.

Adult↗
PubMed1990

Identification of a linear epitope on the fusion glycoprotein of respiratory syncytial virus.

The fusion glycoprotein of the Edinburgh strain of respiratory syncytial (RS) virus was cloned from infected cell mRNA. A full-length clone was subjected to sequence analysis, and compared with other strains of RS virus. The inferred primary amino acid sequence was used to generate a nested set of overlapping peptides spanning the mature protein. Peptides were synthesized on polyethylene pins and examined for their reactivity towards high titre human antisera. Decameric peptides spanning the highly conserved region between amino acids Lys and Ala (positions 470 to 490), reacted strongly with the sera and a detailed study with hexameric peptides located the epitope to FPSDEF, at positions 483 to 488. Replacement synthesis analysis revealed that Pro at position 484 and Glu at 487 were critical components of the binding site and could not be substituted.

Amino Acid Sequence↗
PubMed1989

Changing standards.

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Cultural Characteristics↗
PubMed1986

Pathogenic mechanisms of organisms virulent to the eye.

Gonococci possess long range adhesins in the form of pili permitting initial contact with conjunctival cells. Subsequently sticky surface protein (Protein II) bonds the gonococcus close to the host cell surface. Damage is mediated both by the intracellular uptake and the introduction of pores in the host cell membrane. Unlike gonococci, pseudomonas can only attach to damaged cells but once the eye is invaded a wide range of enzymes and toxins leads to rapid tissue destruction. The mechanisms by which Chlamydia trachomatis induce trachoma are ill-understood but involve intense antigenic stimulation. Methods are now available to investigate the antigenic structure of chlamydia at sub-molecular levels; such an approach is required for the development of a vaccine against trachoma.

Antigens, Bacterial↗
PubMed1986

Age related IgG subclass response to respiratory syncytial virus fusion protein in infected infants.

Respiratory syncytial virus (RSV) fusion protein was purified by immunoaffinity chromatography using a mouse monoclonal antibody coupled to Affi-gel 10. The fusion protein was homogeneous by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and free of other detectable viral or cellular protein. The purified fusion protein was used in a quantitative enzyme-linked immunosorbent assay (ELISA) to measure the age-related antibody response to this protein in infected infants. The four IgG subclasses, IgA and IgM levels were determined for infants under 6 months of age, infants aged 6 months to 1 year and infants aged 1 year and over. Most infants over 6 months of age showed marked increases in both IgG1 and IgG3 antibodies with poor or negligible response with IgG2 and IgG4. By contrast infants under 6 months failed to respond by the production of IgG antibodies, although increases in IgA and IgM levels were observed. These data may explain the failure of primary RSV infections to induce protective immunity and have implications for the strategic use of attenuated RSV vaccines.

Aging↗
PubMed1979

Variations in surface protein composition associated with virulence properties in opacity types of Neisseria gonorrhoeae.

The biological properties of a series of opacity variants of Neisseria gonorrhoeae P9 have been examined. A novel protein, designated protein IId* (mol. wt 28 850), was identified within the set of heat-modifiable surface proteins previously reported. All variants producing extra outer membrane proteins were less sensitive to the bactericidal action of serum than the prototype transparent strain, with protein IIa* (mol. wt 28 500) being associated with increased resistance. The production of a different protein, protein II* (mol. wt 29 000), was correlated with resistance to low molecular weight antimicrobial agents (penicillin, fusidic acid, Cu2+, Zn2+). Increased adhesion to human buccal epithelial cells was demonstrated in all variants that produced extra surface proteins. These variants did not show increased binding to hexyl- and phenyl-substituted Sepharose gels suggesting that hydrophobic interaction was not responsible for their cohesive properties. The prototype strain lacking additional proteins demonstrated the greatest binding to erythrocytes, indicating that adhesion to buccal cells and red blood cells is mediated by different mechanisms. One variant producing protein IIa* showed increased association with leukocytes, whereas another producing protein IIb* showed decreased association with leukocytes. These results show that the heat-modifiable surface proteins are important virulence attributes of the gonococcus: this must be considered in the selection of strains for vaccine trials.

Anti-Bacterial Agents↗
PubMed1978

A solid phase radiommunoassay on hydrophobic membrane filters: detection of antibodies to gonococcal surface antigens.

A solid phase radioimmunoassay (SPRIA) has been developed for detection of IgG antibodies to gonococcal outer membrane components. Gonococcal antigens were immobilized on a solid support by covalent coupling to CNBr-activated Sepharose in the presence of the detergent Triton X-100. Binding of specific antibody to the Sepharose-antigen complex was detected using radiolabelled Protein A as the antiglobulin. Protein A was labelled by radioacetylation with tritiated acetic anhydride, yielding a product of high specific activity and high stability. No detectable loss of activity was observed over a ten month period. The entire assay was performed on Mitex teflon hydrophobic membrane filters which held the Sepharose beads and aqueous supernatant as a discrete drop of liquid. The supernatants and incubation media were easily and rapidly removed from the beads by suction on a specially designed manifold system. This procedure removed the need for repeated and time-consuming centrifugations. Titres were obtained graphically from double log plots of cpm bound versus antiserum dilution by extrapolation of the straight line to a point corresponding to twice the control level of radioactivity binding. The assay proved to be a very reliable and simple procedure for the detection of IgG antibodies to gonococcal surface antigens.

Antibody Specificity↗
PubMed1978

The surface properties of Neisseria gonorrhoeae: determinants of susceptibility to antibody complement killing.

Monovalent rabbit antisera were prepared to highly purified gonococcal lipopolysaccharide (LPS), to pili and to two major purified outer envelope proteins. All these antisera were free from significant specific IgM antibody and were standardized to 4 microgram specific IgG antibody per test, permitting accurate comparisons between the different gonococcal surface antigens as triggers of the complement-dependent bactericidal reaction. LPS was the most effective antigen at inducing a bactericidal response to homologous and heterologous gonococci, followed by the two individual outer envelope proteins. Pili were relatively ineffective. Strain P9 gonococci grown in vivo or which possessed a 'capsule' in vitro were more resistant to serum killing than the non-capsulated parent strain. One highly susceptible strain, F62, which was killed by complement in the absence of any LPS antibody, was able to directly activate complement by the alternative pathway.

Antibodies↗
PubMed1977

The bacteriology of the cervix and uterus.

Fifty hysterectomy specimens were studied by a multiple biopsy technique to determine the normal bacteriological flora of the uterine cavity and cervical canal. All the uterine cavities and 23 of the cervical canals were sterile. The lower half of the other 27 cervical canals contained organisms similar to those found in the vaginal vault.

Adult↗
PubMed1975

Studies on lipopolysaccharides isolated from strains of Neisseria gonorrhoeae.

Lipopolysaccharides, extracted by phenol-water from five strains fo Neisseria gonorrhoeae, were purified by treatment with ribonuclease followed by multiple washes. These preparations were fatal to mice when administered in submicrogram amounts with actinomycin D, the LD50 values varying from 4 to 16 mug/kg. Analyses showed that all preparations contained glucose, galactose, glucosamine, heptose, 2-keto-3-deoxyoctonic acid and phosphate. All the lipopolysaccharides contained the same fatty acids, namely beta-OH-10:0, beta-OH-12:0, beta-OH-14:0, 12:0, 14:0,16:0, 16:1, 18:0 and 18:1. We were unable to detect significant differences between the lipopolysaccharides of virulent and avirulent gonococci or between penicillin-sensitive and resistant strains. Gonococcal lipopolysaccharides appeared to lack O-antigen side chains.

Animals↗