Cultivation of Ehrlichia chaffeensis in mouse embryo, Vero, BGM, and L929 cells and study of Ehrlichia-induced cytopathic effect and plaque formation

Abstract

We successfully propagated Ehrlichia chaffeensis in mouse embryo, Vero, BGM, and L929 cells inoculated with host cell-free ehrlichiae, indicating that E. chaffeensis is capable of entry, survival, and growth in a relatively wide range of cell types derived from different species. We demonstrated rapid adaptation of E. chaffeensis in these cell lines, so that typical morulae could be detected as early as 5 days after inoculation. E. chaffeensis-induced cytopathic effect with different morphological characteristics in mouse embryo, Vero, and L929 cells. The earliest cytopathic effect appeared in untreated and irradiated mouse embryo cells at 4 days postinoculation. As the infected foci gradually expanded, the center of the foci showed necrotic cells with pyknotic nuclei and degraded morulae. E. chaffeensis caused cell lysis in untreated and irradiated L929 cells, with formation of distinct, round macroscopic plaques at 18 days postinoculation. In untreated and irradiated Vero cells, E. chaffeensis produced infected foci composed of loosely interwoven necrotic cells, spaces of detached cells, cells filled with morulae, and uninfected cells, resulting in characteristic reticular foci. Irradiated cells generally contained many large morulae and presented larger cytopathic foci. DH82 and BGM cells did not develop obvious cytopathic foci under the conditions employed. The findings reported herein offer the opportunity to study the pathogenic mechanism of cell injury by E. chaffeensis, the basis for quantification of infectious E. chaffeensis, improved approaches for recovery of ehrlichiae from human patients and tick hosts, and additional methods for cultivation of E. chaffeensis for molecular analysis.