The Type II Protein Secretion System of Legionella pneumophila Promotes Growth at Low Temperatures

Abstract

The gram-negative bacterium Legionella pneumophila grows in both natural and man-made water systems and in the mammalian lung as a facultative intracellular parasite. The PilD prepilin peptidase of L. pneumophila promotes type IV pilus biogenesis and type II protein secretion. Whereas pili enhance adherence, Legionella type II secretion is critical for intracellular growth and virulence. Previously, we observed that pilD transcript levels are greater in legionellae grown at 30 versus 37°C. Using a new pilD::lacZ fusion strain, we now show that pilD transcriptional initiation increases progressively as L. pneumophila is grown at 30, 25, and 17°C. Legionella pilD mutants also had a dramatically reduced ability to grow in broth and to form colonies on agar at the lower temperatures. Whereas strains specifically lacking type IV pili were not defective for lowtemperature growth, mutations in type II secretion (lsp) genes greatly impaired the capacity of L. pneumophila to form colonies at 25, 17, and 12°C. Indeed, the lsp mutants were completely unable to grow at 12°C. The growth defect of the pilD and lsp mutants was complemented by reintroduction of the corresponding intact gene. Interestingly, the lsp mutants displayed improved growth at 25°C when plated next to a streak of wild-type but not mutant bacteria, implying that a secreted, diffusible factor promotes low-temperature growth. Mutants lacking either the known secreted acid phosphatases, lipases, phospholipase C, lysophospholipase A, or protease grew normally at 25°C, suggesting the existence of a critical, yet-to-be-defined exoprotein(s). In summary, these data document, for the first time, that L. pneumophila replicates at temperatures below 20°C and that a bacterial type II protein secretion system facilitates growth at low temperatures.

Transcriptional profiling of Legionella pneumophila biofilm cells and the influence of iron on biofilm formation

“…pillar-and mushroom-like structures) formed at 25 u C remained more stably attached than filamentous, mycelial-mat-like biofilms formed at 37 and 42 u C. Also, Mampel et al (2006) reported that biofilm formation at 37 u C was 30 % lower than at 23 and 30 u C. Furthermore, we have shown that adhesion is highest for late stationary phase cells. This is expected, as L. pneumophila shows increased expression of the flagellum and of type IV pili, both of which have been described in other bacteria to be involved in adhesion (O'Toole & Kolter, 1998;Pratt & Kolter, 1998;Watnick & Kolter, 1999), in stationary phase during low-temperature growth (Heuner et al, 1999;Soderberg et al, 2004). Because stationary phase mimics the transmissive phase in which bacteria are released from amoebae (Molofsky & Swanson, 2004), our observations suggest that bacteria that exit host cells in natural lowtemperature environments are in an optimal physiological state to adhere to biofilms.…”

lbtAandlbtBAre Required for Production of theLegionella pneumophilaSiderophore Legiobactin

“…Since L. pneumophila survives and replicates at temperatures ranging from 4 to 63°C (33,42,73) and since siderophore expression is elevated at lower temperatures in some other aquatic bacteria (14), strain 130b was grown in deferrated CDM at 25°C and 37°C, and supernatants were tested in the CAS assay (Fig. 2).…”

Compromised Outer Membrane Integrity in Vibrio cholerae Type II Secretion Mutants

“…The outer membrane alterations might be responsible for the slowgrowth phenotype noted for T2S mutants of V. cholerae (39,64,77). Although no outer membrane changes have been reported for T2S mutants of Legionella pneumophila, they also display reduced growth rates under certain growth conditions (82).…”