Project description:Synthetic microbial consortia represent a new frontier for synthetic biology given that they can solve more complex problems than monocultures. However, most attempts to co-cultivate these artificial communities fail because of the ‘‘winner-takes-all’’ in nutrients competition. In soil, multiple species can coexist with a spatial organization. Inspired by nature, here we show that an engineered spatial segregation method can assemble stable consortia with both flexibility and precision. We create microbial swarmbot consortia (MSBC) by encapsulating subpopulations with polymeric microcapsules. The crosslinked structure of microcapsules fences microbes, but allows the transport of small molecules and proteins. MSBC method enables the assembly of various synthetic communities and the precise control over the subpopulations. These capabilities can readily modulate the division of labor and communication. Our work integrates the synthetic biology and material science to offer new insights into consortia assembly and server as foundation to diverse applications from biomanufacturing to engineered photosynthesis.
Project description:The principles governing acquisition and interspecies exchange of nutrients in microbial communities and how those exchanges impact community productivity are poorly understood. Here, we examine energy and macronutrient acquisition in unicyanobacterial consortia for which species-resolved genome information exists for all members, allowing us to use multi-omic approaches to predict species’ abilities to acquire resources and examine expression of resource-acquisition genes during succession. Metabolic reconstruction indicated that a majority of heterotrophic community members lacked the genes required to directly acquire the inorganic nutrients provided in culture medium, suggesting high metabolic interdependency. The sole primary producer in consortium UCC-O, cyanobacterium Phormidium sp. OSCR, displayed declining expression of energy harvest, carbon fixation, and nitrate and sulfate reduction proteins but sharply increasing phosphate transporter expression over 28 days. Most heterotrophic members likewise exhibited signs of phosphorus starvation during succession. Though similar in their responses to phosphorus limitation, heterotrophs displayed species-specific expression of nitrogen acquisition genes. These results suggest niche partitioning around nitrogen sources may structure the community when organisms directly compete for limited phosphate. Such niche complementarity around nitrogen sources may increase community diversity and productivity in phosphate-limited phototrophic communities.
Project description:Microbial autotroph-heterotroph interactions influence biogeochemical cycles on a global scale, but the diversity and complexity of natural systems and their intractability to in situ manipulation make it challenging to elucidate the principles governing these interactions. The study of assembling phototrophic biofilm communities provides a robust means to identify such interactions and evaluate their contributions to the recruitment and maintenance of phylogenetic and functional diversity overtime. To examine primary succession in phototrophic communities, we isolated two unicyanobacterial consortia from the microbial mat in HotLake, Washington, characterizing the membership and metabolic function of each consortium. We then analyzed the spatial structures and quantified the community compositions of their assembling biofilms. The consortia retained the same suite of heterotrophic species, identified as abundant members of the mat and assigned to Alphaproteobacteria, Gammaproteobacteria, and Bacteroidetes. Autotroph growth rates dominated early in assembly, yielding to increasing heterotroph growth rates late in succession. The two consortia exhibited similar assembly patterns, with increasing relative abundances of members from Bacteroidetes and Alphaproteobacteria concurrent with decreasing relative abundances of those from Gamma proteobacteria. Despite these similarities at higher taxonomic levels, the relative abundances of individual heterotrophic species were substantially different in the developing consortial biofilms. This suggests that, although similar niches are created by the cyanobacterial metabolisms, the resulting webs of autotroph-heterotroph and heterotroph-heterotroph interactions are specific to each primary producer. The relative simplicity and tractability of the Hot Lake unicyanobacterial consortia make them useful model systems for deciphering interspecies interactions and assembly principles relevant to natural microbial communities.
Project description:Microbial communities colonize plant tissues and contribute to host function. How these communities form and how individual members contribute to shaping the microbial community are not well understood. Synthetic microbial communities, where defined individual isolates are combined, can serve as valuable model systems for uncovering the organizational principles of communities. Using genome-defined organisms, systematic analysis by computationally-based network reconstruction can lead to mechanistic insights and the metabolic interactions between species. In this study, 10 bacterial strains isolated from the Populus deltoides rhizosphere were combined and passaged in two different media environments to form a stable microbial community. The membership and relative abundances of the strains stabilized after around 5 growth cycles and resulted in just a few dominant strains. To unravel the underlying metabolic interactions, the KBase platform was used for constructing community-level models and for elucidating the metabolic processes involved in shaping the microbial communities. These analyses were complemented by growth curves of the individual isolates, pairwise interaction screens, and metaproteomics of the community. Flux balance analysis was used to model the metabolic potential in the microbial community and identify potential metabolic exchanges among the component species. Revealing the mechanisms of interaction among plant-associated microorganisms will provide insights into strategies for engineering microbial communities that can potentially increase plant growth and disease resistance. Further, deciphering the membership and metabolic potentials of a bacterial community will enable the design of synthetic co-cultures with desired biological functions.
Project description:The interrelationships between our diets and the structure and operations of our gut microbial communities are poorly understood. A model microbial community of ten sequenced human gut bacteria was introduced into gnotobiotic mice and changes in the abundance of each species were measured in response to randomized perturbations of four defined ingredients in the host diet. From the responses, we developed a statistical model that predicted over 50% of the variation in species abundance in response to the diet perturbations and were able to identify which factors in the diet best explained the changes seen for each community member. The community’s transcriptional response was driven by the absolute abundance of each species, as diet ingredient concentrations were not associated with significant changes in the transcription of individual community members.
Project description:Chemical signaling in the plant microbiome can have drastic effects on microbial community structure, and on host growth and development. Previously, we demonstrated that the auxin metabolic signal interference performed by the bacterial genus Variovorax via a novel auxin degradation locus was essential for maintaining stereotypic root development in an ecologically-relevant bacterial synthetic community. Here, we dissect the Variovorax auxin degradation locus to define the genes necessary and sufficient for indole-3-acetic acid (IAA) degradation and signal interference. We determine the crystal structures and binding properties of the operon’s MarR-family repressor with IAA and other auxins. We identify auxin-degradation operons across the bacterial tree of life and define two distinct types based on gene content and metabolic products: iac-like and iad-like. We solve the structures of MarRs from representatives of each auxin degradation operon type, establishing that each have distinct IAA binding pockets. Comparison of representative IAA degrading strains from diverse bacterial genera show that while all degrade IAA, only strains containing iad-like auxin degrading operons interfere with auxin signaling in a complex synthetic community context. This suggests that iad-like operon containing strains, including Variovorax species, play a key ecological role in modulating auxins in the plant microbiome.
Project description:Known as “The Oriental Botanic Garden” and the natural gene bank of biological species, Shennongjia is one of the most biologically diverse areas in China and a member of UNESCO's World Network of Biosphere Reserves. The macro-organism resources of shennongjia have been deeply explored. However, the microbial community structure was scarcely detected. In this study, we aim to detedect the microbial community along six sites of Shennonajia Mountain and explore the major controlling factor in shaping microbial community with a microarray-based metagenomics tool named GeoChip 4.2.
Project description:mRNA sequencing in bacteria is challenging due to the abundance of ribosomal rRNA that cannot be easily removed prior to sequencing. While commercially available kits target specific rRNA sequences found in defined lists of common bacterial species, they are frequently inefficient when applied to other divergent species, including those from environmental isolates. Similar to the commercial kits, other common techniques for rRNA depletion typically employ large probe sets that tile full-length rRNA sequences; however, such approaches are both time consuming and expensive when applied to multiple species or complex consortia of non-model microbes. To overcome these limitations, we present EMBR-seq+, which employs less than twenty target oligonucleotides per rRNA molecule, and builds upon our previous rRNA depletion approach, EMBR-seq, through the addition of an RNase H depletion step, to achieve rRNA removal efficiencies of up to 99%. First, we applied EMBR-seq+ to monocultures of Escherichia coli, Geobacter metallireducens, and Fibrobacter succinogenes strain UWB7 to deplete rRNA to approximately 1-7% of the sequencing reads, demonstrating that the new method can be easily extended to diverse bacterial species. Further, in more complex co-cultures between F. succinogenes strain UWB7 and anerobic fungal species, we applied EMBR-seq+ to deplete both bacterial and fungal rRNA, with an approximately 4-fold improved bacterial rRNA depletion efficiency compared to a previous report using a commercial kit, thereby showing that the method can be effectively translated to non-model microbial mixtures. Notably, we also demonstrate that for microbial species with poorly annotated genomes and unknown rRNA sequences, the RNase H depletion component of EMBR-seq+ enables rapid iterations in probe design without requiring to start experiments from total RNA each time, and was key for depleting fungal rRNA to enrich the bacterial mRNA readout in co-cultures. Finally, efficient depletion of rRNA enabled systematic quantification of the reprogramming of the bacterial transcriptome when cultured in the presence of the anerobic fungi, Anaeromyces robustus and Caecomyces churrovis. We observed that F. succinogenes strain UWB7 transcribes nearly 200 carbohydrate-active enzyme (CAZyme) genes in both monoculture and co-culture conditions, with several lignocellulose-degrading CAZymes downregulated in the presence of an anerobic gut fungus. This finding is consistent with the premise that bacteria and fungi specialize in different aspects of biomass breakdown, such that the presence of one regulates the CAZyme production of the other. This also supports previous findings that the fungi release excess reducing sugars in the supernatant, which benefits other members of the microbial community. Thus EMBR-seq+ provides a new and detailed perspective of a rumen microbiome model system by dramatically improving the efficiency of mRNA sequencing, and more generally also enables high-throughput, cost-effective and rapid quantification of the transcriptome to gain functional insights into less-studied and non-model microbial systems.
Project description:Known as M-bM-^@M-^\The Oriental Botanic GardenM-bM-^@M-^] and the natural gene bank of biological species, Shennongjia is one of the most biologically diverse areas in China and a member of UNESCO's World Network of Biosphere Reserves. The macro-organism resources of shennongjia have been deeply explored. However, the microbial community structure was scarcely detected. In this study, we aim to detedect the microbial community along six sites of Shennonajia Mountain and explore the major controlling factor in shaping microbial community with a microarray-based metagenomics tool named GeoChip 4.2. Seventy-three samples were collected from six sites along the Shennongjia Mountain, with 5-15 replicates in every site