A hierarchy of metabolite exchanges in metabolic models of microbial species and communities.

The metabolic network of an organism can be analyzed as a constraint-based model. This analysis can be biased, optimizing an objective such as growth rate, or unbiased, aiming to describe the full feasible space of metabolic fluxes through pathway analysis or random flux sampling. In particular, pathway analysis can decompose the flux space into fundamental and formally defined metabolic pathways. Unbiased methods scale poorly with network size due to combinatorial explosion, but a promising approach to improve scalability is to focus on metabolic subnetworks, e.g., cells' metabolite exchanges with each other and the environment, rather than the full metabolic networks. Here, we applied pathway enumeration and flux sampling to metabolite exchanges in microbial species and a microbial community, using models ranging from central carbon metabolism to genome-scale and focusing on pathway definitions that allow direct targeting of subnetworks such as metabolite exchanges (elementary conversion modes, elementary flux patterns, and minimal pathways). Enumerating growth-supporting metabolite exchanges, we found that metabolite exchanges from different pathway definitions were related through a hierarchy, and we show that this hierarchical relationship between pathways holds for metabolic networks and subnetworks more generally. Metabolite exchange frequencies, defined as the fraction of pathways in which each metabolite was exchanged, were similar across pathway definitions, with a few specific exchanges explaining large differences in pathway counts. This indicates that biological interpretation of predicted metabolite exchanges is robust to the choice of pathway definition, and it suggests strategies for more scalable pathway analysis. Our results also signal wider biological implications, facilitating detailed and interpretable analysis of metabolite exchanges and other subnetworks in fields such as metabolic engineering and synthetic biology.

SALARECON connects the Atlantic salmon genome to growth and feed efficiency.

Atlantic salmon (Salmo salar) is the most valuable farmed fish globally and there is much interest in optimizing its genetics and rearing conditions for growth and feed efficiency. Marine feed ingredients must be replaced to meet global demand, with challenges for fish health and sustainability. Metabolic models can address this by connecting genomes to metabolism, which converts nutrients in the feed to energy and biomass, but such models are currently not available for major aquaculture species such as salmon. We present SALARECON, a model focusing on energy, amino acid, and nucleotide metabolism that links the Atlantic salmon genome to metabolic fluxes and growth. It performs well in standardized tests and captures expected metabolic (in)capabilities. We show that it can explain observed hypoxic growth in terms of metabolic fluxes and apply it to aquaculture by simulating growth with commercial feed ingredients. Predicted limiting amino acids and feed efficiencies agree with data, and the model suggests that marine feed efficiency can be achieved by supplementing a few amino acids to plant- and insect-based feeds. SALARECON is a high-quality model that makes it possible to simulate Atlantic salmon metabolism and growth. It can be used to explain Atlantic salmon physiology and address key challenges in aquaculture such as development of sustainable feeds.

Model-based integration of genomics and metabolomics reveals SNP functionality in Mycobacterium tuberculosis.

Human tuberculosis is caused by members of the Mycobacterium tuberculosis complex (MTBC) that vary in virulence and transmissibility. While genome-wide association studies have uncovered several mutations conferring drug resistance, much less is known about the factors underlying other bacterial phenotypes. Variation in the outcome of tuberculosis infection and diseases has been attributed primarily to patient and environmental factors, but recent evidence indicates an additional role for the genetic diversity among MTBC clinical strains. Here, we used metabolomics to unravel the effect of genetic variation on the strain-specific metabolic adaptive capacity and vulnerability. To define the functionality of single-nucleotide polymorphisms (SNPs) systematically, we developed a constraint-based approach that integrates metabolomic and genomic data. Our model-based predictions correctly classify SNP effects in pyruvate kinase and suggest a genetic basis for strain-specific inherent baseline susceptibility to the antibiotic para-aminosalicylic acid. Our method is broadly applicable across microbial life, opening possibilities for the development of more selective treatment strategies.

Effects of Yeast Species and Processing on Intestinal Health and Transcriptomic Profiles of Atlantic Salmon (Salmo salar) Fed Soybean Meal-Based Diets in Seawater.

The objective of the current study was to examine the effects of yeasts on intestinal health and transcriptomic profiles from the distal intestine and spleen tissue of Atlantic salmon fed SBM-based diets in seawater. Cyberlindnera jadinii (CJ) and Wickerhamomyces anomalus (WA) yeasts were heat-inactivated with spray-drying (ICJ and IWA) or autolyzed at 50 °C for 16 h (ACJ and AWA), followed by spray-drying. Six diets were formulated, one based on fishmeal (FM), a challenging diet with 30% soybean meal (SBM) and four other diets containing 30% SBM and 10% of each of the four yeast fractions (i.e., ICJ, ACJ, IWA and AWA). The inclusion of CJ yeasts reduced the loss of enterocyte supranuclear vacuolization and reduced the population of CD8α labeled cells present in the lamina propria of fish fed the SBM diet. The CJ yeasts controlled the inflammatory responses of fish fed SBM through up-regulation of pathways related to wound healing and taurine metabolism. The WA yeasts dampened the inflammatory profile of fish fed SBM through down-regulation of pathways related to toll-like receptor signaling, C-lectin receptor, cytokine receptor and signal transduction. This study suggests that the yeast species, Cyberlindnera jadinii and Wickerhamomyces anomalus are novel high-quality protein sources with health-beneficial effects in terms of reducing inflammation associated with feeding plant-based diets to Atlantic salmon.

Integrating host and microbiome biology using holo-omics.

Holo-omics is the use of omics data to study a host and its inherent microbiomes - a biological system known as a "holobiont". A microbiome that exists in such a space often encounters habitat stability and in return provides metabolic capacities that can benefit their host. Here we present an overview of beneficial host-microbiome systems and propose and discuss several methodological frameworks that can be used to investigate the intricacies of the many as yet undefined host-microbiome interactions that influence holobiont homeostasis. While this is an emerging field, we anticipate that ongoing methodological advancements will enhance the biological resolution that is necessary to improve our understanding of host-microbiome interplay to make meaningful interpretations and biotechnological applications.

Nucleobase adducts bind MR1 and stimulate MR1-restricted T cells.

MR1T cells are a recently found class of T cells that recognize antigens presented by the major histocompatibility complex-I-related molecule MR1 in the absence of microbial infection. The nature of the self-antigens that stimulate MR1T cells remains unclear, hampering our understanding of their physiological role and therapeutic potential. By combining genetic, pharmacological, and biochemical approaches, we found that carbonyl stress and changes in nucleobase metabolism in target cells promote MR1T cell activation. Stimulatory compounds formed by carbonyl adducts of nucleobases were detected within MR1 molecules produced by tumor cells, and their abundance and antigenicity were enhanced by drugs that induce carbonyl accumulation. Our data reveal carbonyl-nucleobase adducts as MR1T cell antigens. Recognizing cells under carbonyl stress allows MR1T cells to monitor cellular metabolic changes with physiological and therapeutic implications.

Extracting functionally accurate context-specific models of Atlantic salmon metabolism.

Constraint-based models (CBMs) are used to study metabolic network structure and function in organisms ranging from microbes to multicellular eukaryotes. Published CBMs are usually generic rather than context-specific, meaning that they do not capture differences in reaction activities, which, in turn, determine metabolic capabilities, between cell types, tissues, environments, or other conditions. Only a subset of a CBM's metabolic reactions and capabilities are likely to be active in any given context, and several methods have therefore been developed to extract context-specific models from generic CBMs through integration of omics data. We tested the ability of six model extraction methods (MEMs) to create functionally accurate context-specific models of Atlantic salmon using a generic CBM (SALARECON) and liver transcriptomics data from contexts differing in water salinity (life stage) and dietary lipids. Three MEMs (iMAT, INIT, and GIMME) outperformed the others in terms of functional accuracy, which we defined as the extracted models' ability to perform context-specific metabolic tasks inferred directly from the data, and one MEM (GIMME) was faster than the others. Context-specific versions of SALARECON consistently outperformed the generic version, showing that context-specific modeling better captures salmon metabolism. Thus, we demonstrate that results from human studies also hold for a non-mammalian animal and major livestock species.

Effect of yeast species and processing on intestinal microbiota of Atlantic salmon (Salmo salar) fed soybean meal-based diets in seawater.

BACKGROUND: Yeasts are gaining attention as alternative ingredients in aquafeeds. However, the impact of yeast inclusion on modulation of intestinal microbiota of fish fed plant-based ingredients is limited. Thus, the present study investigates the effects of yeast and processing on composition, diversity and predicted metabolic capacity of gut microbiota of Atlantic salmon smolt fed soybean meal (SBM)-based diet. Two yeasts, Cyberlindnera jadinii (CJ) and Wickerhamomyces anomalus (WA), were produced in-house and processed by direct heat-inactivation with spray-drying (ICJ and IWA) or autolyzed at 50 °C for 16 h, followed by spray-drying (ACJ and AWA). In a 42-day feeding experiment, fish were fed one of six diets: a fishmeal (FM)-based diet, a challenging diet with 30% SBM and four other diets containing 30% SBM and 10% of each of the four yeast products (i.e., ICJ, ACJ, IWA and AWA). Microbial profiling of digesta samples was conducted using 16S rRNA gene sequencing, and the predicted metabolic capacities of gut microbiota were determined using genome-scale metabolic models. RESULTS: The microbial composition and predicted metabolic capacity of gut microbiota differed between fish fed FM diet and those fed SBM diet. The digesta of fish fed SBM diet was dominated by members of lactic acid bacteria, which was similar to microbial composition in the digesta of fish fed the inactivated yeasts (ICJ and IWA diets). Inclusion of autolyzed yeasts (ACJ and AWA diets) reduced the richness and diversity of gut microbiota in fish. The gut microbiota of fish fed ACJ diet was dominated by the genus Pediococcus and showed a predicted increase in mucin O-glycan degradation compared with the other diets. The gut microbiota of fish fed AWA diet was highly dominated by the family Bacillaceae. CONCLUSIONS: The present study showed that dietary inclusion of FM and SBM differentially modulate the composition and predicted metabolic capacity of gut microbiota of fish. The inclusion of inactivated yeasts did not alter the modulation caused by SBM-based diet. Fish fed ACJ diet increased relative abundance of Pediococcus, and mucin O-glycan degradation pathway compared with the other diets.

Modulation of Atlantic salmon (Salmo salar) gut microbiota composition and predicted metabolic capacity by feeding diets with processed black soldier fly (Hermetia illucens) larvae meals and fractions.

BACKGROUND: Black soldier fly (Hermetia illucens) is a promising insect species to use as a novel ingredient in fish feeds. Black soldier fly larvae consists of three major fractions, namely protein, lipid, and exoskeleton. These fractions contain bioactive compounds that can modulate the gut microbiota in fish such as antimicrobial peptides, lauric acid, and chitin. However, it is not certain how, or which fractions of black solider fly would affect gut microbiota in fish. In the present study, black soldier fly larvae were processed into three different meals (full-fat, defatted and de-chitinized) and two fractions (oil and exoskeleton), and included in diets for Atlantic salmon (Salmo salar). Atlantic salmon pre-smolts were fed with these diets in comparison with a commercial-like control diet for eight weeks to investigate the effects of insect meals and fractions on the composition and predicted metabolic capacity of gut microbiota. The gut microbiota was profiled by 16S rRNA gene sequencing, and the predicted metabolic capacities of gut microbiota were determined using genome-scale metabolic models. RESULTS: The inclusion of insect meals and fractions decreased abundance of Proteobacteria and increased abundance of Firmicutes in salmon gut. The diets that contained insect chitin, i.e., insect meals or exoskeleton diets, increased abundance of chitinolytic bacteria including lactic acid bacteria and Actinomyces in salmon gut, with fish fed full-fat meal diet showing the highest abundances. The diets that contained insect lipids, i.e., insect meals and oil diets enriched Bacillaceae in fish gut. The fish fed diets containing full-fat insect meal had a unique gut microbiota composition dominated by beneficial lactic acid bacteria and Actinomyces, and showed a predicted increase in mucin degradation compared to the other diets. CONCLUSIONS: The present results showed that the dietary inclusion of insect meals and fractions can differently modulate the composition and predicted metabolic capacity of gut microbiota in Atlantic salmon pre-smolts. The use of full-fat black soldier fly larvae meal in diets for salmon is more favorable for beneficial modulation of gut microbiota than larvae processed by separation of lipid or exoskeleton fractions.

The Spleen as a Target to Characterize Immunomodulatory Effects of Down-Stream Processed Cyberlindnera jadinii Yeasts in Atlantic Salmon Exposed to a Dietary Soybean Meal Challenge.

Aquaculture feeds have changed dramatically from being largely based on fishmeal (FM) towards increased use of plant protein sources, which could impact the fish's immune response. In order to characterize immunomodulatory properties of novel functional ingredients, this study used four diets, one based on FM, a challenging diet with 40% soybean meal (SBM), and two diets containing 40% SBM with 5% of Cyberlindnera jadinii yeast exposed to different down-stream processing conditions: heat-inactivated (ICJ) or autolysation (ACJ). The immunomodulatory effects of the diets were analyzed in the spleen of Atlantic salmon after 37 days of feeding, using a transcriptomic evaluation by RNA sequencing (RNA-seq) and the detection of specific immunological markers at the protein level through indirect Enzyme-linked Immunosorbent Assay (indirect ELISA). The results showed that SBM (compared to FM) induced a down-regulation of pathways related to ion binding and transport, along with an increase at the protein level of pro-inflammatory cytokines such as tumor necrosis factor alpha (TNFα) and interferon gamma (IFNγ). On the other hand, while ICJ (compared to FM-group) maintain the inflammatory response associated with SBM, with higher levels of TNFα and IFNγ, and with an upregulation of creatine kinase activity and phosphagen metabolic process, the inclusion of ACJ was able to modulate the response of Atlantic salmon compared to fish fed the SBM-diet by the activation of biological pathways related to endocytosis, Pattern recognition receptor (PPRs)-signal transduction and transporter activity. In addition, ACJ was also able to control the pro-inflammatory profile of SBM, increasing Interleukin 10 (IL-10) levels and decreasing TNFα production, triggering an immune response similar to that of fish fed an FM-based diet. Finally, we suggest that the spleen is a good candidate to characterize the immunomodulatory effects of functional ingredients in Atlantic salmon. Moreover, the inclusion of ACJ in fish diets, with the ability to control inflammatory processes, could be considered in the formulation of sustainable salmon feed.

Microbial network disturbances in relapsing refractory Crohn's disease.

Inflammatory bowel diseases (IBD) can be broadly divided into Crohn's disease (CD) and ulcerative colitis (UC) from their clinical phenotypes. Over 150 host susceptibility genes have been described, although most overlap between CD, UC and their subtypes, and they do not adequately account for the overall incidence or the highly variable severity of disease. Replicating key findings between two long-term IBD cohorts, we have defined distinct networks of taxa associations within intestinal biopsies of CD and UC patients. Disturbances in an association network containing taxa of the Lachnospiraceae and Ruminococcaceae families, typically producing short chain fatty acids, characterize frequently relapsing disease and poor responses to treatment with anti-TNF-α therapeutic antibodies. Alterations of taxa within this network also characterize risk of later disease recurrence of patients in remission after the active inflamed segment of CD has been surgically removed.

Publisher Correction: Microbial network disturbances in relapsing refractory Crohn's disease.

Owing to an error during typesetting, a number of references were deleted from the Methods reference list. This altered all of the references in the Methods section and some of the references in Extended Data Fig. 5, making them inaccurate. References 121-134 were added back into the Methods reference list, and the references in the Methods section and in Extended Data Fig. 5 were renumbered accordingly. The error has been corrected in the PDF and HTML versions of this article.

BioBrick assembly standards and techniques and associated software tools.

The BioBrick idea was developed to introduce the engineering principles of abstraction and standardization into synthetic biology. BioBricks are DNA sequences that serve a defined biological function and can be readily assembled with any other BioBrick parts to create new BioBricks with novel properties. In order to achieve this, several assembly standards can be used. Which assembly standards a BioBrick is compatible with, depends on the prefix and suffix sequences surrounding the part. In this chapter, five of the most common assembly standards will be described, as well as some of the most used assembly techniques, cloning procedures, and a presentation of the available software tools that can be used for deciding on the best method for assembling of different BioBricks, and searching for BioBrick parts in the Registry of Standard Biological Parts database.