RESUMO
Microbes interact with the world around them at the chemical level. However, directly examining the chemical exchange between microbes and microbes and their environment, at ecological scales, i.e., the scale of a single bacterial cell or small groups of cells, remains a key challenge. Here we address this obstacle by presenting a methodology that enables matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) of bacterial microcolonies. By combining optimized sample preparation with subatmospheric pressure MALDI, we demonstrate that chemical output from groups of as few as â¼50 cells can be visualized with MALDI-IMS. Application of this methodology to Bacillus subtilis and Streptomyces coelicolor revealed heterogeneity in chemical output across microcolonies and asymmetrical metabolite production when cells grew within physiological gradients produced by Medicago sativa roots. Taken together, these results indicate that MALDI-IMS can readily visualize metabolites made by very small assemblages of bacterial cells and that even these small groups of cells can differentially produce metabolites in response to local chemical gradients.
Assuntos
Bacillus subtilis/metabolismo , Metaboloma/fisiologia , Peptídeos Cíclicos/metabolismo , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Streptomyces coelicolor/metabolismo , Bacillus subtilis/química , Bacillus subtilis/crescimento & desenvolvimento , Medicago sativa/microbiologia , Raízes de Plantas/microbiologia , Prótons , Percepção de Quorum/fisiologia , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/instrumentação , Streptomyces coelicolor/química , Streptomyces coelicolor/crescimento & desenvolvimentoRESUMO
Some insects form symbioses in which actinomycetes provide defense against pathogens by making antimicrobials. The range of chemical strategies employed across these associations, and how these strategies relate to insect lifestyle, remains underexplored. We assessed subsocial passalid beetles of the species Odontotaenius disjunctus, and their frass (fecal material), which is an important food resource within their galleries, as a model insect/actinomycete system. Through chemical and phylogenetic analyses, we found that O. disjunctus frass collected across eastern North America harbored multiple lineages of Streptomyces and diverse antimicrobials. Metabolites detected in frass displayed synergistic and antagonistic inhibition of a fungal entomopathogen, Metarhizium anisopliae, and multiple streptomycete isolates inhibited this pathogen when co-cultivated directly in frass. These findings support a model in which the lifestyle of O. disjunctus accommodates multiple Streptomyces lineages in their frass, resulting in a rich repertoire of antimicrobials that likely insulates their galleries against pathogenic invasion.
Assuntos
Anti-Infecciosos/análise , Anti-Infecciosos/metabolismo , Besouros/microbiologia , Streptomyces/química , Streptomyces/genética , Animais , Anti-Infecciosos/farmacologia , Fezes/microbiologia , Fungos/efeitos dos fármacos , América do Norte , Filogenia , Streptomyces/classificação , Streptomyces/metabolismo , SimbioseRESUMO
Microbiomes associated with various plant structures often contain members with the potential to make specialized metabolites, e.g., molecules with antibacterial, antifungal, or siderophore activities. However, when and where microbes associated with plants produce specialized metabolites, and the potential role of these molecules in mediating intramicrobiome interactions, is not well understood. Root nodules of legume plants are organs devoted to hosting symbiotic bacteria that fix atmospheric nitrogen and have recently been shown to harbor a relatively simple accessory microbiome containing members with the ability to produce specialized metabolites in vitro On the basis of these observations, we sought to develop a model nodule microbiome system for evaluating specialized microbial metabolism in planta Starting with an inoculum derived from field-grown Medicago sativa nodules, serial passaging through gnotobiotic nodules yielded a simplified accessory community composed of four members: Brevibacillus brevis, Paenibacillus sp., Pantoea agglomerans, and Pseudomonas sp. Some members of this community exhibited clear cooperation in planta, while others were antagonistic and capable of disrupting cooperation between other partners. Using matrix-assisted laser desorption ionization-imaging mass spectrometry, we found that metabolites associated with individual taxa had unique distributions, indicating that some members of the nodule community were spatially segregated. Finally, we identified two families of molecules produced by B. brevisin planta as the antibacterial tyrocidines and a novel set of gramicidin-type molecules, which we term the britacidins. Collectively, these results indicate that in addition to nitrogen fixation, legume root nodules are likely also sites of active antimicrobial production.
Assuntos
Bactérias/metabolismo , Brevibacillus/metabolismo , Interações Microbianas , Microbiota , Nódulos Radiculares de Plantas/microbiologia , Bactérias/classificação , Brevibacillus/genética , Medicago sativa/microbiologia , Nitrogênio/metabolismo , Fixação de Nitrogênio , SimbioseRESUMO
Microbes occupy diverse habitats, forming interconnected, dynamic communities. Elucidating the principles of microbial community function is a grand challenge for microbiology, and it will entail experiments that engage microbiomes across multiple levels of complexity. For example, community-level hypotheses often require testing at the mechanistic and/or genetic levels, while mechanistic relationships require community-level evaluation to understand their importance in context. In this Perspective, we articulate the need for model microbiome systems that enable experimentation in both community and reductionist frameworks, with an emphasis on understanding the role of specialized metabolites in microbial communities. We consider essential criteria for developing such model microbiome systems and discuss potential future models that address the ecology of specialized metabolism.