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P J Watt

Publications and source records attributed to P J Watt.

At least 19 recordsLinked to original sources
PubMed1995

Extent and kinetics of genetic change in the omp1 gene of Chlamydia trachomatis in two villages with endemic trachoma.

Variants of Chlamydia trachomatis in two Gambian villages with hyperendemic trachoma were analyzed by omp1-based polymerase chain reaction and sequencing from conjunctival swabs. Samples collected over a 22-month period included a complete cross-sectional study of each village. Overall, 4 genovar A and 4 B variants were characterized by point mutations in the omp1 gene, resulting in changes in the inferred amino acid sequence. Two genovar A and 2 B variants accounted for 87% of the total ocular chlamydial infection in both villages. Although some flux in the prevalence of individual variants was observed overtime, their overall distribution remained remarkably stable. There was no evidence of major antigenic shift arising from recombination events at the omp1 locus as described for genital tract infection. These results indicate that omp1 variation in these two trachoma-hyperendemic communities is limited and unlikely to hamper development of trachoma vaccines based on the major outer membrane protein.

Amino Acid Sequence↗
PubMed1993

Immunogenicity and pathogenicity of a triple temperature-sensitive modified respiratory syncytial virus in adult volunteers.

The immunogenicity and pathogenicity of a strain of respiratory syncytial (RS) virus modified by sequential induction of three temperature-sensitive (ts) mutations have been evaluated by intranasal administration to 22 adult volunteers. This modified virus, a triple ts mutant designated ts1C, was derived from a double mutant ts1B evaluated in a previous trial. The original isolate (strain RSS-2) and all its derivatives were propagated throughout in human diploid cells in a specially assigned laboratory. The triple mutant ts1C is unable to multiply in MRC-5 cells at 37 degrees C and above. Following nasal administration of ts1C, immune responses were observed in volunteers with low pre-existing neutralizing antibody titres. The ability of mutant ts1C to induce upper respiratory tract disease in adults was greatly diminished in comparison with the non-ts wild-type virus, but not markedly more so than a previously tested double ts mutant (ts1B) which replicates at 37 degrees C. Mutant ts1C, however, may have greater potential as a live vaccine in view of its inherently greater genetic stability.

Adult↗
PubMed1993

Genotyping ocular strains of Chlamydia trachomatis by single-tube nested PCR.

A single-tube nested PCR able to discriminate between ocular infection with Chlamydia trachomatis of serovar A, B, or C is described. The method uses genotype-specific primers labeled with different fluorochromes, "drop-in/drop-out" PCR amplification, and product analysis on an Applied Biosystems 373A DNA Sequencer using GENESCAN 672 software. The system readily detected mixed infection with serotypes A and B within the same eye. This strategy provides a rapid genotyping method applicable to a wide variety of epidemiological studies.

Base Sequence↗
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↗
PubMed1984

Antigenic and structural variation in the major nucleocapsid protein of respiratory syncytial virus.

Antigenic and structural variation in the major nucleocapsid protein, VPN41, from different strains of respiratory syncytial (RS) virus was observed using a combination of monoclonal antibodies and two-dimensional peptide mapping. Limited trypsin treatment of intact nucleocapsids produced two peptide fragments Mr 27K and Mr 14K. Two monoclonal antibodies, N1 and N2, reactive with primary sequence epitopes located on intact nucleocapsids also reacted with either the 27K fragment (N2) or the 14K fragment (N1). Competitive radioimmunoassay studies using N1 and N2 antibodies revealed two discrete antigenic groups among the seven human strains of RS virus examined. A bovine strain of RS virus, although antigenically similar to the human strain of RS virus, was placed in a separate group. Two-dimensional peptide mapping of 125I-labelled VPN41 purified by SDS-PAGE revealed extensive structural homology between all strains. However, several unique tryptic/chymotryptic peptides supported the grouping obtained with the monoclonal antibodies.

Animals↗
PubMed1983

Antibodies to respiratory syncytial virus polypeptides and their significance in human infection.

The human antibody response to respiratory syncytial (RS) virus infection was investigated using radioimmunoprecipitation analysis (RIPA). A total of nine RS virus-specific proteins, VP200, VGP95, VP68, VGP48, VPN41, VP35, VP27, VP23 and VGP20 were identified by comparing 35S- or 3H-labelled extracts of infected and uninfected HEp-2 cells, and by radioimmunoprecipitation using a hyperimmune human serum. Three glycopeptides, VGP95, VGP48 and VGP20, were identified by incorporation of [3H]glucosamine, and two of these (VGP48 and VGP20) were assumed to be part of a single disulphide-bonded polypeptide since they were precipitated by a monoclonal antibody raised against a surface protein. Human serum antibodies to three major RS virus proteins, VGP95, VGP48/VGP20 and VPN41 were measured by RIPA using radioiodinated RS virus antigens. Sera from a group of mothers whose babies escaped RS virus infection during a local epidemic showed increased antibody levels to VPN41 when compared to sera from mothers whose babies had become infected with RS virus within the first 6 months of life. In infants who remained uninfected with RS virus during the first 12 months of life the maternal gift of antibody decayed to about 50% at 3 months with traces of antibodies detected in a few sera at 12 months. The antibody levels detected in the sera of infants less than 3 months old convalescent from primary RS virus infection did not exceed the mean levels present in the serum of uninfected babies. Infants between the ages of 6 and 12 months were able to mount an IgG response to VPN41 and VGP48 but, unlike adults and older children, a particularly striking finding was their failure to produce antibodies to VGP95.

Animals↗
PubMed1983

Monoclonal antibodies to gonococcal pili: studies on antigenic determinants on pili from variants of strain P9.

Several hybrid cell lines producing monoclonal antibodies against gonococcal pili have been derived. The reactivity of each antibody against variant pili of strain P9 was determined by ELISA. Using type-specific as well as broadly cross-reacting antibodies labelled with 125I, the capacity of unlabelled hybridoma antibodies to inhibit their attachment to various pilus types was determined. Some antibodies competed significantly and were judged to react at the same or closely positioned antigenic sites, whereas others bound non-competitively to the same pilus. These studies suggest the existence of distinct antigenic sites on gonococcal pili: a common antigenic region shared by all P9 pilus types and a type-specific region. Both antigenic sites contain more than one epitope.

Animals↗
PubMed1982

Effect of anti-pilus antibodies on survival of gonococci within guinea pig subcutaneous chambers.

Guinea pigs were immunized with either alpha or beta pili from Neisseria gonorrhoeae P9 before the inoculation of subcutaneous chambers with a mixture of variants having alpha, beta, gamma, or delta pili. Animals immunized with alpha pili showed significant protection, but those immunized with beta pili did not. The degree of protection could be correlated with the cross-reactivity of the antibodies produced to the heterologous pilus types.

Animals↗
PubMed1981

Bacteriological colonisation of uterine cavity: role of tailed intrauterine contraceptive device.

Intrauterine contraceptive devices (IUCDs) are thought to cause pelvic inflammatory disease by allowing vaginal bacteria to pass into the uterus along the tail of the device. In this study the uterine cavities of 22 women using an IUCD were examined by a multiple biopsy technique. All five uteruses with a tailless IUCD were sterile but 15 out of 17 with a tailed device contained bacteria. The bacteria had not reached the fundus and most were commensals. The bacteria were not introduced by insertion of the IUCD as bacteria were present in several cases long after insertion. No differences in bacterial count were found between monofilamentous and multifilamentous devices. Bacteria were cultured from only four devices, which suggested that the bacteria adhere to the endometrium and not to the device. The bacteria in the cavity represent interference by the tail with the protective mechanisms of the uterus, which explains the increase in pelvic inflammatory disease in IUCD users.

Adult↗
PubMed1981

The preparation and properties of alpha and beta pili from variants of Neisseria gonorrhoeae P9.

The alpha and beta type pili produced by variants of Neisseria gonorrhoeae P9 were isolated and characterized. The two types of pili showed clear differences in morphology, buoyant density (alpha, 1.292 g ml-1; beta, 1.282 g ml-1) and isoelectric point (alpha, pI 5.2; beta, pI 4.3). Amino acid analysis of alpha and beta pili showed that their overall composition was similar with the exception that beta pili had a higher content of glutamate and alanine residues. A high degree of structural homology was seen in two-dimensional peptide maps of tryptic hydrolysates of alpha and beta pili. The molecular differences between alpha and beta pili were reflected in their antigenic activity and in their ability to attach to human cells. The binding of alpha pili to buccal epithelial cells was pH dependent with a maximum binding of 49% at pH 6.5, whereas the attachment of beta pili showed no pH optimum. The binding of pili to erythrocytes differed from binding to buccal epithelial cells in that attachment of both pili was identical with no pH optimum. Data from attachment of pili to human cells suggest that erythrocytes lack receptors for binding gonococcal pili.

Amino Acids↗
PubMed1981

The role of outer membrane proteins in the survival of Neisseria gonorrhoeae P9 within guinea-pig subcutaneous chambers.

Guinea-pig subcutaneous chambers were infected with a mixture of gonococcal variants of defined outer membrane protein profile. Survival within the chambers was a two-stage process. The initial advantage conferred by the lack of opacity-related outer membrane protein was transient and survivors were replaced by opaque colonial variants. Amongst these survivors were variants which produced opacity-related proteins (IId, IIe and IIf) not present in the initial inoculum. Thus, outer membrane protein composition is an important factor in survival in vivo.

Animals↗
PubMed1980

The cohesive properties of variants of Neisseria gonorrhoeae strain P9: specific pilus-mediated and non-specific interactions.

The cohesive properties of virulent pilated Neisseria gonorrhoeae strain P9 (P++) have been compared with those of a non-pilated isogenic variant (P-) possessing the same outer membrane components. The binding of P++ gonococci to buccal epithelial cells was dependent on pH, with an optimum at pH 6.5 to 7.0 . This adhesion was markedly inhibited by treatment of the buccal epithelial cells with a neuraminidase/exoglycosidase mixture. In contrast, the binding of P++ gonococci to erythrocytes was unaffected by pH. A possible explanation is that pili bind to a carbohydrate receptor present on buccal epithelial cells but lacking on erythrocytes. The adhesion of P- gonococci to erythrocytes and to buccal epithelial cells was unaffected by pH but enhanced by treatment of the cells with neuaminidase or periodate. Presumably, neuraminic acid residues on host cell surface carbohydrates inhibit adhesion. The finding that P- gonococci bind to amphipathic gels suggests hydrophobic interactions as a possible non-specific mechanism attaching P- gonococci to host cell surfaces.

Erythrocytes↗