The effect of Legionella pneumophila infection on the encystation of Acanthamoeba castellanii

Acanthamoeba is a genus of free-living amoebae commonly found in soil, water, and other habitats. This organism undergoes 2 distinct stages in its life cycle, the trophozoite and the cyst. Under adverse conditions, trophozoites transform into cysts, which are notably resistant to harsh physical and chemical conditions. Infected by Legionella pneumophila has been shown to decrease the number of cysts in its host Acanthamoeba species, although the mechanisms responsible for this effect remain poorly understood. In this study, A. castellanii was co-cultured with either L. pneumophila or Escherichia coli to assess the impact on encystation and to explore the genes involved in this process. Following a 72-h encystation induction period, it was observed that Acanthamoeba infected with Legionella exhibited a 45.8% reduction in cyst formation compared to the control group. In contrast, Acanthamoeba that phagocytosed E. coli showed a 21.7% decrease. To identify the genes involved in this phenomenon, real-time PCR analysis was conducted on 20 genes known to be upregulated during encystation. This analysis was performed to verify their expression patterns at 24, 48, and 72 h. Notably, ten genes, including cyst-specific protein 21, glycosyltransferase, RSNARE, and cellulose synthase, did not exhibit increased expression in Legionella-infected Acanthamoeba. However, these genes showed elevated expression levels in both the control group and the bacteria-phagocytosed Acanthamoeba. This suggests that several cellular processes, including cell wall formation, are inhibited in Acanthamoeba infected with Legionella, resulting in reduced encystation.