The growth and nine subcultivations of the experimental Nichols strain of pathogenic Treponema pallidum were successfully accomplished in cultured baby hamster kidney tissue cells (BHK-21) using serum-free media. The number (motile and non-motile cells occurring extracellulary) of cell generations generally increased with each subcultivation, the largest increases (greater than 3-0) occurring in subcultures 4, 5, 7, 8, and 9. Also, the number of motile cells decreased with each subcultivation. Virulent organisms were demonstrated in subcultures 1, 4, 5, 7, and 8, and their numbers estimated with the use of standards established by the animal inoculation titration of counted numbers of organisms freshly harvested from experimental infections; the estimated number of virulent organisms was higher than or equal to the counted cultivated treponemes injected, which may be attributable to uncounted virulent spirochaetes occurring intracellularly.
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