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J H Pearce

Publications and source records attributed to J H Pearce.

At least 37 records · Page 2Linked to original sources
PubMed1989

Ultrastructural studies on the intracellular fate of Chlamydia psittaci (strain guinea pig inclusion conjunctivitis) and Chlamydia trachomatis (strain lymphogranuloma venereum 434): modulation of intracellular events and relationship with endocytic mechanism.

Previous observations on the highly infectious LGV strain 434 of Chlamydia trachomatis and the guinea pig inclusion conjunctivitis (GPIC) strain of C. psittaci (which requires centrifugation of inocula with host cell monolayers for maximum infectivity) indicated that infectivity differences were expressed, not at entry, but at an intracellular stage affecting multiplication. Centrifugation increased the potential of internalized chlamydiae to undergo productive infection. Here, analysis of the intracellular fate of chlamydiae by ultrastructural methods indicates that strain GPIC exhibits two patterns of behaviour depending on the mode of inoculation. Strain GPIC showed limited entry, with 47% of intracellular organisms becoming associated with thorotrast-labelled lysosomes, following static incubation with monolayers. In contrast, with centrifugation, entry was not limited and association with lysosomes was reduced to 12%; strain 434 behaved similarly but independently of the mode of inoculation. The different results for strain GPIC correlated with distinct entry mechanisms. Entry during static incubation was unimpaired either by treatment with cytochalasin D or by temperature reduction to 20 degrees C, suggesting that it was pinocytic. Entry during centrifugation was markedly impaired by both treatments, suggesting that it was phagocytic. The data lead to two novel conclusions: first, that chlamydiae can apparently enter cells by both pinocytic and phagocytic mechanisms; second, that the entry mechanism influences intracellular fate. It is suggested that entry mechanism is linked to selection of the vesicle membrane forming around the internalizing chlamydiae. This, in turn, may influence both intracellular translocation and subsequent inhibition or promotion of multiplication of the internalized parasite.

Animals↗
PubMed1987

Association of Chlamydia trachomatis with mammalian and cultured insect cells lacking putative chlamydial receptors.

The association of radiolabelled Chlamydia trachomatis (strains DK-20 and 434) with cultured cell monolayers has been examined. Previously, N-acetyl neuraminic acid and N-acetyl-glucosamine had been suggested to be involved in the association of certain strains of C. trachomatis with cultured cell monolayers. We have now studied the association of non-LGV (DK-20) and LGV (434) strains of C. trachomatis with cultured cells lacking these putative receptor components. Comparison of the association of strain DK-20 with McCoy or HeLa cells (containing sialic acids), with neuraminidase-treated McCoy or HeLa or with Aedes aegypti cells (all lacking sialic acids), surprisingly, showed enhanced binding to cells lacking the putative receptor component. Similarly, comparison of the association of chlamydial strain 434 with BHK-21 wild-type (containing N-linked N-acetylglucosamine) and BHK-21 RicR14 mutant cells (lacking this component) showed enhanced binding to the mutant cells.

Aedes↗
PubMed1987

Regulation of Chlamydia psittaci (strain guinea pig inclusion conjunctivitis) growth in McCoy cells by amino acid antagonism.

Chlamydiae have amino acid requirements for growth in tissue culture as defined by those amino acids whose individual omission from the growth medium prevents chlamydial multiplication. We have tested the hypothesis that this inhibition of growth arises as a result of antagonism between particular amino acids such that inhibition occurs when the concentration of one amino acid is reduced in the presence of the antagonist amino acid at high concentration. Using the Chlamydia psittaci strain guinea pig inclusion conjunctivitis (GPIC), in the presence of cycloheximide, the requirement for valine was abrogated by the simultaneous omission of isoleucine, that for phenylalanine by simultaneous omission of tryptophan and that for leucine by simultaneous omission of isoleucine plus valine. The antagonism shown between leucine and isoleucine plus valine appears to be unique among bacteria. In the absence of cycloheximide, GPIC had an additional need for tryptophan, tyrosine and isoleucine; these amino acid requirements were shown for both infected McCoy, HeLa and BHK cells. The results are consistent with a mechanism for regulation of parasite growth which depends on the balance of amino acid concentrations in the extracellular environment.

Amino Acids↗
PubMed1985

Influence of cysteine deprivation on chlamydial differentiation from reproductive to infective life-cycle forms.

The effects of omission of individual amino acids from growth medium on the differentiation of Chlamydia trachomatis DK-20 (serotype E) during infection of cycloheximide-treated McCoy cells are described. As judged by inclusion body staining with acridine orange, omission of cysteine from the medium severely retarded differentiation of reproductive reticulate body (RB) to infective elementary body (EB) forms. The effect appeared specific to cysteine in that omission of other amino acids had little or no effect on differentiation once RBs appeared. On restoration of cysteine, culture infectivity increased and inclusions contained organisms which, by cytochemical and morphological criteria, were differentiating to infective forms, indicating that cysteine deprivation did not irreversibly inhibit differentiation. Impairment of RB to EB differentiation in cysteine-less medium was also observed for three strains of Chlamydia psittaci and 10 other strains of C. trachomatis. It is suggested that the effect arises via the biosynthetic requirement for cysteine for provision of three cysteine-rich proteins, whose synthesis and insertion into the outer membrane have previously been shown to accompany RB to EB differentiation of C. psittaci 6BC and C. trachomatis 434 (serotype L2). Synthesis of cysteine-rich outer membrane proteins during differentiation may thus be common to all chlamydiae.

Amino Acids↗
PubMed1984

Structural and polypeptide differences between envelopes of infective and reproductive life cycle forms of Chlamydia spp.

Significant differences in cysteine-containing proteins and detergent-related solubility properties were observed between outer membrane protein complexes of reproductive (reticulate body) and infective (elementary body) forms of Chlamydia psittaci (6BC). Elementary bodies harvested at 48 h postinfection possessed a 40-kilodalton major outer membrane protein and three extraordinarily cysteine-rich outer membrane proteins of 62, 59, and 12 kilodaltons, all of which were not solubilized by sodium dodecyl sulfate in the absence of thiol reagents. Intracellularly dividing reticulate bodies harvested at 21 h postinfection were severely deficient in the cysteine-rich proteins but possessed almost as much major outer membrane protein as did the elementary bodies. Most of the major outer membrane protein of reticulate bodies was solubilized by sodium dodecyl sulfate and was present in envelopes as monomers, although a proportion formed disulfide-cross-linked oligomers. By 21 to 24 h postinfection, reticulate bodies commenced synthesis of the cysteine-rich proteins which were found in outer membranes as disulfide-cross-linked complexes. The outer membranes of reticulate bodies of Chlamydia trachomatis (LGV434) also were found to be deficient in cysteine-rich proteins and to be more susceptible to dissociation in sodium dodecyl sulfate than were outer membranes of elementary bodies.

Alkylation↗
PubMed1983

Differential amino acid utilization by Chlamydia psittaci (strain guinea pig inclusion conjunctivitis) and its regulatory effect on chlamydial growth.

The effect of omission of individual amino acids from growth medium on the multiplication of Chlamydia psittaci (strain guinea pig inclusion conjunctivitis) in cycloheximide-treated McCoy cells has been examined. Marked differences were observed in the amounts of particular amino acids required for normal chlamydial multiplication: omission of either leucine, phenylalanine or valine completely inhibited multiplication, whereas absence of any one of another 10 amino acids had no effect on numbers of cells infected. Threshold concentrations of 80, 80 and approx. 8 nmol ml-1 for leucine, valine and phenylalanine, respectively, were needed for normal chlamydial multiplication. These requirements could not be related either to unusually high content in the whole organism, to degradation in the medium, or, from studies with leucine, to deficient association of leucine with host cells. Leucine deprivation at late stages of the developmental cycle also appeared to regulate multiplication. Possible mechanisms responsible for these effects are discussed.

Amino Acids↗
PubMed1983

Amino acid requirements of strains of Chlamydia trachomatis and C. psittaci growing in McCoy cells: relationship with clinical syndrome and host origin.

The effects of omission of individual amino acids from growth medium on the multiplication of a range of Chlamydia trachomatis and C. psittaci strains in cycloheximide-treated McCoy cells have been assessed. Differences in requirements were revealed which for C. trachomatis strains correlated with clinical syndrome and for C. psittaci with host origin. All 11 strains of C. trachomatis examined showed a requirement for addition of histidine to the medium; this was not shown by any of four C. psittaci strains. Among the strains of C. trachomatis, three from cases of trachoma, representing serotypes A, B and C, showed a distinctive requirement for the addition of tryptophan to the medium, whilst six strains of oculogenital origin, representing serotypes D-I, exhibited no requirement for tryptophan or methionine; a lymphogranuloma venereum and a 'fast variant' strain both showed a requirement for methionine. Of the four C. psittaci strains from different hosts, three showed distinct patterns of amino acid requirements. All chlamydiae required the addition of valine to medium and the majority required leucine, phenylalanine and also glutamine.

Amino Acids↗
PubMed1981

Interaction of chlamydiae with host cells and mucous surfaces.

For chlamydiae, as obligate intracellular parasites, attachment to and ingestion by host cells are essential steps in reproduction. Their attachment site appears to be heat-sensitive; it has not been correlated with any morphological entity. Antibody blocks chlamydial attachment to cells and, for certain chlamydia psittaci and chlamydia trachomatis strains which are highly infective for cell cultures, N-acetylglucosamine appears to contribute to cell receptor specificity. Sialic acid residues have been suggested as receptors for other C. trachomatis strains. The guinea-pig inclusion conjunctivitis strain of C. psittaci becomes associated with the conjunctiva during incubation of inoculated tissue fragments in vitro. However, although antibody from tears neutralizes infectivity of this strain in vivo, association of the organism with tissue fragments is not inhibited, suggesting that antibody neutralization in vivo is not mediated by prevention of attachment to cells. Chlamydial infectivity for cell monolayers is greatly increased by centrifugation. The process is temperature-dependent and involves cooperative interactions between direction force and the pressure generated during centrifugation. Enhanced infectivity appears to result from changes induced in the cell surface. These changes may favour nonspecific interactions in attachment, since antibody inhibition of infectivity on static cell monolayers is overcome by centrifugation.

Acetylglucosamine↗
PubMed1980

Inapparent chlamydial infection in the urogenital tract of guinea-pigs.

Microbiologically inapparent urogenital infection appeared to be induced in male guinea-pigs inoculated intra-urethrally with low doses of the guinea-pig inclusion conjunctivitis strain (GP-IC) of Chlamydia psittaci. This state was indicated by the ability of inoculated animals to donate eye infection to normal animals caged with them. Donors failed to develop overt urogenital infection throughout the period of transmission judged by both absence of infected cells in urethral scrapings and failure to isolate GP-IC in cell culture; however, inoculation of donors with 5-iododeoxyuridine led to transient appearance of infectivity in scrapings. In distinction from overtly infected animals, donors failed to develop serum antibody and remained susceptible to urethral challenge with larger doses of GP-IC. Animals that had recovered from overt urethral infection were resistant to challenge and appeared unable to transmit eye infection.

Animals↗
PubMed1979

Modulation by centrifugation of cell susceptibility to chlamydial infection.

Enhancement of chlamydial infection of cell monolayers by centrifugation was shown to depend on induced cell surface changes. Evidence for this came from analysis of two forms of organism attachment which take place during centrifugation. In 'productive binding', organisms attached to cells and then entered and infected them. In 'unproductive binding', organisms became attached to cells but were not ingested. These organisms could be stripped from the cells by treatment with trypsin and could then infect fresh monolayers. Measurement of attachment kinetics during centrifugation showed that cells passed through three different susceptibility states. Only productive binding occurred in the first 20 min; cells then entered a refractory state during which no attachment took place At about 45 min, attachment recommenced but this allowed only unproductive binding. Induced movement of cell surface structures may enhance infection by promoting specific or non-specific interactions. Failure of ingestion may result from insufficient cell 'receptors' for circumferential binding of the whole chlamydial surface so that engulfment cannot take place.

Cell Membrane↗
PubMed1979

Host modification of chlamydiae: differential infectivity for cell monolayers of chlamydiae grown in eggs and monolayers.

Cell monolayer-grown chlamydiae (CGO) differed from egg-grown organisms (EGO) in their increased spontaneous infectivity relative to centrifuge-assisted infectivity for monolayers. For each population spontaneous: centrifuge-assisted infectivity ratios were constant over a wide dose range. Spontaneous infection increased linearly with time and could not be exhausted from either population by prolonged adsorption; there was no change in infectivity ratios in residual supernatants. Further, one passage of EGO through monolayers gave CGO with stable infectivity properties not increased by further cell passage yet reverting on a single passage in eggs. Spontaneous infection of monolayers with EGO gave progeny with the same infectivity ratios as monolayers infected with EGO by centrifugation. The change in properties following EGO infection of monolayers occurred prior to natural release from cells. We conclude that EGO and CGO are two phenotypically distinct, homogeneous populations. The two infection modes are not properties of subpopulations within EGO and CGO. The relationship of these observations on chlamydiae to other possible host-imposed phenomena is considered.

Animals↗
PubMed1979

Host modification of chlamydiae: presence of an egg antigen on the surface of chlamydiae grown in the chick embryo.

Egg-grown chlamydiae (EGO) have a yolk sac antigen assoicated with their surface which is absent from cell monolayer-grown organisms (CGO). EGO infectivity was specifically neutralized by rabbit antiserum to normal yolk sac; CGO infectivity, before or after incubation with normal yolk sac material, was not neutralized. Treatment of EGO with Clostridium welchii culture filtrate, containing phospholipase C, abolished spontaneous infectivity for monolayers and neutralization by anti-yolk sac antiserum but did not affect centrifuge-assisted infectivity. The possible significance of host antigen on the chlamydial surface is considered.

Animals↗
PubMed1979

Differential neutralization of spontaneous and centrifuge-assisted chlamydial infectivity.

Neutralization by specific antibody of a fast-killing variant strain of Chlamydia trachomatis, which showed high spontaneous infectivity for cell monolayers, was examined. It appeared that in spontaneous infection antibody-treated chlamydiae were neutralized by inhibition of attachment to cells. Centrifugation imposed a different effect: infection was inhibited at some step at or subsequent to attachment.

Antibodies, Bacterial↗
PubMed1976

Infectivity titration of guinea-pig inclusion conjunctivitis agent in irradiated McCoy cells.

Estimation of the infectivity of the agent of guinea-pig inclusion conjunctivitis for irradiated McCoy cells, assayed as inclusion-forming units, was influenced by the age of cells after irradiation, the maturation time of the inclusions, the centrifugal force and the centrifugation temperature. Agent passaged through irradiated McCoy cells or guinea-pig conjunctivae showed a greater capacity to infect irradiated McCoy cells without centrifuging than agent grown in a chick embryo. The nature of the change and the mechanism of infectivity enhancement by centrifuging are discussed.

Animals↗