Norris S J, Edmondson D G
Infect Immun. 1986 Sep;53(3):534-9. doi: 10.1128/iai.53.3.534-539.1986.
Limited multiplication of Treponema pallidum subsp. pallidum (Nichols strain) can be obtained in the presence of Sf1Ep rabbit epithelial cell cultures, but continuous culture has not yet been achieved. In the system currently employed, growth is exponential for the first 10 to 15 days of culturing, after which multiplication and the percentage of motile organisms decrease. In an effort to identify culture conditions which may adversely affect treponemal viability and growth, eight culture parameters were monitored over a 12-day period of incubation. Several of these parameters, including pH, redox potential, dissolved oxygen concentration, and glucose levels were found to change dramatically during the course of incubation, indicating that they may be responsible for the cessation of treponemal multiplication. The feasibility of extending the period of growth by subculturing was also investigated. In preparation for planned serial subcultivation experiments, several subculture procedures were tested and found to be effective in allowing the transfer of T. pallidum from 3-day-old primary cultures to secondary cultures without loss of motility or growth potential. Increases of up to 55-fold were observed in secondary cultures, but increased growth due to subculturing was not a consistent finding. Use of subculture intervals of greater than or equal to 6 days resulted in a progressive decrease in treponemal multiplication in secondary cultures, although retention of motility was extended in the subcultures compared with motility in the primary cultures. These results indicate that the lack of continued multiplication of T. pallidum in subcultures is not due to damage to the treponemes during subculture. Prolonged multiplication of T. pallidum may be obtained through the stabilization of culture conditions by either performing subcultures at regular intervals or by medium replacement techniques. It was also found that primary T. pallidum cultures could be established by using as the inoculum treponemes that had been stored at -70 degrees C in a medium containing 15% glycerol.
在Sf1Ep兔上皮细胞培养物存在的情况下，梅毒螺旋体苍白亚种（Nichols株）可实现有限增殖，但尚未实现连续培养。在目前使用的系统中，培养的前10至15天生长呈指数增长，之后增殖以及活动生物体的百分比会下降。为了确定可能对梅毒螺旋体活力和生长产生不利影响的培养条件，在12天的孵育期内监测了八个培养参数。发现其中几个参数，包括pH值、氧化还原电位、溶解氧浓度和葡萄糖水平在孵育过程中发生了显著变化，表明它们可能是梅毒螺旋体增殖停止的原因。还研究了通过传代培养延长生长周期的可行性。在准备计划中的连续传代培养实验时，测试了几种传代培养程序，发现它们能有效地将3日龄原代培养物中的梅毒螺旋体转移至二代培养物，而不会丧失活力或生长潜力。在二代培养物中观察到增殖增加高达55倍，但传代培养导致的生长增加并非一致的结果。使用大于或等于6天的传代间隔会导致二代培养物中梅毒螺旋体增殖逐渐减少，尽管与原代培养物中的活力相比，传代培养物中的活力保留时间延长。这些结果表明，梅毒螺旋体在传代培养中缺乏持续增殖并非由于传代培养过程中对螺旋体的损伤。通过定期传代培养或培养基更换技术稳定培养条件，可能实现梅毒螺旋体的长期增殖。还发现，通过使用保存在含15%甘油培养基中于-70℃保存的梅毒螺旋体作为接种物，可以建立梅毒螺旋体原代培养物。
