Regulation, Integrase-Dependent Excision, and Horizontal Transfer of Genomic Islands in Legionella pneumophila

Abstract

Legionella pneumophila is a Gram-negative freshwater agent which multiplies in specialized nutrient-rich vacuoles of amoebae. When replicating in human alveolar macrophages, Legionella can cause Legionnaires' disease. Recently, we identified a new type of conjugation/type IVA secretion system (T4ASS) in L. pneumophila Corby (named trb-tra). Analogous versions of trb-tra are localized on the genomic islands Trb-1 and Trb-2. Both can exist as an episomal circular form, and Trb-1 can be transferred horizontally to other Legionella strains by conjugation. In our current work, we discovered the importance of a site-specific integrase (Int-1, lpc2818) for the excision and conjugation process of Trb-1. Furthermore, we identified the genes lvrRABC (lpc2813 to lpc2816) to be involved in the regulation of Trb-1 excision. In addition, we demonstrated for the first time that a Legionella genomic island (LGI) of L. pneumophila Corby (LpcGI-2) encodes a functional type IV secretion system. The island can be transferred horizontally by conjugation and is integrated site specifically into the genome of the transconjugants. LpcGI-2 generates three different episomal forms. The predominant episomal form, form A, is generated integrase dependently (Lpc1833) and transferred by conjugation in a pilT-dependent manner. Therefore, the genomic islands Trb-1 and LpcGI-2 should be classified as integrative and conjugative elements (ICEs). Coculture studies of L. pneumophila wild-type and mutant strains revealed that the int-1 and lvrRABC genes (located on Trb-1) as well as lpc1833 and pilT (located on LpcGI-2) do not influence the in vivo fitness of L. pneumophila in Acanthamoeba castellanii.

Growth-related Metabolism of the Carbon Storage Poly-3-hydroxybutyrate in Legionella pneumophila

“…Intracellular Replication (Infection) Assay in A. castellanii and U937 Cells-The intracellular multiplication assays were carried out at a growth temperature of 37°C as described earlier (19,39).…”

csrT Represents a New Class of csrA -Like Regulatory Genes Associated with Integrative Conjugative Elements of Legionella pneumophila

“…Previously, we identified distinct families of csrA-like genes based on their amino acid sequence similarity (22). Likewise, Legionella ICEs encode several different lineages of T4SSs (4,26,(41)(42)(43). Therefore, we investigated whether particular lineages of ICE-encoded T4SSs were genetically linked to a distinct csrA paralog.…”

“…The ICE Trb-1 mobile element of L. pneumophila strain Corby also encodes a csrA paralog, lvrC (26). Mobility of ICE Trb-1 is repressed in laboratory conditions, as deletion of the lvrRABC regulatory region significantly increases the number of episomal ICE Trb-1 copies (26).…”

Legionella oakridgensis ATCC 33761 genome sequence and phenotypic characterization reveals its replication capacity in amoebae

“…oakridgensis proliferated both inside membrane structures (endoplasmic reticulum) and in the cytoplasm (Takekawa et al, 2012 showed no increased fitness in co-culture with U937 cells (Lautner et al, 2013). Yet we can only hypothesize, if the plasmids, the identified genomic islands or genes (e.g.…”

“…pneumophila Corby (lpc2190 to lpc2314, Lautner et al, 2013). The island seems to be integrated into the tRNA Met gene and we yet identified three integrases, several cation efflux systems (helABC, copA, cepABC, and ctpA), the putative T4SS encoding region and the regulatory operon lvrRABC.…”