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© 2015 Taketani et al.The capsule from Bacteroides, a common gut symbiont, has long been a model system for studying the molecular mechanisms of host-symbiont interactions. The Bacteroides capsule is thought to consist of an array of phase-variable polysac

Background Fiberoptic bronchoscopy (FOB) is used to directly visualize the lumen of the trachea, as well as proximal and distal airways. It can be used to diagnose or treat abnormalities within or adjacent to these airways. However, it is an invasive procedure that may lead to several complications. Minimum-intensity projection (MinIP) in high-resolution computed tomography (CT) of the lungs is particularly useful because multiple lung diseases may present with reduced CT attenuation values. Patients and methods In this study, we included 115 patients admitted to the Chest Department, Mansoura University, Egypt. All patients were subjected to complete history taking, thorough physical examination, routine laboratory investigations, in addition to chest radiograph. CT and FOB were ordered for all cases. Results CT examination of the studied cases revealed mass in 43.5%, delayed resolved pneumonia in 30.4%, and collapse in 13% of cases. Furthermore, either of cavity, narrowing, or collapse was present in 4.3% of cases. Lesions were detected in the left lung in 43.5%, right lung in was 30.4%, and both lungs in 13% of cases. The sensitivity and specificity of MinIP was 69.4 and 100%, respectively, compared with FOB. The positive predictive value was 100%, whereas the negative predictive value was 71.4%, with an accuracy of 82.6%. The sensitivity of MinIP in the diagnosis was different according to the nature of the lesion. We found significant differences when comparing conventional CT with MinIP, especially in collapse, nodules, delayed resolved pneumonia, cavity, and narrowing. However, such a significant difference was not detected in the diagnosis of mass lesions. Conclusion We conclude that MinIP CT is a very important technique for detection of endobronchial lesion compared with conventional CT, and we recommend use of this technique when an endobronchial lesion is suspected.

The modular architecture of nonribosomal peptide synthetases (NRPSs) has inspired efforts to study their evolution and engineering. In this study, we analyze in detail a unique family of NRPSs from the defensive intracellular bacterial symbiont, Candidatus Endobryopsis kahalalidifaciens ( Ca. E. kahalalidifaciens). We show that intensive and indiscriminate recombination events erase trivial sequence covariations induced by phylogenetic relatedness, revealing nonmodular functional constraints and clear recombination units. Moreover, we reveal unique substrate specificity determinants for multiple enzymatic domains, allowing us to accurately predict and experimentally discover the products of an orphan NRPS in Ca. E. kahalalidifaciens directly from environmental samples of its algal host. Finally, we expanded our analysis to 1,531 diverse NRPS pathways and revealed similar functional constraints to those observed in Ca. E. kahalalidifaciens' NRPSs. Our findings reveal the sequence bases of genetic exchange, functional constraints, and substrate specificity in Ca. E. kahalalidifaciens' NRPSs, and highlight them as a uniquely primed system for diversifying evolution., (© The Author(s) 2024. Published by Oxford University Press on behalf of National Academy of Sciences.)

Marsupial neonates are born with immature immune systems, making them vulnerable to pathogens. While neonates receive maternal protection, they can also independently combat pathogens, though the mechanisms remain unknown. Using the sugar glider (Petaurus breviceps) as a model, we investigated immunological defense strategies of marsupial neonates. Cathelicidins, a family of antimicrobial peptides expanded in the genomes of marsupials, are highly expressed in developing neutrophils. Sugar glider cathelicidins reside in two genomic clusters and their coordinated expression is achieved by enhancer sharing within clusters and long-range physical interactions between clusters. These cathelicidins modulate immune responses and have potent antimicrobial effects, sufficient to provide protection in a mouse model of sepsis. Lastly, cathelicidins have a complex evolutionary history, where marsupials and monotremes are the only tetrapods that retained two cathelicidin clusters. Thus, cathelicidins are critical mediators of marsupial immunity, and their evolution reflects the life history-specific immunological needs of these animals.

Background: Alveolar periosteoplasty during primary repair of cleft lip is still a topic of debate due to the fear exacerbating maxillary retrusion. The authors present their experience with early closure of alveolar clefts. The study aims to analyze gingivoperiosteoplasty (GPP) by comparing the use of locoregional flaps versus distant grafts for reconstruction of the lower layer of the primary palate cleft., Methods: Seventeen infants underwent 22 alveolar cleft repairs. After nasal floor repair by nasal mucoperiosteum, the oral layer was repaired by maxillary or gingival periosteal flaps in seven patients with alveolar clefts less than or equal to 6 mm wide, and tibial or pericranial periosteal grafts in 10 patients with wider clefts. At teenage years, crossbites in three flap-GPP and three graft-GPP patients were compared with nine older adolescents without primary GPP., Results: Alveolar clefts were perfectly sealed. Radiographs during the early 3 postoperative years showed new bone formation more posteriorly extended in patients who underwent graft-GPP. Teeth eruption and alveolar rigidity at the mixed dentition age eliminated the need for secondary bone grafting. The anterior crossbites in adolescent patients ranged between -2 and -14 mm; crossing was relatively smaller in patients with a younger age and without cleft palate. The mean crossbite was 7.2 mm in the six teenagers and 9.6 mm in the control cases., Conclusions: Graft-GPP may be a good alternative to flap-GPP, particularly for wide alveolar cleft repair. Maxillary retrusion is aggravated in patients with cleft palate and older age at assessment. GPP may not increase crossbite., Competing Interests: The authors have no financial interest to declare in relation to the content of this article., (Copyright © 2024 The Authors. Published by Wolters Kluwer Health, Inc. on behalf of The American Society of Plastic Surgeons.)

Phragmanthera austroarabica (Loranthaceae), a semi-parasitic plant, is well known for its high content of polyphenols that are responsible for its antioxidant and anti-inflammatory activities. Gallic acid, catechin, and methyl gallate are bioactive metabolites of common occurrence in the family of Loranthaceae. Herein, the concentrations of these bioactive metabolites were assessed using high-performance thin layer chromatography (HPTLC). Methyl gallate, catechin, and gallic acid were scanned at 280 nm. Their concentrations were assessed as 14.5, 6.5 and 43.6 mg/g of plant dry extract, respectively. Phragmanthera austroarabica extract as well as the three pure compounds were evaluated regarding the cytotoxic activity. The plant extract exhibited promising cytotoxic activity against MDA-MB-231 breast cells with the IC 50 value of 19.8 μg/mL while the tested pure compounds displayed IC 50 values in the range of 21.26-29.6 μg/mL. For apoptosis investigation, P. austroarabica induced apoptotic cell death by 111-fold change and necrosis by 9.31-fold change. It also activated the proapoptotic genes markers and inhibited the antiapoptotic gene, validating the apoptosis mechanism. Moreover, in vivo studies revealed a significant reduction in the breast tumor volume and weight in solid Ehrlich carcinoma (SEC) mice. The treatment of SEC mice with P. austroarabica extract improved both hematological and biochemical parameters with amelioration in the liver and kidney histopathology to near normal. Taken together, P. austroarabica extract exhibited promising anti-cancer activity through an apoptosis-induction.

Great progress has been made in understanding gut microbiomes' products and their effects on health and disease. Less attention, however, has been given to the inputs that gut bacteria consume. Here, we quantitatively examine inputs and outputs of the mouse gut microbiome, using isotope tracing. The main input to microbial carbohydrate fermentation is dietary fiber and to branched-chain fatty acids and aromatic metabolites is dietary protein. In addition, circulating host lactate, 3-hydroxybutyrate, and urea (but not glucose or amino acids) feed the gut microbiome. To determine the nutrient preferences across bacteria, we traced into genus-specific bacterial protein sequences. We found systematic differences in nutrient use: most genera in the phylum Firmicutes prefer dietary protein, Bacteroides dietary fiber, and Akkermansia circulating host lactate. Such preferences correlate with microbiome composition changes in response to dietary modifications. Thus, diet shapes the microbiome by promoting the growth of bacteria that preferentially use the ingested nutrients., Competing Interests: Declaration of interests J.D.R. is a member of the Rutgers Cancer Institute of New Jersey and the University of Pennsylvania Diabetes Research Center; a co-founder and stockholder in Empress Therapeutics and Serien Therapeutics; and an advisor and stockholder in Agios Pharmaceuticals, Bantam Pharmaceuticals, Colorado Research Partners, Rafael Pharmaceuticals, Barer Institute, and L.E.A.F. Pharmaceuticals. M.S.D. is a member of the scientific advisory boards of DeepBiome Therapeutics and VastBiome., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Since the discovery of the kahalalide family of marine depsipeptides in 1993, considerable work has been done to develop these compounds as new and biologically distinct anti-cancer agents. Clinical trials and laboratory research have yielded a wealth of data that indicates tolerance of kahalalides in healthy cells and selective activity against diseased cells. Currently, two molecules have attracted the greates level of attention, kahalalide F (KF) and isokahalalide F (isoKF, Irvalec, PM 02734, elisidepsin). Both compounds were originally isolated from the sarcoglossan mollusk Elysia rufescens but due to distinct structural characteristics it has been hypothesized and recently shown that the ultimate origin of the molecules is microbial. The search for their true source has been a subject of considerable research in the anticipation of finding new analogs and a culturable expression system that can produce sufficient material through fermentation to be industrially relevant., (Copyright © 2022. Published by Elsevier Masson SAS.)

The human microbiome encodes a large repertoire of biochemical enzymes and pathways, most of which remain uncharacterized. Here, using a metagenomics-based search strategy, we discovered that bacterial members of the human gut and oral microbiome encode enzymes that selectively phosphorylate a clinically used antidiabetic drug, acarbose 1,2 , resulting in its inactivation. Acarbose is an inhibitor of both human and bacterial α-glucosidases 3 , limiting the ability of the target organism to metabolize complex carbohydrates. Using biochemical assays, X-ray crystallography and metagenomic analyses, we show that microbiome-derived acarbose kinases are specific for acarbose, provide their harbouring organism with a protective advantage against the activity of acarbose, and are widespread in the microbiomes of western and non-western human populations. These results provide an example of widespread microbiome resistance to a non-antibiotic drug, and suggest that acarbose resistance has disseminated in the human microbiome as a defensive strategy against a potential endogenous producer of a closely related molecule., (© 2021. The Author(s), under exclusive licence to Springer Nature Limited.)

Plasmids are autonomous genetic elements that can be exchanged between microorganisms via horizontal gene transfer (HGT). Despite the central role they play in antibiotic resistance and modern biotechnology, our understanding of plasmids' natural ecology is limited. Recent experiments have shown that plasmids can spread even when they are a burden to the cell, suggesting that natural plasmids may exist as parasites. Here, we use mathematical modeling to explore the ecology of such parasitic plasmids. We first develop models of single plasmids and find that a plasmid's population dynamics and optimal infection strategy are strongly determined by the plasmid's HGT mechanism. We then analyze models of co-infecting plasmids and show that parasitic plasmids are prone to a "tragedy of the commons" in which runaway plasmid invasion severely reduces host fitness. We propose that this tragedy of the commons is averted by selection between competing populations and demonstrate this effect in a metapopulation model. We derive predicted distributions of unique plasmid types in genomes-comparison to the distribution of plasmids in a collection of 17,725 genomes supports a model of parasitic plasmids with positive plasmid-plasmid interactions that ameliorate plasmid fitness costs or promote the invasion of new plasmids., (© 2021. The Author(s).)

Pseudomonas aeruginosa is a significant threat in both healthcare and industrial biofouling. Surface attachment of P. aeruginosa is particularly problematic as surface association induces virulence and is necessary for the ensuing process of biofilm formation, which hampers antibiotic treatments. Previous efforts have searched for dispersal agents of mature biofilm collectives, but there are no known factors that specifically disperse individual surface-attached P. aeruginosa. In this study, we develop a quantitative single-cell surface-dispersal assay and use it to show that P. aeruginosa itself produces factors that can stimulate its dispersal. Through bioactivity-guided fractionation, mass spectrometry, and nuclear magnetic resonance, we elucidated the structure of one such factor, 2-methyl-4-hydroxyquinoline (MHQ). MHQ is an alkyl quinolone with a previously unknown activity and is synthesized by the PqsABC enzymes. Pure MHQ is sufficient to disperse P. aeruginosa, but the dispersal activity of natural P. aeruginosa conditioned media requires additional factors. Whereas other alkyl quinolones have been shown to act as antibiotics or membrane depolarizers, MHQ lacks these activities and known antibiotics do not induce dispersal. In contrast, we show that MHQ inhibits the activity of Type IV Pili (TFP) and that TFP targeting can explain its dispersal activity. Our work thus identifies single-cell surface dispersal as a new activity of P. aeruginosa-produced small molecules, characterizes MHQ as a promising dispersal agent, and establishes TFP inhibition as a viable mechanism for P. aeruginosa dispersal., Competing Interests: Conflict of interest A provisional patent describing MHQ as a potential therapeutic in surface dispersal has been filed., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Microbial natural products constitute a wide variety of chemical compounds, many which can have antibiotic, antiviral, or anticancer properties that make them interesting for clinical purposes. Natural product classes include polyketides (PKs), nonribosomal peptides (NRPs), and ribosomally synthesized and post-translationally modified peptides (RiPPs). While variants of biosynthetic gene clusters (BGCs) for known classes of natural products are easy to identify in genome sequences, BGCs for new compound classes escape attention. In particular, evidence is accumulating that for RiPPs, subclasses known thus far may only represent the tip of an iceberg. Here, we present decRiPPter (Data-driven Exploratory Class-independent RiPP TrackER), a RiPP genome mining algorithm aimed at the discovery of novel RiPP classes. DecRiPPter combines a Support Vector Machine (SVM) that identifies candidate RiPP precursors with pan-genomic analyses to identify which of these are encoded within operon-like structures that are part of the accessory genome of a genus. Subsequently, it prioritizes such regions based on the presence of new enzymology and based on patterns of gene cluster and precursor peptide conservation across species. We then applied decRiPPter to mine 1,295 Streptomyces genomes, which led to the identification of 42 new candidate RiPP families that could not be found by existing programs. One of these was studied further and elucidated as a representative of a novel subfamily of lanthipeptides, which we designate class V. The 2D structure of the new RiPP, which we name pristinin A3 (1), was solved using nuclear magnetic resonance (NMR), tandem mass spectrometry (MS/MS) data, and chemical labeling. Two previously unidentified modifying enzymes are proposed to create the hallmark lanthionine bridges. Taken together, our work highlights how novel natural product families can be discovered by methods going beyond sequence similarity searches to integrate multiple pathway discovery criteria., Competing Interests: I have read the journal's policy and the authors of this manuscript have the following competing interests: P.C. is currently an employee of Verily Life Sciences. M.H. is currently an employee of LifeMine Therapeutics. M.S.D. is a member of the Scientific Advisory Board of DeepBiome Therapeutics. M.A.F. is a cofounder and director of Federation Bio. M.H.M. is on the scientific advisory board of Hexagon Bio and co-founder of Design Pharmaceuticals.

The human gut microbiome harbors hundreds of bacterial species with diverse biochemical capabilities. Dozens of drugs have been shown to be metabolized by single isolates from the gut microbiome, but the extent of this phenomenon is rarely explored in the context of microbial communities. Here, we develop a quantitative experimental framework for mapping the ability of the human gut microbiome to metabolize small molecule drugs: Microbiome-Derived Metabolism (MDM)-Screen. Included are a batch culturing system for sustained growth of subject-specific gut microbial communities, an ex vivo drug metabolism screen, and targeted and untargeted functional metagenomic screens to identify microbiome-encoded genes responsible for specific metabolic events. Our framework identifies novel drug-microbiome interactions that vary between individuals and demonstrates how the gut microbiome might be used in drug development and personalized medicine., Competing Interests: Declaration of Interests M.S.D. is a member of the scientific advisory board of DeepBiome Therapeutics. A patent is being filed by Princeton University for the use of quantitative MDM-Screen to measure inter-individual variability in drug metabolism., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Consumption of glucosinolates, pro-drug-like metabolites abundant in Brassica vegetables, has been associated with decreased risk of certain cancers. Gut microbiota have the ability to metabolize glucosinolates, generating chemopreventive isothiocyanates. Here, we identify a genetic and biochemical basis for activation of glucosinolates to isothiocyanates by Bacteroides thetaiotaomicron, a prominent gut commensal species. Using a genome-wide transposon insertion screen, we identified an operon required for glucosinolate metabolism in B. thetaiotaomicron. Expression of BT2159-BT2156 in a non-metabolizing relative, Bacteroides fragilis, resulted in gain of glucosinolate metabolism. We show that isothiocyanate formation requires the action of BT2158 and either BT2156 or BT2157 in vitro. Monocolonization of mice with mutant BtΔ2157 showed reduced isothiocyanate production in the gastrointestinal tract. These data provide insight into the mechanisms by which a common gut bacterium processes an important dietary nutrient., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Thousands of natural products have been identified from cultured microorganisms, yet evidence of their production in the environment has proven elusive. Technological advances in mass spectrometry, combined with public databases, now make it possible to address this disparity by detecting compounds directly from environmental samples. Here, we used adsorbent resins, tandem mass spectrometry, and next-generation sequencing to assess the metabolome of marine sediments and its relationship to bacterial community structure. We identified natural products previously reported from cultured bacteria, providing evidence they are produced in situ , and compounds of anthropogenic origin, suggesting this approach can be used as an indicator of environmental impact. The bacterial metabolite staurosporine was quantified and shown to reach physiologically relevant concentrations, indicating that it may influence sediment community structure. Staurosporine concentrations were correlated with the relative abundance of the staurosporine-producing bacterial genus Salinispora and production confirmed in strains cultured from the same location, providing a link between compound and candidate producer. Metagenomic analyses revealed numerous biosynthetic gene clusters related to indolocarbazole biosynthesis, providing evidence for noncanonical sources of staurosporine and a path forward to assess the relationships between natural products and the organisms that produce them. Untargeted environmental metabolomics circumvents the need for laboratory cultivation and represents a promising approach to understanding the functional roles of natural products in shaping microbial community structure in marine sediments. IMPORTANCE Natural products are readily isolated from cultured bacteria and exploited for useful purposes, including drug discovery. However, these compounds are rarely detected in the environments from which the bacteria are obtained, thus limiting our understanding of their ecological significance. Here, we used environmental metabolomics to directly assess chemical diversity in marine sediments. We identified numerous metabolites and, in one case, isolated strains of bacteria capable of producing one of the compounds detected. Coupling environmental metabolomics with community and metagenomic analyses provides opportunities to link compounds and producers and begin to assess the complex interactions mediated by specialized metabolites in marine sediments., (Copyright © 2019 American Society for Microbiology.)

Brine, the historically known food additive salt solution, has been widely used as a pickling media to preserve flavor or enhance food aroma, appearance, or other qualities. The influence of pickling, using brine, on the aroma compounds and the primary and secondary metabolite profile in onion bulb Allium cepa red cv. and lemon fruit Citrus limon was evaluated using multiplex metabolomics technologies. In lemon, pickling negatively affected its key odor compound "citral", whereas monoterpene hydrocarbons limonene and γ-terpinene increased in the pickled product. Meanwhile, in onion sulphur rearrangement products appeared upon storage, i.e., 3,5-diethyl-1,2,4-trithiolane. Profiling of the polar secondary metabolites in lemon fruit via ultra-performance liquid chromatography coupled to MS annotated 37 metabolites including 18 flavonoids, nine coumarins, five limonoids, and two organic acids. With regard to pickling impact, notable and clear separation among specimens was observed with an orthogonal projections to least squares-discriminant analysis (OPLS-DA) score plot for the lemon fruit model showing an enrichment of limonoids and organic acids and that for fresh onion bulb showing an abundance of flavonols and saponins. In general, the pickling process appeared to negatively impact the abundance of secondary metabolites in both onion and lemon, suggesting a decrease in their food health benefits.

The gut microbiota modulate host biology in numerous ways, but little is known about the molecular mediators of these interactions. Previously, we found a widely distributed family of nonribosomal peptide synthetase gene clusters in gut bacteria. Here, by expressing a subset of these clusters in Escherichia coli or Bacillus subtilis, we show that they encode pyrazinones and dihydropyrazinones. At least one of the 47 clusters is present in 88% of the National Institutes of Health Human Microbiome Project (NIH HMP) stool samples, and they are transcribed under conditions of host colonization. We present evidence that the active form of these molecules is the initially released peptide aldehyde, which bears potent protease inhibitory activity and selectively targets a subset of cathepsins in human cell proteomes. Our findings show that an approach combining bioinformatics, synthetic biology, and heterologous gene cluster expression can rapidly expand our knowledge of the metabolic potential of the microbiota while avoiding the challenges of cultivating fastidious commensals., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Unlabelled: The capsule from Bacteroides, a common gut symbiont, has long been a model system for studying the molecular mechanisms of host-symbiont interactions. The Bacteroides capsule is thought to consist of an array of phase-variable polysaccharides that give rise to subpopulations with distinct cell surface structures. Here, we report the serendipitous discovery of a previously unknown surface structure in Bacteroides thetaiotaomicron: a surface layer composed of a protein of unknown function, BT1927. BT1927, which is expressed in a phase-variable manner by ~1:1,000 cells in a wild-type culture, forms a hexagonally tessellated surface layer. The BT1927-expressing subpopulation is profoundly resistant to complement-mediated killing, due in part to the BT1927-mediated blockade of C3b deposition. Our results show that the Bacteroides surface structure is capable of a far greater degree of structural variation than previously known, and they suggest that structural variation within a Bacteroides species is important for productive gut colonization., Importance: Many bacterial species elaborate a capsule, a structure that resides outside the cell wall and mediates microbe-microbe and microbe-host interactions. Species of Bacteroides, the most abundant genus in the human gut, produce a capsule that consists of an array of polysaccharides, some of which are known to mediate interactions with the host immune system. Here, we report the discovery of a previously unknown surface structure in Bacteroides thetaiotaomicron. We show that this protein-based structure is expressed by a subset of cells in a population and protects Bacteroides from killing by complement, a component of the innate immune system. This novel surface layer protein is conserved across many species of the genus Bacteroides, suggesting an important role in colonization and host immune modulation., (Copyright © 2015 Taketani et al.)

Genetic tools to engineer a prominent member of the human gut microbiome represent initial steps toward cell-based diagnostics and therapeutics., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Several recent studies describe the influence of the gut microbiota on host brain and behavior. However, the mechanisms responsible for microbiota-nervous system interactions are largely unknown. Using a combination of genetics, biochemistry, and crystallography, we identify and characterize two phylogenetically distinct enzymes found in the human microbiome that decarboxylate tryptophan to form the β-arylamine neurotransmitter tryptamine. Although this enzymatic activity is exceedingly rare among bacteria more broadly, analysis of the Human Microbiome Project data demonstrate that at least 10% of the human population harbors at least one bacterium encoding a tryptophan decarboxylase in their gut community. Our results uncover a previously unrecognized enzymatic activity that can give rise to host-modulatory compounds and suggests a potential direct mechanism by which gut microbiota can influence host physiology, including behavior., (Copyright © 2014 Elsevier Inc. All rights reserved.)

In complex biological systems, small molecules often mediate microbe-microbe and microbe-host interactions. Using a systematic approach, we identified 3,118 small-molecule biosynthetic gene clusters (BGCs) in genomes of human-associated bacteria and studied their representation in 752 metagenomic samples from the NIH Human Microbiome Project. Remarkably, we discovered that BGCs for a class of antibiotics in clinical trials, thiopeptides, are widely distributed in genomes and metagenomes of the human microbiota. We purified and solved the structure of a thiopeptide antibiotic, lactocillin, from a prominent member of the vaginal microbiota. We demonstrate that lactocillin has potent antibacterial activity against a range of Gram-positive vaginal pathogens, and we show that lactocillin and other thiopeptide BGCs are expressed in vivo by analyzing human metatranscriptomic sequencing data. Our findings illustrate the widespread distribution of small-molecule-encoding BGCs in the human microbiome, and they demonstrate the bacterial production of drug-like molecules in humans. PAPERCLIP:, (Copyright © 2014 Elsevier Inc. All rights reserved.)

Natural products (secondary metabolites) found in marine invertebrates are often thought to be produced by resident symbiotic bacteria, and these products appear to play a major role in the symbiotic interaction of bacteria and their hosts. In these animals, there is extensive variation, both in chemistry and in the symbiotic bacteria that produce them. Here, we sought to answer the question of what factors underlie chemical variation in the ocean. As a model, we investigated the colonial tunicate Lissoclinum patella because of its rich and varied chemistry and its broad geographic range. We sequenced mitochondrial cytochrome c oxidase 1 (COXI) genes, and found that animals classified as L. patella fall into three phylogenetic groups that may encompass several cryptic species. The presence of individual natural products followed the phylogenetic relationship of the host animals, even though the compounds are produced by symbiotic bacteria that do not follow host phylogeny. In sum, we show that cryptic populations of animals underlie the observed chemical diversity, suggesting that the host controls selection for particular secondary metabolite pathways. These results imply novel approaches to obtain chemical diversity from the oceans, and also demonstrate that the diversity of marine natural products may be greatly impacted by cryptic local extinctions.

Defining the functional status of host-associated microbial ecosystems has proven challenging owing to the vast number of predicted genes within the microbiome and relatively poor understanding of community dynamics and community-host interaction. Metabolomic approaches, in which a large number of small molecule metabolites can be defined in a biological sample, offer a promising avenue to 'fingerprint' microbiota functional status. Here, we examined the effects of the human gut microbiota on the fecal and urinary metabolome of a humanized (HUM) mouse using an optimized ultra performance liquid chromatography-mass spectrometry-based method. Differences between HUM and conventional mouse urine and fecal metabolomic profiles support host-specific aspects of the microbiota's metabolomic contribution, consistent with distinct microbial compositions. Comparison of microbiota composition and metabolome of mice humanized with different human donors revealed that the vast majority of metabolomic features observed in donor samples are produced in the corresponding HUM mice, and individual-specific features suggest 'personalized' aspects of functionality can be reconstituted in mice. Feeding the mice a defined, custom diet resulted in modification of the metabolite signatures, illustrating that host diet provides an avenue for altering gut microbiota functionality, which in turn can be monitored via metabolomics. Using a defined model microbiota consisting of one or two species, we show that simplified communities can drive major changes in the host metabolomic profile. Our results demonstrate that metabolomics constitutes a powerful avenue for functional characterization of the intestinal microbiota and its interaction with the host.

The switch from culture-based enumeration to deep sequencing has enabled microbial community composition to be profiled en masse. In a new article, Maurice et al. (2013) report the use of fluorescence-activated cell sorting (FACS) to perform a high-throughput analysis of gut microbiota community function., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Secondary metabolites are ubiquitous in bacteria, but by definition, they are thought to be nonessential. Highly toxic secondary metabolites such as patellazoles have been isolated from marine tunicates, where their exceptional potency and abundance implies a role in chemical defense, but their biological source is unknown. Here, we describe the association of the tunicate Lissoclinum patella with a symbiotic α-proteobacterium, Candidatus Endolissoclinum faulkneri, and present chemical and biological evidence that the bacterium synthesizes patellazoles. We sequenced and assembled the complete Ca. E. faulkneri genome, directly from metagenomic DNA obtained from the tunicate, where it accounted for 0.6% of sequence data. We show that the large patellazoles biosynthetic pathway is maintained, whereas the remainder of the genome is undergoing extensive streamlining to eliminate unneeded genes. The preservation of this pathway in streamlined bacteria demonstrates that secondary metabolism is an essential component of the symbiotic interaction.

The relationship between tunicates and the uncultivated cyanobacterium Prochloron didemni has long provided a model symbiosis. P. didemni is required for survival of animals such as Lissoclinum patella and also makes secondary metabolites of pharmaceutical interest. Here, we present the metagenomes, chemistry, and microbiomes of four related L. patella tunicate samples from a wide geographical range of the tropical Pacific. The remarkably similar P. didemni genomes are the most complex so far assembled from uncultivated organisms. Although P. didemni has not been stably cultivated and comprises a single strain in each sample, a complete set of metabolic genes indicates that the bacteria are likely capable of reproducing outside the host. The sequences reveal notable peculiarities of the photosynthetic apparatus and explain the basis of nutrient exchange underlying the symbiosis. P. didemni likely profoundly influences the lipid composition of the animals by synthesizing sterols and an unusual lipid with biofuel potential. In addition, L. patella also harbors a great variety of other bacterial groups that contribute nutritional and secondary metabolic products to the symbiosis. These bacteria possess an enormous genetic potential to synthesize new secondary metabolites. For example, an antitumor candidate molecule, patellazole, is not encoded in the genome of Prochloron and was linked to other bacteria from the microbiome. This study unveils the complex L. patella microbiome and its impact on primary and secondary metabolism, revealing a remarkable versatility in creating and exchanging small molecules.

Ribosomal peptide natural products are ubiquitous, yet relatively few tools exist to predict structures and clone new pathways. Cyanobactin ribosomal peptides are found in ~30% of all cyanobacteria, but the connection between gene sequence and structure was not defined, limiting the rapid identification of new compounds and pathways. Here, we report discovery of four orphan cyanobactin gene clusters by genome mining and an additional pathway by targeted cloning, which represented a tyrosine O-prenylating biosynthetic pathway. Genome mining enabled discovery of five cyanobactins, including peptide natural products from Spirulina supplements. A phylogenetic model defined four cyanobactin genotypes, which explain the synthesis of multiple cyanobactin structural classes and help direct pathway cloning and structure prediction efforts. These strategies were applied to DNA isolated from a mixed cyanobacterial bloom containing cyanobactins., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

The complex evolution of secondary metabolism is important in biology, drug development, and synthetic biology. To examine this problem at a fine scale, we compared the genomes and chemistry of 24 strains of uncultivated cyanobacteria, Prochloron didemni, that live symbiotically with tropical ascidians and that produce natural products isolated from the animals. Although several animal species were obtained along a >5500 km transect of the Pacific Ocean, P. didemni strains are >97% identical across much of their genomes, with only a few exceptions concentrated in secondary metabolism. Secondary metabolic gene clusters were sporadically present or absent in identical genomic locations with no consistent pattern of co-occurrence. Discrete mutations were observed, leading to new chemicals that we isolated from animals. Functional cassettes encoding diverse chemicals are exchanged among a single population of symbiotic P. didemni that spans the tropical Pacific, providing the host animals with a varying arsenal of secondary metabolites.

Ascidians (tunicates; sea squirts) are sources of diverse, bioactive natural products, one of which is an approved drug and many of which are potent drug leads. It has been shown that symbiotic bacteria living with ascidians produce some of the bioactive compounds isolated from whole animals, and indirect evidence strongly implicates symbiotic bacteria in the synthesis of many others. However, for the majority the producing organism has not been identified. In cases where a symbiotic origin has been definitively assigned, the resulting data lead to improved paths to drug discovery and development from marine animals. This review traces evidence for symbiotic production where such evidence exists and describes the strengths and limitations of that evidence., (Copyright © 2010 Elsevier Ltd. All rights reserved.)