Unsustained multiplication of treponema pallidum (nichols virulent strain) in vitro in the presence of oxygen

Abstract

Treponema pallidum (Nichols virulent strain) was incubated with or without oxygen, using a modified medium supplemented with reduced glutathione and a variety of nutrients (PRNFjo-B). Twoto fourfold increases in treponemal numbers were observed in cultures without mammalian cells within 96 h of incubation under 5 to 6% oxygen. Treponemal motility and multiplication were maintained more satisfactorily in cultures that were diluted and transferred daily, using an equal volume of fresh medium. Treponemes incubated without oxygen did not significantly increase in number. Virulent microorganisms were detected for at least 96 h in the cell-free system. In the presence of 3 to 4% oxygen, two-to fivefold increases in treponemal numbers were observed in the supernatant fluids of cultures containing human prepuce cells after 48 to 120 h at 35°C. Without oxygen, treponemal numbers rarely approached a threefold increase. Virulent treponemes were detected by the rabbit skin lesion test after at least 120 h in vitro. Regardless of the system of incubation, increases in treponemal numbers could not be sustained for longer than 120 h, and treponemal virulence decreased as a function of time in vitro.

Further studies on replication of virulent Treponema pallidum in tissue cultures of Sf1Ep cells

“…In survival studies of T. pallidum in tissue culture, consideration had to be given to temperatures that would favor growth of the tissue cultures as well as the treponemes. Thus, temperatures between 33 to 36°C have been previously utilized (2,5,8). In the present studies, the optimum temperature for replication of T. pallidum was found to be between 33 and 35°C.…”

Virulence and Adhesion of the Treponema pallidum Nichols Strain Simultaneously Decrease in a Continuous-Infection New Zealand White Rabbit Model

“…It has been reported that the level of T. pallidum virulence decreased after culture in vitro . ,,− Besides, few research studies on the virulence of T. pallidum after passage in vivo have been reported.…”

In vitro cultivation of Treponema pallidum: independent confirmation

“…Their system utilized a combination of several important elements which had been previously investigated in their own laboratory and by others, including (i) monolayer cultures of a slow-growing mammalian cell line, SflEp cottontail rabbit epithelial cells (6, 10, 14); (ii) a modified tissue culture medium with 20% fetal bovine serum (FBS) and 0.63 mM dithiothreitol (DTT) to reduce oxygen toxicity (2,6,8,9,12); (iii) a small quantity (1/30th of the total medium volume) of infected testes extract ( 6) and (iv) incubation at 33°C under a microaerobic atmosphere containing 1.5% oxygen (1, 3,6,12,13). Because of the importance of this finding for future T. pallidum research and past failures in repeating reported cultivations, the techniques of Fieldsteel et al were duplicated in an attempt to independently corroborate their results.…”