GSE125876
Symbiont-mediated defense against Legionella pneumophila in amoebae
Legionella pneumophila are important opportunistic pathogens for which environmental reservoirs such as protists are crucial for the infection of humans. Free-living amoebae are considered key hosts providing nutrients and shelter for highly efficient intracellular proliferation of L. pneumophila, which eventually leads to lysis of the amoeba host cell. Yet, the significance of other bacterial players for L. pneumophila ecology is poorly understood. In this study we used a ubiquitous amoeba and their bacterial endosymbiont to investigate the impact of this common association on L. pneumophila infection. We demonstrate that Acanthamoeba castellanii harboring the chlamydial symbiont Protochlamydia amoebophila were able to erase L. pneumophila and, in contrast to symbiont-free amoebae, survived the infection and were able to resume growth. Environmental amoeba isolates harboring P. amoebophila were equally well-protected, and fresh environmental isolates of L. pneumophila were equally well-erased, suggesting ecological relevance of this symbiont-mediated protection. We further show that protection was not mediated by impaired L. pneumophila uptake. Instead, we observed reduced virulence of L. pneumophila released from symbiont-containing amoebae that is strongly supported by transcriptome data. Interference with transition to the transmissive phase is thus likely the basis for this protection. Finally, our data indicate that the defensive response of amoebae harboring P. amoebophila leaves the amoebae with superior fitness reminiscent of immunological memory. Given that mutualistic associations between bacteria and amoebae are widely distributed, P. amoebophila and potentially other amoeba endosymbionts could be key elements in shaping environmental survival, abundance and virulence of this important pathogen thereby affecting frequency of human infection. Overall design: RNA-Seq of A. castellanii Neff amoebae infected with L. pneumophila Paris either in the presence of the amoeba endosymbiont P. amoebophila UWE25, or without the endosymbiont; in either case, 3 time points during L. pneumophila were sequenced: 24 hours post L. pneumophila infection (hpi), 96 hpi, and bacteria released at 96 hpi (extracellular bacteria); all in biological triplicates; in addition, amoebae only containing the endosymbionts at 24 hpi (biological duplicates) and 96 hpi (biological duplicates) were sequenced; in all conditions, total RNA (i.e. simulanteously from all organisms present) was extracted, DNase-treated, rRNA-depleted, libraries prepared, and sequenced on an Illumina HiSeq 2500;