Mutualistic Cross-feeding Research Articles

Establishment of synthetic microbial consortia with Corynebacterium glutamicum and Pseudomonas putida: Design, construction, and application to production of γ-glutamylisopropylamide and l-theanine.

Microbial synthetic consortia are a promising alternative to classical monoculture for biotechnological applications and fermentative processes. Their versatile use offers advantages in the degradation of complex substrates, the allocation of the metabolic burden between individual partners, or the division of labour in energy utilisation, substrate supply or product formation. Here, stable synthetic consortia between the two industrially relevant production hosts, Pseudomonas putida KT2440 and Corynebacterium glutamicum ATCC13032, were established for the first time. By applying arginine auxotrophy/overproduction and/or formamidase-based utilisation of the rare nitrogen source formamide, different types of interaction were realised, such as commensal relationships (+/0 and 0/+) and mutualistic cross-feeding (+/+). These consortia did not only show stable growth but could also be used for fermentative production of the γ-glutamylated amines theanine and γ-glutamyl-isopropylamide (GIPA). The consortia produced up to 2.8 g L-1 of GIPA and up to 2.6 g L-1 of theanine, a taste-enhancing constituent of green tea leaves. Thus, the advantageous approach of using synthetic microbial consortia for fermentative production of value-added compounds was successfully demonstrated.

Combining SIMS and mechanistic modelling to reveal nutrient kinetics in an algal-bacterial mutualism.

Microbial communities are of considerable significance for biogeochemical processes, for the health of both animals and plants, and for biotechnological purposes. A key feature of microbial interactions is the exchange of nutrients between cells. Isotope labelling followed by analysis with secondary ion mass spectrometry (SIMS) can identify nutrient fluxes and heterogeneity of substrate utilisation on a single cell level. Here we present a novel approach that combines SIMS experiments with mechanistic modelling to reveal otherwise inaccessible nutrient kinetics. The method is applied to study the onset of a synthetic mutualistic partnership between a vitamin B12-dependent mutant of the alga Chlamydomonas reinhardtii and the B12-producing, heterotrophic bacterium Mesorhizobium japonicum, which is supported by algal photosynthesis. Results suggest that an initial pool of fixed carbon delays the onset of mutualistic cross-feeding; significantly, our approach allows the first quantification of this expected delay. Our method is widely applicable to other microbial systems, and will contribute to furthering a mechanistic understanding of microbial interactions.

Mutualistic cross-feeding in microbial systems generates bistability via an Allee effect

In microbial ecosystems, species not only compete for common resources but may also display mutualistic interactions as a result from metabolic cross-feeding. Such mutualism can lead to bistability. Depending on the initial population sizes, species will either survive or go extinct. Various phenomenological models have been suggested to describe bistability in mutualistic systems. However, these models do not account for interaction mediators such as nutrients. In contrast, nutrient-explicit models do not provide an intuitive understanding of what causes bistability. Here, we reduce a theoretical nutrient-explicit model of two mutualistic cross-feeders in a chemostat, uncovering an explicit relation to a growth model with an Allee effect. We show that the dilution rate in the chemostat leads to bistability by turning a weak Allee effect into a strong Allee effect. This happens as long as there is more production than consumption of cross-fed nutrients. Thanks to the explicit relationship of the reduced model with the underlying experimental parameters, these results allow to predict the biological conditions that sustain or prevent the survival of mutualistic species.

Metabolic Signaling and Spatial Interactions in the Oral Polymicrobial Community

Oral supra- and subgingival biofilms are complex communities in which hundreds of bacteria, viruses, and fungi reside and interact. In these social environments, microbes compete and cooperate for resources, such as living space and nutrients. The metabolic activities of bacteria can transform their microenvironment and dynamically influence the fitness and growth of cohabitating organisms. Biofilm communities are temporally and spatially organized largely due to cell-to-cell communication, which promotes synergistic interactions. Metabolic interactions maintain biofilm homeostasis through mutualistic cross-feeding, metabolic syntrophy, and cross-respiration. These interactions include reciprocal metabolite exchanges that promote the growth of physiologically compatible bacteria, processive catabolism of complex substrates, and unidirectional interactions that are globally important for the polymicrobial community. Additionally, oral bacterial interactions can lead to detoxification of oxidative compounds, which will provide protection to the community at large. It has also been established that specific organisms provide terminal electron acceptors to partner species that result in a shift from fermentation to respiration, thus increasing ATP yields and improving fitness. Indeed, many interspecies relationships are multidimensional, and the net outcome can be spatially and temporally dependent. Cross-kingdom interactions also occur as oral yeast are antagonistic to some oral bacteria, while numerous mutualistic interactions contribute to yeast-bacterial colonization, fitness in the oral community, and the pathogenesis of caries. Consideration of this social environment reveals behaviors and phenotypes that are not apparent through the study of microbes in isolation. Here, we provide a comprehensive overview of the metabolic interactions that shape the oral microbial community.

Recipient-Biased Competition for an Intracellularly Generated Cross-Fed Nutrient Is Required for Coexistence of Microbial Mutualists.

ABSTRACTMany mutualistic microbial relationships are based on nutrient cross-feeding. Traditionally, cross-feeding is viewed as being unidirectional, from the producer to the recipient. This is likely true when a producer’s waste, such as a fermentation product, has value only for a recipient. However, in some cases the cross-fed nutrient holds value for both the producer and the recipient. In such cases, there is potential for nutrient reacquisition by producer cells in a population, leading to competition against recipients. Here, we investigated the consequences of interpartner competition for cross-fed nutrients on mutualism dynamics by using an anaerobic coculture pairing fermentative Escherichia coli and phototrophic Rhodopseudomonas palustris. In this coculture, E. coli excretes waste organic acids that provide a carbon source for R. palustris. In return, R. palustris cross-feeds E. coli ammonium (NH4+), a compound that both species value. To explore the potential for interpartner competition, we first used a kinetic model to simulate cocultures with varied affinities for NH4+ in each species. The model predicted that interpartner competition for NH4+ could profoundly impact population dynamics. We then experimentally tested the predictions by culturing mutants lacking NH4+ transporters in both NH4+ competition assays and mutualistic cocultures. Both theoretical and experimental results indicated that the recipient must have a competitive advantage in acquiring cross-fed NH4+ to sustain the mutualism. This recipient-biased competitive advantage is predicted to be crucial, particularly when the communally valuable nutrient is generated intracellularly. Thus, the very metabolites that form the basis for mutualistic cross-feeding can also be subject to competition between mutualistic partners.

Privatization of cooperative benefits stabilizes mutualistic cross-feeding interactions in spatially structured environments.

Metabolic cross-feeding interactions are ubiquitous in natural microbial communities. However, it remains generally unclear whether the production and exchange of metabolites incurs fitness costs to the producing cells and if so, which ecological mechanisms can facilitate a cooperative exchange of metabolites among unrelated individuals. We hypothesized that positive assortment within structured environments can maintain mutualistic cross-feeding. To test this, we engineered Acinetobacter baylyi and Escherichia coli to reciprocally exchange essential amino acids. Interspecific coculture experiments confirmed that non-cooperating types were selectively favoured in spatially unstructured (liquid culture), yet disfavoured in spatially structured environments (agar plates). Both an individual-based model and experiments with engineered genotypes indicated that a segregation of cross-feeders and non-cooperating auxotrophs stabilized cooperative cross-feeding in spatially structured environments. Chemical imaging confirmed that auxotrophs were spatially excluded from cooperative benefits. Together, these results demonstrate that cooperative cross-feeding between different bacterial species is favoured in structured environments such as bacterial biofilms, suggesting this type of interactions might be common in natural bacterial communities.

Anoxic Conditions Promote Species-Specific Mutualism between Gut Microbes In Silico.

The human gut is inhabited by thousands of microbial species, most of which are still uncharacterized. Gut microbes have adapted to each other's presence as well as to the host and engage in complex cross feeding. Constraint-based modeling has been successfully applied to predicting microbe-microbe interactions, such as commensalism, mutualism, and competition. Here, we apply a constraint-based approach to model pairwise interactions between 11 representative gut microbes. Microbe-microbe interactions were computationally modeled in conjunction with human small intestinal enterocytes, and the microbe pairs were subjected to three diets with various levels of carbohydrate, fat, and protein in normoxic or anoxic environments. Each microbe engaged in species-specific commensal, parasitic, mutualistic, or competitive interactions. For instance, Streptococcus thermophilus efficiently outcompeted microbes with which it was paired, in agreement with the domination of streptococci in the small intestinal microbiota. Under anoxic conditions, the probiotic organism Lactobacillus plantarum displayed mutualistic behavior toward six other species, which, surprisingly, were almost entirely abolished under normoxic conditions. This finding suggests that the anoxic conditions in the large intestine drive mutualistic cross feeding, leading to the evolvement of an ecosystem more complex than that of the small intestinal microbiota. Moreover, we predict that the presence of the small intestinal enterocyte induces competition over host-derived nutrients. The presented framework can readily be expanded to a larger gut microbial community. This modeling approach will be of great value for subsequent studies aiming to predict conditions favoring desirable microbes or suppressing pathogens.