Genome-Scale Metabolic Model Driven Design of a Defined Medium for Campylobacter jejuni M1cam

Abstract

Campylobacter jejuni, the most frequent cause of food-borne bacterial gastroenteritis, is a fastidious organism when grown in the laboratory. Oxygen is required for growth, despite the presence of the metabolic mechanism for anaerobic respiration. Amino acid auxotrophies are variably reported and energy metabolism can occur through several electron donor/acceptor combinations. Overall, the picture is one of a flexible, but vulnerable metabolism. To understand Campylobacter metabolism, we have constructed a fully curated, metabolic model for the reference organism M1 (our variant is M1cam) and validated it through laboratory experiments. Our results show that M1cam is auxotrophic for methionine, niacinamide, and pantothenate. There are complete biosynthesis pathways for all amino acids except methionine and it can produce energy, but not biomass, in the absence of oxygen. M1cam will grow in DMEM/F-12 defined media but not in the previously published Campylobacter specific defined media tested. Using the model, we identified potential auxotrophies and substrates that may improve growth. With this information, we designed simple defined media containing inorganic salts, the auxotrophic substrates, L-methionine, niacinamide, and pantothenate, pyruvate and additional amino acids L-cysteine, L-serine, and L-glutamine for growth enhancement. Our defined media supports a 1.75-fold higher growth rate than Brucella broth after 48 h at 37 • C and sustains the growth of other Campylobacter jejuni strains. This media can be used to design reproducible assays that can help in better understanding the adaptation, stress resistance, and the virulence mechanisms of this pathogen. We have shown that with a well-curated metabolic model it is possible to design a media to grow this fastidious organism. This has implications for the investigation of new Campylobacter species defined through metagenomics, such as C. infans.

Boosting Biomass Quantity and Quality by Improved Mixotrophic Culture of the Diatom Phaeodactylum tricornutum

“…The genome-encoded metabolic potential, as a genome-scale metabolic model (GSM), has been previously exploited to define optimal conditions, design growth media, and predict metabolic fluxes under different genetic and environmental conditions (Kim et al, 2017 ; Pan and Reed, 2018 ; Gu et al, 2019 ; Ong et al, 2020 ; Tejera et al, 2020 ). Its analysis enables an investigation of metabolic behavior of the whole system and has been utilized to inform experimental design and to provide a rationale for experimental observations (McCloskey et al, 2013 ; Villanova et al, 2017 ; Mishra et al, 2018 ; van der Ark et al, 2018 ).…”

Genome-Scale Metabolic Modelling Approach to Understand the Metabolism of the Opportunistic Human Pathogen Staphylococcus epidermidis RP62A

“…The model was constructed on the basis of the BioCyc Pathway/Genome Database (PGDB) [19,20] for S. epidermidis RP62A, version 20.1, using the ScrumPy metabolic modelling package [21] and in a modular fashion, using the approach described by Tejera et al [18], Ahmad et al [22], and was comprised of the following:…”

“…Model curation and theoretical validation to ensure the stoichiometric and energetic consistency of the whole model was carried out using the approaches described in Tejera et al [18], Gevorgyan et al [28], Poolman et al [29]. This is an iterative process: if an individual problem is identified (e.g., synthesis of ATP with no mass flow) modifications to individual reactions are corrected (e.g., by correcting the reversibility and/or directionality of a reaction) and the corrections are used to generate a new version of the model.…”

“…Genome-scale metabolic models (GSMs) describe the network of reactions that are assumed to be present in a given organism, typically derived from an annotated genome, and may be used in a variety of endeavours, including the identification of novel pathways [13], metabolic engineering strategies [14], identification of potential drug targets [15,16], and the identification of minimal growth requirements (and design of minimal media) [17,18].…”

Substrate Utilisation and Energy Metabolism in Non-Growing Campylobacter jejuni M1cam

“…Different species require different conditions that include temperatures of 37 to 42 °C and specific atmospheric conditions, including a gas mix of 5–10% O , 5–10% CO and 80–85% N 2 , with some strains also requiring H [ 5 ]. Individual isolates, including different strains of C. jejuni , also show varied substrate auxotrophies, making the investigation of nutrient requirements for growth necessary but difficult [ 6 , 7 , 8 ]. Despite these strict growth requirements, C. jejuni can be isolated from a wide range of environmental niches, including water, soil, food, and milk [ 4 ].…”