RESUMO
To maintain a symbiotic relationship between the host and its resident intestinal microbiota, appropriate mucosal T cell responses to commensal antigens must be established. Mice acquire both IgG and IgA maternally; the former has primarily been implicated in passive immunity to pathogens while the latter mediates host-commensal mutualism. Here, we report the surprising observation that mice generate T cell-independent and largely Toll-like receptor (TLR)-dependent IgG2b and IgG3 antibody responses against their gut microbiota. We demonstrate that maternal acquisition of these antibodies dampens mucosal T follicular helper responses and subsequent germinal center B cell responses following birth. This work reveals a feedback loop whereby T cell-independent, TLR-dependent antibodies limit mucosal adaptive immune responses to newly acquired commensal antigens and uncovers a broader function for maternal IgG.
Assuntos
Animais Recém-Nascidos/imunologia , Microbioma Gastrointestinal , Imunidade nas Mucosas , Imunoglobulina A/imunologia , Imunoglobulina G/imunologia , Leite Humano/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Animais , Animais Recém-Nascidos/microbiologia , Linfócitos B/imunologia , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Transdução de Sinais , Organismos Livres de Patógenos Específicos , Receptores Toll-Like/imunologiaRESUMO
The animal foregut is the first tissue to encounter ingested food, bacteria, and viruses. We characterized the adult Drosophila foregut using transcriptomics to better understand how it triages consumed items for digestion or immune response and manages resources. Cell types were assigned and validated using GFP-tagged and Gal4 reporter lines. Foregut-associated neuroendocrine cells play a major integrative role by coordinating gut activity with nutrition, the microbiome, and circadian cycles; some express clock genes. Multiple epithelial cell types comprise the proventriculus, the central foregut organ that secretes the peritrophic matrix (PM) lining the gut. Analyzing cell types synthesizing individual PM layers revealed abundant mucin production close to enterocytes, similar to the mammalian intestinal mucosa. The esophagus and salivary gland express secreted proteins likely to line the esophageal surface, some of which may generate a foregut commensal niche housing specific gut microbiome species. Overall, our results imply that the foregut coordinates dietary sensing, hormonal regulation, and immunity in a manner that has been conserved during animal evolution.
Assuntos
Líquidos Corporais , Drosophila , Animais , Células Epiteliais , Contagem de Células , Estado Nutricional , MamíferosRESUMO
A longstanding goal of biology is to identify the key genes and species that critically impact evolution, ecology, and health. Network analysis has revealed keystone species that regulate ecosystems and master regulators that regulate cellular genetic networks. Yet these studies have focused on pairwise biological interactions, which can be affected by the context of genetic background and other species present, generating higher-order interactions. The important regulators of higher-order interactions are unstudied. To address this, we applied a high-dimensional geometry approach that quantifies epistasis in a fitness landscape to ask how individual genes and species influence the interactions in the rest of the biological network. We then generated and also reanalyzed 5-dimensional datasets (two genetic, two microbiome). We identified key genes (e.g., the rbs locus and pykF) and species (e.g., Lactobacilli) that control the interactions of many other genes and species. These higher-order master regulators can induce or suppress evolutionary and ecological diversification by controlling the topography of the fitness landscape. Thus, we provide a method and mathematical justification for exploration of biological networks in higher dimensions.
Assuntos
Microbiota , Microbiota/genética , Epistasia Genética , Evolução BiológicaRESUMO
Observational studies reveal substantial variability in microbiome composition across individuals. Targeted studies in gnotobiotic animals underscore this variability by showing that some bacterial strains colonize deterministically, while others colonize stochastically. While some of this variability can be explained by external factors like environmental, dietary, and genetic differences between individuals, in this paper we show that for the model organism Drosophila melanogaster, interactions between bacteria can affect the microbiome assembly process, contributing to a baseline level of microbiome variability even among isogenic organisms that are identically reared, housed, and fed. In germ-free flies fed known combinations of bacterial species, we find that some species colonize more frequently than others even when fed at the same high concentration. We develop an ecological technique that infers the presence of interactions between bacterial species based on their colonization odds in different contexts, requiring only presence/absence data from two-species experiments. We use a progressive sequence of probabilistic models, in which the colonization of each bacterial species is treated as an independent stochastic process, to reproduce the empirical distributions of colonization outcomes across experiments. We find that incorporating context-dependent interactions substantially improves the performance of the models. Stochastic, context-dependent microbiome assembly underlies clinical therapies like fecal microbiota transplantation and probiotic administration and should inform the design of synthetic fecal transplants and dosing regimes.
Assuntos
Bactérias/classificação , Drosophila melanogaster/microbiologia , Microbiota , Animais , Fenômenos Fisiológicos Bacterianos/genética , Modelos Biológicos , Especificidade da Espécie , Processos EstocásticosRESUMO
Cell- and tissue-level processes often occur across days or weeks, but few imaging methods can capture such long timescales. Here, we describe Bellymount, a simple, noninvasive method for longitudinal imaging of the Drosophila abdomen at subcellular resolution. Bellymounted animals remain live and intact, so the same individual can be imaged serially to yield vivid time series of multiday processes. This feature opens the door to longitudinal studies of Drosophila internal organs in their native context. Exploiting Bellymount's capabilities, we track intestinal stem cell lineages and gut microbial colonization in single animals, revealing spatiotemporal dynamics undetectable by previously available methods.
Assuntos
Anatomia Transversal/métodos , Drosophila/anatomia & histologia , Microbioma Gastrointestinal , Microscopia Intravital/métodos , Vísceras/anatomia & histologia , Fatores Etários , Animais , Drosophila/microbiologia , Intestinos/anatomia & histologia , Intestinos/diagnóstico por imagem , Imagem Óptica/métodos , Vísceras/diagnóstico por imagemRESUMO
Gut bacteria can affect key aspects of host fitness, such as development, fecundity, and lifespan, while the host, in turn, shapes the gut microbiome. However, it is unclear to what extent individual species versus community interactions within the microbiome are linked to host fitness. Here, we combinatorially dissect the natural microbiome of Drosophila melanogaster and reveal that interactions between bacteria shape host fitness through life history tradeoffs. Empirically, we made germ-free flies colonized with each possible combination of the five core species of fly gut bacteria. We measured the resulting bacterial community abundances and fly fitness traits, including development, reproduction, and lifespan. The fly gut promoted bacterial diversity, which, in turn, accelerated development, reproduction, and aging: Flies that reproduced more died sooner. From these measurements, we calculated the impact of bacterial interactions on fly fitness by adapting the mathematics of genetic epistasis to the microbiome. Development and fecundity converged with higher diversity, suggesting minimal dependence on interactions. However, host lifespan and microbiome abundances were highly dependent on interactions between bacterial species. Higher-order interactions (involving three, four, and five species) occurred in 13-44% of possible cases depending on the trait, with the same interactions affecting multiple traits, a reflection of the life history tradeoff. Overall, we found these interactions were frequently context-dependent and often had the same magnitude as individual species themselves, indicating that the interactions can be as important as the individual species in gut microbiomes.
Assuntos
Microbioma Gastrointestinal/fisiologia , Trato Gastrointestinal/microbiologia , Interações entre Hospedeiro e Microrganismos/fisiologia , Interações Microbianas/fisiologia , Microbiota/fisiologia , Animais , Bactérias/isolamento & purificação , Biodiversidade , Drosophila melanogaster , Epistasia Genética , Fertilidade , Microbioma Gastrointestinal/genética , Vida Livre de Germes , Interações entre Hospedeiro e Microrganismos/genética , Longevidade , Interações Microbianas/genética , Microbiota/genética , Fenótipo , ReproduçãoRESUMO
The concept of genetic epistasis defines an interaction between two genetic loci as the degree of non-additivity in their phenotypes. A fitness landscape describes the phenotypes over many genetic loci, and the shape of this landscape can be used to predict evolutionary trajectories. Epistasis in a fitness landscape makes prediction of evolutionary trajectories more complex because the interactions between loci can produce local fitness peaks or troughs, which changes the likelihood of different paths. While various mathematical frameworks have been proposed to investigate properties of fitness landscapes, Beerenwinkel et al. (Stat Sin 17(4):1317-1342, 2007a) suggested studying regular subdivisions of convex polytopes. In this sense, each locus provides one dimension, so that the genotypes form a cube with the number of dimensions equal to the number of genetic loci considered. The fitness landscape is a height function on the coordinates of the cube. Here, we propose cluster partitions and cluster filtrations of fitness landscapes as a new mathematical tool, which provides a concise combinatorial way of processing metric information from epistatic interactions. Furthermore, we extend the calculation of genetic interactions to consider interactions between microbial taxa in the gut microbiome of Drosophila fruit flies. We demonstrate similarities with and differences to the previous approach. As one outcome we locate interesting epistatic information on the fitness landscape where the previous approach is less conclusive.
Assuntos
Bactérias/genética , Evolução Biológica , Drosophila/genética , Drosophila/microbiologia , Epistasia Genética , Aptidão Genética , Microbiota , Animais , Bactérias/classificação , Genótipo , Modelos Genéticos , Mutação , FenótipoAssuntos
Drosophila , Microbiota , Animais , Dieta , Drosophila melanogaster , Trato GastrointestinalRESUMO
Cilia and flagella are microtubule-based organelles that protrude from the cell body. Ciliary assembly requires intraflagellar transport (IFT), a motile system that delivers cargo from the cell body to the flagellar tip for assembly. The process controlling injections of IFT proteins into the flagellar compartment is, therefore, crucial to ciliogenesis. Extensive biochemical and genetic analyses have determined the molecular machinery of IFT, but these studies do not explain what regulates IFT injection rate. Here, we provide evidence that IFT injections result from avalanche-like releases of accumulated IFT material at the flagellar base and that the key regulated feature of length control is the recruitment of IFT material to the flagellar base. We used total internal reflection fluorescence microscopy of IFT proteins in live cells to quantify the size and frequency of injections over time. The injection dynamics reveal a power-law tailed distribution of injection event sizes and a negative correlation between injection size and frequency, as well as rich behaviors such as quasiperiodicity, bursting, and long-memory effects tied to the size of the localized load of IFT material awaiting injection at the flagellar base, collectively indicating that IFT injection dynamics result from avalanche-like behavior. Computational models based on avalanching recapitulate observed IFT dynamics, and we further show that the flagellar Ras-related nuclear protein (Ran) guanosine 5'-triphosphate (GTP) gradient can in theory act as a flagellar length sensor to regulate this localized accumulation of IFT. These results demonstrate that a self-organizing, physical mechanism can control a biochemically complex intracellular transport pathway.
Assuntos
Chlamydomonas reinhardtii/fisiologia , Cílios/fisiologia , Transporte Biológico Ativo , Chlamydomonas reinhardtii/genética , Flagelos/fisiologia , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Cinesinas/genética , Cinesinas/metabolismo , Microscopia de Fluorescência , Microscopia de Vídeo , Modelos Biológicos , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismoRESUMO
Cells control organelle size with great precision and accuracy to maintain optimal physiology, but the mechanisms by which they do so are largely unknown. Cilia and flagella are simple organelles in which a single measurement, length, can represent size. Maintenance of flagellar length requires an active transport process known as intraflagellar transport, and previous measurements suggest that a length-dependent feedback regulates intraflagellar transport. But the question remains: how is a length-dependent signal produced to regulate intraflagellar transport appropriately? Several conceptual models have been suggested, but testing these models quantitatively requires that they be cast in mathematical form. Here, we derive a set of mathematical models that represent the main broad classes of hypothetical size-control mechanisms currently under consideration. We use these models to predict the relation between length and intraflagellar transport, and then compare the predicted relations for each model with experimental data. We find that three models-an initial bolus formation model, an ion current model, and a diffusion-based model-show particularly good agreement with available experimental data. The initial bolus and ion current models give mathematically equivalent predictions for length control, but fluorescence recovery after photobleaching experiments rule out the initial bolus model, suggesting that either the ion current model or a diffusion-based model is more likely correct. The general biophysical principles of the ion current and diffusion-based models presented here to measure cilia and flagellar length can be generalized to measure any membrane-bound organelle volume, such as the nucleus and endoplasmic reticulum.
Assuntos
Chlamydomonas/fisiologia , Cílios , Modelos Biológicos , Cílios/fisiologia , Difusão , Flagelos/fisiologia , Recuperação de Fluorescência Após Fotodegradação , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Íons/metabolismo , Microscopia de Fluorescência , Movimento , Tamanho das Organelas , Especificidade da EspécieRESUMO
Gut bacteria are prevalent throughout the Metazoa and form complex microbial communities associated with food breakdown, nutrient provision and disease prevention. How hosts acquire and maintain a consistent bacterial flora remains mysterious even in the best-studied animals, including humans, mice, fishes, squid, bugs, worms and flies. This essay visits the evidence that hosts have co-evolved relationships with specific bacteria and that some of these relationships are supported by specialized physical niches that select, sequester and maintain microbial symbionts. Genetics approaches could uncover the mechanisms for recruiting and maintaining the stable and consistent members of the microbiome. This article is part of the theme issue 'Sculpting the microbiome: how host factors determine and respond to microbial colonization'.
Assuntos
Microbioma Gastrointestinal , Microbiota , Humanos , Animais , Camundongos , Bactérias , Peixes/microbiologiaRESUMO
The optimisation of synthetic and natural microbial communities has vast potential for emerging applications in medicine, agriculture and industry. Realising this goal is contingent on a close correlation between theory, experiments, and the real world. Although the temporal pattern of resource supply can play a major role in microbial community assembly, resource dynamics are commonly treated inconsistently in theoretical and experimental research. Here we explore how the composition of communities varies under continuous resource supply, typical of theoretical approaches, versus pulsed resource supply, typical of experiments. Using simulations of classical resource competition models, we show that community composition diverges rapidly between the two regimes, with almost zero overlap in composition once the pulsing interval stretches beyond just four hours. The implication for the rapidly growing field of microbial community optimisation is that the resource supply regime must be tailored to the community being optimised. As such, we argue that resource supply dynamics should be considered both a constraint in the design of novel microbial communities and as a tuning mechanism for the optimisation of pre-existing communities like those found in the human gut.
Assuntos
Ecossistema , Microbiota , Modelos Biológicos , HumanosRESUMO
Encryption makes information available only to those with the decoding key. We propose that microbes, living in a chemical environment, encrypt nutrients, thereby making them available only to those with the decoding enzymes, such as their kin. Examples of encrypted nutrients include cobamides, which are expensive to make and valuable for microbial fitness. Furthermore, we propose that hosts encrypt nutrients to encourage desirable colonizers. For instance, plant root exudates and breast milk oligosaccharides encourage beneficial microbes.
Assuntos
Cobamidas , Sideróforos , Humanos , Nutrientes , PolissacarídeosRESUMO
The intestines of animals are colonized by commensal microbes, which impact host development, health, and behavior. Precise quantification of colonization is essential for studying the complex interactions between host and microbe both to validate the microbial composition and study its effects. Drosophila melanogaster, which has a low native microbial diversity and is economical to rear with defined microbial composition, has emerged as a model organism for studying the gut microbiome. Analyzing the microbiome of an individual organism requires identification of which microbial species are present and quantification of their absolute abundance. This article presents a method for the analysis of a large number of individual fly microbiomes. The flies are prepared in 96-well plates, enabling the handling of a large number of samples at once. Microbial abundance is quantified by plating up to 96 whole fly homogenates on a single agar plate in an array of spots and then counting the colony forming units (CFUs) that grow in each spot. This plating system is paired with an automated CFU quantification platform, which incorporates photography of the plates, differentiation of fluorescent colonies, and automated counting of the colonies using an ImageJ plugin. Advantages are that (i) this method is sensitive enough to detect differences between treatments, (ii) the spot plating method is as accurate as traditional plating methods, and (iii) the automated counting process is accurate and faster than manual counting. The workflow presented here enables high-throughput quantification of CFUs in a large number of replicates and can be applied to other microbiology study systems including in vitro and other small animal models.
Assuntos
Drosophila , Microbioma Gastrointestinal , Animais , Contagem de Colônia Microbiana , Drosophila melanogaster , Células-TroncoRESUMO
The intestine is responsible for efficient absorption and packaging of dietary lipids before they enter the circulatory system. This review provides a comprehensive overview of how intestinal enterocytes from diverse model organisms absorb dietary lipid and subsequently secrete the largest class of lipoproteins (chylomicrons) to meet the unique needs of each animal. We discuss the putative relationship between diet and metabolic disease progression, specifically Type 2 Diabetes Mellitus. Understanding the molecular response of intestinal cells to dietary lipid has the potential to undercover novel therapies to combat metabolic syndrome.
Assuntos
Diabetes Mellitus Tipo 2 , Metabolismo dos Lipídeos , Animais , Humanos , Metabolismo dos Lipídeos/fisiologia , Absorção Intestinal , Intestinos , Gorduras na Dieta/metabolismoRESUMO
In exponential population growth, variability in the timing of individual division events and environmental factors (including stochastic inoculation) compound to produce variable growth trajectories. In several stochastic models of exponential growth we show power-law relationships that relate variability in the time required to reach a threshold population size to growth rate and inoculum size. Population-growth experiments in E. coli and S. aureus with inoculum sizes ranging between 1 and 100 are consistent with these relationships. We quantify how noise accumulates over time, finding that it encodes-and can be used to deduce-information about the early growth rate of a population.
Assuntos
Escherichia coli , Staphylococcus aureus , Modelos Biológicos , Processos Estocásticos , Densidade DemográficaRESUMO
How, when, and why organisms age are fascinating issues that can only be fully addressed by adopting an evolutionary perspective. Consistently, the main evolutionary theories of ageing, namely the Mutation Accumulation theory, the Antagonistic Pleiotropy theory, and the Disposable Soma theory, have formulated stimulating hypotheses that structure current debates on both the proximal and ultimate causes of organismal ageing. However, all these theories leave a common area of biology relatively under-explored. The Mutation Accumulation theory and the Antagonistic Pleiotropy theory were developed under the traditional framework of population genetics, and therefore are logically centred on the ageing of individuals within a population. The Disposable Soma theory, based on principles of optimising physiology, mainly explains ageing within a species. Consequently, current leading evolutionary theories of ageing do not explicitly model the countless interspecific and ecological interactions, such as symbioses and host-microbiomes associations, increasingly recognized to shape organismal evolution across the Web of Life. Moreover, the development of network modelling supporting a deeper understanding on the molecular interactions associated with ageing within and between organisms is also bringing forward new questions regarding how and why molecular pathways associated with ageing evolved. Here, we take an evolutionary perspective to examine the effects of organismal interactions on ageing across different levels of biological organisation, and consider the impact of surrounding and nested systems on organismal ageing. We also apply this perspective to suggest open issues with potential to expand the standard evolutionary theories of ageing.
Assuntos
Envelhecimento , Evolução Biológica , Humanos , Envelhecimento/genéticaRESUMO
Non-mammalian model organisms have been essential for our understanding of the mechanisms that control development, disease, and physiology, but they are underutilized in pharmacological and toxicological phenotypic screening assays due to their low throughput in comparison with cell-based screens. To increase the utility of using Drosophila melanogaster in screening, we designed the Whole Animal Feeding FLat (WAFFL), a novel, flexible, and complete system for feeding, monitoring, and assaying flies in a high-throughput format. Our 3D printed system is compatible with inexpensive and readily available, commercial 96-well plate consumables and equipment. Experimenters can change the diet at will during the experiment and video record for behavior analysis, enabling precise dosing, measurement of feeding, and analysis of behavior in a 96-well plate format.
Assuntos
Ração Animal , Drosophila melanogaster , Animais , Drosophila melanogaster/fisiologia , Ensaios de Triagem em Larga EscalaRESUMO
Lactobacilli and Acetobacter sp. are commercially important bacteria that often form communities in natural fermentations, including food preparations, spoilage, and in the digestive tract of the fruit fly Drosophila melanogaster. Communities of these bacteria are widespread and prolific, despite numerous strain-specific auxotrophies, suggesting they have evolved nutrient interdependencies that regulate their growth. The use of a chemically-defined medium (CDM) supporting the growth of both groups of bacteria would facilitate the identification of the molecular mechanisms for the metabolic interactions between them. While numerous CDMs have been developed that support specific strains of lactobacilli or Acetobacter, there has not been a medium formulated to support both genera. We developed such a medium, based on a previous CDM designed for growth of lactobacilli, by modifying the nutrient abundances to improve growth yield. We further simplified the medium by substituting casamino acids in place of individual amino acids and the standard Wolfe's vitamins and mineral stocks in place of individual vitamins and minerals, resulting in a reduction from 40 to 8 stock solutions. These stock solutions can be used to prepare several CDM formulations that support robust growth of numerous lactobacilli and Acetobacters. Here, we provide the composition and several examples of its use, which is important for tractability in dissecting the genetic and metabolic basis of natural bacterial species interactions.