title: Fluorescent membrane markers elucidate the association of Borrelia burgdorferi with tick cell lines.
abstract: 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.
authors.affiliation: Universidade Federal Rural do Rio de Janeiro, Universidade Federal Rural do Rio de Janeiro, Laboratório de Doenças Parasitárias, Instituto de Veterinária, Seropédica, RJ , Brasil, Laboratório de Doenças Parasitárias, Instituto de Veterinária, Universidade Federal Rural do Rio de Janeiro, Seropédica, RJ, Brasil.
authors.forename: R C
authors.initials: RC
authors.lastname: Teixeira
authors.affiliation: Universidade Federal Rural do Rio de Janeiro, Universidade Federal Rural do Rio de Janeiro, Laboratório de Doenças Parasitárias, Instituto de Veterinária, Seropédica, RJ , Brasil, Laboratório de Doenças Parasitárias, Instituto de Veterinária, Universidade Federal Rural do Rio de Janeiro, Seropédica, RJ, Brasil.
authors.forename: B A
authors.initials: BA
authors.lastname: Baêta
authors.affiliation: Fundação Oswaldo Cruz, Fundação Oswaldo Cruz, Instituto Oswaldo Cruz, Rio de Janeiro, RJ , Brasil, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, RJ, Brasil.
authors.forename: J S
authors.initials: JS
authors.lastname: Ferreira
authors.affiliation: Fundação Oswaldo Cruz, Fundação Oswaldo Cruz, Instituto Oswaldo Cruz, Rio de Janeiro, RJ , Brasil, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, RJ, Brasil.
authors.forename: R C
authors.initials: RC
authors.lastname: Medeiros
authors.affiliation: Fundação Oswaldo Cruz, Fundação Oswaldo Cruz, Instituto Oswaldo Cruz, Rio de Janeiro, RJ , Brasil, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, RJ, Brasil.
authors.forename: C M
authors.initials: CM
authors.lastname: Maya-Monteiro
authors.affiliation: Fundação Oswaldo Cruz, Fundação Oswaldo Cruz, Instituto Oswaldo Cruz, Rio de Janeiro, RJ , Brasil, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, RJ, Brasil.
authors.forename: F A
authors.initials: FA
authors.lastname: Lara
authors.affiliation: The Pirbright Institute, The Pirbright Institute, The Tick Cell Biobank, Pirbright , UK, The Tick Cell Biobank, The Pirbright Institute, Pirbright, UK.
authors.forename: L
authors.initials: L
authors.lastname: Bell-Sakyi
authors.affiliation: Universidade Federal Rural do Rio de Janeiro, Universidade Federal Rural do Rio de Janeiro, Laboratório de Doenças Parasitárias, Instituto de Veterinária, Seropédica, RJ , Brasil, Laboratório de Doenças Parasitárias, Instituto de Veterinária, Universidade Federal Rural do Rio de Janeiro, Seropédica, RJ, Brasil.
authors.forename: A H
authors.initials: AH
authors.lastname: Fonseca
mesh_terms: D000818:Animals; D025065:Borrelia burgdorferi / Q000302:isolation & purification; D002460:Cell Line; D002478:Cells, Cultured; D003470:Culture Media; D005434:Flow Cytometry / Q000379:methods; D005456:Fluorescent Dyes; D018613:Microscopy, Confocal / Q000379:methods; D009930:Organic Chemicals; D010587:Phagocytosis; D015203:Reproducibility of Results; D013144:Spirochaetales / Q000302:isolation & purification; D013194:Staining and Labeling / Q000379:methods*; D017282:Tick-Borne Diseases / Q000382:microbiology; D013987:Ticks / Q000166:cytology* / Q000382:microbiology*; D013997:Time Factors
chemical_list: D003470:Culture Media; D005456:Fluorescent Dyes; D009930:Organic Chemicals; C070080:PKH 26; C451241:PKH67