Biology subjects

Hirning, A. J.

Publications and source records attributed to Hirning, A. J..

4 recordsLinked to original sources

bioRxiv2025

Long-term homeostasis in microbial consortia via auxotrophic cross-feeding

Synthetic microbial consortia are collections of multiple strains or species of engineered organisms living in a shared ecosystem. Because they can separate metabolic tasks among different strains, synthetic microbial consortia have myriad applications in developing biomaterials, biomanufacturing, and biotherapeutics. However, synthetic consortia often require burdensome control mechanisms to ensure that the members of the community remain at the correct proportions. This is especially true in continuous culture systems in which slight differences in growth rates can lead to extinctions. Here, we present a simple method for controlling consortia proportions using cross-feeding in continuous auxotrophic co-culture. We use mutually auxotrophic E. coli with different essential gene deletions and regulate the growth rates of members of the consortium via cross-feeding of the missing nutrients in each strain. We demonstrate precise regulation of the co-culture steady-state ratio by exogenous addition of the missing nutrients. We also model the co-cultures behavior using a system of ordinary differential equations that enable us to predict its response to changes in nutrient concentrations. Our work provides a powerful tool for consortia proportion control with minimal metabolic costs to the constituent strains.

synthetic biology↗

bioRxiv2023

Indirect enrichment of desirable, but less fit phenotypes, from a synthetic microbial community using microdroplet confinement

Spatial structure within microbial communities can provide nearly limitless opportunities for social interactions and are an important driver for evolution. As metabolites are often molecular signals, metabolite diffusion within microbial communities can affect the composition and dynamics of the community in a manner that can be challenging to deconstruct. We used encapsulation of a synthetic microbial community within microdroplets to investigate the effects of spatial structure and metabolite diffusion on population dynamics and to examine the effects of cheating by one member of the community. The synthetic community was comprised of three strains: a Producer that makes the diffusible quorum sensing molecule (N-(3-Oxododecanoyl)-L-homoserine lactone, C12-oxo-HSL) or AHL; a Receiver that is killed by AHL and a Non-Producer or cheater that benefits from the extinction of the Receivers, but without the costs associated with the AHL synthesis. We demonstrate that despite rapid diffusion of AHL between microdroplets, the spatial structure imposed by the microdroplets allow a more efficient but transient enrichment of more rare and slower growing Producer subpopulations. Eventually, the Non-Producer population drove the Producers to extinction. By including fluorescence-activated microdroplet sorting and providing sustained competition by the Receiver strain, we demonstrate a strategy for indirect enrichment of a rare and unlabeled Producer. The ability to screen and enrich metabolite Producers from a much larger population under conditions of rapid diffusion provides an important framework for the development of applications in synthetic ecology and biotechnology. Abstract Figure O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=124 SRC="FIGDIR/small/523444v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@81dcd9org.highwire.dtl.DTLVardef@1ca30f9org.highwire.dtl.DTLVardef@1553cc3org.highwire.dtl.DTLVardef@1085821_HPS_FORMAT_FIGEXP M_FIG C_FIG

synthetic biology↗

bioRxiv2022

Tunable dynamics in a multi-straintranscriptional pulse generator

A major challenge in synthetic biology is the manipulation of engineered gene circuits toward a specified behavior. This challenge becomes more difficult as synthetic systems become more complex by incorporating additional genes or strains. Here we demonstrate that circuit dynamics can be tuned in synthetic consortia through the manipulation of strain fractions within the community. To do this, we constructed a microbial consortium comprised of three strains of engineered Escherichia coli that, when co-cultured, use homoserine lactone (HSL) mediated intercellular signaling to create a multi-strain incoherent type-1 feedforward loop (I1-FFL). Like naturally occurring I1-FFL motifs in gene networks, this engineered microbial consortium acts as a pulse generator of gene expression. We demonstrated that the amplitude of the pulse can be easily tuned by adjusting the relative population fractions of the strains. We created a mathematical model for the temporal dynamics of the microbial consortium and, using this model, identified population fractions that produced desired pulse characteristics. Our work demonstrates that intercellular gene circuits can be effectively tuned simply by adjusting the starting fractions of each strain type.

synthetic biology↗

bioRxiv2019

Spatiotemporal dynamics of synthetic microbial consortia in microfluidic devices

Synthetic microbial consortia consist of two or more engineered strains that grow together and share the same resources. When intercellular signaling pathways are included in the engineered strains, close proximity of the microbes can generate complex dynamic behaviors that are difficult to obtain using a single strain. However, when a consortium is not cultured in a well-mixed environment the constituent strains passively compete for space as they grow and divide, complicating cell-cell signaling. Here, we explore the temporal dynamics of the spatial distribution of consortia co-cultured in microfluidic devices. To do this, we grew two different strains of Escherichia coli in microfluidic devices with cell-trapping regions (traps) of several different designs. We found that the size and shape of the traps are critical determinants of spatiotemporal dynamics. In small traps, cells can easily signal one another but the relative proportion of each strain within the trap can fluctuate wildly. In large traps, the relative ratio of strains is stabilized, but intercellular signaling can be hindered by distances between cells. This presents a trade-off between the trap size and the effectiveness of intercellular signaling, which can be mitigated by controlling the initial seeding of cells in the large trap. These results show how synthetic microbial consortia behave in microfluidic traps and provide a method to help remedy the spatial heterogeneity inherent to different trap geometries.

synthetic biology↗