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S Graves

Publications and source records attributed to S Graves.

At least 55 records · Page 3Linked to original sources
PubMed1984

Susceptibility of Treponema pallidum to the toxic products of oxygen reduction and the non-treponemal nature of its catalase.

We examined the sensitivity of Treponema pallidum (Nichols strain) to toxic products of oxygen reduction. T pallidum was sensitive to hydrogen peroxide at concentrations similar to those to which obligate anaerobes are sensitive. Accelerated death of T pallidum occurred at hydrogen peroxide concentrations below 50 mumol/l. Agents protective against hydrogen peroxide and the hydroxyl free radical (catalase, peroxidase, and mannitol) significantly enhanced treponemal survival in vitro under all three conditions of aerobiosis tested--that is, air, 3% oxygen, and 3% oxygen in conjunction with a reduced medium. Superoxide dismutase (which provides protection against superoxide radicals) did not enhance treponemal survival in normal media. When superoxide radicals were generated in the medium by means of a xanthine and xanthine oxidase system, however, the enzyme did protect T pallidum. A possible toxic involvement of singlet oxygen was also indicated by enhanced treponemal survival in air in the presence of histidine. Extracts of T pallidum from infected rabbit testes showed catalase activity which, on polyacrylamide gel electrophoresis, had the same relative mobility as purified rabbit catalase. The treponemal catalase activity was neutralised by anti rabbit catalase antiserum (raised in guinea pigs). This confirmed that the catalase was of rabbit origin and not an endogenous enzyme of T pallidum. We discuss the relation of these results to the obligate parasitism of T pallidum.

Animals↗
PubMed1984

Inhibitory effect of syphilitic rabbit serum on DNA synthesis in rabbit cells in vitro.

A previously described toxic factor associated with Treponema pallidum (Nichols) and found in extracts of syphilitic rabbit testes has now also been detected in syphilitic rabbit serum. The toxic factor, which inhibits DNA synthesis in baby rabbit genital organ (BRGO) cells in vitro, is present in rabbit serum up to 30 days after infection with T pallidum.

Animals↗
PubMed1984

Oxygen toxicity in Treponema pallidum: deoxyribonucleic acid single-stranded breakage induced by low doses of hydrogen peroxide.

The effect of hydrogen peroxide on Treponema pallidum was investigated. The in vitro loss of virulence (as measured by rabbit inoculation) of T. pallidum was accelerated by as low as 100 microM hydrogen peroxide in the complex maintenance medium used. Higher doses led to rapidly accelerated death with 500 microM hydrogen peroxide causing sterilization of the medium within 3 to 4 h. Since hydrogen peroxide is known to cause single-stranded breaks in DNA, the effect of hydrogen peroxide on the treponemal genome was examined. Extensive breakage was caused by 100 microM hydrogen peroxide as determined on alkaline sucrose gradients. A limit was reached at 250 microM and above. Single-stranded breaks could be demonstrated as early as 5-10 min after exposure to hydrogen peroxide when the treponemes were exposed to 250 microM hydrogen peroxide; accelerated death was evident by 2 h past exposure demonstrating that DNA breakage was preceding death. Treponemal death caused by penicillin did not result in DNA breakage. The repair-proficient bacterium Escherichia coli K-12 was compared with T. pallidum. It required 10-100 times more hydrogen peroxide to cause various levels of breakage. Escherichia coli K-12 rapidly repaired DNA breakage once hydrogen peroxide was removed by addition of catalase. Treponema pallidum, in comparison, showed little or no repair in vitro. Addition of catalase or dithiothreitol to the medium protected against all but a low level of breakage; this may reflect on the ability of catalase and reducing agents to protect T. pallidum against oxygen toxicity in vitro.

DNA Repair↗
PubMed1983

Effect of serum concentration and metabolic inhibitors on the attachment of Treponema pallidum to rabbit cells.

The effect of host-cell metabolism on the attachment of Treponema pallidum to mammalian cells in vitro was studied. The growth of baby rabbit genital organ (BRGO) cells was enhanced by increasing the concentration of serum ("serum shift-up") in the growth medium. Cells starved for 24 h in serum-free medium showed a burst of DNA synthesis when shifted to fresh medium containing 20% serum. In aerobic conditions, they were much more heavily coated with attached T. pallidum than cells shifted to 20% serum after maintenance at serum concentrations of 2.5%, 5% or 10%. This effect was very pronounced during the first few hours of co-incubation. In microaerophilic conditions, the extent of T. pallidum adherence also paralleled the increase in DNA synthesis by BRGO cells. Cycloheximide and methotrexate greatly inhibited DNA and protein synthesis in BRGO cells, but did not affect the motility of T. pallidum. When BRGO cell metabolism was inhibited by these two drugs, attachment of T. pallidum was significantly decreased. These results indicate that T. pallidum attaches best to actively growing BRGO cells in tissue culture. This may explain the apparently preferential parasitism of actively growing tissues by T. pallidum in syphilis in man.

Adhesiveness↗
PubMed1983

Inhibition of macromolecular synthesis in cultured rabbit cells by Treponema pallidum (Nichols).

Treponema pallidum partially inhibited the synthesis of DNA, RNA, and protein by rabbit cells in vitro. The inhibition of DNA synthesis was proportional to treponemal concentration and persisted during the period of exposure to T. pallidum. The toxic effect was not dependent on treponemal metabolism or on whole treponemes, since heat- and penicillin-killed treponemes and a cell-free sonicate of treponemes had similar toxicities. The toxic factor(s) was also detected in extracts of syphilitic rabbit testes but not in extracts of normal rabbit testes or testes inflamed by chemical means. The T. pallidum-derived toxic material had a molecular weight greater than 20,000 as determined by dialysis. Protein and DNA synthesis were most rapidly inhibited; RNA synthesis continued at normal rates for up to 2 h after exposure to treponemes. Protein synthesis or a necessary precursor of protein synthesis appeared to be the primary target of the T. pallidum toxin(s).

Animals↗
PubMed1983

Factors affecting the attachment of Treponema pallidum to mammalian cells in vitro.

Attachment of Treponema pallidum (Nichols) to mammalian cells is probably the first step in the pathogenesis of syphilis. It may also be important for the multiplication of T pallidum in vitro. When factors affecting the attachment of T pallidum to mammalian cells in vitro were studied significantly greater numbers of treponemes were found to attach to baby rabbit genital organ (BRGO) cells than to five other mammalian cell lines. When attached to BRGO cells T pallidum survived longer in vitro than unattached treponemes. Eagle's minimal essential medium was superior to three other culture media in increasing attachment and maintaining the survival of treponemes. Dithiothreitol (0.25-1.0 mmol/l) had no effect on the attachment of T pallidum to BRGO cells. Anaerobic conditions were superior to microaerophilic conditions, and the latter were superior to aerobic conditions for the attachment and survival of T pallidum to BRGO cells. Within the range of concentrations tested the number of treponemes attached to the BRGO cells was directly dependent on the concentrations of viable treponemes in the inoculum. Greater numbers of treponemes attached to actively metabolising BRGO cells than to quiescent or slowly growing cells.

Animals↗
PubMed1983

Effect of syphilitic rabbit sera taken at different periods after infection on treponemal motility, treponemal attachment to mammalian cells in vitro, and treponemal infection in rabbits.

The time course of antibody synthesis during syphilis was studied in experimentally infected rabbits. A rapid antibody response was seen; the rabbits became positive in both the rapid plasma reagin (RPR) test and Treponema pallidum haemagglutination assay (TPHA) by nine days after infection. Treponemal immobilising antibodies were also seen as early as nine days after infection. Antibody inhibition of treponemal attachment to baby rabbit genital organ (BRGO) cells in culture occurred with immune sera taken 30 days after infection but not earlier. When T pallidum was mixed with immune syphilitic rabbit sera taken at different stages of the infection and used to infect normal rabbits the rabbits became partially resistant to T pallidum only when the treponemes were mixed with sera taken at least 30 days after syphilitic infection. This appearance correlated well with the development of antibodies which blocked attachment of T pallidum to host cells. These antibodies may be involved in the resistance to reinfection which develops in syphilis as the disease progresses.

Animals↗
PubMed1983

The role of respiratory protection on increased survival of Treponema pallidum (Nichols) when cocultivated with mammalian cells in vitro.

The ability of mammalian cells in tissue culture to protect against oxygen toxicity for Treponema pallidum was examined. Addition of catalase to the incubation medium enhanced T. pallidum survival when co-incubation was carried out under aerobic conditions. When co-incubation was carried out under 3% oxygen, catalase had no enhancing effect on survival despite the fact it was still highly stimulatory when T. pallidum was incubated under 3% oxygen in the same medium with no tissue culture cells present. Inactivation of the catalase present endogenously in the mammalian cells by the addition of the catalase inhibitor 3-amino-1,2,4-triazole largely eliminated the enhancing effect of mammalian cells on the survival of T. pallidum under 3% oxygen. Increasing the oxygen consumption of the host mammalian cells with 0.1 mM 2,4-dinitrophenol enhanced T. pallidum under both aerobic and microaerobic conditions; a much greater effect was seen under aerobic conditions. The results indicated that mammalian cells offer significant protection against toxic oxygen reduction products for T. pallidum in vitro under microaerobic conditions.

2,4-Dinitrophenol↗
PubMed1982

Relationship between direct-potentiometric and flame-photometric measurement of sodium in blood.

The relationship between direct-potentiometric and flame-photometric measurements of sodium has been studied in human sera and other, simpler systems. When water content was varied by using an inert substance (silica), de-ionized sera, or de-ionized individual proteins, the percentage that the flame photometric values were of the direct potentiometric Na+ values was identical to the measured water content. However, in 35 patients the percentage of Na+ values was 99.1% and the water content was 92.0%, a discrepancy of 7.1%. De-ionization of sera removed this discrepancy, apparently because of the lower pH of the de-ionized sera. The percentage of flame-photometric to direct-potentiometric Na+ values varied as a function of pH in pooled sera; in lyophilized and reconstituted sera; and in de-ionized, lyophilized, and reconstituted sera. Four explanations for the discrepancy between the percentage of Na+ values and the water content are discussed: a calibration problem, a measurement artifact, Na+ binding, and water binding. At this time there are no definitive data concerning which explanation is correct. We expect that our finding of a pH dependency for the percentage of measured Na+ values can be used to develop model systems to elucidate the mechanism producing the discrepancy between the percentage of Na+ values and the water content.

Bicarbonates↗
PubMed1981

Sequential changes in susceptibility to Treponema pallidum of rabbits previously infected with Treponema paraluis-cuniculi.

Rabbits immunised with virulent Treponema paraluis-cuniculi were challenged intradermally with graded doses of Treponema pallidum at three, five, seven, 12, and 30 months to ascertain the level of protection to T pallidum at various intervals after immunisation.Rabbits challenged at three months after immunisation showed no protection against T pallidum and developed syphilitic lesions significantly faster than the control rabbits, which suggests that the former rabbits were immunosuppressed. Some protection was evident at five and seven months after immunisation, as fewer inoculation sites developed syphilitic lesions with challenges of 10(3), 10(2), and 10 T pallidum and lesions developed significantly slower with 10(6) challenge. Two rabbits showed significant protection at 12 months after immunisation but a third, presumably still immunosuppressed, developed lesions significantly faster than the control rabbits after challenge. At 30 months after immunisation one rabbit was completely protected and developed no lesions after challenge; the other rabbit showed only partial protection against challenge with 10(4), 10(3), and 10(2) but complete protection against challenge with 10 T pallidum.T paraluis-cuniculi appeared to induce a state of immunosuppression by three months after infection; in one rabbit this may have been 12 months. In most immunised rabbits, however, limited cross-protection to low challenge doses of T pallidum developed by five months and was also detectable at seven and 12 months. Only one rabbit was completely resistant to challenge with 10(4)T pallidum after 30 months and another was only partly immune. Thus, T paraluis-cuniculi infection does not produce a rapid pronounced cross-immunity to T pallidum in rabbits, which may thus limit its usefulness as a vaccine against syphilis.

Animals↗
PubMed1981

Experimental infection of man with rabbit-virulent Treponema paraluis-cuniculi.

Virulent Treponema paraluis-cuniculi was inoculated intradermally into the arm of a human volunteer and into the shaved backs of 10 rabbits. An identical, but heat-killed, preparation was inoculated into the opposite arm of the volunteer as control. A superficial and transient infection developed in the volunteer, shown by a small zone of erythema that persisted for 24 days. The control preparation caused a smaller zone of erythema that disappeared after five days. A very poor immune response was detected by standard serological tests for syphilis. The inoculated rabbits developed lesions about six days after infection and seroconverted by 84 days. The poor antitreponemal antibody response to T paraluis-cuniculi infection in the volunteer suggests that this naturally attenuated treponeme may not be suitable as a vaccine against infection with t pallidum in humans.

Animals↗
PubMed1981

Redox potential and survival of virulent Treponema pallidum under microaerophilic conditions.

A strongly reduced culture medium, capable of maintaining the virulence of Treponema pallidum (Nichols) for several days, was exposed to an atmosphere of 3% oxygen in nitrogen for 2-3 days before inoculation with T pallidum. By using various volumes of medium in uniform tubes a range of redox potentials (Ecal) from -94 mV to -325 mV was produced depending on the surface area-to-volume ratios of the medium. The anaerobic medium had an Ecal value of -387 mV. The medium was inoculated with T pallidum and incubated in an atmosphere of 3% oxygen. The survival of treponemes at different redox potentials was monitored by observing the retention of motility and by measuring the latent period of infection after inoculation of the cultures into the shaved backs of rabbits. Under these conditions T pallidum survived longest at low (electronegative) redox potential. An inverse linear relationship was observed between the redox potential of the culture medium and the survival of T pallidum, as measured by the time required for a 90% reduction of virulent organisms. No optimum redox potential was detected, the most electronegative medium (-325 mV, Ecal) giving the best survival.

Bacteriological Techniques↗
PubMed1980

Rate of clearance of virulent Treponema pallidum (Nichols) from the blood stream of normal, Mycobacterium bovis BCG-treated, and immune syphilitic rabbits.

The rate of clearance of virulent Treponema pallidum (Nichols) from the blood stream of normal rabbits and rabbits previously treated with Mycobacterium bovis BCG was similar, there being treponemes still circulating 8 h after intravenous inoculation. In contrast, immune syphilitic rabbits cleared the virulent treponemes within 1 to 2 hours. Rabbits with passive humoral immunity to T. pallidum (after the transfer of 70 ml of immune serum) showed a similar clearance rate to that of the immune rabbits. Rabbits previously treated with BCG and with passive humoral immunity did not show a synergistic enhanced clearance rate, it being similar to that of immune rabbits.

Animals↗
PubMed1980

Susceptibility of rabbits venereally infected with Treponema paraluis-cuniculi to superinfections with Treponema pallidum.

Three female rabbits, venereally infected with Treponema paraluis-cuniculi between five and eight months previously, had treponeme-containing genital lesions and positive results to serological tests for syphilis. These rabbits and four normal female rabbits were challenged with Treponema pallidum intradermally on the shaved back with triplicate doses of 10(4), 10(3), 10(2), and 10 treponemes (all at different sites). Significantly fewer syphilitic lesions developed in the rabbits previously infected with T paraluis-cuniculi (4/36) compared with the control rabbits (37/48), and the mean size of the lesions (diameter of induration) was significantly less (4 mm compared with 10 mm). It appears that previous venereal infection of female rabbits with T paraluis-cuniculi induced a level of cross-immunity against infection with T pallidum. Protection was however not complete.

Animals↗
PubMed1979

Optimum concentration of dissolved oxygen for the survival of virulent Treponema pallidum under conditions of low oxidation-reduction potential.

A maintenance medium with a low oxidation-reduction (redox) potential, when gently bubbled with 5% oxygen in nitrogen or with air for various periods of time, gave a range of dissolved oxygen concentrations between 1.6 and 5.8 micrograms/l. Virulent Treponema pallidum (Nichols strain) inoculated into these media were assayed 24 and 48 hours later for motility and virulence and were compared with samples taken at zero time. Virulent T. pallidum survived best in the presence of 2.4 micrograms/l dissolved oxygen over a 48-hour period, which corresponded to a gaseous mixture of 3% oxygen in nitrogen. Higher concentrations of oxygen did not give significantly different results from anaerobic conditions over this period. Thus, until it can be grown in vitro, T. pallidum would appear to be a microaerophilic bacterium.

Anaerobiosis↗
PubMed1979

Susceptibility of rabbits to Treponema pallidum after infection with Mycobacterium bovis.

Rabbits stimulated with Mycobacterium bovis (strain BCG) one month before challenge with Treponema pallidum (Nichols) did not show any modification in their development of syphilitic lesions. A second infection with BCG given at the same time and at the same intradermal site as the T. pallidum challenge also failed to protect the rabbits against syphilis. Thus the non-specific activation of cell-mediated immunity by BCG does not appear to protect rabbits against T. pallidum infection even when both activation and challenge take place in the dermis. The role of the macrophage in syphilis remains obscure.

Animals↗