Counting Legionella cells within single amoeba host cells

Abstract

Here we present the first attempt to quantify Legionella pneumophila cell numbers within individual amoeba hosts that may be released into engineered water systems. The maximum numbers of culturable L. pneumophila cells grown within Acanthamoeba polyphaga and Naegleria fowleri were 1,348 (mean, 329) and 385 (mean, 44) CFU trophozoite(-1), respectively.

Fig 1

FSC and SSC profiles of L. pneumophila-infected amoeba trophozoites. Plots are representative of three independent experiments. A. polyphaga-infected cells (a) and N. fowleri-infected cells (b) at 0 and 48 h after legionella infection. Individual amoeba cells were sorted from a single region (indicated in red) for L. pneumophila CFU enumeration based on forward scatter (FSC-A) and side scatter (SSC-A). Images in insets are representative of sorted cells within the gated region and microscopically verified (magnification, ×400) for intracellular infection at 48 h. Infected cells and uninfected cysts/trophozoites are indicated by black and white arrows, respectively. Black bars, 50 μm.