Ecological modelling approaches for predicting emergent properties in microbial communities

Nature Ecology & Evolution, Journal Year: 2022, Volume and Issue: 6(7), P. 855 - 865

Published: May 16, 2022

Language: Английский

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Metabolic exchanges are ubiquitous in natural microbial communities

Nature Microbiology, Journal Year: 2023, Volume and Issue: 8(12), P. 2244 - 2252

Published: Nov. 23, 2023

Language: Английский

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Cyanobacteria as cell factories for the photosynthetic production of sucrose

Frontiers in Microbiology, Journal Year: 2023, Volume and Issue: 14

Published: Feb. 14, 2023

Biofuels and other biologically manufactured sustainable goods are growing in popularity demand. Carbohydrate feedstocks required for industrial fermentation processes have traditionally been supplied by plant biomass, but the large quantities to produce replacement commodity products may prevent long-term feasibility of this approach without alternative strategies sugar feedstocks. Cyanobacteria under consideration as potential candidates production carbohydrate feedstocks, with potentially lower land water requirements relative plants. Several cyanobacterial strains genetically engineered export significant sugars, especially sucrose. Sucrose is not only naturally synthesized accumulated cyanobacteria a compatible solute tolerate high salt environments, also an easily fermentable disaccharide used many heterotrophic bacteria carbon source. In review, we provide comprehensive summary current knowledge endogenous sucrose synthesis degradation pathways. We summarize genetic modifications that found increase secretion. Finally, consider state synthetic microbial consortia rely on sugar-secreting strains, which co-cultivated alongside microbes able directly convert sugars into higher-value compounds (e.g., polyhydroxybutyrates, 3-hydroxypropionic acid, or dyes) single-pot reaction. recent advances reported such cyanobacteria/heterotroph co-cultivation perspective future developments likely realize their bioindustrial potential.

Language: Английский

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A model-driven approach to upcycling recalcitrant feedstocks in Pseudomonas putida by decoupling PHA production from nutrient limitation

Cell Reports, Journal Year: 2024, Volume and Issue: 43(4), P. 113979 - 113979

Published: March 22, 2024

Bacterial polyhydroxyalkanoates (PHAs) have emerged as promising eco-friendly alternatives to petroleum-based plastics since they are synthesized from renewable resources and offer exceptional properties. However, their production is limited the stationary growth phase under nutrient-limited conditions, requiring customized strategies costly two-phase bioprocesses. In this study, we tackle these challenges by employing a model-driven approach reroute carbon flux remove regulatory constraints using synthetic biology. We construct collection of Pseudomonas putida-overproducing strains at expense lignin-related compounds growth-coupling approaches. PHA was successfully achieved during phase, resulting in up 46% PHA/cell dry weight while maintaining balanced carbon-to-nitrogen ratio. Our additionally validated an upcycling scenario enzymatically hydrolyzed polyethylene terephthalate feedstock. These findings potential revolutionize address global plastic crisis overcoming complexities traditional

Language: Английский

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Toward merging bottom–up and top–down model-based designing of synthetic microbial communities

Current Opinion in Microbiology, Journal Year: 2022, Volume and Issue: 69, P. 102169 - 102169

Published: June 25, 2022

Language: Английский

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Adaptive Laboratory Evolution of Microorganisms: Methodology and Application for Bioproduction

Microorganisms, Journal Year: 2022, Volume and Issue: 11(1), P. 92 - 92

Published: Dec. 29, 2022

Adaptive laboratory evolution (ALE) is a useful experimental methodology for fundamental scientific research and industrial applications to create microbial cell factories. By using ALE, cells are adapted the environment that researchers set based on their objectives through serial transfer of populations in batch cultivations or continuous cultures fitness (i.e., growth) under such an increases. Then, omics analyses evolved mutants, including genome sequencing, transcriptome, proteome metabolome analyses, performed. It expected can understand evolutionary adaptation processes, applications, microorganisms exhibit increased carbon source availability, stress tolerance, production target compounds analysis data. In this review article, methodologies ALE introduced. Moreover, application creation as factories has also been

Language: Английский

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Case Studies in the Assessment of Microbial Fitness: Seemingly Subtle Changes Can Have Major Effects on Phenotypic Outcomes

Journal of Molecular Evolution, Journal Year: 2023, Volume and Issue: 91(3), P. 311 - 324

Published: Feb. 8, 2023

Abstract Following the completion of an adaptive evolution experiment, fitness evaluations are routinely conducted to assess magnitude adaptation. In doing so, proper consideration should be given when determining appropriate methods as trade-offs may exist between accuracy and throughput. Here, we present three instances in which small changes framework or execution significantly impacted outcomes. The first case illustrates that discrepancies conclusions can arise depending on approach evaluating fitness, culture vessel used, sampling method. second reveals variations environmental conditions occur associated with material. Specifically, these subtle greatly affect microbial physiology leading pH distorting measurements. Finally, last reports heterogeneity CFU formation time result inaccurate conclusions. Based each case, considerations recommendations presented for future experiments.

Language: Английский

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Mutagenesis techniques for evolutionary engineering of microbes – exploiting CRISPR-Cas, oligonucleotides, recombinases, and polymerases

Trends in Microbiology, Journal Year: 2024, Volume and Issue: 32(9), P. 884 - 901

Published: March 15, 2024

The natural process of evolutionary adaptation is often exploited as a powerful tool to obtain microbes with desirable traits. For industrial microbes, engineering used generate variants that show increased yields or resistance stressful environments, thus obtaining superior microbial cell factories. However, even in large populations, the supply beneficial mutations typically low, which implies improved time-consuming and inefficient. To overcome this limitation, different techniques have been developed boost mutation rates. While some these methods simply increase overall rate across genome, others use recent developments DNA synthesis, synthetic biology, CRISPR-Cas control type location mutations. This review summarizes most important field model microorganisms. It discusses how both vitro vivo approaches can genetic diversity host, special emphasis on for optimization metabolic pathways precision fermentation.

Language: Английский

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Microbial production of riboflavin: Biotechnological advances and perspectives

Metabolic Engineering, Journal Year: 2021, Volume and Issue: 68, P. 46 - 58

Published: Sept. 2, 2021

Language: Английский

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Adaptive laboratory evolution of microbial co‐cultures for improved metabolite secretion

Molecular Systems Biology, Journal Year: 2021, Volume and Issue: 17(8)

Published: Aug. 1, 2021

Article9 August 2021Open Access Source DataTransparent process Adaptive laboratory evolution of microbial co-cultures for improved metabolite secretion Dimitrios Konstantinidis orcid.org/0000-0002-2134-6823 Structural and Computational Biology Unit, European Molecular Laboratory, Heidelberg, Germany Faculty Biosciences, Heidelberg University, Search more papers by this author Filipa Pereira orcid.org/0000-0002-0557-8480 Eva-Maria Geissen orcid.org/0000-0002-0423-7019 Kristina Grkovska orcid.org/0000-0002-3784-494X Eleni Kafkia orcid.org/0000-0001-9550-4487 Medical Research Council Toxicology Cambridge, UK Paula Jouhten orcid.org/0000-0003-1075-7448 VTT Technical Centre Finland Ltd, Espoo, Yongkyu Kim orcid.org/0000-0002-3336-6741 Saravanan Devendran orcid.org/0000-0002-1540-5241 Michael Zimmermann orcid.org/0000-0002-5797-3589 Kiran Raosaheb Patil Corresponding Author [email protected] orcid.org/0000-0002-6166-8640 Information Konstantinidis1,2, Pereira1,†, Geissen1, Grkovska1, Kafkia1,3, Jouhten4, Kim1,†, Devendran1, Zimmermann1 *,1,3 1Structural 2Faculty 3Medical 4VTT †Present address: Life Science Institute, University Michigan, Ann Arbor, USA Brain Korea Institute Technology, Seoul, South *Corresponding author. Tel: +44 1223 3 35640; E-mail: Systems (2021)17:e10189https://doi.org/10.15252/msb.202010189 PDFDownload PDF article text main figures. Peer ReviewDownload a summary the editorial decision including letters, reviewer comments responses to feedback. ToolsAdd favoritesDownload CitationsTrack CitationsPermissions ShareFacebookTwitterLinked InMendeleyWechatReddit Figures & Info Abstract has proven highly effective obtaining microorganisms with enhanced capabilities. Yet, method is inherently restricted traits that are positively linked cell fitness, such as nutrient utilization. Here, we introduce coevolution obligatory mutualistic communities improving fitness-costly metabolites through natural selection. In strategy, metabolic cross-feeding connects target metabolite, despite its cost secretor, survival proliferation entire community. We thus co-evolved wild-type lactic acid bacteria engineered auxotrophic Saccharomyces cerevisiae in synthetic growth medium leading bacterial isolates two B-group vitamins, viz., riboflavin folate. The increased production was specific targeted vitamin, evident also milk, complex environment naturally contains vitamins. Genomic, proteomic metabolomic analyses evolved bacteria, combination flux balance analysis, showed altered regulation towards supply vitamin precursors. Together, our findings demonstrate how metabolism adapts lifestyle exchange. SYNOPSIS shows can be used improvement secretion. Mutualistic exerts selection pressure compound. Lactic were yeast secretion; retained monocultures milk. Multi-omics analysis strains uncovered cellular networks adapt lifestyle. Introduction long-term Escherichia coli experiment (Lenski, 2017) highlighted under well-controlled conditions gain fundamental insights into adaptive processes. These experiments have helped, example, gauging predictability evolutionary outcomes (McDonald, 2019) divergence between fitness trajectories mutation rates clonal asexual populations (Maddamsetti et al, 2015). now well-established tool development biotechnological characteristics (Dragosits Mattanovich, 2013). applications include adaptation harsh (Wallace-Salinas Gorwa-Grauslund, 2013; Stella 2019), substrate utilization (Zhou 2012) boosting metabolically (Portnoy 2011; Tenaillon, 2018). Further, enabled studying emergence stability interspecies interactions antagonism (Koskella Brockhurst, 2014) exchange (Mee 2014; Harcombe studies primarily focus on establishing models cross-feeding, less emphasis molecular basis adaptations. while amino common (Machado 2021) been (Wintermute Silver, 2010), involving other nutrients well studied. approach does not require prior knowledge genetic elements underlying trait wish improve. Thus, applied arbitrarily organisms amenable engineering. particularly attractive when use due legislative or consumer preference considerations, example fermented food products (Burgess 2006). requirement applying evolution, whether organisms, interest correlated (Winkler While minimal underlines elegance success it underscores limited applicability impose toll To enable keeping advantages offered here exert This makes producer. Consider community members wherein each partner depends one essential metabolites. Secretion these will directly coupled both members. Any all compounds, their costs secretors, then subjected via selecting overall growth. tested concept consisting (LAB) produce vitamins (riboflavin folate) When grown nitrogen excess, secretes acids which LAB (Ponomarova 2017). yeast-lactic satisfies obligate mutualism. strains, engineered, products, folate, relevant applications. Beyond direct industrial relevance, study establishes proof feasibility using communities. Results Coevolution selects known secretors (Hugenholtz Smid, 2002). started identifying Lactobacillus plantarum strain from isolate collection could secrete riboflavin. As S. prototrophic riboflavin, auxotrophy deleting RIB4 RIB5 genes. double deletion mutant ability support L. chemically defined medium. survive absence (Fig 1A). Figure 1. A an cultured together experiments. Vitamin secreting selected based presence required biosynthetic genes genome. periodically transferred new media, fitness. Performance twelve during experiment. originated same parental co-culture. Shown optical density measurements before transfer. Individual shown blue-shaded lines; dark blue line average Riboflavin estimates (n = 134) 5, biological replicates). fluorescence intensity values (440-nm excitation /520-nm emission) supernatants collected after 72 h culturing supplemented Extracellular intracellular levels LC-MS Levels extracellular flavins (FMN + FAD), data available online figure. Data 1 [msb202010189-sup-0003-SDataFig1.zip] Download figure PowerPoint next performed serial transfer derived cultures. Non-shaking facilitate microaerobic preferred strain. non-shaking condition expected check and/or dominance cheater cells (i.e. profit goods but do contribute return) facilitating spatial organization, generally favours co-operators over cheaters (Stump experiment, cultures fresh media whenever reached stationary stage Following 25 transfers, maximal had sevenfold 1B). Only collapsed (after approximately 20 passages), indicating partners remain robust. From populations, isolated planatarum (134 isolates) (6 further characterization. phenotype 134 (circa 160 generations) estimated measuring cell-free supernatant. Notably, 60% secretion, up 5.6-fold increase comparison 1C). Four values, non-improved (D5), verified ultra-performance liquid chromatography (UPLC; Appendix Table S1). amount produced strain, medium, quantified chromatography–mass spectrometry (LC-MS) analysis. secreted 42 ± 16.4 ng/ml isolates, B4 (isolate B 4), 61.1 (3.2-fold increase), another one, E6 E 6), circa 426 208.7 (10-fold increase) 1D). addition investigated Flavin coenzymes, i.e. FAD FMN, constitute biologically active forms well. at similar 1E, S1), FMN undetectable samples. obtain comprehensive picture capacity analysed accumulation flavin coenzymes. exhibited considerable (144 55.1 ng/ml; 14.4-fold strain; two-tailed t-test, P < 0.05) Isolate accumulated 27 14.9 (2.7-fold comparable being observed contrast E6, (C2, C 2, G7, G 7) did show (C2: 60.9 20.7 ng/ml, G7: 18.2 3.2 ng/ml). However, higher amounts (1.7 1.9-fold C2 respectively; 11 6.7 15 16.6 Fig 1E). resulted positive biosynthesis. Our setting links desired compound whole community, even if productivity imposes burden individual Indeed, producing phenotype, strong decrease rate (B4: 0.05, E6: 0.05; 2A). mimicked co-culture availability 2017), strain's OD600 value significantly decreased (two-tailed 0.05), 2B). Additionally, grew better than either only "negative control" D5 production; S1, 2C). monitor closer species dynamics occurred flow cytometry containing Δrib4:rib5 constitutively express red protein (RFP), isolates. Sorting B4, supported 1.5- 3.5-fold 2D), still number 2D). Per OD, somewhat increased, albeit variation across replicates EV1A). ratio change > nature relation whereby tightly EV1B). By provided resources partner, which, turn, bacteria. 2. Effect maximum vertical horizontal lines mark respectively supplementation. Growth kinetics four co-cultured replicates; grey bars mean SD). Yeast (RFP-positive events fluorescence-activated sorting, Methods) panel C. Samples 84 conditioned 2 [msb202010189-sup-0004-SDataFig2.zip] Click expand EV1. Evaluation associated structure Bacterial sorting (FACS) co-cultures. 2C inoculation counts normalized sample time. P-values unpaired t-test. numbers RFP-positive (showed FACS A. Representative image fixed 2C) (RFP positive), seen wide-field microscope, different time points kinetics. scale bar bottom left corner microscope equal 6 μm. aqueous suspension (PBS) attach (auto-aggregation—left), they exhibit mixed (co-aggregation—right). n SD. Estimation light absorbance 570 nm dye bound biofilm monoculture (left), (right) Bar heights average; dots points; 4 replicates. Direct contact necessary co-aggregate form biofilms (Kawarai 2007; Arroyo-López 2012). therefore set out investigate dependent cell–cell Microscopic observation formation extended aggregates EV1C D). compared aggregation forming determine surface promote contact. Evolved aggregate monoculture, aggregating less, independently yeast. Nevertheless, differences small statistically significant EV1E). Furthermore, three tendency corresponding EV1F). indicate there no independence sufficiently vice versa. 20% 40% 2E). Similarly, considerably 2F). displayed 16 49% density, EV2A B). (Appendix S1A) S1B), (˜34% average). Both confirm intended lack aggregation. over-secreting emerged population able enhance supplying vitamin. At time, capability assimilate resource resulting net impact compensated for, efficient EV2. (Δrib4:rib5) yeast-L. respectively. B. C–H. Amino (LC-MS analysis) samples areas curve (AUC) peaks. Regulation pathway precursor successful coevolution, identify changes Genome sequencing reveal any mutations along operon, identified single nucleotide polymorphisms (SNPs) various regulate transcription uptake S2). Evolution reported (Turkarslan fine-tuning regulatory optimizing usage partners. Since none link biosynthesis, decided analyse

Language: Английский

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