@article{gentry_collins_panitchpakdi_belda-ferre_stewart_terrazas_lu_zuffa_yan_avila-pacheco_et al._2024, title={Reverse metabolomics for the discovery of chemical structures from humans}, volume={626}, ISSN={["1476-4687"]}, DOI={10.1038/s41586-023-06906-8}, abstractNote={Determining structure and phenotypic context of molecules detected in untargeted metabolomics experiments remains challenging. Here, we present reverse metabolomics as a discovery strategy, where MS/MS spectra are acquired from newly synthesized compounds and searched for in public metabolomics data to uncover phenotypic associations. To demonstrate the concept, we broadly synthesized and explored multiple classes of metabolites in humans – N-acyl amides, fatty acid esters of hydroxy fatty acids, bile acid esters and conjugated bile acids. Using repository scale analysis,15,16 we discovered that some conjugated bile acids were associated with inflammatory bowel disease (IBD). Using four distinct human IBD cohorts for validation, we found that Glu, Ile/Leu, Phe, Thr, Trp and Tyr conjugated cholic acids were elevated in Crohn’s disease. Several of these compounds and related structures were shown to affect pathways associated with inflammatory bowel disease, such as interferon-gamma production in CD4+ T cells40 and PXR agonism.41 Bacteria belonging to the Bifidobacterium, Clostridium, and Enterococcus genera were able to produce these bile amidates when cultured. Because searching repositories with MS/MS spectra has only recently become possible, reverse metabolomics approach can now be employed as a general strategy to discover other molecules from human and animal ecosystems.}, number={7998}, journal={NATURE}, author={Gentry, Emily C. and Collins, Stephanie L. and Panitchpakdi, Morgan and Belda-Ferre, Pedro and Stewart, Allison K. and Terrazas, Marvic Carrillo and Lu, Hsueh-han and Zuffa, Simone and Yan, Tingting and Avila-Pacheco, Julian and et al.}, year={2024}, month={Feb} }
 @article{foley_walker_stewart_o'flaherty_gentry_patel_beaty_allen_pan_simpson_et al._2023, title={Bile salt hydrolases shape the bile acid landscape and restrict Clostridioides difficile growth in the murine gut}, volume={3}, ISSN={["2058-5276"]}, DOI={10.1038/s41564-023-01337-7}, abstractNote={Bile acids (BAs) mediate the crosstalk between human and microbial cells and influence diseases including Clostridioides difficile infection (CDI). While bile salt hydrolases (BSHs) shape the BA pool by deconjugating conjugated BAs, the basis for their substrate selectivity and impact on C. difficile remain elusive. Here we survey the diversity of BSHs in the gut commensals Lactobacillaceae, which are commonly used as probiotics, and other members of the human gut microbiome. We structurally pinpoint a loop that predicts BSH preferences for either glycine or taurine substrates. BSHs with varying specificities were shown to restrict C. difficile spore germination and growth in vitro and colonization in pre-clinical in vivo models of CDI. Furthermore, BSHs reshape the pool of microbial conjugated bile acids (MCBAs) in the murine gut, and these MCBAs can further restrict C. difficile virulence in vitro. The recognition of conjugated BAs by BSHs defines the resulting BA pool, including the expansive MCBAs. This work provides insights into the structural basis of BSH mechanisms that shape the BA landscape and promote colonization resistance against C. difficile.}, journal={NATURE MICROBIOLOGY}, author={Foley, Matthew H. and Walker, Morgan E. and Stewart, Allison K. and O'Flaherty, Sarah and Gentry, Emily C. and Patel, Shakshi and Beaty, Violet V. and Allen, Garrison and Pan, Meichen and Simpson, Joshua B. and et al.}, year={2023}, month={Mar} }
 @article{stewart_foley_dougherty_mcgill_gulati_gentry_hagey_dorrestein_theriot_dodds_et al._2023, title={Using Multidimensional Separations to Distinguish Isomeric Amino Acid-Bile Acid Conjugates and Assess Their Presence and Perturbations in Model Systems}, volume={95}, ISSN={["1520-6882"]}, DOI={10.1021/acs.analchem.3c03057}, abstractNote={Bile acids play key roles in nutrient uptake, inflammation, signaling, and microbiome composition. While previous bile acid analyses have primarily focused on profiling 5 canonical primary and secondary bile acids and their glycine and taurine amino acid-bile acid (AA-BA) conjugates, recent studies suggest that many other microbial conjugated bile acids (or MCBAs) exist. MCBAs are produced by the gut microbiota and serve as biomarkers, providing information about early disease onset and gut health. Here we analyzed 8 core bile acids synthetically conjugated with 22 proteinogenic and nonproteogenic amino acids totaling 176 MCBAs. Since many of the conjugates were isomeric and only 42 different m/z values resulted from the 176 MCBAs, a platform coupling liquid chromatography, ion mobility spectrometry, and mass spectrometry (LC-IMS-MS) was used for their separation. Their molecular characteristics were then used to create an in-house extended bile acid library for a combined total of 182 unique compounds. Additionally, ∼250 rare bile acid extracts were also assessed to provide additional resources for bile acid profiling and identification. This library was then applied to healthy mice dosed with antibiotics and humans having fecal microbiota transplantation (FMT) to assess the MCBA presence and changes in the gut before and after each perturbation.}, number={41}, journal={ANALYTICAL CHEMISTRY}, author={Stewart, Allison K. and Foley, Matthew H. and Dougherty, Michael K. and Mcgill, Sarah K. and Gulati, Ajay S. and Gentry, Emily C. and Hagey, Lee R. and Dorrestein, Pieter C. and Theriot, Casey M. and Dodds, James N. and et al.}, year={2023}, month={Oct}, pages={15357–15366} }
 @article{doyle_odenkirk_stewart_nelson_baker_cruz_2022, title={Assessing the Fate of Dissolved Organic Compounds in Landfill Leachate and Wastewater Treatment Systems}, volume={11}, ISSN={["2690-0637"]}, url={https://doi.org/10.1021/acsestwater.2c00320}, DOI={10.1021/acsestwater.2c00320}, abstractNote={Landfill leachate and municipal wastewater are major sources of chemical pollutants that contaminate our drinking water sources. Evaluating the dissolved organic chemical composition in wastewater treatment plants is therefore essential to understand how the discharge impacts the environment, wildlife, and human health. In this study, we utilized a nontargeted analysis method coupling liquid chromatography and tandem mass spectrometry (LC-MS/MS) to analyze chemical features at different points along two landfill leachate treatment plants (LLTPs) and two municipal wastewater treatment plants (WWTPs) in the Southeastern United States. Significant feature differences were observed for the WWTPs where activated sludge clarification was employed versus the LLTPs utilizing reverse osmosis. Specifically, even though both LLTPs had the largest number of features in their influent water, their effluent following reverse osmosis yielded a lower number of features than the WWTPs. Additionally, the clarification processes of each WWTP exhibited different efficiencies as chemical disinfection removed more features than UV disinfection. Feature identification was then made using the LC, MS, and MS/MS information. Analysis of the identified molecules showed that lipids were the most effectively removed from all plants, while alkaloid and organic nitrogen compounds were the most recalcitrant.}, journal={ACS ES&T WATER}, author={Doyle, Michael G. and Odenkirk, Melanie T. and Stewart, Allison K. and Nelson, Jacob P. and Baker, Erin S. and Cruz, Florentino}, year={2022}, month={Nov} }
 @article{manley_phan_stewart_mosley_xue_cha_bai_lightfoot_rucker_collins_et al._2022, title={Self-sacrificial tyrosine cleavage by an Fe:Mn oxygenase for the biosynthesis of para-aminobenzoate in Chlamydia trachomatis}, volume={119}, ISSN={["1091-6490"]}, url={http://dx.doi.org/10.1073/pnas.2210908119}, DOI={10.1073/pnas.2210908119}, abstractNote={Significance As a precursor to the essential vitamin tetrahydrofolate, para-aminobenzoate (pABA) is needed for nucleotide and amino acid synthesis. Chlamydia trachomatis, the causative agent of the sexually transmitted infection, lacks the canonical pabABC biosynthetic pathway and instead utilizes a novel stratagem for pABA synthesis that is catalyzed by the enzyme Chlamydia protein associating with death domains (CADD). Originally assigned as a diiron enzyme, we report high in vitro activity relies on the stoichiometric addition of both Fe and Mn. Furthermore, CADD is shown to activate O2 to self-sacrificially produce pABA from Tyr27. This work addresses the unusual mechanism by which Chlamydiae produce pABA, which is a prime target for the development of therapeutic agents to treat infections.}, number={39}, journal={PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA}, publisher={Proceedings of the National Academy of Sciences}, author={Manley, Olivia M. and Phan, Han N. and Stewart, Allison K. and Mosley, Dontae A. and Xue, Shan and Cha, Lide and Bai, Hongxia and Lightfoot, Veda C. and Rucker, Pierson A. and Collins, Leonard and et al.}, year={2022}, month={Sep} }
 @article{lin_lehman_stewart_panny_bracci_wright_paige_strangman_kehn-hall_2021, title={Homoseongomycin, a compound isolated from marine actinomycete bacteria K3-1, is a potent inhibitor of encephalitic alphaviruses}, volume={191}, ISSN={["1872-9096"]}, DOI={10.1016/j.antiviral.2021.105087}, abstractNote={Marine microorganisms have been a resource for novel therapeutic drugs for decades. In addition to anticancer drugs, the drug acyclovir, derived from a marine sponge, is FDA-approved for the treatment of human herpes simplex virus-1 infections. Most alphaviruses that are infectious to terrestrial animals and humans, such as Venezuelan and eastern equine encephalitis viruses (VEEV and EEEV), lack efficient antiviral drugs and it is imperative to develop these remedies. To push the discovery and development of anti-alphavirus compounds forward, this study aimed to isolate and screen for potential antiviral compounds from cultured marine microbes originating from the marine environment. Compounds from marine microbes were of interest as they are prolific producers of bioactive compounds across the spectrum of human diseases and infections. Homoseongomycin, an actinobacteria isolated from a marine sponge displayed impressive activity against VEEV from a total of 76 marine bioactive products. The 50% effective concentration (EC50) for homoseongomycin was 8.6 μM for suppressing VEEV TC-83 luciferase reporter virus replication. Homoseongomycin was non-toxic up to 50 μM and partially rescued cells from VEEV induced cell death. Homoseongomycin exhibited highly efficient antiviral activity with a reduction of VEEV infectious titers by 8 log10 at 50 μM. It also inhibited EEEV replication with an EC50 of 1.2 μM. Mechanism of action studies suggest that homoseongomycin affects both early and late stages of the viral life cycle. Cells treated with 25 μM of homoseongomycin had a ~90% reduction in viral entry. In comparison, later stages showed a more robust reduction in infectious titers (6 log10) and VEEV extracellular viral RNA levels (4 log10), but a lesser impact on intracellular viral RNA levels (1.5 log10). In sum, this work demonstrates that homoseongomycin is a potential anti-VEEV and anti-EEEV compound due to its low cytotoxicity and potent antiviral activity.}, journal={ANTIVIRAL RESEARCH}, author={Lin, Shih-Chao and Lehman, Caitlin W. and Stewart, Allison K. and Panny, Lauren and Bracci, Nicole and Wright, Jeffrey L. C. and Paige, Mikell and Strangman, Wendy K. and Kehn-Hall, Kylene}, year={2021}, month={Jun} }
 @article{foley_o'flaherty_allen_rivera_stewart_barrangou_theriot_2021, title={Lactobacillus bile salt hydrolase substrate specificity governs bacterial fitness and host colonization}, volume={118}, ISSN={["1091-6490"]}, url={https://doi.org/10.1073/pnas.2017709118}, DOI={10.1073/pnas.2017709118}, abstractNote={Significance The transformation of bile acids (BAs) by the gut microbiota is increasingly recognized as an important factor shaping host health. The prerequisite step of BA metabolism is carried out by bile salt hydrolases (BSHs), which are encoded by select gut and probiotic bacteria. Despite their prevalence, the utility of harboring a bsh is unclear. Here, we investigate the role of BSHs encoded by Lactobacillus acidophilus and Lactobacillus gasseri. We show that BA type and BSH substrate preferences affect in vitro and in vivo growth of both species. These findings contribute to a mechanistic understanding of bacterial survival in various BA-rich niches and inform future efforts to leverage BSHs as a therapeutic tool for manipulating the gut microbiota. Primary bile acids (BAs) are a collection of host-synthesized metabolites that shape physiology and metabolism. BAs transit the gastrointestinal tract and are subjected to a variety of chemical transformations encoded by indigenous bacteria. The resulting microbiota-derived BA pool is a mediator of host–microbiota interactions. Bacterial bile salt hydrolases (BSHs) cleave the conjugated glycine or taurine from BAs, an essential upstream step for the production of deconjugated and secondary BAs. Probiotic lactobacilli harbor a considerable number and diversity of BSHs; however, their contribution to Lactobacillus fitness and colonization remains poorly understood. Here, we define and compare the functions of multiple BSHs encoded by Lactobacillus acidophilus and Lactobacillus gasseri. Our genetic and biochemical characterization of lactobacilli BSHs lend to a model of Lactobacillus adaptation to the gut. These findings deviate from previous notions that BSHs generally promote colonization and detoxify bile. Rather, we show that BSH enzymatic preferences and the intrinsic chemical features of various BAs determine the toxicity of these molecules during Lactobacillus growth. BSHs were able to alter the Lactobacillus transcriptome in a BA-dependent manner. Finally, BSHs were able to dictate differences in bacterial competition in vitro and in vivo, defining their impact on BSH-encoding bacteria within the greater gastrointestinal tract ecosystem. This work emphasizes the importance of considering the enzymatic preferences of BSHs alongside the conjugated/deconjugated BA–bacterial interaction. These results deepen our understanding of the BA–microbiome axis and provide a framework to engineer lactobacilli with improved bile resistance and use probiotics as BA-altering therapeutics.}, number={6}, journal={PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA}, publisher={Proceedings of the National Academy of Sciences}, author={Foley, Matthew H. and O'Flaherty, Sarah and Allen, Garrison and Rivera, Alissa J. and Stewart, Allison K. and Barrangou, Rodolphe and Theriot, Casey M.}, year={2021}, month={Feb} }
 @article{rock_st armour_horman_phillips_ruis_stewart_jima_muddiman_stapleton_patisaul_2020, title={Effects of Prenatal Exposure to a Mixture of Organophosphhate Flame Ritardants on Placental Gene Expression and Serotonergic Innervaion in the Fetal Rat Brain}, volume={176}, ISSN={["1096-0929"]}, DOI={10.1093/toxsci/kfaa046}, abstractNote={There is a growing need to understand the potential neurotoxicity of organophosphate ester flame retardants (OPFRs) and plasticizers because use and, consequently, human exposure, is rapidly expanding. We have previously shown in rats that developmental exposure to the commercial FR mixture Firemaster® 550 (FM 550), which contains OPFRs, results in sex-specific behavioral effects, and identified the placenta as a potential target of toxicity. The placenta is a critical coordinator of fetal growth and neurodevelopment, and a source of neurotransmitters (NTs) for the developing brain. We have shown in rats and humans that FRs accumulate in placental tissue, and induce functional changes, including altered neurotransmitter (NT) production. Here we sought to establish if OPFRs (triphenyl phosphate, TPHP, and a mixture of isopropylated triarylphosphate isomers, ITPs) alter placental function and fetal forebrain development, with disruption of tryptophan (Trp) metabolism as a primary pathway of interest. Wistar rat dams were orally exposed to OPFRs (0, 500, 1,000, or 2,000 μg/day) or a serotonin (5-HT) agonist (5-MT) for 14 days during gestation and placenta and fetal forebrain tissues collected for analysis by transcriptomics and metabolomics. Relative abundance of genes responsible for the transport and synthesis of placental 5-HT were disrupted, and multiple neuroactive metabolites in the 5-HT and kynurenine (Kyn) metabolic pathways were upregulated. Additionally, 5-HTergic projections were significantly longer in the fetal forebrains of exposed males. These findings suggest that OPFRs have the potential to impact the 5-HTergic system in the fetal forebrain by disrupting placental Trp metabolism.}, number={1}, journal={TOXICOLOGICAL SCIENCES}, author={Rock, Kylie D. and St Armour, Genevieve and Horman, Brian and Phillips, Allison and Ruis, Matthew and Stewart, Allison K. and Jima, Dereje and Muddiman, David C. and Stapleton, Heather M. and Patisaul, Heather B.}, year={2020}, month={Jul}, pages={203–223} }