Project description:The rate, timing, and mode of species dispersal is recognized as a key driver of the structure and function of communities of macroorganisms, and may be one ecological process that determines the diversity of microbiomes. Many previous studies have quantified the modes and mechanisms of bacterial motility using monocultures of a few model bacterial species. But most microbes live in multispecies microbial communities, where direct interactions between microbes may inhibit or facilitate dispersal through a number of physical (e.g., hydrodynamic) and biological (e.g., chemotaxis) mechanisms, which remain largely unexplored. Using cheese rinds as a model microbiome, we demonstrate that physical networks created by filamentous fungi can impact the extent of small-scale bacterial dispersal and can shape the composition of microbiomes. From the cheese rind of Saint Nectaire, we serendipitously observed the bacterium Serratia proteamaculans actively spreads on networks formed by the fungus Mucor. By experimentally recreating these pairwise interactions in the lab, we show that Serratia spreads on actively growing and previously established fungal networks. The extent of symbiotic dispersal is dependent on the fungal network: diffuse and fast-growing Mucor networks provide the greatest dispersal facilitation of the Serratia species, while dense and slow-growing Penicillium networks provide limited dispersal facilitation. Fungal-mediated dispersal occurs in closely related Serratia species isolated from other environments, suggesting that this bacterial-fungal interaction is widespread in nature. Both RNA-seq and transposon mutagenesis point to specific molecular mechanisms that play key roles in this bacterial-fungal interaction, including chitin utilization and flagellin biosynthesis. By manipulating the presence and type of fungal networks in multispecies communities, we provide the first evidence that fungal networks shape the composition of bacterial communities, with Mucor networks shifting experimental bacterial communities to complete dominance by motile Proteobacteria. Collectively, our work demonstrates that these strong biophysical interactions between bacterial and fungi can have community-level consequences and may be operating in many other microbiomes.

Project description:The intracellular bacterial pathogen Legionella pneumophila causes an inflammatory pneumonia called Legionnaires’ Disease. For virulence, L. pneumophila requires a Dot/Icm type IV secretion system that translocates bacterial effectors to the host cytosol. L. pneumophila lacking the Dot/Icm system is recognized by Toll-like receptors (TLRs), leading to a canonical NF-κB-dependent transcriptional response. In addition, L. pneumophila expressing a functional Dot/Icm system potently induces unique transcriptional targets, including proinflammatory genes such as Il23a and Csf2. Here we demonstrate that this Dot/Icm-dependent response, which we term the effector-triggered response (ETR), requires five translocated bacterial effectors that inhibit host protein synthesis. Upon infection of macrophages with virulent L. pneumophila, these five effectors caused a global decrease in host translation, thereby preventing synthesis of IκB, an inhibitor of the NF-κB transcription factor. Thus, macrophages infected with wildtype L. pneumophila exhibited prolonged activation of NF-κB, which was associated with transcription of ETR target genes such as Il23a and Csf2. L. pneumophila mutants lacking the five effectors still activated TLRs and NF-κB, but because the mutants permitted normal IκB synthesis, NF-κB activation was more transient and was not sufficient to fully induce the ETR. L. pneumophila mutants expressing enzymatically inactive effectors were also unable to fully induce the ETR, whereas multiple compounds or bacterial toxins that inhibit host protein synthesis via distinct mechanisms recapitulated the ETR when administered with TLR ligands. Previous studies have demonstrated that the host response to bacterial infection is induced primarily by specific microbial molecules that activate TLRs or cytosolic pattern recognition receptors. Our results add to this model by providing a striking illustration of how the host immune response to a virulent pathogen can also be shaped by pathogen-encoded activities, such as inhibition of host protein synthesis. Three-condition experiment: macrophages left uninfected (negative control), or infected with wildtype Legionella pneumophila, or the mutant Δ5, which lacks five bacterial effectors involved in inhibition of host protein synthesis (lgt1, lgt2, lgt3, sidI, sidL) (two experimental conditions). Biological replicates: two, independently infected, harvested, and hybridized to arrays. One technical replicate per array.

Project description:The intracellular bacterial pathogen Legionella pneumophila causes an inflammatory pneumonia called Legionnaires’ Disease. For virulence, L. pneumophila requires a Dot/Icm type IV secretion system that translocates bacterial effectors to the host cytosol. L. pneumophila lacking the Dot/Icm system is recognized by Toll-like receptors (TLRs), leading to a canonical NF-κB-dependent transcriptional response. In addition, L. pneumophila expressing a functional Dot/Icm system potently induces unique transcriptional targets, including proinflammatory genes such as Il23a and Csf2. Here we demonstrate that this Dot/Icm-dependent response, which we term the effector-triggered response (ETR), requires five translocated bacterial effectors that inhibit host protein synthesis. Upon infection of macrophages with virulent L. pneumophila, these five effectors caused a global decrease in host translation, thereby preventing synthesis of IκB, an inhibitor of the NF-κB transcription factor. Thus, macrophages infected with wildtype L. pneumophila exhibited prolonged activation of NF-κB, which was associated with transcription of ETR target genes such as Il23a and Csf2. L. pneumophila mutants lacking the five effectors still activated TLRs and NF-κB, but because the mutants permitted normal IκB synthesis, NF-κB activation was more transient and was not sufficient to fully induce the ETR. L. pneumophila mutants expressing enzymatically inactive effectors were also unable to fully induce the ETR, whereas multiple compounds or bacterial toxins that inhibit host protein synthesis via distinct mechanisms recapitulated the ETR when administered with TLR ligands. Previous studies have demonstrated that the host response to bacterial infection is induced primarily by specific microbial molecules that activate TLRs or cytosolic pattern recognition receptors. Our results add to this model by providing a striking illustration of how the host immune response to a virulent pathogen can also be shaped by pathogen-encoded activities, such as inhibition of host protein synthesis. Four-condition experiment: macrophages left uninfected (negative control), or infected with wildtype Legionella pneumophila, the flagellin-deficient mutant ΔflaA, or the secretion-deficient mutant ΔdotA (three experimental conditions). Biological replicates: two, independently infected, harvested, and hybridized to arrays. One to two technical replicates per array, as indicated in file titles.

Project description:The intracellular bacterial pathogen Legionella pneumophila causes an inflammatory pneumonia called Legionnaires’ Disease. For virulence, L. pneumophila requires a Dot/Icm type IV secretion system that translocates bacterial effectors to the host cytosol. L. pneumophila lacking the Dot/Icm system is recognized by Toll-like receptors (TLRs), leading to a canonical NF-κB-dependent transcriptional response. In addition, L. pneumophila expressing a functional Dot/Icm system potently induces unique transcriptional targets, including proinflammatory genes such as Il23a and Csf2. Here we demonstrate that this Dot/Icm-dependent response, which we term the effector-triggered response (ETR), requires five translocated bacterial effectors that inhibit host protein synthesis. Upon infection of macrophages with virulent L. pneumophila, these five effectors caused a global decrease in host translation, thereby preventing synthesis of IκB, an inhibitor of the NF-κB transcription factor. Thus, macrophages infected with wildtype L. pneumophila exhibited prolonged activation of NF-κB, which was associated with transcription of ETR target genes such as Il23a and Csf2. L. pneumophila mutants lacking the five effectors still activated TLRs and NF-κB, but because the mutants permitted normal IκB synthesis, NF-κB activation was more transient and was not sufficient to fully induce the ETR. L. pneumophila mutants expressing enzymatically inactive effectors were also unable to fully induce the ETR, whereas multiple compounds or bacterial toxins that inhibit host protein synthesis via distinct mechanisms recapitulated the ETR when administered with TLR ligands. Previous studies have demonstrated that the host response to bacterial infection is induced primarily by specific microbial molecules that activate TLRs or cytosolic pattern recognition receptors. Our results add to this model by providing a striking illustration of how the host immune response to a virulent pathogen can also be shaped by pathogen-encoded activities, such as inhibition of host protein synthesis.

Dataset Information

Antimicrobial peptides of the apidaecin family inhibit the bacterial ribosome by multimodal mechanisms involving three different sites on the large ribosomal subunit

Publications

Multimodal binding and inhibition of bacterial ribosomes by the antimicrobial peptides Api137 and Api88.

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