Relevant Genes Linked to Virulence Are Required for Salmonella Typhimurium to Survive Intracellularly in the Social Amoeba Dictyostelium discoideum

Riquelme, Sebastián A.; Varas, Macarena; Valenzuela, Camila; Velozo, P.; Chahin, Nicolás; Aguilera, Paulina Morales; Sabag, Andrea; Labra, Bayron; Álvarez, Sergio A.; Chávez, Francisco P.; Santiviago, Carlos A.

doi:10.3389/fmicb.2016.01305

Cited by 36 publications

(55 citation statements)

References 83 publications

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“…Autophagy and related processes have emerged as important innate immune defense mechanisms against intracellular bacterial infections, and several pathogens, including Salmonella, are known to have evolved mechanisms to counteract the host autophagy response [4][5][6][8][9][10][11][12][74][75][76]. Macrophages are the key cell type responsible for dissemination of Salmonella during systemic typhoid disease.…”

Section: Discussionmentioning

confidence: 99%

Macrophages target Salmonella by Lc3-associated phagocytosis in a systemic infection model

et al. 2019

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Innate immune defense against intracellular pathogens, like Salmonella, relies heavily on the autophagy machinery of the host. This response is studied intensively in epithelial cells, the target of Salmonella during gastrointestinal infections. However, little is known of the role that autophagy plays in macrophages, the predominant carriers of this pathogen during systemic disease. Here we utilize a zebrafish embryo model to study the interaction of S. enterica serovar Typhimurium with the macroautophagy/autophagy machinery of macrophages in vivo. We show that phagocytosis of live but not heat-killed Salmonella triggers recruitment of the autophagy marker GFP-Lc3 in a variety of patterns labeling tight or spacious bacteria-containing compartments, also revealed by electron microscopy. Neutrophils display similar GFP-Lc3 associations, but genetic modulation of the neutrophil/macrophage balance and ablation experiments show that macrophages are critical for the defense response. Deficiency of atg5 reduces GFP-Lc3 recruitment and impairs host resistance, in contrast to atg13 deficiency, indicating that Lc3-Salmonella association at this stage is independent of the autophagy preinitiation complex and that macrophages target Salmonella by Lc3associated phagocytosis (LAP). In agreement, GFP-Lc3 recruitment and host resistance are impaired by deficiency of Rubcn/Rubicon, known as a negative regulator of canonical autophagy and an inducer of LAP. We also found strict dependency on NADPH oxidase, another essential factor for LAP. Both Rubcn and NADPH oxidase are required to activate a Salmonella biosensor for reactive oxygen species inside infected macrophages. These results identify LAP as the major host protective autophagy-related pathway responsible for macrophage defense against Salmonella during systemic infection.

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Section: Discussionmentioning

confidence: 99%

Macrophages target Salmonella by Lc3-associated phagocytosis in a systemic infection model

et al. 2019

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“…We anticipated that these mutants would be contained in a vacuolar compartment unable to support the intracellular survival of the pathogen. Therefore, we performed infection assays in D. discoideum under conditions that have been described by our group (27, 28) ( Figure 7 ). We found that the number of internalized bacteria was similar between the different strains ( Figure 7A ), indicating that the deletion of genes sopB and sifA does not affect the uptake of S .…”

Section: Resultsmentioning

confidence: 99%

SopB- and SifA-dependent shaping of theSalmonella-containing vacuole proteome in the social amoebaDictyostelium discoideum

Valenzuela

Gil

Urrutia

et al. 2019

Preprint

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word count: 212 Abstract 22The ability of Salmonella to survive and replicate within mammalian host cells involves the 23 generation of a membranous compartment known as the Salmonella-containing vacuole 24 (SCV). Salmonella employs a number of effector proteins that are injected into host cells for 25 SCV formation using its type-three secretion systems encoded in SPI-1 and SPI-2 (T3SSSPI-1 26 and T3SSSPI-2, respectively). Recently, we reported that S. Typhimurium requires T3SSSPI-1 27 and T3SSSPI-2 to survive in the model amoeba Dictyostelium discoideum. Despite these 28 findings, the involved effector proteins have not been identified yet. Therefore, we evaluated 29 the role of two major S. Typhimurium effectors SopB and SifA during D. discoideum 30 intracellular niche formation. First, we established that S. Typhimurium resides in a vacuolar 31 compartment within D. discoideum. Next, we isolated SCVs from amoebae infected with wild 32 type or the DsopB and DsifA mutant strains of S. Typhimurium, and we characterized the 33 composition of this compartment by quantitative proteomics. This comparative analysis 34 suggests that S. Typhimurium requires SopB and SifA to modify the SCV proteome in order 35 to generate a suitable intracellular niche in D. discoideum. Accordingly, we observed that 36 SopB and SifA are needed for intracellular survival of S. Typhimurium in this organism. Thus, 37 our results provide insight into the mechanisms employed by Salmonella to survive 38 intracellularly in phagocytic amoebae. 39 40 Importance 41The molecular mechanisms involved in Salmonella survival to predation by phagocytic 42 amoebae, such as D. discoideum, remains poorly understood. Although we established that 43 S. Typhimurium requires two specialized type-three secretion systems to survive in D. 44 discoideum, no effector protein has been implicated in this process so far. Here, we 45 confirmed the presence of a membrane-bound compartment containing S. Typhimurium in D. 46 discoideum, and purified the D. discoideum SCV to characterize the associated proteome. In 47 doing so, we established a key role for effector proteins SopB and SifA in remodeling the 48 protein content of the SCV that ultimately allow the intracellular survival of S. Typhimurium in 49 D. discoideum. We also discuss similarities and differences with the proteomes of the human 50 SCV. These findings contribute to unravel the mechanisms used by Salmonella to survive in 51 the environment exploiting phagocytic amoebae as a reservoir.52 55 Bacteria from the Salmonella genus infect warm-blooded animals targeting the 56 gastrointestinal tract. Particular serotypes, such as Salmonella enterica serovar Typhimurium 57 (S. Typhimurium), represent major leading causes of gastroenteritis in humans in developing 58 countries (1). Overall, Salmonella infections account for over 150,000 deaths annually in the 59 world, most of them associated with foodborne infections (1). 60Among the essential genes required for Salmonella virulence, pathogenicity islands SPI-1 61...

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“…To be able to understand this wide range of hosts and the diversity of virulence strategies, there is a need for establishing more models for studying mechanisms required for infecting diverse hosts. is was successfully accomplished using D. discoideum as a host model to study the pathogenesis of Klebsiella, Legionella, Pseudomonas, and Salmonella infections [21][22][23]36].…”

Section: Discussionmentioning

confidence: 99%

“…However, due to limited tools and antigens to study intracellular lifestyle in free-living amoeba, such as A. castellanii and H. vermiformis, D. discoideum has been used as a model organism to study phagocytosis, cell motility, and virulence factors for many bacterial pathogens, such as Pseudomonas, Legionella, Mycobacterium, Salmonella, and Klebsiella [18][19][20][21][22][23]. Dictyostelium has been established as a model organism for studying the life cycle of the fish pathogen, F. noatunensis, but it has not been established for studying the strains of Francisella that cause the disease in humans [24][25][26].…”

Section: Introductionmentioning

confidence: 99%

“…Dictyostelium has been established as a model organism for studying the life cycle of the fish pathogen, F. noatunensis, but it has not been established for studying the strains of Francisella that cause the disease in humans [24][25][26]. It has been shown that Salmonella requires O-antigen and the type VI secretion system (T6SS) for the survival within amoebae [22]. Francisella uses the T6SS to avoid lysosomal fusion within the macrophages [27].…”

Section: Introductionmentioning

confidence: 99%

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Optimisation of External Factors for the Growth of Francisella novicida within Dictyostelium discoideum

Kelava

Marečić

Fućak

et al. 2020

BioMed Research International

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The amoeba Dictyostelium discoideum has been used as a model organism to study host-pathogen interaction in many intracellular bacteria. Francisella tularensis is a Gram-negative, highly infectious bacterium that causes the zoonotic disease tularemia. The bacterium is able to replicate in different phagocytic and nonphagocytic cells including mammalian, amoebae, and arthropod cells. The aim of this study was to determine the optimal temperature and infection dose in the interaction of Francisella novicida with D. discoideum in order to establish a model of Francisella infection in the social amoeba. The amoeba cells were infected with a different multiplicity of infection (5, 10, and 100) and incubated at different temperatures (22, 25, 27, 30, and 37°C). The number of intracellular bacteria within D. discoideum, as well as cytotoxicity, was determined at 2, 4, 24, 48, and 72 hours after infection. Our results showed that the optimal temperature for Francisella intracellular replication within amoeba is 30°C with the MOI of 10. We can conclude that this MOI and temperature induced the optimal growth of bacteria in Dictyostelium with low cytotoxicity.

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Relevant Genes Linked to Virulence Are Required for Salmonella Typhimurium to Survive Intracellularly in the Social Amoeba Dictyostelium discoideum

Cited by 36 publications

References 83 publications

Macrophages target Salmonella by Lc3-associated phagocytosis in a systemic infection model

Macrophages target Salmonella by Lc3-associated phagocytosis in a systemic infection model

SopB- and SifA-dependent shaping of theSalmonella-containing vacuole proteome in the social amoebaDictyostelium discoideum

Optimisation of External Factors for the Growth of Francisella novicida within Dictyostelium discoideum

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