Influence of Acanthamoeba castellanii on Intracellular Growth of Different Legionella Species in Human Monocytes

Abstract

[2449][2450][2451][2452][2453][2454][2455][2456] 1996). In this study, we introduce an in vitro coculture model of legionellae, Mono Mac 6 cells (MM6) and Acanthamoeba castellanii, using a cell culture chamber system which separates both cell types by a microporous polycarbonate membrane impervious to bacteria, amoebae, and human cells. Whereas L. pneumophila has shown a maximal 4-log-unit multiplication within MM6, which could not be further increased by coculture with Acanthamoeba castellanii, significantly enhanced replication of L. gormanii, L. micdadei, L. steigerwaltii, L. longbeachae, and L. dumoffii was seen after coculture with amoebae. This effect was seen only with uninfected amoebae, not with Legionella-infected amoebae. The supporting effect for intracellular multiplication in MM6 could be reproduced in part by addition of a cell-free coculture supernatant obtained from a coincubation experiment with uninfected A. castellanii and Legionella-infected MM6, suggesting that amoeba-derived effector molecules are involved in this phenomenon. This coculture model allows investigations of molecular and biochemical mechanisms which are responsible for the enhancement of intracellular multiplication of legionellae in monocytic cells after interaction with amoebae.In 1980, Rowbotham published the first report on intracellular multiplication of Legionella pneumophila within Acanthamoeba spp. and Naegleria spp. (33). Thereafter, several reports described the replication of Legionella culture isolates from clinical samples within protozoa isolated from the presumed source of infection (2, 6, 21-23, 29, 34, 36, 45). Intracellular growth within protozoa enhances the ability of L. pneumophila to infect human monocytes (18), induces phenotypic modulation (1, 4), and causes resistance to chemical disinfectants, biocides, and antibiotics (3, 5).Inhalation of legionellae packaged in amoebae results in the induction of more-severe clinical cases of legionellosis (31,33). This speculation was supported by a recently published mouse model of coinhalation of L. pneumophila and Hartmannella vermiformis. Coinhalation with H. vermiformis significantly enhanced the intrapulmonary growth of L. pneumophila, resulting in greater mortality than that from inhalation of legionellae alone (7). Intrapulmonary growth of mutant strains of L. pneumophila with reduced virulence for H. vermiformis but maintained virulence for monocytes was not significantly enhanced by coinhalation (8). Intrapulmonary growth of L. pneumophila was significantly greater in mice inoculated with L. pneumophila-infected H. vermiformis than in mice inoculated with an equivalent number of bacteria or coinoculated with L. pneumophila and uninfected H. vermiformis (9). The mechanism of intrapulmonary growth enhancement of legionellae by amoebae remains to be determined. We therefore established an in vitro coculture model of the Mono Mac 6 cell line (MM6), Acanthamoeba castellanii, and Legionella species with different intracellular growth rates within MM6 (30) in ...

Current and Emerging Legionella Diagnostics for Laboratory and Outbreak Investigations

“…However, it soon became clear that the unique physiology of legionellae was primarily adapted for survival and replication within numerous protozoan genera, including Acanthamoeba, Naegleria, Hartmannella, and Tetrahymena (24,53,54), and secondarily as a freeliving or biofilm-associated aquatic bacterium. Their association with amoebae, in the vegetative or cyst form, may induce virulent bacterial phenotypes, assist in distribution, and provide protection from harsh or bactericidal environmental conditions, such as excessive heat and chlorine (55)(56)(57)(58)(59)(60)(61)(62)(63)(64)(65).…”

Temperature-Regulated Formation of Mycelial Mat-Like Biofilms by Legionella pneumophila

“…Second, L. pneumophila may be inherently more virulent than other species. However, only a few aspects of its pathogenicity e.g., cytopathogenicity and intracellular multiplication, have currently been shown to be enhanced, compared to non-L. pneumophila species (1,23,30,42), providing no strong support for this hypothesis. Third, L. pneumophila may be easier to transmit than other Legionella species.…”

Novel Lysophospholipase A Secreted by Legionella pneumophila

“…For example, since cell swelling may be essential for bacterial multiplication in host cells and since glycerophosphorylcholine, a reaction product of LPLA, causes cell swelling, LPLA is suspected to contribute to Legionella intracellular growth (11,19,22). Furthermore, LPLA might protect L. pneumophila from toxic lysophosphatidylcholine that is generated by action of PLA on surfactant (12,30).…”