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The parental environment of C. elegans can have lasting effects on progeny development and immunity. Vitamin B12 exposure in C. elegans has been shown to accelerate development and protect against pathogenic bacteria. Here, we show that parental exposure to dietary vitamin B12 or vitamin B12-producing bacteria results in offspring with accelerated growth that persists for a single generation. During infection with the microsporidian Nematocida parisii, the offspring of worms fed vitamin B12 diets have better reproductive fitness but similar infection levels, suggesting increased tolerance to microsporidian infection. Vitamin B12-induced intergenerational growth acceleration and N. parisii tolerance is dependent upon the methionine biosynthesis pathway. Offspring from vitamin B12-exposed parents are protected from pathogenic Pseudomonas vranovensis and this protection is mediated through methionine biosynthesis and propionyl-CoA breakdown pathways. Our results show how parental microbial diet impacts progeny development through the transfer of vitamin B12 which results in accelerated growth and pathogen tolerance.
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We report the magnetic behavior of the hybrid perovskites [Gua]Mn1-xFe2x/3â¡x/3(HCOO)3 (0 ≤ x ≤ 0.88), showing that vacancy ordering drives bulk ferrimagnetism for x > 0.6. The behavior is rationalized in terms of a simple microscopic model of percolation-induced ferrimagnetism. Monte Carlo simulations driven by this model reproduce the experimental dependence of magnetic susceptibility on x and show that, at intermediate compositions, domains of short-range vacancy order lead to the emergence of local magnetization. Our results open up a new avenue for the design of multiferroic hybrid perovskites.
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Shigella flexneri is a human-adapted pathovar of Escherichia coli that can invade the intestinal epithelium, causing inflammation and bacillary dysentery. Although an important human pathogen, the host response to S. flexneri has not been fully described. Zebrafish larvae represent a valuable model for studying human infections in vivo. Here, we use a Shigella-zebrafish infection model to generate mRNA expression profiles of host response to Shigella infection at the whole-animal level. Immune response-related processes dominate the signature of early Shigella infection (6â h post-infection). Consistent with its clearance from the host, the signature of late Shigella infection (24â h post-infection) is significantly changed, and only a small set of immune-related genes remain differentially expressed, including acod1 and gpr84. Using mutant lines generated by ENU, CRISPR mutagenesis and F0 crispants, we show that acod1- and gpr84-deficient larvae are more susceptible to Shigella infection. Together, these results highlight the power of zebrafish to model infection by bacterial pathogens and reveal the mRNA expression of the early (acutely infected) and late (clearing) host response to Shigella infection.
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Disentería Bacilar , Animales , Humanos , Disentería Bacilar/genética , Shigella flexneri/genética , Shigella flexneri/metabolismo , Pez Cebra/genética , Pez Cebra/microbiología , Inflamación/microbiología , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
Pel exopolysaccharide biosynthetic loci are phylogenetically widespread biofilm matrix determinants in bacteria. In Pseudomonas aeruginosa, Pel is crucial for cell-to-cell interactions and reducing susceptibility to antibiotic and mucolytic treatments. While genes encoding glycoside hydrolases have long been linked to biofilm exopolysaccharide biosynthesis, their physiological role in biofilm development is unclear. Here we demonstrate that the glycoside hydrolase activity of P. aeruginosa PelA decreases adherent biofilm biomass and is responsible for generating the low molecular weight secreted form of the Pel exopolysaccharide. We show that the generation of secreted Pel contributes to the biomechanical properties of the biofilm and decreases the virulence of P. aeruginosa in Caenorhabditis elegans and Drosophila melanogaster. Our results reveal that glycoside hydrolases found in exopolysaccharide biosynthetic systems can help shape the soft matter attributes of a biofilm and propose that secreted matrix components be referred to as matrix associated to better reflect their influence.
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Biopelículas , Glicósido Hidrolasas , Polisacáridos Bacterianos , Pseudomonas aeruginosa , Animales , Fenómenos Biomecánicos , Drosophila melanogaster/microbiología , Glicósido Hidrolasas/genética , Pseudomonas aeruginosa/fisiología , Virulencia , Caenorhabditis elegans/microbiologíaRESUMEN
Argonaute (AGO) proteins associate with small RNAs to direct their effector function on complementary transcripts. The nematode Caenorhabditis elegans contains an expanded family of 19 functional AGO proteins, many of which have not been fully characterized. In this work, we systematically analyzed every C. elegans AGO using CRISPR-Cas9 genome editing to introduce GFP::3xFLAG tags. We have characterized the expression patterns of each AGO throughout development, identified small RNA binding complements, and determined the effects of ago loss on small RNA populations and developmental phenotypes. Our analysis indicates stratification of subsets of AGOs into distinct regulatory modules, and integration of our data led us to uncover novel stress-induced fertility and pathogen response phenotypes due to ago loss.
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Proteínas de Caenorhabditis elegans , Animales , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Interferencia de ARN , Proteínas Argonautas/genética , Proteínas Argonautas/metabolismo , ARN Interferente Pequeño/metabolismo , Redes Reguladoras de GenesRESUMEN
Microsporidia are a large phylum of obligate intracellular parasites that infect an extremely diverse range of animals and protists. In this chapter, we review what is currently known about microsporidia host specificity and what factors influence microsporidia infection. Extensive sampling in nature from related hosts has provided insight into the host range of many microsporidia species. These field studies have been supported by experiments conducted in controlled laboratory environments which have helped to demonstrate host specificity. Together, these approaches have revealed that, while examples of generalist species exist, microsporidia specificity is often narrow, and species typically infect one or several closely related hosts. For microsporidia to successfully infect and complete their life cycle within a compatible host, several steps must occur, including spore germination, host cell invasion, and proliferation of the parasite within the host tissue. Many factors influence infection, including temperature, seasonality, nutrient availability, and the presence or absence of microbes, as well as the developmental stage, sex, and genetics of the host. Several studies have identified host genomic regions that influence resistance to microsporidia, and future work is likely to uncover molecular mechanisms of microsporidia host specificity in more detail.
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Microsporidios , Microsporidiosis , Animales , Especificidad del Huésped/genética , Estadios del Ciclo de Vida/genética , Microsporidios/genética , Microsporidiosis/genéticaRESUMEN
Inherited immunity describes how some animals can pass on the "memory" of a previous infection to their offspring. This can boost pathogen resistance in their progeny and promote survival. While inherited immunity has been reported in many invertebrates, the mechanisms underlying this epigenetic phenomenon are largely unknown. The infection of Caenorhabditis elegans by the natural microsporidian pathogen Nematocida parisii results in the worms producing offspring that are robustly resistant to microsporidia. The present protocol describes the study of intergenerational immunity in the simple and genetically tractable N. parisii -C. elegans infection model. The current article describes methods for infecting C. elegans and generating immune-primed offspring. Methods are also given for assaying resistance to microsporidia infection by staining for microsporidia and visualizing infection by microscopy. In particular, inherited immunity prevents host cell invasion by microsporidia, and fluorescence in situ hybridization (FISH) can be used to quantify invasion events. The relative amount of microsporidia spores produced in the immune-primed offspring can be quantified by staining the spores with a chitin-binding dye. To date, these methods have shed light on the kinetics and pathogen specificity of inherited immunity, as well as the molecular mechanisms underlying it. These techniques, alongside the extensive tools available for C. elegans research, will enable important discoveries in the field of inherited immunity.
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Caenorhabditis elegans , Microsporidiosis , Animales , Caenorhabditis elegans/genética , Hibridación Fluorescente in SituRESUMEN
Despite reports of parental exposure to stress promoting physiological adaptations in progeny in diverse organisms, there remains considerable debate over the significance and evolutionary conservation of such multigenerational effects. Here, we investigate four independent models of intergenerational adaptations to stress in Caenorhabditis elegans - bacterial infection, eukaryotic infection, osmotic stress, and nutrient stress - across multiple species. We found that all four intergenerational physiological adaptations are conserved in at least one other species, that they are stress -specific, and that they have deleterious tradeoffs in mismatched environments. By profiling the effects of parental bacterial infection and osmotic stress exposure on progeny gene expression across species, we established a core set of 587 genes that exhibited a greater than twofold intergenerational change in expression in response to stress in C. elegans and at least one other species, as well as a set of 37 highly conserved genes that exhibited a greater than twofold intergenerational change in expression in all four species tested. Furthermore, we provide evidence suggesting that presumed adaptive and deleterious intergenerational effects are molecularly related at the gene expression level. Lastly, we found that none of the effects we detected of these stresses on C. elegans F1 progeny gene expression persisted transgenerationally three generations after stress exposure. We conclude that intergenerational responses to stress play a substantial and evolutionarily conserved role in regulating animal physiology and that the vast majority of the effects of parental stress on progeny gene expression are reversible and not maintained transgenerationally.
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Adaptación Biológica , Caenorhabditis elegans , Evolución Molecular , Animales , Caenorhabditis elegans/genética , Caenorhabditis elegans/microbiología , Caenorhabditis elegans/parasitología , Caenorhabditis elegans/fisiología , Estado Nutricional , Presión OsmóticaRESUMEN
The Caenorhabditis elegans genome encodes nineteen functional Argonaute proteins that use 22G-RNAs, 26G-RNAs, miRNAs or piRNAs to regulate target transcripts. Only one Argonaute is essential under normal laboratory conditions: CSR-1. While CSR-1 has been studied widely, nearly all studies have overlooked the fact that the csr-1 locus encodes two isoforms. These isoforms differ by an additional 163 amino acids present in the N-terminus of CSR-1a. Using CRISPR-Cas9 genome editing to introduce GFP::3xFLAG into the long (CSR-1a) and short (CSR-1b) isoforms, we found that CSR-1a is expressed during spermatogenesis and in several somatic tissues, including the intestine. CSR-1b is expressed constitutively in the germline. small RNA sequencing of CSR-1 complexes shows that they interact with partly overlapping sets of 22G-RNAs. Phenotypic analyses reveal that the essential functions of csr-1 described in the literature coincide with CSR-1b, while CSR-1a plays tissue specific functions. During spermatogenesis, CSR-1a integrates into an sRNA regulatory network including ALG-3, ALG-4 and WAGO-10 that is necessary for fertility at 25°C. In the intestine, CSR-1a silences immunity and pathogen-responsive genes, and its loss results in improved survival from the pathogen Pseudomonas aeruginosa. Our findings functionally distinguish the CSR-1 isoforms and highlight the importance of studying each AGO isoform independently.
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Proteínas de Caenorhabditis elegans/fisiología , Caenorhabditis elegans/genética , Espermatogénesis/genética , Alelos , Animales , Caenorhabditis elegans/embriología , Caenorhabditis elegans/crecimiento & desarrollo , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Femenino , Fertilidad , Expresión Génica , Masculino , Mutación , Oocitos/metabolismo , Complejo de la Endopetidasa Proteasomal/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Isoformas de Proteínas/fisiología , ARN Pequeño no Traducido/metabolismo , Espermatozoides/metabolismoRESUMEN
Parental infection can result in the production of offspring with enhanced immunity phenotypes. Critically, the mechanisms underlying inherited immunity are poorly understood. Here, we show that Caenorhabditis elegans infected with the intracellular microsporidian parasite N. parisii produce progeny that are resistant to microsporidia infection. We determine the kinetics of the response and show that intergenerational immunity prevents host-cell invasion by Nematocida parisii and enhances survival to the bacterial pathogen Pseudomonas aeruginosa We demonstrate that immunity is induced by the parental transcriptional response to infection, which can be mimicked through maternal somatic depletion of PALS-22 and the retinoblastoma protein ortholog, LIN-35. We find that other biotic and abiotic stresses (viral infection and cadmium exposure) that induce a similar transcriptional response as microsporidia also induce immunity in progeny. Together, our results reveal how a parental transcriptional signal can be induced by distinct stimuli and protect offspring against multiple classes of pathogens.
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Pathogens are abundant and drive evolution of host immunity. Whilst immune memory is classically associated with adaptive immunity, studies in diverse species now show that priming of innate immune defences can also protect against secondary infection. Remarkably, priming may also be passed on to progeny to enhance pathogen resistance and promote survival in future generations. Phenotypic changes that occur independent of DNA sequence underlie both 'within-generation' priming and 'multigenerational' priming. However, the molecular mechanisms responsible for these phenomena are still poorly understood. Caenorhabditis elegans is a simple and genetically tractable model organism that has enabled key advances in immunity and environmental epigenetics. Using both natural and human pathogens, researchers have uncovered numerous examples of innate immune priming in this animal. Viral infection models have provided key evidence for a conserved antiviral RNA silencing mechanism that is inherited in progeny. Bacterial infection models have explored mechanisms of within-generation and multigenerational priming that span chromatin modification and transcriptional changes, small RNA pathways, maternal provisioning and pathogen avoidance strategies. Together, these studies are providing novel insight into the immune reactivity of the genome and have important consequences for our understanding of health and evolution. In this review, we present the current evidence for learned protection against pathogens in C. elegans, discuss the significance and limitations of these findings and highlight important avenues of future investigation.
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Infecciones Bacterianas/inmunología , Caenorhabditis elegans/inmunología , Inmunidad Innata/inmunología , Memoria Inmunológica/inmunología , Virosis/inmunología , Animales , Infecciones Bacterianas/genética , Infecciones Bacterianas/microbiología , Caenorhabditis elegans/microbiología , Caenorhabditis elegans/virología , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/inmunología , Proteínas de Caenorhabditis elegans/metabolismo , Regulación de la Expresión Génica/inmunología , Interacciones Huésped-Parásitos/inmunología , Inmunidad Innata/genética , Memoria Inmunológica/genética , Transducción de Señal/genética , Transducción de Señal/inmunología , Virosis/genética , Virosis/virologíaRESUMEN
In the following pages, we share a collection of photos, drawings, and mixed-media creations, most of them especially made for this JoN issue, manifesting C. elegans researchers' affection for their model organism and the founders of the field. This is a celebration of our community's growth, flourish, spread, and bright future. Descriptions provided by the contributors, edited for space. 1.
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Caenorhabditis elegans , Medicina en las Artes , Animales , Literatura Moderna , Medicina en la Literatura , Microscopía , InvestigadoresRESUMEN
Although primarily conceptualized as a disorder of phonological awareness, developmental dyslexia is often associated with broader problems perceiving and attending to transient or rapidly-moving visual stimuli. However, the extent to which such visual deficits represent the cause or the consequence of dyslexia remains contentious, and very little research has examined the relative contributions of phonological, visual, and other variables to reading performance more broadly. We measured visual sensitivity to global motion (GM) and global form (GF), performance on various language and other cognitive tasks believed to be compromised in dyslexia (phonological awareness, processing speed, and working memory), together with a range of social and demographic variables often omitted in previous research, such as age, gender, non-verbal intelligence, and socio-economic status in an unselected sample (nâ¯=â¯132) of children aged 6-11.5â¯yrs from two different primary schools in Edinburgh, UK. We found that: (i) Mean GM sensitivity (but not GF) was significantly lower in poor readers (medium effect size); (ii) GM sensitivity accounted for only 3% of the variance in reading scores; (iii) GM sensitivity deficits were observed in only 16% of poor readers; (iv) the best predictors of reading performance were phonological awareness, non-verbal intelligence, and socio-economic status, suggesting the importance of controlling for these in future studies of vision and reading. These findings suggest that developmental dyslexia is unlikely to represent a single category of neurodevelopmental disorder underpinned by lower-level deficits in visual motion processing.
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Cognición/fisiología , Dislexia/fisiopatología , Percepción de Movimiento/fisiología , Lectura , Percepción Visual/fisiología , Niño , Estudios Transversales , Femenino , Humanos , Masculino , Memoria a Corto Plazo/fisiología , Factores SocioeconómicosRESUMEN
Emergency granulopoiesis is a hematopoietic program of stem cell-driven neutrophil production used to counteract immune cell exhaustion following infection. Shigella flexneri is a Gram-negative enteroinvasive pathogen controlled by neutrophils. In this study, we use a Shigella-zebrafish (Danio rerio) infection model to investigate emergency granulopoiesis in vivo We show that stem cell-driven neutrophil production occurs in response to Shigella infection and requires macrophage-independent signaling by granulocyte colony-stimulating factor (Gcsf). To test whether emergency granulopoiesis can function beyond homoeostasis to enhance innate immunity, we developed a reinfection assay using zebrafish larvae that have not yet developed an adaptive immune system. Strikingly, larvae primed with a sublethal dose of Shigella are protected against a secondary lethal dose of Shigella in a type III secretion system (T3SS)-dependent manner. Collectively, these results highlight a new role for emergency granulopoiesis in boosting host defense and demonstrate that zebrafish larvae can be a valuable in vivo model to investigate innate immune memory.IMPORTANCEShigella is an important human pathogen of the gut. Emergency granulopoiesis is the enhanced production of neutrophils by hematopoietic stem and progenitor cells (HSPCs) upon infection and is widely considered a homoeostatic mechanism for replacing exhausted leukocytes. In this study, we developed a Shigella-zebrafish infection model to investigate stem cell-driven emergency granulopoiesis. We discovered that zebrafish initiate granulopoiesis in response to Shigella infection, via macrophage-independent signaling of granulocyte colony-stimulating factor (Gcsf). Strikingly, larvae primed with a sublethal dose of Shigella are protected against a secondary lethal dose of Shigella in a type III secretion system (T3SS)-dependent manner. Taken together, we show that zebrafish infection can be used to capture Shigella-mediated stem cell-driven granulopoiesis and provide a new model system to study stem cell biology in vivo Our results also highlight the potential of manipulating stem cell-driven granulopoiesis to boost innate immunity and combat infectious disease.
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Coinfección/inmunología , Modelos Animales de Enfermedad , Disentería Bacilar/microbiología , Leucopoyesis , Neutrófilos/inmunología , Shigella flexneri/fisiología , Animales , Coinfección/microbiología , Coinfección/fisiopatología , Disentería Bacilar/inmunología , Disentería Bacilar/fisiopatología , Femenino , Humanos , Larva/inmunología , Larva/microbiología , Macrófagos/inmunología , Masculino , Neutrófilos/citología , Pez Cebra/inmunología , Pez Cebra/microbiologíaRESUMEN
Polycyclic aromatic hydrocarbons such as benzo[a]pyrene (BaP) can induce cytochrome P450 1A1 (CYP1A1) via a p53-dependent mechanism. The effect of different p53-activating chemotherapeutic drugs on CYP1A1 expression, and the resultant effect on BaP metabolism, was investigated in a panel of isogenic human colorectal HCT116 cells with differing TP53 status. Cells that were TP53(+/+), TP53(+/-) or TP53(-/-) were treated for up to 48â¯h with 60⯵M cisplatin, 50⯵M etoposide or 5⯵M ellipticine, each of which caused high p53 induction at moderate cytotoxicity (60-80% cell viability). We found that etoposide and ellipticine induced CYP1A1 in TP53(+/+) cells but not in TP53(-/-) cells, demonstrating that the mechanism of CYP1A1 induction is p53-dependent; cisplatin had no such effect. Co-incubation experiments with the drugs and 2.5⯵M BaP showed that: (i) etoposide increased CYP1A1 expression in TP53(+/+) cells, and to a lesser extent in TP53(-/-) cells, compared to cells treated with BaP alone; (ii) ellipticine decreased CYP1A1 expression in TP53(+/+) cells in BaP co-incubations; and (iii) cisplatin did not affect BaP-mediated CYP1A1 expression. Further, whereas cisplatin and etoposide had virtually no influence on CYP1A1-catalysed BaP metabolism, ellipticine treatment strongly inhibited BaP bioactivation. Our results indicate that the underlying mechanisms whereby etoposide and ellipticine regulate CYP1A1 expression must be different and may not be linked to p53 activation alone. These results could be relevant for smokers, who are exposed to increased levels of BaP, when prescribing chemotherapeutic drugs. Beside gene-environment interactions, more considerations should be given to potential drug-environment interactions during chemotherapy.
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Benzo(a)pireno/farmacología , Cisplatino/farmacología , Neoplasias Colorrectales/metabolismo , Citocromo P-450 CYP1A1/biosíntesis , Elipticinas/farmacología , Etopósido/farmacología , Proteína p53 Supresora de Tumor/metabolismo , Activación Metabólica , Benzo(a)pireno/farmacocinética , Carcinógenos/farmacocinética , Carcinógenos/farmacología , Supervivencia Celular/efectos de los fármacos , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Citocromo P-450 CYP1A1/metabolismo , Citocromo P-450 CYP3A/biosíntesis , Citocromo P-450 CYP3A/metabolismo , Aductos de ADN/metabolismo , Daño del ADN , Elipticinas/farmacocinética , Inducción Enzimática/efectos de los fármacos , Genes p53 , Células HCT116 , Humanos , Proteína p53 Supresora de Tumor/deficiencia , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Shigella flexneri, a Gram-negative enteroinvasive pathogen, causes inflammatory destruction of the human intestinal epithelium. Infection by S. flexneri has been well-studied in vitro and is a paradigm for bacterial interactions with the host immune system. Recent work has revealed that components of the cytoskeleton have important functions in innate immunity and inflammation control. Septins, highly conserved cytoskeletal proteins, have emerged as key players in innate immunity to bacterial infection, yet septin function in vivo is poorly understood. Here, we use S. flexneri infection of zebrafish (Danio rerio) larvae to study in vivo the role of septins in inflammation and infection control. We found that depletion of Sept15 or Sept7b, zebrafish orthologs of human SEPT7, significantly increased host susceptibility to bacterial infection. Live-cell imaging of Sept15-depleted larvae revealed increasing bacterial burdens and a failure of neutrophils to control infection. Strikingly, Sept15-depleted larvae present significantly increased activity of Caspase-1 and more cell death upon S. flexneri infection. Dampening of the inflammatory response with anakinra, an antagonist of interleukin-1 receptor (IL-1R), counteracts Sept15 deficiency in vivo by protecting zebrafish from hyper-inflammation and S. flexneri infection. These findings highlight a new role for septins in host defence against bacterial infection, and suggest that septin dysfunction may be an underlying factor in cases of hyper-inflammation.
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Disentería Bacilar/inmunología , Inmunidad Innata/inmunología , Septinas/metabolismo , Animales , Modelos Animales de Enfermedad , Disentería Bacilar/microbiología , Interacciones Huésped-Patógeno/inmunología , Humanos , Inflamación/inmunología , Inflamación/microbiología , Mucosa Intestinal/microbiología , Larva/metabolismo , Neutrófilos/metabolismo , Neutrófilos/microbiología , Shigella flexneri , Pez CebraRESUMEN
Bdellovibrio bacteriovorus are predatory bacteria that invade and kill a range of Gram-negative bacterial pathogens in natural environments and in vitro [1, 2]. In this study, we investigated Bdellovibrio as an injected, antibacterial treatment in vivo, using zebrafish (Danio rerio) larvae infected with an antibiotic-resistant strain of the human pathogen Shigella flexneri. When injected alone, Bdellovibrio can persist for more than 24 hr in vivo yet exert no pathogenic effects on zebrafish larvae. Bdellovibrio injection of zebrafish containing a lethal dose of Shigella promotes pathogen killing, leading to increased zebrafish survival. Live-cell imaging of infected zebrafish reveals that Shigella undergo rounding induced by the invasive predation from Bdellovibrio in vivo. Furthermore, Shigella-dependent replication of Bdellovibrio was captured inside the zebrafish larvae, indicating active predation in vivo. Bdellovibrio can be engulfed and ultimately eliminated by host neutrophils and macrophages, yet have a sufficient dwell time to prey on pathogens. Experiments in immune-compromised zebrafish reveal that maximal therapeutic benefits of Bdellovibrio result from the synergy of both bacterial predation and host immunity, but that in vivo predation contributes significantly to the survival outcome. Our results demonstrate that successful antibacterial therapy can be achieved via the host immune system working together with bacterial predation by Bdellovibrio. Such cooperation may be important to consider in the fight against antibiotic-resistant infections in vivo.
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Antibiosis , Bdellovibrio/fisiología , Disentería Bacilar/inmunología , Disentería Bacilar/microbiología , Shigella flexneri/fisiología , Animales , Inmunidad Celular , Inmunidad Innata , Larva/inmunología , Larva/microbiología , Pez CebraRESUMEN
Septins, cytoskeletal proteins with well-characterised roles in cytokinesis, form cage-like structures around cytosolic Shigella flexneri and promote their targeting to autophagosomes. However, the processes underlying septin cage assembly, and whether they influence S. flexneri proliferation, remain to be established. Using single-cell analysis, we show that the septin cages inhibit S. flexneri proliferation. To study mechanisms of septin cage assembly, we used proteomics and found mitochondrial proteins associate with septins in S. flexneri-infected cells. Strikingly, mitochondria associated with S. flexneri promote septin assembly into cages that entrap bacteria for autophagy. We demonstrate that the cytosolic GTPase dynamin-related protein 1 (Drp1) interacts with septins to enhance mitochondrial fission. To avoid autophagy, actin-polymerising Shigella fragment mitochondria to escape from septin caging. Our results demonstrate a role for mitochondria in anti-Shigella autophagy and uncover a fundamental link between septin assembly and mitochondria.
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Autofagia , Mitocondrias/metabolismo , Septinas/metabolismo , Shigella/fisiología , Proteínas de Ciclo Celular/metabolismo , Línea Celular , Proteínas del Citoesqueleto/metabolismo , Humanos , Dinámicas Mitocondriales , Proteínas Mitocondriales/metabolismo , Modelos Biológicos , Unión ProteicaRESUMEN
Nucleotide-signaling pathways are found in all kingdoms of life and are utilized to coordinate a rapid response to external stimuli. The stringent response alarmones guanosine tetra- (ppGpp) and pentaphosphate (pppGpp) control a global response allowing cells to adapt to starvation conditions such as amino acid depletion. One more recently discovered signaling nucleotide is the secondary messenger cyclic diadenosine monophosphate (c-di-AMP). Here, we demonstrate that this signaling nucleotide is essential for the growth of Staphylococcus aureus, and its increased production during late growth phases indicates that c-di-AMP controls processes that are important for the survival of cells in stationary phase. By examining the transcriptional profile of cells with high levels of c-di-AMP, we reveal a significant overlap with a stringent response transcription signature. Examination of the intracellular nucleotide levels under stress conditions provides further evidence that high levels of c-di-AMP lead to an activation of the stringent response through a RelA/SpoT homologue (RSH) enzyme-dependent increase in the (p)ppGpp levels. This activation is shown to be indirect as c-di-AMP does not interact directly with the RSH protein. Our data extend this interconnection further by showing that the S. aureus c-di-AMP phosphodiesterase enzyme GdpP is inhibited in a dose-dependent manner by ppGpp, which itself is not a substrate for this enzyme. Altogether, these findings add a new layer of complexity to our understanding of nucleotide signaling in bacteria as they highlight intricate interconnections between different nucleotide-signaling networks.
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Fosfatos de Dinucleósidos/metabolismo , Guanosina Pentafosfato/metabolismo , Guanosina Tetrafosfato/metabolismo , Transducción de Señal , Staphylococcus aureus/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Western Blotting , División Celular/genética , Perfilación de la Expresión Génica , Regulación Bacteriana de la Expresión Génica , Viabilidad Microbiana/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , Staphylococcus aureus/genética , Staphylococcus aureus/crecimiento & desarrolloRESUMEN
We sought to determine the extent to which red-green, colour-opponent mechanisms in the human visual system play a role in the perception of drifting luminance-modulated targets. Contrast sensitivity for the directional discrimination of drifting luminance-modulated (yellow-black) test sinusoids was measured following adaptation to isoluminant red-green sinusoids drifting in either the same or opposite direction. When the test and adapt stimuli drifted in the same direction, large sensitivity losses were evident at all test temporal frequencies employed (1-16 Hz). The magnitude of the loss was independent of temporal frequency. When adapt and test stimuli drifted in opposing directions, large sensitivity losses were evident at lower temporal frequencies (1-4 Hz) and declined with increasing temporal frequency. Control studies showed that this temporal-frequency-dependent effect could not reflect the activity of achromatic units. Our results provide evidence that chromatic mechanisms contribute to the perception of luminance-modulated motion targets drifting at speeds of up to at least 32 degrees s(-1). We argue that such mechanisms most probably lie within a parvocellular-dominated cortical visual pathway, sensitive to both chromatic and luminance modulation, but only weakly selective for the direction of stimulus motion.