Borrelia burgdorferi in Tick Cell Culture: Growth and Cellular Adherence

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Abstract
Borrelia burgdorferi, strain 297, was incubated with tick cell lines isolated from Rhipicephalus appendiculatus (Neumann), R. sanguineus (Latreille), Anocentor nitens (Neumann), Dermacentor variabilis (Say), and Boophilus microplus (Canestrini). A tick cell culture medium, L-15B, was modified by the addition of gelatin and N'-acetylglucosamine (L-15B/S) to permit the cocultivation of B. burgdorferi and tick cells. Spirochetes continuously passaged axenically in L-15B/S had longer population doubling times (27.1 ± 4.5 h) than those grown in BSK medium (11.7 ± 2.2 h), which was unsuitable for tick cells. Growth of spirochetes cocultured with five of six lines did not change, but when B. burgdorferi was incubated with R. sanguineus cells, the spirochetes disappeared and could not be detected after 1 wk. Spirochetes bound themselves to tick cells by one end within 30 min after being added to cultures. Not all spirochetes attached to tick cells nor did all cells of a given line bind spirochetes. B. burgdorferi had the greatest affinity for embryonic R. appendiculatus cells and the least for D. variabilis cells. Scanning electron microscopy revealed numerous intertwined, coiled spirochetes attached to foci at the surface of R. appendiculatus cells, apparently causing the formation of round bodies. Fewer spirochetes attached to A. nitens cells. High spirochete concentrations (>107 per ml) elicited cytopathic effects—loss of surface membrane extensions, rounding up, detachment, and lysis. This system should be applicable in the study of tick cell-spirochete interactions and the analysis of spirochete tropism and binding to tick cells.