| 1. |
- Abdel-Haleem, Alyaa M., et al.
(författare)
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Integrated Metabolic Modeling, Culturing, and Transcriptomics Explain Enhanced Virulence of Vibrio cholerae during Coinfection with Enterotoxigenic Escherichia coli
- 2020
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Ingår i: mSystems. - 2379-5077. ; 5:5
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Tidskriftsartikel (refereegranskat)abstract
- Gene essentiality is altered during polymicrobial infections. Nevertheless, most studies rely on single-species infections to assess pathogen gene essentiality. Here, we use genome-scale metabolic models (GEMs) to explore the effect of coinfection of the diarrheagenic pathogen Vibrio cholerae with another enteric pathogen, enterotoxigenic Escherichia coli (ETEC). Model predictions showed that V. cholerae metabolic capabilities were increased due to ample cross-feeding opportunities enabled by ETEC. This is in line with increased severity of cholera symptoms known to occur in patients with dual infections by the two pathogens. In vitro co-culture systems confirmed that V. cholerae growth is enhanced in cocultures relative to single cultures. Further, expression levels of several V. cholerae metabolic genes were significantly perturbed as shown by dual RNA sequencing (RNAseq) analysis of its cocultures with different ETEC strains. A decrease in ETEC growth was also observed, probably mediated by nonmetabolic factors. Single gene essentiality analysis predicted conditionally independent genes that are essential for the pathogen's growth in both single-infection and coinfection scenarios. Our results reveal growth differences that are of relevance to drug targeting and efficiency in polymicrobial infections. IMPORTANCE Most studies proposing new strategies to manage and treat infections have been largely focused on identifying druggable targets that can inhibit a pathogen's growth when it is the single cause of infection. In vivo, however, infections can be caused by multiple species. This is important to take into account when attempting to develop or use current antibacterials since their efficacy can change significantly between single infections and coinfections. In this study, we used genome-scale metabolic models (GEMs) to interrogate the growth capabilities of Vibrio cholerae in single infections and coinfections with enterotoxigenic E. coli (ETEC), which cooccur in a large fraction of diarrheagenic patients. Coinfection model predictions showed that V. cholerae growth capabilities are enhanced in the presence of ETEC relative to V. cholerae single infection, through cross-fed metabolites made available to V. cholerae by ETEC. In vitro, cocultures of the two enteric pathogens further confirmed model predictions showing an increased growth of V. cholerae in coculture relative to V. cholerae single cultures while ETEC growth was suppressed. Dual RNAseq analysis of the cocultures also confirmed that the transcriptome of V. cholerae was distinct during coinfection compared to single-infection scenarios where processes related to metabolism were significantly perturbed. Further, in silico gene-knockout simulations uncovered discrepancies in gene essentiality for V. cholerae growth between single infections and coinfections. Integrative model-guided analysis thus identified druggable targets that would be critical for V. cholerae growth in both single infections and coinfections; thus, designing inhibitors against those targets would provide a broader spectrum of coverage against cholera infections.
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| 2. |
- Alalam, Hanna, et al.
(författare)
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A Genetic Trap in Yeast for Inhibitors of SARS-CoV-2 Main Protease
- 2021
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Ingår i: MSYSTEMS. - 2379-5077. ; 6:6
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Tidskriftsartikel (refereegranskat)abstract
- The ongoing COVID-19 pandemic urges searches for antiviral agents that can block infection or ameliorate its symptoms. Using dissimilar search strategies for new antivirals will improve our overall chances of finding effective treatments. Here, we have established an experimental platform for screening of small molecule inhibitors of the SARS-CoV-2 main protease in Saccharomyces cerevisiae cells, genetically engineered to enhance cellular uptake of small molecules in the environment. The system consists of a fusion of the Escherichia coli toxin MazF and its antitoxin MazE, with insertion of a protease cleavage site in the linker peptide connecting the MazE and MazF moieties. Expression of the viral protease confers cleavage of the MazEF fusion, releasing the MazF toxin from its antitoxin, resulting in growth inhibition. In the presence of a small molecule inhibiting the protease, cleavage is blocked and the MazF toxin remains inhibited, promoting growth. The system thus allows positive selection for inhibitors. The engineered yeast strain is tagged with a fluorescent marker protein, allowing precise monitoring of its growth in the presence or absence of inhibitor. We detect an established main protease inhibitor by a robust growth increase, discernible down to 1 mM. The system is suitable for robotized large-scale screens. It allows in vivo evaluation of drug candidates and is rapidly adaptable for new variants of the protease with deviant site specificities. IMPORTANCE The COVID-19 pandemic may continue for several years before vaccination campaigns can put an end to it globally. Thus, the need for discovery of new antiviral drug candidates will remain. We have engineered a system in yeast cells for the detection of small molecule inhibitors of one attractive drug target of SARS-CoV-2, its main protease, which is required for viral replication. The ability to detect inhibitors in live cells brings the advantage that only compounds capable of entering the cell and remain stable there will score in the system. Moreover, because of its design in yeast cells, the system is rapidly adaptable for tuning the detection level and eventual modification of the protease cleavage site in the case of future mutant variants of the SARSCoV-2 main protease or even for other proteases.
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| 3. |
- Alalam, Hanna, et al.
(författare)
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A High-Throughput Method for Screening for Genes Controlling Bacterial Conjugation of Antibiotic Resistance.
- 2020
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Ingår i: mSystems. - 2379-5077. ; 5:6
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Tidskriftsartikel (refereegranskat)abstract
- The rapid horizontal transmission of antibiotic resistance genes on conjugative plasmids between bacterial host cells is a major cause of the accelerating antibiotic resistance crisis. There are currently no experimental platforms for fast and cost-efficient screening of genetic effects on antibiotic resistance transmission by conjugation, which prevents understanding and targeting conjugation. We introduce a novel experimental framework to screen for conjugation-based horizontal transmission of antibiotic resistance between >60,000 pairs of cell populations in parallel. Plasmid-carrying donor strains are constructed in high-throughput. We then mix the resistance plasmid-carrying donors with recipients in a design where only transconjugants can reproduce, measure growth in dense intervals, and extract transmission times as the growth lag. As proof-of-principle, we exhaustively explore chromosomal genes controlling F-plasmid donation within Escherichia coli populations, by screening the Keio deletion collection in high replication. We recover all seven known chromosomal gene mutants affecting conjugation as donors and identify many novel mutants, all of which diminish antibiotic resistance transmission. We validate nine of the novel genes' effects in liquid mating assays and complement one of the novel genes' effect on conjugation (rseA). The new framework holds great potential for exhaustive disclosing of candidate targets for helper drugs that delay resistance development in patients and societies and improve the longevity of current and future antibiotics. Further, the platform can easily be adapted to explore interspecies conjugation, plasmid-borne factors, and experimental evolution and be used for rapid construction of strains.IMPORTANCE The rapid transmission of antibiotic resistance genes on conjugative plasmids between bacterial host cells is a major cause of the accelerating antibiotic resistance crisis. There are currently no experimental platforms for fast and cost-efficient screening of genetic effects on antibiotic resistance transmission by conjugation, which prevents understanding and targeting conjugation. We introduce a novel experimental framework to screen for conjugation-based horizontal transmission of antibiotic resistance between >60,000 pairs of cell populations in parallel. As proof-of-principle, we exhaustively explore chromosomal genes controlling F-plasmid donation within E. coli populations. We recover all previously known and many novel chromosomal gene mutants that affect conjugation efficiency. The new framework holds great potential for rapid screening of compounds that decrease transmission. Further, the platform can easily be adapted to explore interspecies conjugation, plasmid-borne factors, and experimental evolution and be used for rapid construction of strains.
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| 4. |
- Allen, Lisa Zeigler, et al.
(författare)
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The Baltic Sea Virome : Diversity and Transcriptional Activity of DNA and RNA Viruses
- 2017
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Ingår i: mSystems. - 2379-5077. ; 2:1
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Tidskriftsartikel (refereegranskat)abstract
- Metagenomic and metatranscriptomic data were generated from size-fractionated samples from 11 sites within the Baltic Sea and adjacent marine waters of Kattegat and freshwater Lake Tornetrask in order to investigate the diversity, distribution, and transcriptional activity of virioplankton. Such a transect, spanning a salinity gradient from freshwater to the open sea, facilitated a broad genome-enabled investigation of natural as well as impacted aspects of Baltic Sea viral communities. Taxonomic signatures representative of phages within the widely distributed order Caudovirales were identified with enrichments in lesser-known families such as Podoviridae and Siphoviridae. The distribution of phage reported to infect diverse and ubiquitous heterotrophic bacteria (SAR11 clades) and cyanobacteria (Synechococcus sp.) displayed population-level shifts in diversity. Samples from higher-salinity conditions (>14 practical salinity units [PSU]) had increased abundances of viruses for picoeukaryotes, i.e., Ostreococcus. These data, combined with host diversity estimates, suggest viral modulation of diversity on the whole-community scale, as well as in specific prokaryotic and eukaryotic lineages. RNA libraries revealed single-stranded DNA (ssDNA) and RNA viral populations throughout the Baltic Sea, with ssDNA phage highly represented in Lake Tornetrask. Further, our data suggest relatively high transcriptional activity of fish viruses within diverse families known to have broad host ranges, such as Nodoviridae (RNA), Iridoviridae (DNA), and predicted zoonotic viruses that can cause ecological and economic damage as well as impact human health. IMPORTANCE Inferred virus-host relationships, community structures of ubiquitous ecologically relevant groups, and identification of transcriptionally active populations have been achieved with our Baltic Sea study. Further, these data, highlighting the transcriptional activity of viruses, represent one of the more powerful uses of omics concerning ecosystem health. The use of omics-related data to assess ecosystem health holds great promise for rapid and relatively inexpensive determination of perturbations and risk, explicitly with regard to viral assemblages, as no single marker gene is suitable for widespread taxonomic coverage.
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| 5. |
- Amador-García, Ahinara, et al.
(författare)
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Extending the Proteomic Characterization of Candida albicans Exposed to Stress and Apoptotic Inducers through Data-Independent Acquisition Mass Spectrometry
- 2021
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Ingår i: mSystems. - 2379-5077. ; 6:5
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Tidskriftsartikel (refereegranskat)abstract
- Candida albicans is a commensal fungus that causes systemic infections in immunosuppressed patients. In order to deal with the changing environment during commensalism or infection, C. albicans must reprogram its proteome. Characterizing the stress-induced changes in the proteome that C. albicans uses to survive should be very useful in the development of new antifungal drugs. We studied the C. albicans global proteome after exposure to hydrogen peroxide (H2O2) and acetic acid (AA), using a data-independent acquisition mass spectrometry (DIA-MS) strategy. More than 2,000 C. albicans proteins were quantified using an ion library previously constructed using data-dependent acquisition mass spectrometry (DDA-MS). C. albicans responded to treatment with H2O2 with an increase in the abundance of many proteins involved in the oxidative stress response, protein folding, and proteasome-dependent catabolism, which led to increased proteasome activity. The data revealed a previously unknown key role for Prn1, a protein similar to pirins, in the oxidative stress response. Treatment with AA resulted in a general decrease in the abundance of proteins involved in amino acid biosynthesis, protein folding, and rRNA processing. Almost all proteasome proteins declined, as did proteasome activity. Apoptosis was observed after treatment with H2O2 but not AA. A targeted proteomic study of 32 proteins related to apoptosis in yeast supported the results obtained by DIA-MS and allowed the creation of an efficient method to quantify relevant proteins after treatment with stressors (H2O2, AA, and amphotericin B). This approach also uncovered a main role for Oye32, an oxidoreductase, suggesting this protein as a possible apoptotic marker common to many stressors. IMPORTANCE Fungal infections are a worldwide health problem, especially in immunocompromised patients and patients with chronic disorders. Invasive candidiasis, caused mainly by C. albicans, is among the most common fungal diseases. Despite the existence of treatments to combat candidiasis, the spectrum of drugs available is limited. For the discovery of new drug targets, it is essential to know the pathogen response to different stress conditions. Our study provides a global vision of proteomic remodeling in C. albicans after exposure to different agents, such as hydrogen peroxide, acetic acid, and amphotericin B, that can cause apoptotic cell death. These results revealed the significance of many proteins related to oxidative stress response and proteasome activity, among others. Of note, the discovery of Prn1 as a key protein in the defense against oxidative stress as well the increase in the abundance of Oye32 protein when apoptotic process occurred point them out as possible drug targets.
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| 6. |
- Bellieny-Rabelo, Daniel, et al.
(författare)
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Horizontally Acquired Quorum-Sensing Regulators Recruited by the PhoP Regulatory Network Expand the Host Adaptation Repertoire in the Phytopathogen Pectobacterium brasiliense
- 2020
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Ingår i: mSystems. - Washington : American Society for Microbiology. - 2379-5077. ; 5:1
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Tidskriftsartikel (refereegranskat)abstract
- In this study, we examine the impact of transcriptional network rearrangements driven by horizontal gene acquisition in PhoP and SlyA regulons using as a case study a phytopathosystem comprised of potato tubers and the soft-rot pathogen Pectobacterium brasiliense 1692 (Pb1692). Genome simulations and statistical analyses uncovered the tendency of PhoP and SlyA networks to mobilize lineage-specific traits predicted as horizontal gene transfer at late infection, highlighting the prominence of regulatory network rearrangements in this stage of infection. The evidence further supports the circumscription of two horizontally acquired quorum-sensing regulators (carR and expR1) by the PhoP network. By recruiting carR and expR1, the PhoP network also impacts certain host adaptation- and bacterial competition-related systems, seemingly in a quorum sensing-dependent manner, such as the type VI secretion system, carbapenem biosynthesis, and plant cell wall-degrading enzymes (PCWDE) like cellulases and pectate lyases. Conversely, polygalacturonases and the type III secretion system (T3SS) exhibit a transcriptional pattern that suggests quorum-sensing-independent regulation by the PhoP network. This includes an uncharacterized novel phage-related gene family within the T3SS gene cluster that has been recently acquired by two Pectobacterium species. The evidence further suggests a PhoP-dependent regulation of carbapenem- and PCWDE-encoding genes based on the synthesized products' optimum pH. The PhoP network also controls slyA expression in planta, which seems to impact carbohydrate metabolism regulation, especially at early infection, when 76.2% of the SlyA-regulated genes from that category also require PhoP to achieve normal expression levels.IMPORTANCE: Exchanging genetic material through horizontal transfer is a critical mechanism that drives bacteria to efficiently adapt to host defenses. In this report, we demonstrate that a specific plant-pathogenic species (from the Pectobacterium genus) successfully integrated a population density-based behavior system (quorum sensing) acquired through horizontal transfer into a resident stress-response gene regulatory network controlled by the PhoP protein. Evidence found here underscores that subsets of bacterial weaponry critical for colonization, typically known to respond to quorum sensing, are also controlled by PhoP. Some of these traits include different types of enzymes that can efficiently break down plant cell walls depending on the environmental acidity level. Thus, we hypothesize that PhoP's ability to elicit regulatory responses based on acidity and nutrient availability fluctuations has strongly impacted the fixation of its regulatory connection with quorum sensing. In addition, another global gene regulator, known as SlyA, was found under the PhoP regulatory network. The SlyA regulator controls a series of carbohydrate metabolism-related traits, which also seem to be regulated by PhoP. By centralizing quorum sensing and slyA under PhoP scrutiny, Pectobacterium cells added an advantageous layer of control over those two networks that potentially enhances colonization efficiency.
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| 7. |
- Benkwitz-Bedford, Sam, et al.
(författare)
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Machine Learning Prediction of Resistance to Subinhibitory Antimicrobial Concentrations from Escherichia coli Genomes.
- 2021
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Ingår i: mSystems. - 2379-5077. ; 6:4
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Tidskriftsartikel (refereegranskat)abstract
- Escherichia coli is an important cause of bacterial infections worldwide, with multidrug-resistant strains incurring substantial costs on human lives. Besides therapeutic concentrations of antimicrobials in health care settings, the presence of subinhibitory antimicrobial residues in the environment and in clinics selects for antimicrobial resistance (AMR), but the underlying genetic repertoire is less well understood. Here, we used machine learning to predict the population doubling time and cell growth yield of 1,407 genetically diverse E. coli strains expanding under exposure to three subinhibitory concentrations of six classes of antimicrobials from single-nucleotide genetic variants, accessory gene variation, and the presence of known AMR genes. We predicted cell growth yields in the held-out test data with an average correlation (Spearman's ρ) of 0.63 (0.36 to 0.81 across concentrations) and cell doubling times with an average correlation of 0.59 (0.32 to 0.92 across concentrations), with moderate increases in sample size unlikely to improve predictions further. This finding points to the remaining missing heritability of growth under antimicrobial exposure being explained by effects that are too rare or weak to be captured unless sample size is dramatically increased, or by effects other than those conferred by the presence of individual single-nucleotide polymorphisms (SNPs) and genes. Predictions based on whole-genome information were generally superior to those based only on known AMR genes and were accurate for AMR resistance at therapeutic concentrations. We pinpointed genes and SNPs determining the predicted growth and thereby recapitulated many known AMR determinants. Finally, we estimated the effect sizes of resistance genes across the entire collection of strains, disclosing the growth effects for known resistance genes in each individual strain. Our results underscore the potential of predictive modeling of growth patterns from genomic data under subinhibitory concentrations of antimicrobials, although the remaining missing heritability poses a challenge for achieving the accuracy and precision required for clinical use. IMPORTANCE Predicting bacterial growth from genome sequences is important for a rapid characterization of strains in clinical diagnostics and to disclose candidate novel targets for anti-infective drugs. Previous studies have dissected the relationship between bacterial growth and genotype in mutant libraries for laboratory strains, yet no study so far has examined the predictive power of genome sequence in natural strains. In this study, we used a high-throughput phenotypic assay to measure the growth of a systematic collection of natural Escherichia coli strains and then employed machine learning models to predict bacterial growth from genomic data under nontherapeutic subinhibitory concentrations of antimicrobials that are common in nonclinical settings. We found a moderate to strong correlation between predicted and actual values for the different collected data sets. Moreover, we observed that the known resistance genes are still effective at sublethal concentrations, pointing to clinical implications of these concentrations.
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| 8. |
- Bergenholm, David, 1987, et al.
(författare)
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Reconstruction of a Global Transcriptional Regulatory Network for Control of Lipid Metabolism in Yeast by Using Chromatin Immunoprecipitation with Lambda Exonuclease Digestion
- 2018
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Ingår i: mSystems. - 2379-5077. ; 3:4
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Tidskriftsartikel (refereegranskat)abstract
- To build transcription regulatory networks, transcription factor binding must be analyzed in cells grown under different conditions because their responses and targets differ depending on environmental conditions. We performed wholegenome analysis of the DNA binding of five Saccharomyces cerevisiae transcription factors involved in lipid metabolism, Ino2, Ino4, Hap1, Oaf1, and Pip2, in response to four different environmental conditions in chemostat cultures, which allowed us to keep the specific growth rate constant. Chromatin immunoprecipitation with lambda exonuclease digestion (ChIP-exo) enabled the detection of binding events at a high resolution. We discovered a large number of unidentified targets and thus expanded functions for each transcription factor (e.g., glutamate biosynthesis as a target of Oaf1 and Pip2). Moreover, condition-dependent binding of transcription factors in response to cell metabolic state (e.g., differential binding of Ino2 between fermentative and respiratory metabolic conditions) was clearly suggested. Combining the new binding data with previously published data from transcription factor deletion studies revealed the high complexity of the transcriptional regulatory network for lipid metabolism in yeast, which involves the combinatorial and complementary regulation by multiple transcription factors. We anticipate that our work will provide insights into transcription factor binding dynamics that will prove useful for the understanding of transcription regulatory networks.
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| 9. |
- Bontemps, Zelia, et al.
(författare)
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Stochastic and deterministic assembly processes of microbial communities in relation to natural attenuation of black stains in Lascaux Cave
- 2024
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Ingår i: mSystems. - : American Society for Microbiology. - 2379-5077. ; 9:2
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Tidskriftsartikel (refereegranskat)abstract
- Community assembly processes are complex and understanding them represents a challenge in microbial ecology. Here, we used Lascaux Cave as a stable, confined environment to quantify the importance of stochastic vs deterministic processes during microbial community dynamics across the three domains of life in relation to an anthropogenic disturbance that had resulted in the side-by-side occurrence of a resistant community (unstained limestone), an impacted community (present in black stains), and a resilient community (attenuated stains). Metabarcoding data showed that the microbial communities of attenuated stains, black stains, and unstained surfaces differed, with attenuated stains being in an intermediate position. We found four scenarios to explain community response to disturbance in stable conditions for the three domains of life. Specifically, we proposed the existence of a fourth, not-documented yet scenario that concerns the always-rare microbial taxa, where stochastic processes predominate even after disturbance but are replaced by deterministic processes during post-disturbance recovery. This suggests a major role of always-rare taxa in resilience, perhaps because they might provide key functions required for ecosystem recovery.IMPORTANCEThe importance of stochastic vs deterministic processes in cave microbial ecology has been a neglected topic so far, and this work provided an opportunity to do so in a context related to the dynamics of black-stain alterations in Lascaux, a UNESCO Paleolithic cave. Of particular significance was the discovery of a novel scenario for always-rare microbial taxa in relation to disturbance, in which stochastic processes are replaced later by deterministic processes during post-disturbance recovery, i.e., during attenuation of black stains. The importance of stochastic vs deterministic processes in cave microbial ecology has been a neglected topic so far, and this work provided an opportunity to do so in a context related to the dynamics of black-stain alterations in Lascaux, a UNESCO Paleolithic cave. Of particular significance was the discovery of a novel scenario for always-rare microbial taxa in relation to disturbance, in which stochastic processes are replaced later by deterministic processes during post-disturbance recovery, i.e., during attenuation of black stains.
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| 10. |
- Borchert, Erik, et al.
(författare)
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Deciphering a Marine Bone-Degrading Microbiome Reveals a Complex Community Effort
- 2021
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Ingår i: mSystems. - 2379-5077 .- 2379-5077. ; 6:1
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Tidskriftsartikel (refereegranskat)abstract
- The marine bone biome is a complex assemblage of macro- and microor- ganisms; however, the enzymatic repertoire to access bone-derived nutrients remains unknown. The bone matrix is a composite material made up mainly of organic colla- gen and inorganic hydroxyapatite. We conducted field experiments to study microbial assemblages that can use organic bone components as nutrient source. Bovine and turkey bones were deposited at 69 m depth in a Norwegian fjord (Byfjorden, Bergen). Metagenomic sequence analysis was used to assess the functional potential of micro- bial assemblages from bone surface and the bone-eating worm Osedax mucofloris, which is a frequent colonizer of whale falls and known to degrade bone. The bone microbiome displayed a surprising taxonomic diversity revealed by the examination of 59 high-quality metagenome-assembled genomes from at least 23 bacterial families. Over 700 genes encoding enzymes from 12 relevant enzymatic families pertaining to collagenases, peptidases, and glycosidases putatively involved in bone degradation were identified. Metagenome-assembled genomes (MAGs) of the class Bacteroidia con- tained the most diverse gene repertoires. We postulate that demineralization of inor- ganic bone components is achieved by a timely succession of a closed sulfur biogeo- chemical cycle between sulfur-oxidizing and sulfur-reducing bacteria, causing a drop in pH and subsequent enzymatic processing of organic components in the bone sur- face communities. An unusually large and novel collagen utilization gene cluster was retrieved from one genome belonging to the gammaproteobacterial genus Colwellia. IMPORTANCE Bones are an underexploited, yet potentially profitable feedstock for biotechnological advances and value chains, due to the sheer amounts of residues produced by the modern meat and poultry processing industry. In this metagenomic study, we decipher the microbial pathways and enzymes that we postulate to be involved in bone degradation in the marine environment. We here demonstrate the interplay between different bacterial community members, each supplying different enzymatic functions with the potential to cover an array of reactions relating to the degradation of bone matrix components. We identify and describe a novel gene cluster for collagen utilization, which is a key function in this unique environment. We propose that the interplay between the different microbial taxa is necessary to achieve the complex task of bone degradation in the marine environment.
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| 11. |
- Cabrol, Léa, et al.
(författare)
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Redox gradient shapes the abundance and diversity of mercury-methylating microorganisms along the water column of the Black Sea
- 2023
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Ingår i: mSystems. - : American Society for Microbiology. - 2379-5077. ; 8:4
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Tidskriftsartikel (refereegranskat)abstract
- In the global context of seawater deoxygenation triggered by climate change and anthropogenic activities, changes in redox gradients impacting biogeochemical transformations of pollutants, such as mercury, become more likely. Being the largest anoxic basin worldwide, with high concentrations of the potent neurotoxic methylmercury (MeHg), the Black Sea is an ideal natural laboratory to provide new insights about the link between dissolved oxygen concentration and hgcAB gene-carrying (hgc+) microorganisms involved in the formation of MeHg. We combined geochemical and microbial approaches to assess the effect of vertical redox gradients on abundance, diversity, and metabolic potential of hgc+ microorganisms in the Black Sea water column. The abundance of hgcA genes [congruently estimated by quantitative PCR (qPCR) and metagenomics] correlated with MeHg concentration, both maximal in the upper part of the anoxic water. Besides the predominant Desulfobacterales, hgc+ microorganisms belonged to a unique assemblage of diverse—previously underappreciated—anaerobic fermenters from Anaerolineales, Phycisphaerae (characteristic of the anoxic and sulfidic zone), Kiritimatiellales, and Bacteroidales (characteristic of the suboxic zone). The metabolic versatility of Desulfobacterota differed from strict sulfate reduction in the anoxic water to reduction of various electron acceptors in the suboxic water. Linking microbial activity and contaminant concentration in environmental studies is rare due to the complexity of biological pathways. In this study, we disentangle the role of oxygen in shaping the distribution of Hg-methylating microorganisms consistently with MeHg concentration, and we highlight their taxonomic and metabolic niche partitioning across redox gradients, improving the prediction of the response of marine communities to the expansion of oxygen-deficient zones. IMPORTANCE Methylmercury (MeHg) is a neurotoxin detected at high concentrations in certain marine ecosystems, posing a threat to human health. MeHg production is mainly mediated by hgcAB gene-carrying (hgc+) microorganisms. Oxygen is one of the main factors controlling Hg methylation; however, its effect on the diversity and ecology of hgc+ microorganisms remains unknown. Under the current context of seawater deoxygenation, mercury cycling is expected to be disturbed. Here, we show the strong effect of oxygen gradients on the distribution of potential Hg methylators. In addition, we show for the first time the significant contribution of a unique assemblage of potential fermenters from Anaerolineales, Phycisphaerae, and Kiritimatiellales to Hg methylation, stratified in different redox niches along the Black Sea gradient. Our results considerably expand the known taxonomic diversity and ecological niches prone to the formation of MeHg and contribute to better apprehend the consequences of oxygen depletion in seawater.
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| 12. |
- Casey, John R., et al.
(författare)
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Adaptive Evolution of Phosphorus Metabolism in Prochlorococcus
- 2016
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Ingår i: mSystems. - : AMER SOC MICROBIOLOGY. - 2379-5077. ; 1:6
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Tidskriftsartikel (refereegranskat)abstract
- Inorganic phosphorus is scarce in the eastern Mediterranean Sea, where the high-light-adapted ecotype HLI of the marine picocyanobacterium Prochlorococcus marinus thrives. Physiological and regulatory control of phosphorus acquisition and partitioning has been observed in HLI both in culture and in the field; however, the optimization of phosphorus metabolism and associated gains for its phosphorus-limited-growth (PLG) phenotype have not been studied. Here, we reconstructed a genome-scale metabolic network of the HLI axenic strain MED4 (iJC568), consisting of 568 metabolic genes in relation to 794 reactions involving 680 metabolites distributed in 6 subcellular locations. iJC568 was used to quantify metabolic fluxes under PLG conditions, and we observed a close correspondence between experimental and computed fluxes. We found that MED4 has minimized its dependence on intracellular phosphate, not only through drastic depletion of phosphorus-containing biomass components but also through network-wide reductions in phosphate-reaction participation and the loss of a key enzyme, succinate dehydrogenase. These alterations occur despite the stringency of having relatively few pathway redundancies and an extremely high proportion of essential metabolic genes (47%; defined as the percentage of lethal in silico gene knockouts). These strategies are examples of nutrient-controlled adaptive evolution and confer a dramatic growth rate advantage to MED4 in phosphorus-limited regions. IMPORTANCE Microbes are known to employ three basic strategies to compete for limiting elemental resources: (i) cell quotas may be adjusted by alterations to cell physiology or by substitution of a more plentiful resource, (ii) stressed cells may synthesize high-affinity transporters, and (iii) cells may access more costly sources from internal stores, by degradation, or by petitioning other microbes. In the case of phosphorus, a limiting resource in vast oceanic regions, the cosmopolitan cyanobacterium Prochlorococcus marinus thrives by adopting all three strategies and a fourth, previously unknown strategy. By generating a detailed model of its metabolism, we found that strain MED4 has evolved a way to reduce its dependence on phosphate by minimizing the number of enzymes involved in phosphate transformations, despite the stringency of nearly half of its metabolic genes being essential for survival. Relieving phosphorus limitation, both physiologically and throughout intermediate metabolism, substantially improves phosphorus-specific growth rates.
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| 13. |
- Cheng, Liqin, et al.
(författare)
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A MicroRNA Gene Panel Predicts the Vaginal Microbiota Composition
- 2021
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Ingår i: mSystems. - : American Society for Microbiology. - 2379-5077. ; 6:3
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Tidskriftsartikel (refereegranskat)abstract
- The vaginal microbiota plays an essential role in vaginal health. The vaginas of many reproductive-age women are dominated by one of the Lactobacillus species. However, the vaginas of a large number of women are characterized by the colonization of several other anaerobes. Notably, some women with the non-Lactobacillus-dominated vaginal microbiota develop bacterial vaginosis, which has been correlated with sexually transmitted infections and other adverse outcomes. However, interactions and mechanisms linking the vaginal microbiota to host response are still under investigation. There are studies suggesting a link between human microRNAs and gut microbiota, but limited analysis has been carried out on the interplay of microRNAs and vaginal microbiota. In this study, we performed a microRNA expression array profiling on 67 vaginal samples from young Swedish women. MicroRNAs were clustered into distinct groups according to vaginal microbiota composition. Interestingly, 182 microRNAs were significantly elevated in their expression in the non-Lactobacillus-dominated community, suggesting an antagonistic relationship between Lactobacillus and microRNAs. Of the elevated microRNAs, 10 microRNAs displayed excellent diagnostic potential, visualized by receiver operating characteristics analysis. We further validated our findings in 34 independent samples where expression of top microRNA candidates strongly separated the Lactobacillus-dominated community from the non-Lactobacillus-dominated community in the vaginal microbiota. Notably, the Lactobacillus crispatus-dominated community showed the most profound differential microRNA expression compared with the non-Lactobacillus-dominated community. In conclusion, we demonstrate a strong relationship between the vaginal microbiota and numerous genital microRNAs, which may facilitate a deeper mechanistic interplay in this biological niche. IMPORTANCE Vaginal microbiota is correlated with women's health, where a non-Lactobacillus-dominated community predisposes women to a higher risk of disease, including human papillomavirus (HPV). However, the molecular relationship between the vaginal microbiota and host is largely unexplored. In this study, we investigated a link between the vaginal microbiota and host microRNAs in a group of young women. We uncovered an inverse correlation of the expression of microRNAs with the abundance of Lactobacillus species in the vaginal microbiota. Particularly, the expression of microRNA miR-23a-3p and miR-130a-3p, displaying significantly elevated levels in non-Lactobacillus-dominated communities, predicted the bacterial composition of the vaginal microbiota in an independent validation group. Since targeting of microRNAs is explored in the clinical setting, our results warrant investigation of whether microRNA modulation could be used for treating vaginosis recurrence and vaginosis-related diseases. Conversely, commensal bacteria could be used for treating diseases with microRNA aberrations.
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| 14. |
- Chowdhury, Sounak, et al.
(författare)
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Streptococcus pyogenes Forms Serotype- and Local Environment-Dependent Interspecies Protein Complexes
- 2021
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Ingår i: mSystems. - 2379-5077. ; 6:5, s. 1-19
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Tidskriftsartikel (refereegranskat)abstract
- Streptococcus pyogenes is known to cause both mucosal and systemic infections in humans. In this study, we used a combination of quantitative and structural mass spectrometry techniques to determine the composition and structure of the interaction network formed between human plasma proteins and the surfaces of different S. pyogenes serotypes. Quantitative network analysis revealed that S. pyogenes forms serotype-specific interaction networks that are highly dependent on the domain arrangement of the surface-attached M protein. Subsequent structural mass spectrometry analysis and computational modeling of one of the M proteins, M28, revealed that the network structure changes across different host microenvironments. We report that M28 binds secretory IgA via two separate binding sites with high affinity in saliva. During vascular leakage mimicked by increasing plasma concentrations in saliva, the binding of secretory IgA was replaced by the binding of monomeric IgA and C4b-binding protein (C4BP). This indicates that an upsurge of C4BP in the local microenvironment due to damage to the mucosal membrane drives the binding of C4BP and monomeric IgA to M28. These results suggest that S. pyogenes has evolved to form microenvironment-dependent host-pathogen protein complexes to combat human immune surveillance during both mucosal and systemic infections. IMPORTANCE Streptococcus pyogenes (group A Streptococcus [GAS]), is a human-specific Gram-positive bacterium. Each year, the bacterium affects 700 million people globally, leading to 160,000 deaths. The clinical manifestations of S. pyogenes are diverse, ranging from mild and common infections like tonsillitis and impetigo to life-threatening systemic conditions such as sepsis and necrotizing fasciitis. S. pyogenes expresses multiple virulence factors on its surface to localize and initiate infections in humans. Among all these expressed virulence factors, the M protein is the most important antigen. In this study, we perform an in-depth characterization of the human protein interactions formed around one of the foremost human pathogens. This strategy allowed us to decipher the protein interaction networks around different S. pyogenes strains on a global scale and to compare and visualize how such interactions are mediated by M proteins.
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| 15. |
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| 16. |
- El-Shehawy, Rehab, et al.
(författare)
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Microbiota-Dependent and -Independent Production of L-Dopa in the Gut of Daphnia magna
- 2021
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Ingår i: mSystems. - 2379-5077. ; 6:6
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Tidskriftsartikel (refereegranskat)abstract
- Host-microbiome interactions are essential for the physiological and ecological performance of the host, yet these interactions are challenging to identify. Neurotransmitters are commonly implicated in these interactions, but we know very little about the mechanisms of their involvement, especially in invertebrates. Here, we report a peripheral catecholamine (CA) pathway involving the gut microbiome of the model species Daphnia magna. We demonstrate the following: (i) tyrosine hydroxylase and Dopa (3,4-dihydroxyphenylalanine) decarboxylase enzymes are present in the gut wall; (ii) Dopa decarboxylase gene is expressed in the gut by the host, and its expression follows the molt cycle peaking after ecdysis; (iii) biologically active L-Dopa, but not dopamine, is present in the gut lumen; (iv) gut bacteria produce L-Dopa in a concentration-dependent manner when provided l-tyrosine as a substrate. Impinging on gut bacteria involvement in host physiology and ecologically relevant traits, we suggest L-Dopa as a communication agent in the host-microbiome interactions in daphnids and, possibly, other crustaceans.IMPORTANCE Neurotransmitters are commonly implicated in host-microbiome communication, yet the molecular mechanisms of this communication remain largely elusive. We present novel evidence linking the gut microbiome to host development and growth via neurotransmitter L-Dopa in Daphnia, the established model species in ecology and evolution. We found that both Daphnia and its gut microbiome contribute to the synthesis of the L-Dopa in the gut. We also identified a peripheral pathway in the gut wall, with a molt stage-dependent dopamine synthesis, linking the gut microbiome to the daphnid development and growth. These findings suggest a central role of L-Dopa in the bidirectional communication between the animal host and its gut bacteria and translating into the ecologically important host traits suitable for subsequent testing of causality by experimental studies.
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| 17. |
- Ellenbogen, Jared B., et al.
(författare)
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Methylotrophy in the Mire : direct and indirect routes for methane production in thawing permafrost
- 2024
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Ingår i: mSystems. - 2379-5077. ; 9:1
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Tidskriftsartikel (refereegranskat)abstract
- While wetlands are major sources of biogenic methane (CH4), our understanding of resident microbial metabolism is incomplete, which compromises the prediction of CH4 emissions under ongoing climate change. Here, we employed genome-resolved multi-omics to expand our understanding of methanogenesis in the thawing permafrost peatland of Stordalen Mire in Arctic Sweden. In quadrupling the genomic representation of the site’s methanogens and examining their encoded metabolism, we revealed that nearly 20% of the metagenome-assembled genomes (MAGs) encoded the potential for methylotrophic methanogenesis. Further, 27% of the transcriptionally active methanogens expressed methylotrophic genes; for Methanosarcinales and Methanobacteriales MAGs, these data indicated the use of methylated oxygen compounds (e.g., methanol), while for Methanomassiliicoccales, they primarily implicated methyl sulfides and methylamines. In addition to methanogenic methylotrophy, >1,700 bacterial MAGs across 19 phyla encoded anaerobic methylotrophic potential, with expression across 12 phyla. Metabolomic analyses revealed the presence of diverse methylated compounds in the Mire, including some known methylotrophic substrates. Active methylotrophy was observed across all stages of a permafrost thaw gradient in Stordalen, with the most frozen non-methanogenic palsa found to host bacterial methylotrophy and the partially thawed bog and fully thawed fen seen to house both methanogenic and bacterial methylotrophic activities. Methanogenesis across increasing permafrost thaw is thus revised from the sole dominance of hydrogenotrophic production and the appearance of acetoclastic at full thaw to consider the co-occurrence of methylotrophy throughout. Collectively, these findings indicate that methanogenic and bacterial methylotrophy may be an important and previously underappreciated component of carbon cycling and emissions in these rapidly changing wetland habitats.
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| 18. |
- Froslev Nielsen, Jens Christian, 1987, et al.
(författare)
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Comparative Transcriptome Analysis Shows Conserved Metabolic Regulation during Production of Secondary Metabolites in Filamentous Fungi
- 2019
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Ingår i: mSystems. - 2379-5077. ; 4:2
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Tidskriftsartikel (refereegranskat)abstract
- Filamentous fungi possess great potential as sources of medicinal bioactive compounds, such as antibiotics, but efficient production is hampered by a limited understanding of how their metabolism is regulated. We investigated the metabolism of six secondary metabolite-producing fungi of the Penicillium genus during nutrient depletion in the stationary phase of batch fermentations and assessed conserved metabolic responses across species using genome-wide transcriptional profiling. A coexpression analysis revealed that expression of biosynthetic genes correlates with expression of genes associated with pathways responsible for the generation of precursor metabolites for secondary metabolism. Our results highlight the main metabolic routes for the supply of precursors for secondary metabolism and suggest that the regulation of fungal metabolism is tailored to meet the demands for secondary metabolite production. These findings can aid in identifying fungal species that are optimized for the production of specific secondary metabolites and in designing metabolic engineering strategies to develop high-yielding fungal cell factories for production of secondary metabolites. IMPORTANCE Secondary metabolites are a major source of pharmaceuticals, especially antibiotics. However, the development of efficient processes of production of secondary metabolites has proved troublesome due to a limited understanding of the metabolic regulations governing secondary metabolism. By analyzing the conservation in gene expression across secondary metabolite-producing fungal species, we identified a metabolic signature that links primary and secondary metabolism and that demonstrates that fungal metabolism is tailored for the efficient production of secondary metabolites. The insight that we provide can be used to develop high-yielding fungal cell factories that are optimized for the production of specific secondary metabolites of pharmaceutical interest.
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| 19. |
- Fu, Xi, et al.
(författare)
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Continental-Scale Microbiome Study Reveals Different Environmental Characteristics Determining Microbial Richness, Composition, and Quantity in Hotel Rooms
- 2020
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Ingår i: mSystems. - : AMER SOC MICROBIOLOGY. - 2379-5077. ; 5:3
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Tidskriftsartikel (refereegranskat)abstract
- Culture-independent microbiome surveys have been conducted in homes, hospitals, schools, kindergartens and vehicles for public transport, revealing diverse microbial distributions in built environments. However, microbiome composition and the associated environmental characteristics have not been characterized in hotel environments. We presented here the first continental-scale microbiome study of hotel rooms (n = 68) spanning Asia and Europe. Bacterial and fungal communities were described by amplicon sequencing of the 16S rRNA gene and internal transcribed spacer (ITS) region and quantitative PCR. Similar numbers of bacterial (4,344) and fungal (4,555) operational taxonomic units were identified in the same sequencing depth, but most fungal taxa showed a restricted distribution compared to bacterial taxa. Aerobic, ubiquitous bacteria dominated the hotel microbiome with compositional similarity to previous samples from building and human nasopharynx environments. The abundance of Aspergillus was negatively correlated with latitude and accounted for -80% of the total fungal load in seven low-latitude hotels. We calculated the association between hotel microbiome and 16 indoor and outdoor environmental characteristics. Fungal composition and absolute quantity showed concordant associations with the same environmental characteristics, including latitude, quality of the interior, proximity to the sea, and visible mold, while fungal richness was negatively associated with heavy traffic (95% confidence interval [CI] = -127.05 to -0.25) and wall-to-wall carpet (95% CI = -47.60 to -3.82). Bacterial compositional variation was associated with latitude, quality of the interior, and floor type, while bacterial richness was negatively associated with recent redecoration (95% CI -179.00 to -44.55) and mechanical ventilation (95% CI = -136.71 to -5.12). IMPORTANCE This is the first microbiome study to characterize the microbiome data and associated environmental characteristics in hotel environments. In this study, we found concordant variation between fungal compositional variation and absolute quantity and discordant variation between community variation/quantity and richness. Our study can be used to promote hotel hygiene standards and provide resource information for future microbiome and exposure studies associated with health effects in hotel rooms.
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| 20. |
- Garcia, Sarahi L., et al.
(författare)
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Freshwater Chlorobia Exhibit Metabolic Specialization among Cosmopolitan and Endemic Populations
- 2021
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Ingår i: mSystems. - : American Society for Microbiology. - 2379-5077. ; 6:3
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Tidskriftsartikel (refereegranskat)abstract
- Photosynthetic bacteria from the class Chlorobia (formerly phylum Chlorobi) sustain carbon fixation in anoxic water columns. They harvest light at extremely low intensities and use various inorganic electron donors to fix carbon dioxide into biomass. Until now, most information on the functional ecology and local adaptations of Chlorobia members came from isolates and merely 26 sequenced genomes that may not adequately represent natural populations. To address these limitations, we analyzed global metagenomes to profile planktonic Chlorobia cells from the oxyclines of 42 freshwater bodies, spanning subarctic to tropical regions and encompassing all four seasons. We assembled and compiled over 500 genomes, including metagenome-assembled genomes (MAGs), single-amplified genomes (SAGs), and reference genomes from cultures, clustering them into 71 metagenomic operational taxonomic units (mOTUs or “species”). Of the 71 mOTUs, 57 were classified within the genus Chlorobium, and these mOTUs represented up to ∼60% of the microbial communities in the sampled anoxic waters. Several Chlorobium-associated mOTUs were globally distributed, whereas others were endemic to individual lakes. Although most clades encoded the ability to oxidize hydrogen, many lacked genes for the oxidation of specific sulfur and iron substrates. Surprisingly, one globally distributed Scandinavian clade encoded the ability to oxidize hydrogen, sulfur, and iron, suggesting that metabolic versatility facilitated such widespread colonization. Overall, these findings provide new insight into the biogeography of the Chlorobia and the metabolic traits that facilitate niche specialization within lake ecosystems.
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| 21. |
- Getz, Eric W., et al.
(författare)
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The AEGEAN-169 clade of bacterioplankton is synonymous with SAR11 subclade V (HIMB59) and metabolically distinct
- 2023
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Ingår i: mSystems. - : American Society for Microbiology. - 2379-5077. ; 8:3
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Tidskriftsartikel (refereegranskat)abstract
- Bacterioplankton of the SAR11 clade are the most abundant marine microorganisms and consist of numerous subclades spanning order-level divergence (Pelagibacterales). The assignment of the earliest diverging subclade V (a.k.a. HIMB59) to the Pelagibacterales is highly controversial, with multiple recent phylogenetic studies placing them completely separate from SAR11. Other than through phylogenomics, subclade V has not received detailed examination due to limited genomes from this group. Here, we assessed the ecogenomic characteristics of subclade V to better understand the role of this group in comparison to the Pelagibacterales. We used a new isolate genome, recently released single-amplified genomes and metagenome-assembled genomes, and previously established SAR11 genomes to perform a comprehensive comparative genomics analysis. We paired this analysis with the recruitment of metagenomes spanning the open ocean, coastal, and brackish systems. Phylogenomics, average amino acid identity, and 16S rRNA gene phylogeny indicate that SAR11 subclade V is synonymous with the ubiquitous AEGEAN-169 clade and support the contention that this group represents a taxonomic family. AEGEAN-169 shared many bulk genome qualities with SAR11, such as streamlining and low GC content, but genomes were generally larger. AEGEAN-169 had overlapping distributions with SAR11 but was metabolically distinct from SAR11 in its potential to transport and utilize a broader range of sugars as well as in the transport of trace metals and thiamin. Thus, regardless of the ultimate phylogenetic placement of AEGEAN-169, these organisms have distinct metabolic capacities that likely allow them to differentiate their niche from canonical SAR11 taxa.
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| 22. |
- Hamilton, Joshua J., et al.
(författare)
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Metabolic Network Analysis and Metatranscriptomics Reveal Auxotrophies and Nutrient Sources of the Cosmopolitan Freshwater Microbial Lineage acI
- 2017
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Ingår i: mSystems. - 2379-5077. ; 2:4
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Tidskriftsartikel (refereegranskat)abstract
- An explosion in the number of available genome sequences obtained through metagenomics and single-cell genomics has enabled a new view of the diversity of microbial life, yet we know surprisingly little about how microbes interact with each other or their environment. In fact, the majority of microbial species remain uncultivated, while our perception of their ecological niches is based on reconstruction of their metabolic potential. In this work, we demonstrate how the “seed set framework,†which computes the set of compounds that an organism must acquire from its environment (E. Borenstein, M. Kupiec, M. W. Feldman, and E. Ruppin, Proc Natl Acad Sci U S A 105:14482–14487, 2008, https://doi.org/10.1073/pnas.0806162105 ), enables computational analysis of metabolic reconstructions while providing new insights into a microbe’s metabolic capabilities, such as nutrient use and auxotrophies. We apply this framework to members of the ubiquitous freshwater actinobacterial lineage acI, confirming and extending previous experimental and genomic observations implying that acI bacteria are heterotrophs reliant on peptides and saccharides. We also present the first metatranscriptomic study of the acI lineage, revealing high expression of transport proteins and the light-harvesting protein actinorhodopsin. Putative transport proteins complement predictions of nutrients and essential metabolites while providing additional support of the hypothesis that members of the acI are photoheterotrophs.
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| 23. |
- Hossein Khademi, S. M., et al.
(författare)
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Genomic and phenotypic evolution of achromobacter xylosoxidans during chronic airway infections of patients with cystic fibrosis
- 2021
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Ingår i: mSystems. - 2379-5077. ; 6:3
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Tidskriftsartikel (refereegranskat)abstract
- Bacterial pathogens evolve during chronic colonization of the human host by selection for pathoadaptive mutations. One of the emerging and understudied bacterial species causing chronic airway infections in patients with cystic fibrosis (CF) is Achromobacter xylosoxidans. It can establish chronic infections in patients with CF, but the genetic and phenotypic changes associated with adaptation during these infections are not completely understood. In this study, we analyzed the wholegenome sequences of 55 clinical A. xylosoxidans isolates longitudinally collected from the sputum of 6 patients with CF. Four genes encoding regulatory proteins and two intergenic regions showed convergent evolution, likely driven by positive selection for pathoadaptive mutations, across the different clones of A. xylosoxidans. Most of the evolved isolates had lower swimming motility and were resistant to multiple classes of antibiotics, while fewer of the evolved isolates had slower growth or higher biofilm production than the first isolates. Using a genome-wide association study method, we identified several putative genetic determinants of biofilm formation, motility and b-lactam resistance in this pathogen. With respect to antibiotic resistance, we discovered that a combination of mutations in pathoadaptive genes (phoQ and bigR) and two other genes encoding regulatory proteins (spoT and cpxA) were associated with increased resistance to meropenem and ceftazidime. Altogether, our results suggest that genetic changes within regulatory loci facilitate within-host adaptation of A. xylosoxidans and the emergence of adaptive phenotypes, such as antibiotic resistance or biofilm formation. IMPORTANCE A thorough understanding of bacterial pathogen adaptation is essential for the treatment of chronic bacterial infections. One unique challenge in the analysis and interpretation of genomics data is identifying the functional impact of mutations accumulated in the bacterial genome during colonization in the human host. Here, we investigated the genomic and phenotypic evolution of A. xylosoxidans in chronic airway infections of patients with CF and identified several mutations associated with the phenotypic evolution of this pathogen using genome-wide associations. Identification of phenotypes under positive selection and the associated mutations can enlighten the adaptive processes of this emerging pathogen in human infections and pave the way for novel therapeutic interventions.
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| 24. |
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| 25. |
- Hubalek, Valerie, 1981-, et al.
(författare)
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Vitamin and Amino Acid Auxotrophy in Anaerobic Consortia Operating under Methanogenic Conditions
- 2017
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Ingår i: mSystems. - 2379-5077. ; 2:5
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Tidskriftsartikel (refereegranskat)abstract
- Syntrophy among Archaea and Bacteria facilitates the anaerobic degra- dation of organic compounds to CH4 and CO2 . Particularly during aliphatic and aro- matic hydrocarbon mineralization, as in the case of crude oil reservoirs and petroleum-contaminated sediments, metabolic interactions between obligate mutu- alistic microbial partners are of central importance. Using micromanipulation com- bined with shotgun metagenomic approaches, we describe the genomes of complex consortia within short-chain alkane-degrading cultures operating under methano- genic conditions. Metabolic reconstruction revealed that only a small fraction of genes in the metagenome-assembled genomes encode the capacity for fermenta- tion of alkanes facilitated by energy conservation linked to H2 metabolism. Instead, the presence of inferred lifestyles based on scavenging anabolic products and inter- mediate fermentation products derived from detrital biomass was a common fea- ture. Additionally, inferred auxotrophy for vitamins and amino acids suggests that the hydrocarbon-degrading microbial assemblages are structured and maintained by multiple interactions beyond the canonical H2 -producing and syntrophic alkane degrader-methanogen partnership. Compared to previous work, our report points to a higher order of complexity in microbial consortia engaged in anaerobic hydrocar- bon transformation. IMPORTANCE
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