Fluorescent membrane markers elucidate the association of Borrelia burgdorferi with tick cell lines

• Published in: Brazilian journal of medical and biological research Vol. 49; no. 7
• Main Authors: Teixeira, R C, Baêta, B A, Ferreira, J S, Medeiros, R C, Maya-Monteiro, C M, Lara, F A, Bell-Sakyi, L, Fonseca, A H
• Format: Journal Article
• Language: English
• Published: Brazil Associacao Brasileira de Divulgacao Cientifica (ABDC) 01-01-2016; Associação Brasileira de Divulgação Científica
• Subjects: Animals; BIOLOGY; Biomedical Sciences; Borrelia burgdorferi; Borrelia burgdorferi - isolation & purification; Cell Line; Cells, Cultured; Culture Media; Flow cytometry; Flow Cytometry - methods; Fluorescence; Fluorescent Dyes; Fluorescent membrane marker; Ixodidae; MEDICINE, RESEARCH & EXPERIMENTAL; Microscopy, Confocal - methods; Organic Chemicals; Phagocytosis; Reproducibility of Results; Rhipicephalus appendiculatus; Spirochaetales - isolation & purification; Staining and Labeling - methods; Tick cell lines; Tick-Borne Diseases - microbiology; Ticks - cytology; Ticks - microbiology; Time Factors; Tick cell lines; Borrelia burgdorferi; Phagocytosis; Fluorescent membrane marker
• ISSN: 0100-879X; 1414-431X; 1414-431X
• DOI: 10.1590/1414-431X20165211

This study aimed to describe the association of Borrelia burgdorferi s.s. with ixodid tick cell lines by flow cytometry and fluorescence and confocal microscopy. Spirochetes were stained with a fluorescent membrane marker (PKH67 or PKH26), inoculated into 8 different tick cell lines and incubated at 30°C for 24 h. PKH efficiently stained B. burgdorferi without affecting bacterial viability or motility. Among the tick cell lines tested, the Rhipicephalus appendiculatus cell line RA243 achieved the highest percentage of association/internalization, with both high (90%) and low (10%) concentrations of BSK-H medium in tick cell culture medium. Treatment with cytochalasin D dramatically reduced the average percentage of cells with internalized spirochetes, which passed through a dramatic morphological change during their internalization by the host cell as observed in time-lapse photography. Almost all of the fluorescent bacteria were seen to be inside the tick cells. PKH labeling of borreliae proved to be a reliable and valuable tool to analyze the association of spirochetes with host cells by flow cytometry, confocal and fluorescence microscopy.