The Dot/Icm Effector SdhA Is Necessary for Virulence of Legionella pneumophila in Galleria mellonella and A/J Mice
Clare R. HardingCharlotte A. StonehamRalf SchueleinHayley J. NewtonClare V. OatesElizabeth L. HartlandGunnar N. SchroederGad Frankel
44
Citation
44
Reference
10
Related Paper
Citation Trend
Abstract:
Legionella pneumophila is an intracellular bacterium that resides within amoebae and macrophages in a specialized compartment termed the Legionella-containing vacuole (LCV). As well as providing an intracellular niche for replication, the LCV helps to prevent the release of bacterial components into the cytoplasm. Recognition of these components as danger signals by the host activates immune responses leading to clearance of the bacterium. Here, we examined the role of two important virulence factors of L. pneumophila, the potent danger signal flagellin and the translocated Dot/Icm type IVB secretion system effector SdhA, which is crucial to maintain LCV integrity, in the Galleria mellonella infection model. We demonstrate that flagellin expression does not contribute to virulence, replication, or induction of clearance mechanisms. Conversely, SdhA expression is important for virulence. We found that in the absence of SdhA, the LCV in hemocytes showed signs of instability and leakage. Furthermore, in contrast to wild-type L. pneumophila, a ΔsdhA mutant caused a transient depletion of hemocytes and reduced mortality. Analysis of the ΔsdhA mutant in the A/J mouse model also showed a significant replication defect. Together, our data underline the crucial importance of SdhA in infection across different model organisms.Keywords:
SDHA
Legionella
Galleria mellonella
Flagellin
Galleria mellonella
Hemocyte
Cite
Citations (4)
Abstract Flagellin is amongst the most abundant proteins in flagellated bacterial species and constitutes the major building block of the flagellar filament. The proteins FliW and FliS serve in the post-transcriptional control of flagellin and guide the protein to the flagellar type III secretion system (fT3SS), respectively. Here, we present the high-resolution structure of FliS/flagellin heterodimer and show that FliS and FliW bind to opposing interfaces located at the N- and C-termini of flagellin. The FliS/flagellin/FliW heterotrimer is able to interact with FlhA-C suggesting that FliW and FliS are released during flagellin export. After release, FliW and FliS are recycled to execute a new round of post-transcriptional regulation and targeting. Taken together, our study provides a mechanism explaining how FliW and FliS synchronize the production of flagellin with the capacity of the fT3SS to secrete flagellin.
Flagellin
Homeostasis
Cite
Citations (20)
Galleria mellonella (greater wax moth or honeycomb moth) has been introduced as an alternative model to study microbial infections. G. mellonella larvae can be easily and inexpensively obtained in large numbers and are simple to use as they don't require special lab equipment. There are no ethical constraints and their short life cycle makes them ideal for large-scale studies. Although insects lack an adaptive immune response, their innate immune response shows remarkable similarities with the immune response in vertebrates.This review gives a current update of what is known about the immune system of G. mellonella and provides an extensive overview of how G. mellonella is used to study the virulence of Gram-positive and Gram-negative bacteria. In addition, the use of G. mellonella to evaluate the efficacy of antimicrobial agents and experimental phage therapy are also discussed. The review concludes with a critical assessment of the current limitatons of G. mellonella infection models.
Galleria mellonella
Phage therapy
Cite
Citations (596)
Legionella pneumophila, the causative agent of a severe pneumonia named Legionnaires' disease, is an important human pathogen that infects and replicates within alveolar macrophages. Its virulence depends on the Dot/Icm type IV secretion system (T4SS), which is essential to establish a replication permissive vacuole known as the Legionella containing vacuole (LCV). L. pneumophila infection can be modeled in mice however most mouse strains are not permissive, leading to the search for novel infection models. We have recently shown that the larvae of the wax moth Galleria mellonella are suitable for investigation of L. pneumophila infection. G. mellonella is increasingly used as an infection model for human pathogens and a good correlation exists between virulence of several bacterial species in the insect and in mammalian models. A key component of the larvae's immune defenses are hemocytes, professional phagocytes, which take up and destroy invaders. L. pneumophila is able to infect, form a LCV and replicate within these cells. Here we demonstrate protocols for analyzing L. pneumophila virulence in the G. mellonella model, including how to grow infectious L. pneumophila, pretreat the larvae with inhibitors, infect the larvae and how to extract infected cells for quantification and immunofluorescence microscopy. We also describe how to quantify bacterial replication and fitness in competition assays. These approaches allow for the rapid screening of mutants to determine factors important in L. pneumophila virulence, describing a new tool to aid our understanding of this complex pathogen.
Galleria mellonella
Legionella
Cite
Citations (105)
The larvae of the greater wax moth, Galleria mellonella, are pests of active beehives. In infection biology, these larvae are playing a more and more attractive role as an invertebrate host model. Here, we report on the first genome sequence of Galleria mellonella.
Galleria mellonella
Cite
Citations (95)
Анализ и антибактериальная активность фракций гемолимфы иммунизированных личинок Galleria mellonella
В работе исследована антибактериальная активность гемолимфы иммунизированных личинок Galleria mellonella. С помощью ионообменной хроматографии и гель-электрофореза из гемолимфы выделены белковые фракции с антибактериальной активностью. Масс-спектрометрический анализ фракций показал присутствие пептида с массой 5627 Да. По-видимому, данный пептид и определяет антибактериальную активность гемолимфы. Полученные данные позволяют обосновать целесообразность применения препаратов, полученных из личинок Galleria mellonella, в медицине, гомеопатии и терапевтической практике.
Galleria mellonella
Cite
Citations (0)
ABSTRACT Nonvertebrate model hosts represent valuable tools for the study of host-pathogen interactions because they facilitate the identification of bacterial virulence factors and allow the discovery of novel components involved in host innate immune responses. In this report, we determined that the greater wax moth caterpillar Galleria mellonella is a convenient nonmammalian model host for study of the role of the type III secretion system (TTSS) in Pseudomonas aeruginosa pathogenesis. Based on the observation that a mutation in the TTSS pscD gene of P. aeruginosa strain PA14 resulted in a highly attenuated virulence phenotype in G. mellonella , we examined the roles of the four known effector proteins of P. aeruginosa (ExoS, ExoT, ExoU, and ExoY) in wax moth killing. We determined that in P. aeruginosa strain PA14, only ExoT and ExoU play a significant role in G. mellonella killing. Strain PA14 lacks the coding sequence for the ExoS effector protein and does not seem to express ExoY. Moreover, using Δ exoU Δ exoY , Δ exoT Δ exoY , and Δ exoT Δ exoU double mutants, we determined that individual translocation of either ExoT or ExoU is sufficient to obtain nearly wild-type levels of G. mellonella killing. On the other hand, data obtained with a Δ exoT Δ exoU Δ exoY triple mutant and a Δ pscD mutant suggested that additional, as-yet-unidentified P. aeruginosa components of type III secretion are involved in virulence in G. mellonella . A high level of correlation between the results obtained in the G. mellonella model and the results of cytopathology assays performed with a mammalian tissue culture system validated the use of G. mellonella for the study of the P. aeruginosa TTSS.
Galleria mellonella
Cite
Citations (252)
With increasing of the knowledge of resource insect, the exploitation and utilization of the greater wax moth (Galleria mellonella L.) has been focused on, rather than its control. In recent years, G. mellonella has been utilized as an experimental insect in some research of organisms. In this paper, we mainly summarized the utilization of the greater wax moth in the researches of entomopathogenic nematodes, parasitic wasps, Cryptococcus neoformans, defense peptides and antibacterial immunity mechanisms.
Galleria mellonella
Cite
Citations (0)
Flagellin
Pseudomonas syringae
Hypersensitive response
Cite
Citations (18)
There is an increasing number of reports concerning identification of Galleria mellonella immune-relevant peptides and proteins. The large wax moth Galleria mellonella, which is used as a model organism in various fields of medicine and biology, belongs to the order of Lepidoptera. Most of these studies are related to proteins and peptides involved in insect immune system. This review article discusses the properties of the most well-known defense proteins and peptides in the immune response of G. mellonella, which are used in infection studies.
Galleria mellonella
Cite
Citations (0)