Synthetic Microbial Consortia With Programmable Ecological Interactions: Construction And Application

Microorganisms are the oldest,most diverse and widely distributed forms of life on Erath.They are tiny and largely unseen,but are crucial for material cycling and energy flow in both terrestrial and marine ecosystems.In natural settings,microbial individuals rarely live alone but aggregate together to form a complex and dynamic community.They form intricate ecological networks through constantly exchanging matter,energy and information between cells,which are essential for the structure and function of microbial ecosystems.Understanding the overwhelming complexity of ecological networks,however,has been challenging,owing to the intricate context of natural microbial communities and the lack of efficient tools for disentangling species interactions.Recently,synthetic microbial ecology has emerged as a new interdisciplinary frontier,which allows for exploing microbial interaction webs with a fresh angle based on the bottom-up approach.Recent studies have started to employ synthetic microbial consortia to investigate interspecific interactions,but they were confined to generate one or two specific interactions such as cooperation and competition.Evidently,simple synthetic microbial consortia were not sufficient to elucidate natural microbial communities with complex interaction webs.Therefore,there is a need to develop an experimental system that allows for studying microbial consortia with different types of ecological interactions simultaneously and/or microbial communities with complex interaction webs systematically.In this study,we used the engineered strains to assemble microbial communities with different ecological interactions based on constructed modules.And we developed different mathematical models to quantitatively describe microbial dynamics.The specific objectives of the study are as follows:(1)to construct synthetic microbial communities with programmable ecological interactions;(2)to establish frameworks of different microbial kinetic models;(3)to explore how interspecific interactions influence the response of populations to external interference based on synthetic communities.The main results are as follows:1.Engineering microbial communities with different types of ecological interactionsAn artificial microbial experimental system allows for rapidly generating synthetic microbial consortia with different types of ecological interactions.First,based on molecular biology techniques such as the in-frame gene knockout technique and recombinant plasmids,we genetically engineered the model microorganism Escherichia coli to obtain a pair of auxotroph and overproducer strains for an amino acid and a pair of sender and receiver in the quorum sensing system.Then,the engineered mutants were used to construct the metabolism-and quorum sensing-based modules,which were assembled to creat synthetic microbial consortia of commensalism,amensalism,cooperation,competition and predation.2.Exploring the controllability of synthetic microbial communitiesBased on the successful construction of synthetic microbial communities with different type of ecological interactions,we further explored the controllability of the species interactions and their influences on community dynamics.Through the coculture experiments with different initial conditions,we found that: on the one hand,the transitions between exploitation and the types of commensalism,amensalism or competition based on the same paired strains were plausible by tuning the two engineered modules.On the other hand,the synthetic microbial consortia with predatory relationship displayed the unique mode of population dynamics under certain initial inoculation conditions,with distinct patterns from community collapse,predator dominance to prey dominace.3.Constructing and comparing of three different microbial kinetic modelsThree different microbial kinetic models that are generalized Lotka-Volterra,chemical-mediated and integrated models,which are developed based on ordinary differential equations(ODEs).Comparisons between simulated and experimental results indicated that three models could quantitatively capture the experimentally observed population dynamics of synthetic microbial consortia with defined interactions.We also calculated the AIC and BIC to evaluate the performance of models and found that the integrated model had the best performance in describing amensalism and predation scenarios.4.Microbial interactions influence the response of populations to external interferencesWe first performed antibiotic interference experiments to investigate the effect of interspecific interactions on population stability based on the synthetic microbial communities.The results showed that microbes involved in different ecological interactions responsed differently to antibiotic interference.Only the resistance of R-Δarg R increased in amensalism and predatory relationships.And compared to the monoculture,the resistance improved by 167% and 34.6%,respectively.We then combined mathematical models to explore the ecological mechanism by which interactions affected the resistance and found that environmental stress accumulated in the direction of positive interactions and weakened in the direction of negative interactions.In general,we have successfully constructed synthetic microbial communities with programmable ecological interactions;explored the response of microbial communities to external interferences based on the experimental system;and established microbial kinetic models to quantitatively describe the population dynamics of synthetic microbial communities.Our study offers a fresh angle to engineering controllable and manipulable microbial systems for experimentally testing ecological questions with a much greater control and manipulation.