RESUMO
The planarian species Schmidtea mediterranea is a flatworm living in freshwater that is used in the research laboratory as a model to study developmental and regeneration mechanisms, as well as antibacterial mechanisms. However, the cultivable microbial repertoire of the microbes comprising its microbiota remains unknown. Here, we characterized the bacterial constituents of a 10-year-old laboratory culture of planarian species S. mediterranea via culturomics analysis. We isolated 40 cultivable bacterial species, including 1 unidentifiable species. The predominant phylum is Proteobacteria, and the most common genus is Pseudomonas. We discovered that parts of the bacterial flora of the planarian S. mediterranea can be classified as fish pathogens and opportunistic human pathogens.
Assuntos
Bactérias/classificação , Bactérias/crescimento & desenvolvimento , Laboratórios/estatística & dados numéricos , Microbiota , Planárias/microbiologia , Animais , Bactérias/genética , Bactérias/isolamento & purificação , Técnicas de Cultura , RNA Ribossômico 16S/genética , Regeneração , Manejo de EspécimesRESUMO
The powerful regenerative ability of planarians has long been a concern of scientists, but recently, their efficient immune system has attracted more and more attention from researchers. Gamma-interferon-inducible lysosomal thiol reductase (GILT) is related not only to antigen presentation but also to bacteria invasions. But the systematic studies are not yet to be conducted on the relationship between bacterial infection. Our study reveals for the first time that GILT of planarian (DjGILT) plays an essential role in the clearance of Gram-negative bacteria by conducting H2O2 concentration in planarians. In animals that DjGILT was silenced, it persisted for up to 9 days before all bacteria were cleared, compared with 6 days of the control group. When infected with E. coli and V. anguillarum, the level of H2O2 was significantly increased in DjGILT-silenced planarians, and concomitantly, mRNA level of C-type lectin DjCTL, which modulates agglutination and clearance efficiency of invading bacteria, was decreased. Further study showed that the decrease of H2O2 level led to a significant increase in DjCTL transcripts. Collectively, we proposed a mechanism model for the involvement of GILT gene in bacterial elimination. We have for the first time revealed the specific mechanism of GILT in innate immune response against bacterial infection.
Assuntos
Bactérias Gram-Negativas/imunologia , Proteínas de Helminto/imunologia , Interferon gama/farmacologia , Lisossomos/efeitos dos fármacos , Oxirredutases atuantes sobre Doadores de Grupo Enxofre/imunologia , Planárias/imunologia , Sequência de Aminoácidos , Animais , Escherichia coli/imunologia , Escherichia coli/fisiologia , Expressão Gênica/efeitos dos fármacos , Expressão Gênica/imunologia , Bactérias Gram-Negativas/fisiologia , Proteínas de Helminto/classificação , Proteínas de Helminto/genética , Interações Hospedeiro-Patógeno/imunologia , Peróxido de Hidrogênio/imunologia , Peróxido de Hidrogênio/metabolismo , Imunidade Inata/genética , Imunidade Inata/imunologia , Lisossomos/enzimologia , Oxirredutases atuantes sobre Doadores de Grupo Enxofre/genética , Oxirredutases atuantes sobre Doadores de Grupo Enxofre/metabolismo , Filogenia , Planárias/genética , Planárias/microbiologia , Homologia de Sequência de Aminoácidos , Compostos de Sulfidrila/metabolismo , Vibrio/imunologia , Vibrio/fisiologiaRESUMO
Silver nanoparticles (AgNPs) are widely incorporated in household, consumer and medical products. Their unintentional release via wastewaters raises concerns on their environmental impact, particularly for aquatic organisms and their associated bacterial communities. It is known that the microbiome plays an important role in its host's health and physiology, e.g. by producing essential nutrients and providing protection against pathogens. A thorough understanding of the effects of AgNPs on bacterial communities and on their interactions with the host is crucial to fully assess AgNP toxicity on aquatic organisms. Our results indicate that the microbiome of the invertebrate Schmidtea mediterranea, a freshwater planarian, is affected by AgNP exposure at the tested 10 µg/ml concentration. Using targeted amplification of the bacterial 16S rRNA gene V3-V4 region, two independent experiments on the microbiomes of adult worms revealed a consistent decrease in Betaproteobacteriales after AgNP exposure, mainly attributed to a decrease in Curvibacter and Undibacterium. Although developing tissues and organisms are known to be more sensitive to toxic compounds, three independent experiments in regenerating worms showed a less pronounced effect of AgNP exposure on the microbiome, possibly because underlying bacterial community changes during development mask the AgNP induced effect. The presence of a polyvinyl-pyrrolidone (PVP) coating did not significantly alter the outcome of the experiments compared to those with uncoated particles. The observed variation between the different experiments underlines the highly variable nature of microbiomes and emphasises the need to repeat microbiome experiments, within and between physiological states of the animal.
Assuntos
Organismos Aquáticos/efeitos dos fármacos , Betaproteobacteria/efeitos dos fármacos , Nanopartículas Metálicas/toxicidade , Microbiota/efeitos dos fármacos , Planárias/efeitos dos fármacos , Prata/toxicidade , Poluentes Químicos da Água/toxicidade , Animais , Organismos Aquáticos/crescimento & desenvolvimento , Organismos Aquáticos/microbiologia , Betaproteobacteria/genética , Betaproteobacteria/crescimento & desenvolvimento , Nanopartículas Metálicas/química , Microbiota/genética , Planárias/crescimento & desenvolvimento , Planárias/microbiologia , Povidona/química , RNA Ribossômico 16S/genética , Prata/química , Poluentes Químicos da Água/químicaRESUMO
Some animals, such as planaria, can regenerate complex anatomical structures in a process regulated by genetic and biophysical factors, but additional external inputs into regeneration remain to be uncovered. Microbial communities inhabiting metazoan organisms are important for metabolic, immune, and disease processes, but their instructive influence over host structures remains largely unexplored. Here, we show that Aquitalea sp. FJL05, an endogenous commensal bacterium of Dugesia japonica planarians, and one of the small molecules it produces, indole, can influence axial and head patterning during regeneration, leading to regeneration of permanently two-headed animals. Testing the impact of indole on planaria tissues via RNA sequencing, we find that indole alters the regenerative outcomes in planarians through changes in expression to patterning genes, including a downregulation of Wnt pathway genes. These data provide a unique example of the product of a commensal bacterium modulating transcription of patterning genes to affect the host's anatomical structure during regeneration.
Assuntos
Acetobacteraceae/metabolismo , Indóis/metabolismo , Planárias/crescimento & desenvolvimento , Regeneração/genética , Acetobacteraceae/genética , Animais , Cabeça/crescimento & desenvolvimento , Cabeça/microbiologia , Microbiota/genética , Planárias/metabolismo , Planárias/microbiologia , Via de Sinalização Wnt/genéticaRESUMO
Pedobacter schmidteae sp. nov. strain EGT (Collection de Souches de l'Unité des Rickettsie CSUR P6417 = Colección Española de Cultivos Tipo CECT 9771) is a new Pedobacter species isolated from the planarian Schmidtea mediterranea. Schmidtea mediterranea are flatworms living in freshwater and exhibiting an unusual ability to regenerate amputated parts. To date, the gut microbiota of Schmidtea mediterranea remains poorly studied. Here, via the culturomics strategy that consists in using diversified culture conditions, we isolated a new bacterium, strain EG, that we characterized using the taxono-genomics approach that combines phenotypic assays and genome sequencing and analysis. Strain EG exhibits a 16S rRNA sequence similarity of 98.29% with Pedobacter nyackensis strain NWG-II14T, its closest neighbour with standing in nomenclature. It is an aerobic bacterium belonging to the family Sphingobacteriaceae. Colonies are small, round, smooth and transparent. Bacterial cells are Gram-negative, rod-shaped, motile and non-spore-forming bacilli with positive catalase and oxidase activities. The genome sequence is 6,198,518 bp-long with a G + C content of 41.13%, and the Ortho-ANI and dDDH values when compared to P. nyackensis are 77.34% and 21.50%, respectively. Strain EGT exhibits unique characteristics that classify it as the type strain of new bacterial species for which we propose the name Pedobacter schmidteae sp. nov.
Assuntos
Genoma Bacteriano , Pedobacter/patogenicidade , Planárias/microbiologia , Animais , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Catalase/genética , Catalase/metabolismo , Microbiota , Oxirredutases/genética , Oxirredutases/metabolismo , Pedobacter/genética , Pedobacter/metabolismo , RNA Ribossômico 16S/genética , Homologia de SequênciaRESUMO
Candida albicans is one of the most common fungal pathogens of humans. Currently, there are limitations in the evaluation of C. albicans infection in existing animal models, especially in terms of understanding the influence of specific infectious stages of the fungal pathogen on the host. We show that C. albicans infects, grows and invades tissues in the planarian flatworm Schmidtea mediterranea, and that the planarian responds to infection by activating components of the host innate immune system to clear and repair host tissues. We study different stages of C. albicans infection and demonstrate that planarian stem cells increase division in response to fungal infection, a process that is likely evolutionarily conserved in metazoans. Our results implicate MORN2 and TAK1/p38 signaling pathways as possible mediators of the host innate immune response to fungal infection. We propose the use of planarians as a model system to investigate host-pathogen interactions during fungal infections.
Assuntos
Candida albicans/imunologia , Candidíase/imunologia , Interações Hospedeiro-Patógeno/imunologia , Imunidade Inata/imunologia , Planárias/imunologia , Animais , Candida albicans/crescimento & desenvolvimento , Candidíase/microbiologia , Modelos Animais de Doenças , Proteínas Nucleares/metabolismo , Membro 2 do Grupo C da Subfamília 2 de Receptores Nucleares/metabolismo , Planárias/microbiologia , Células-Tronco/citologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
Studies of tissue regeneration and host-pathogen interactions using the model planarian Schmidtea mediterranea have been performed at an experimental temperature of 19 °C. S. mediterranea planarians exposed to 19 °C-32 °C were observed for survival, mobility, feeding and regeneration for three months and elimination of the Staphylococcus aureus pathogen over six days. S. mediterranea planarians died at 30 °C-32 °C after 18 days of observation but tolerated temperatures of 19 °C up to 28 °C with non-significant differences in mobility and feeding behavior. Genetic malleability tested by RNAi feeding was still efficient at 26 °C and 28 °C. Concerning the immune capacity of planarians, we reported an exacerbation of the immune response in worms infected by S. aureus at 26 °C and 28 °C. These observations suggest a temperature modulation of planarian stem cells and illustrate the importance of modulating experimental temperature when using planarians as model organisms to study regeneration and immune response.
Assuntos
Planárias/fisiologia , Regeneração , Aclimatação , Animais , Comportamento Alimentar , Resposta ao Choque Térmico , Imunidade , Planárias/genética , Planárias/imunologia , Planárias/microbiologia , Interferência de RNA , Infecções Estafilocócicas/imunologia , Infecções Estafilocócicas/veterinária , Staphylococcus aureus/imunologia , TemperaturaRESUMO
In recent decades Saccharomyces cerevisiae has proven to be one of the most valuable model organisms of aging research. Pathways such as autophagy or the effect of substances like resveratrol and spermidine that prolong the replicative as well as chronological lifespan of cells were described for the first time in S. cerevisiae. In this study we describe the establishment of an aging reporter that allows a reliable and relative quick screening of substances and genes that have an impact on the replicative lifespan. A cDNA library of the flatworm Dugesia tigrina that can be immortalized by beheading was screened using this aging reporter. Of all the flatworm genes, only one could be identified that significantly increased the replicative lifespan of S.cerevisiae. This gene is the cysteine protease cathepsin L that was sequenced for the first time in this study. We were able to show that this protease has the capability to degrade such proteins as the yeast Sup35 protein or the human α-synuclein protein in yeast cells that are both capable of forming cytosolic toxic aggregates. The degradation of these proteins by cathepsin L prevents the formation of these unfolded protein aggregates and this seems to be responsible for the increase in replicative lifespan.
Assuntos
Catepsina L/metabolismo , Planárias/microbiologia , Saccharomyces cerevisiae/genética , Animais , Catepsina L/genética , DNA Complementar , DNA Fúngico , Regulação Fúngica da Expressão Gênica , Hydra , Longevidade , Saccharomyces cerevisiae/metabolismoRESUMO
Little is known about how organisms exposed to recurrent infections adapt their innate immune responses. Here, we report that planarians display a form of instructed immunity to primo-infection by Staphylococcus aureus that consists of a transient state of heightened resistance to re-infection that persists for approximately 30days after primo-infection. We established the involvement of stem cell-like neoblasts in this instructed immunity using the complementary approaches of RNA-interference-mediated cell depletion and tissue grafting-mediated gain of function. Mechanistically, primo-infection leads to expression of the peptidoglycan receptor Smed-PGRP-2, which in turn promotes Smed-setd8-1 histone methyltransferase expression and increases levels of lysine methylation in neoblasts. Depletion of neoblasts did not affect S. aureus clearance in primo-infection but, in re-infection, abrogated the heightened elimination of bacteria and reduced Smed-PGRP-2 and Smed-setd8-1 expression. Smed-PGRP-2 and Smed-setd8-1 sensitize animals to heightened expression of Smed-p38 MAPK and Smed-morn2, which are downstream components of anti-bacterial responses. Our study reveals a central role of neoblasts in innate immunity against S. aureus to establish a resistance state facilitating Smed-sted8-1-dependent expression of anti-bacterial genes during re-infection.
Assuntos
Proteínas de Transporte/metabolismo , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Planárias/microbiologia , Planárias/fisiologia , Proteínas Metiltransferases/metabolismo , Transdução de Sinais , Staphylococcus aureus/fisiologia , Animais , Proteínas de Transporte/genética , Resistência à Doença/genética , Resistência à Doença/imunologia , Expressão Gênica , Regulação da Expressão Gênica , Proteínas Metiltransferases/genética , Células-Tronco/metabolismo , Células-Tronco/microbiologiaRESUMO
Planarians, which are non-parasitic flatworms, are highly resistant to bacterial infections. To better understand the mechanisms underlying this resistance, we investigated the role of the circadian machinery in the anti-bacterial response of the freshwater planarian Schmidtea mediterranea. We identified Smed-Tim from S. mediterranea as a homolog of the mammalian clock gene Tim. We showed via RNA interference that Smed-Tim is required for the anti-microbial activities of Schmidtea mediterranea against Staphylococcus aureus infection during the light/dark cycle. Indeed, S. aureus infection leads to the expression of Smed-Tim, which in turn promotes Smed-Traf6 and Smed-morn2, but not Smed-p38 MAPK expression, 2 master regulators of planarian anti-microbial responses.
Assuntos
Proteínas de Helminto/imunologia , Proteínas Circadianas Period/imunologia , Planárias/imunologia , Staphylococcus aureus/fisiologia , Animais , Proteínas de Helminto/genética , Luz , Proteínas Circadianas Period/genética , Planárias/genética , Planárias/microbiologia , Planárias/efeitos da radiaçãoRESUMO
Dugesia japonica planarian flatworms are naturally exposed to various microbes but typically survive this challenge. We show that planarians eliminate bacteria pathogenic to Homo sapiens, Caenorhabditis elegans, and/or Drosophila melanogaster and thus represent a model to identify innate resistance mechanisms. Whole-transcriptome analysis coupled with RNAi screening of worms infected with Staphylococcus aureus or Legionella pneumophila identified 18 resistance genes with nine human orthologs, of which we examined the function of MORN2. Human MORN2 facilitates phagocytosis-mediated restriction of Mycobacterium tuberculosis, L. pneumophila, and S. aureus in macrophages. MORN2 promotes the recruitment of LC3, an autophagy protein also involved in phagocytosis, to M. tuberculosis-containing phagosomes and subsequent maturation to degradative phagolysosomes. MORN2-driven trafficking of M. tuberculosis to single-membrane, LC3-positive compartments requires autophagy-related proteins Atg5 and Beclin-1, but not Ulk-1 and Atg13, highlighting the importance of MORN2 in LC3-associated phagocytosis. These findings underscore the value of studying planarian defenses to identify immune factors.
Assuntos
Proteínas de Helminto/imunologia , Legionella pneumophila/fisiologia , Proteínas Associadas aos Microtúbulos/imunologia , Fagocitose , Planárias/imunologia , Planárias/microbiologia , Staphylococcus aureus/fisiologia , Animais , Infecções Bacterianas/genética , Infecções Bacterianas/imunologia , Infecções Bacterianas/microbiologia , Modelos Animais de Doenças , Proteínas de Helminto/genética , Humanos , Legionella pneumophila/imunologia , Proteínas Associadas aos Microtúbulos/genética , Planárias/genética , Staphylococcus aureus/imunologiaRESUMO
Planarians famously can regenerate after decapitation. In this issue, Abnave et al. (2014) find they resist infection by multiple bacterial species pathogenic to humans, Drosophila and C. elegans, including M. tuberculosis. These results identify a conserved gene controlling phagocytosis and establish planarians as a powerful system for analyzing host-pathogen interactions.
Assuntos
Proteínas de Helminto/imunologia , Legionella pneumophila/fisiologia , Proteínas Associadas aos Microtúbulos/imunologia , Fagocitose , Planárias/imunologia , Planárias/microbiologia , Staphylococcus aureus/fisiologia , Animais , HumanosRESUMO
GRP78 (78 kDa glucose-regulated protein) has ubiquitously existed in nearly all organisms from yeast to humans, reflecting the central roles it plays in cell survival. In this report, we isolated and sequenced the full-length cDNA of GRP78 (designated DjGRP78) from the planarian Dugesia japonica. The cDNA is 2121 bp, including an open reading frame (ORF) of 1983 bp encoding a polypeptide of 660 amino acids with three HSP70 family signatures. DjGRP78 contains signal peptides at the N-terminus and a KTEL peptide motif at the C-terminus, which suggests that it localizes in the endoplasmic reticulum (ER). Fluorescent real time RT-PCR was employed to detect the expression pattern of Djgrp78 in response to different stressors. Our results show that heat shock and heavy metals (Hg(2+) and Pb(2+)) induce Djgrp78 expression, but starvation does not. Interestingly, we found that Djgrp78 was up-regulated in planarians with septic tissues, and also verified that it was up-regulated in response to bacterial challenge. Our data indicate that Djgrp78 may be a multifunctional gene, and play important roles in physiological and pathological stress in planarians.
Assuntos
Proteínas de Choque Térmico/metabolismo , Planárias/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Retículo Endoplasmático/metabolismo , Chaperona BiP do Retículo Endoplasmático , Escherichia coli/fisiologia , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/imunologia , Imunidade Inata , Chumbo/metabolismo , Mercúrio/metabolismo , Dados de Sequência Molecular , Filogenia , Planárias/imunologia , Planárias/microbiologia , Estresse FisiológicoRESUMO
Trypsin-like serine proteases are involved in large number of processes, especially in digestive degradation and immune responses. Here, we identify the characterization of a trypsin-like serine protease in planarian, Djtry, which interestingly has the incompletely conserved catalytic triad (K, D, and S). Phylogenetic analysis suggests that Djtry is an ancient type of trypsin-like serine proteases. The spatial and temporal expression patterns of Djtry are shown during regenerating and embryonic development by whole-mount in situ hybridization. Djtry is found to display a tissue specific expression pattern, with a predominant expression detected in whole gut region of intact and regenerating planarian. While the tissue- and stage-specific expression patterns during the embryonic development imply the roles of Djtry involve in yolk degradation and gut formation. Quantitative real-time PCR was carried out to analyze the function of this protease in vivo after planarians were stimulated to a bacterial challenge and food. The results showed that Djtry increased after a bacterial challenge and was basically stable for food. Therefore, the trypsin-like serine protease might be involved in the innate defense reactions against bacterial infection.