mBio最新文献

{"title":"Molecular mechanisms of drug resistance and compensation in SARS-CoV-2 main protease: the interplay between E166 and L50.","authors":"Sarah N Zvornicanin, Ala M Shaqra, Julia Flynn, Heidi Carias Martinez, Weiping Jia, Stephanie Moquin, Dustin Dovala, Daniel N Bolon, Nese Kurt Yilmaz, Celia A Schiffer","doi":"10.1128/mbio.04068-24","DOIUrl":"https://doi.org/10.1128/mbio.04068-24","url":null,"abstract":"<p><p>The SARS-CoV-2 main protease (M^pro) is essential for viral replication and is a primary target for COVID-19 antivirals. Direct-acting antivirals such as nirmatrelvir, the active component of Paxlovid, target the M^pro active site to block viral polyprotein cleavage and thus replication. However, drug resistance mutations at the active site residue Glu166 (E166) have emerged during <i>in vitro</i> selection studies, raising concerns about the durability of current antiviral strategies. Here, we investigate the molecular basis of drug resistance conferred by E166A and E166V mutations against nirmatrelvir and the related PF-00835231, individually and in combination with the distal mutation L50F. We found that E166 mutations reduce nirmatrelvir potency by up to 3,000-fold while preserving substrate cleavage, with catalytic efficiency reduced by only up to twofold. This loss of catalytic efficiency was compensated for by the addition of L50F in the double-mutant variants. We have determined three cocrystal structures of the E166 variants (E166A, E166V, and E166V/L50F) bound to PF-00835231. Comparison of these structures with wild-type enzyme demonstrated that E166 is crucial for dimerization and for shaping the substrate-binding S1 pocket. Our findings highlight the mutability of E166, a prime site for resistance for inhibitors that leverage direct interactions with this position, and the potential emergence of highly resistant and active variants in combination with the compensatory mutation L50F. These insights support the design of inhibitors that target conserved protease features and avoid E166 side-chain interactions to minimize susceptibility to resistance.</p><p><strong>Importance: </strong>Drug resistance remains a great challenge to modern medicine. This study investigates SARS-CoV-2 main protease variants E166A and E166V which confer nirmatrelvir resistance. These variants can retain considerable enzymatic activity through combination with the compensatory mutation L50F. For single- and double-mutant variant enzymes, we assessed catalytic efficiency, measured loss in potency for nirmatrelvir and its analog PF-00835231, and cocrystallized with inhibitors to investigate drug resistance caused by these mutations. Our results contribute toward understanding of molecular mechanisms of resistance and combinations of mutations, which pushes toward resistance-thwarting inhibitor design. These principles also apply broadly to many quickly evolving drug targets in infectious diseases.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0406824"},"PeriodicalIF":5.1,"publicationDate":"2025-04-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143780139","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Microbial solutions for climate change require global partnership.","authors":"J T Lennon, R Rappuoli, D E Bloom, C Brooke, R M Burckhardt, A D Dangour, D Egamberdieva, G K Gronvall, T D Lawley, R Morhard, A Mukhopadhyay, R S Peixoto, P A Silver, V Sperandio, L Y Stein, N K Nguyen","doi":"10.1128/mbio.00778-25","DOIUrl":"https://doi.org/10.1128/mbio.00778-25","url":null,"abstract":"","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0077825"},"PeriodicalIF":5.1,"publicationDate":"2025-04-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143772382","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Dual-use virulence factors of the opportunistic pathogen <i>Chromobacterium haemolyticum</i> mediate hemolysis and colonization.","authors":"Leo Dumjahn, Philipp Wein, Evelyn M Molloy, Kirstin Scherlach, Felix Trottmann, Philippe R Meisinger, Louise M Judd, Sacha J Pidot, Timothy P Stinear, Ingrid Richter, Christian Hertweck","doi":"10.1128/mbio.03605-24","DOIUrl":"https://doi.org/10.1128/mbio.03605-24","url":null,"abstract":"<p><p><i>Chromobacterium haemolyticum</i> is an environmental bacterium that can cause severe and fatal opportunistic infections in humans and animals. Although <i>C. haemolyticum</i> is characterized by its strong β-hemolytic activity, the molecular basis of this phenotype has remained elusive over the more than 15 years since the species was first described. Herein, we report a family of cyclic lipodepsipeptides, the jagaricins, that are responsible for the potent hemolytic activity of <i>C. haemolyticum</i>. Comparative genomics of <i>C. haemolyticum</i> strains revealed a completely conserved gene locus (<i>hml</i>) encoding a nonribosomal peptide synthetase. Metabolic profiling of <i>C. haemolyticum</i> DSM 19808 identified a suite of cyclic lipodepsipeptides as the products, with the three main congeners (jagaricin A-C) being elucidated by a combination of tandem mass spectrometry, chemical derivatization, and nuclear magnetic resonance spectroscopy. Significantly, a <i>C. haemolyticum hml</i> deletion mutant is devoid of hemolytic activity. Moreover, purified jagaricins are hemolytic at low micromolar concentrations in an erythrocyte lysis assay. Further bioassays demonstrated that the cyclic lipodepsipeptides are crucial for the biofilm-forming and swarming behavior of <i>C. haemolyticum</i>. Matrix-assisted laser desorption ionization mass spectrometry imaging showed that primarily jagaricin B and C are involved in these processes <i>in vitro</i>. Our data shed light on the bioactivities of jagaricins, specialized metabolites that likely contribute to both successful niche colonization and the virulence potential of <i>C. haemolyticum</i>.IMPORTANCEDespite the rising incidence of <i>Chromobacterium haemolyticum</i> as a serious opportunistic pathogen, there is limited information on whether the competitive traits that ensure its survival in its freshwater niche also influence host infection. We reveal that <i>C. haemolyticum</i> produces specialized metabolites that not only cause its pronounced hemolytic phenotype but are also crucial for biofilm formation and swarming motility. These results exemplify a case of coincidental evolution, wherein the selective pressures encountered in a primary environmental niche drive the evolution of a trait impacting virulence. This knowledge provides a foundation for the development of antivirulence therapies against the emerging pathogen <i>C. haemolyticum</i>.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0360524"},"PeriodicalIF":5.1,"publicationDate":"2025-04-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143772470","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Structural elements of cyanobacterial co-factor-independent phosphoglycerate mutase that mediate regulation by PirC.","authors":"Tim Orthwein, Janette T Alford, Nathalie Sofie Becker, Phillipp Fink, Karl Forchhammer","doi":"10.1128/mbio.03378-24","DOIUrl":"https://doi.org/10.1128/mbio.03378-24","url":null,"abstract":"<p><p>The 2,3-bisphosphoglycerate-independent phosphoglycerate mutase (iPGAM) has been identified as a regulating key point in the carbon storage metabolism of cyanobacteria. Upon nitrogen starvation, the iPGAM is inhibited by the P_II-interacting regulator PirC, which is released from its interaction partner P_II due to elevated 2-oxoglutarate levels. <i>In silico</i> analysis of 338 different iPGAMs revealed a deep-rooted distinctive evolution of iPGAMs in cyanobacteria. Remarkably, cyanobacterial iPGAMs possess a unique loop structure and an extended C-terminus. Our mass photometry analysis suggests that iPGAM forms a complex with three individual PirC monomers. Biolayer interferometry revealed that the PirC-iPGAM complex is affected by the unique loop and the C-terminal structural elements of iPGAM. A C-terminally truncated iPGAM enzyme showed loss of control by PirC and twofold increased enzymatic activity compared to the iPGAM-WT (wild type), as demonstrated by enzyme assays. By contrast, deleting the loop structure significantly reduced the activity of this variant. Physiological experiments were carried out with different iPGAM variant strains of <i>Synechocystis</i>, in which these structural elements were deleted. The strain expressing the C-terminally truncated iPGAM showed a similar overproduction of polyhydroxybutyrate as deletion of the iPGAM regulator PirC. However, in contrast to the latter, these strains showed higher overall biomass accumulation, making them a better chassis for the production of polyhydroxybutyrate or other valuable substances than the PirC-deficient mutant.IMPORTANCEThe primordial cyanobacteria were responsible for developing oxygenic photosynthesis early in evolution. In the pathways of fixed carbon allocation, the co-factor-independent phosphoglycerate mutase (iPGAM) plays a crucial role by directing the first CO₂ fixation product, 3-phosphoglycerate, toward central anabolic glycolytic-derived pathways. This work reveals a distinct evolution of iPGAM within oxygenic photosynthetic organisms. We have identified two specific segments in cyanobacterial iPGAMs that affect the control of iPGAM activity through its specific interactor protein PirC. This understanding of iPGAM has allowed us to engineer cyanobacterial strains with altered carbon fluxes. Since cyanobacteria can directly convert CO₂ into valuable products, our results demonstrate a novel approach for developing a chassis for biotechnical use.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0337824"},"PeriodicalIF":5.1,"publicationDate":"2025-04-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143772639","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Signaling pathways governing the pathobiological features and antifungal drug resistance of <i>Candida auris</i>.","authors":"Hyunjin Cha, Doyeon Won, Yong-Sun Bahn","doi":"10.1128/mbio.02475-23","DOIUrl":"https://doi.org/10.1128/mbio.02475-23","url":null,"abstract":"<p><p><i>Candida auris</i> is an emerging multidrug-resistant fungal pathogen that poses a significant global health threat. Since its discovery in 2009, <i>C. auris</i> has rapidly spread worldwide, causing severe infections with high mortality rates, particularly in healthcare settings. Its ability to persist in the environment, form biofilms, and resist multiple antifungal drugs underscores the urgent need to understand its pathogenicity mechanisms and associated signaling pathways. Such insights are crucial for elucidating its unique virulence traits and developing targeted therapeutic strategies. Current studies have identified several key pathways involved in its pathogenicity and antifungal drug resistance. The Ras/cAMP/PKA pathway regulates critical virulence factors, including thermotolerance, morphological plasticity, and biofilm formation. The mitogen-activated protein kinase (MAPK) and calcineurin pathways contribute to stress responses and antifungal drug resistance. The regulation of Ace2 and morphogenesis (RAM) pathway influences cell aggregation, while the target of rapamycin (TOR) pathway affects filamentous growth and biofilm development. However, the distinct characteristics of <i>C. auris</i>, such as its rapid environmental spread and clade-specific traits, warrant further investigation into additional signaling pathways. This review provides a comprehensive analysis of known signaling pathways associated with <i>C. auris</i> pathogenicity and antifungal drug resistance, integrating insights from other fungal pathogens. By synthesizing current knowledge and identifying research gaps, this review offers new perspectives on future research directions and potential therapeutic targets against this formidable pathogen.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0247523"},"PeriodicalIF":5.1,"publicationDate":"2025-04-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143772634","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Synergistic induction of PGE2 by oral pathogens and TNF promotes gingival fibroblast-driven stromal-immune cross-talk in periodontitis.","authors":"Elwira Nieboga, Aureliusz Schuster, Dominika M Drapala, Mariia Melnykova, Aleksander Gut, Weronika Lipska, Mateusz Kwitniewski, Marcin Migaczewski, Marta Czesnikiewicz-Guzik, Tomasz Kaczmarzyk, Jan Potempa, Aleksander M Grabiec","doi":"10.1128/mbio.00046-25","DOIUrl":"https://doi.org/10.1128/mbio.00046-25","url":null,"abstract":"<p><p>The interaction between pathogenic microorganisms and stromal cells, in particular fibroblasts, significantly contributes to the pathogenesis of many bacterially driven diseases. In periodontitis, oral pathogens penetrate the epithelial barrier and aggravate ongoing gingival inflammation by promoting the production of inflammatory mediators, such as prostaglandin E2 (PGE2). This study aimed to investigate the functional consequences of the interplay between oral pathogens and a pro-inflammatory environment in the activation of the PGE2 pathway in primary human gingival fibroblasts (GFs). GF infection with <i>Fusobacterium nucleatum</i>, <i>Porphyromonas gingivalis,</i> or <i>Filifactor alocis</i> in the presence of tumor necrosis factor (TNF) led to synergistic induction of cyclooxygenase-2 (COX-2), a key enzyme in the PGE2 synthesis pathway, as well as secretion of PGE2. A similar synergy in COX-2 upregulation was observed upon GF infection with oral pathogens in the presence of IL-1α, IL-1β, and interferon-α (IFN-α). This effect required toll-like receptor-2 (TLR2) and the p38 MAP kinase activation and was specific for fibroblasts as infection of macrophages or keratinocytes with oral pathogens in the proinflammatory environment did not cause synergistic COX-2 induction. Finally, we demonstrated that conditioned media from GFs infected with <i>F. nucleatum</i> under inflammatory conditions amplified the expression of the neutrophil chemokine <i>IL8</i> in macrophages and confirmed that this effect was mediated by synergistic induction of PGE2 in GFs. Collectively, we identify a new mechanism of stromal-immune cross-talk that is driven by synergistic PGE2 induction by oral pathogens and inflammatory cytokines in GFs and may contribute to excessive macrophage activation and neutrophil infiltration in periodontitis.IMPORTANCEPeriodontitis is a highly prevalent, dysbiosis-driven chronic inflammatory disease that not only leads to tooth loss but also is associated with severe systemic diseases. In this work, we describe a novel mechanism responsible for excessive production of PGE2, which is a potent inflammatory mediator that significantly contributes to the pathogenesis of periodontitis. We found that infection of GFs with many species of oral pathogens in the presence of inflammatory cytokines produced by the host leads to synergistic induction of COX-2 expression and PGE2 production. We found that this fibroblast-specific amplification of the COX-2-PGE2 axis by oral pathogens and cytokines is driven by the p38 MAP kinase and promotes enhanced expression of a key neutrophil chemokine by macrophages. These studies have thus enabled the identification of a new mechanism of host-pathogen interactions in periodontitis, improving our understanding of the roles of GFs and their cross-talk with immune cells in disease pathogenesis.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0004625"},"PeriodicalIF":5.1,"publicationDate":"2025-04-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143772643","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Modeling heterogeneity, commitment, and memory of bacterial spore germination.","authors":"William Li, Steven Mednick, Peter Setlow, Yong-Qing Li","doi":"10.1128/mbio.00596-25","DOIUrl":"https://doi.org/10.1128/mbio.00596-25","url":null,"abstract":"<p><p>Spores of bacteria are metabolically dormant, resistant to microbicides, and vectors of food spoilage and diseases and survive for years in their dormant state. Upon exposure to nutrient germinants, spores can rapidly return to life through germination, losing their resistance and becoming easy to kill. Despite extensive research on germination heterogeneity, commitment, and memory, many mechanisms underlying germination of Bacillota spores remain unclear, as a comprehensive mathematical model describing germination characteristics of individual spores is lacking. Woese et al. (PNAS, 59:869, 1968) developed a simple model predicting that time-to-germination of a spore with <i>n</i> active enzymes is proportional to 1/<i>n</i>. Here, we present a novel approach inspired by artificial neural networks to model spore germination, treating it as a decision-making process upon the activation produced by binding of germinants to germinant receptors. Major findings include the following: (i) using a sigmoid activation function to model germination thresholds allows predictions of distributions in time-to-commitment and kinetic germination to be well fitted to experimental observations; (ii) modeling spore commitment and memory after two separate germinant pulses fits well to experimental data of <i>Bacillus</i> spores germinated with L-alanine, and a zero fraction of germinated spores by a second pulse is predicted by loss of memory; and (iii) modeling kinetic CaDPA release from individual spores through SpoVA channels and fitting experimental data of <i>Bacillus cereus</i> spores. This work enhances our understanding of unexplored biophysical intricacies of spore germination, and the use of the model may generate new data.IMPORTANCESpore germination is a crucial process through which spores of bacteria return to life when triggered by germinants, and some spore species cause food spoilage, human diseases, and bioterrorism. Understanding and theoretical predictions of spore germination could facilitate the development of \"germinate to kill\" strategies as spores lose their resistance upon germination. Here, we developed a novel mathematical model to describe the characteristics of spore germination including heterogeneity, commitment, memory, and kinetic CaDPA release using an artificial neural network. This model predicts new aspects of germination such as the retention and loss of memory and the effect of GRs' distribution on germination rate and could be useful in data-driven discoveries to enhance our understanding of germination's biophysical intricacies.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0059625"},"PeriodicalIF":5.1,"publicationDate":"2025-04-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143764283","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Moonlighting enzymes of <i>Borrelia burgdorferi</i>.","authors":"Jorge L Benach","doi":"10.1128/mbio.00279-25","DOIUrl":"https://doi.org/10.1128/mbio.00279-25","url":null,"abstract":"<p><p>Moonlighting enzymes are increasingly recognized in bacteria with dual functions depending on whether they are intracellular or expressed on the surface. Enzymes of the glycolytic pathway are among the most frequently associated with moonlighting functions and lack the signal sequences needed to deliver them to the cell surface. Once these enzymes are on the surface, they perform functions that are associated with pathogenesis and development of infection through interaction with host substrates. One such interaction is adhesion. <i>Borrelia burgdorferi,</i> the etiologic agent of Lyme disease, must encounter a wide number of different tissues and substrates from ticks to mammalian hosts to complete its life cycle and persist. The phosphomannose isomerase of this organism has a moonlighting function, interacting with collagen IV, a main component of the basal lamina. It is abundant in the skin, which is the site of the initial infection of <i>B. burgdorferi</i>.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0027925"},"PeriodicalIF":5.1,"publicationDate":"2025-04-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143764286","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Generation of inositol polyphosphates through a phospholipase C-independent pathway involving carbohydrate and sphingolipid metabolism in <i>Trypanosoma cruzi</i>.","authors":"Mayara S Bertolini, Sabrina E Cline, Miguel A Chiurillo, Brian S Mantilla, Aharon Eidex, Logan P Crowe, Danye Qiu, Henning J Jessen, Adolfo Saiardi, Roberto Docampo","doi":"10.1128/mbio.03318-24","DOIUrl":"https://doi.org/10.1128/mbio.03318-24","url":null,"abstract":"<p><p>Inositol phosphates are involved in a myriad of biological roles and activities such as Ca²⁺ signaling, phosphate homeostasis, energy metabolism, and disease pathogenicity. In <i>Saccharomyces cerevisiae,</i> synthesis of inositol phosphates occurs through the phosphoinositide phospholipase C (PLC)-catalyzed hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP₂) into inositol 1,4,5-trisphosphate (IP₃) and diacylglycerol and further IP₃ phosphorylation by additional kinases that leads to the formation of highly phosphorylated inositol derivatives, known as inositol pyrophosphates. Inositol-tetrakisphosphate 1-kinase (ITPK1) is an enzyme that mediates a PLC-independent inositol polyphosphate synthesis through phosphorylation of inositol monophosphates and other intermediates in the cytosol. In this work, we identified and characterized a <i>Trypanosoma cruzi</i> ITPK1 (TcITPK1) homolog. The ability of TcITPK1 to act as the mediator for this alternative pathway was established through <i>plc1</i>Δ and <i>plc1</i>Δ <i>isc1</i>Δ yeast complementation assays and SAX-HPLC analyses of radioactively labeled inositol. TcITPK1 localizes to the cytosol, and knockout attempts of <i>TcITPK1</i> revealed that only one allele was replaced by the DNA donor cassette at the specific locus, suggesting that <i>null</i> alleles may have lethal effects in epimastigotes. Ablation of <i>T. cruzi</i> phosphoinositide phospholipase C 1 (<i>TcPI-PLC1</i>) affected the synthesis of IP₃ from glucose 6-phosphate but did not affect the synthesis of inositol polyphosphates, while ablation of inositol phosphosphingolipid phospholipase (<i>TcISC1</i>) affected the synthesis of inositol polyphosphates, thus revealing that the PLC-independent pathway using either glucose 6-phosphate or inositol phosphoceramide is involved in the synthesis of inositol polyphosphates, while the PLC-dependent pathway is involved in IP₃ formation needed for Ca²⁺ signaling.</p><p><strong>Importance: </strong>Millions of people are infected with <i>Trypanosoma cruzi,</i> and the current treatment is not satisfactory. Inositol pyrophosphates have been established as important signaling molecules. Our work demonstrates the presence of a phospholipase C-independent pathway for the synthesis of inositol pyrophosphates in <i>T. cruzi</i>. Furthermore, we demonstrate that this pathway starts with the synthesis of inositol monophosphates from glucose 6-phosphate or from inositol phosphoceramide, linking it to carbohydrate and sphingolipid metabolism. The essentiality of the pathway for the survival of <i>T. cruzi</i> infective stages makes it an ideal drug target for treating American trypanosomiasis.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0331824"},"PeriodicalIF":5.1,"publicationDate":"2025-04-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143764277","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Analysis of genetic requirements and nutrient availability for <i>Staphylococcus aureus</i> growth in cystic fibrosis sputum.","authors":"Lauren M Shull, Daniel J Wolter, Dillon E Kunkle, Katherine A Legg, David P Giedroc, Eric P Skaar, Lucas R Hoffman, Michelle L Reniere","doi":"10.1128/mbio.00374-25","DOIUrl":"https://doi.org/10.1128/mbio.00374-25","url":null,"abstract":"<p><p><i>Staphylococcus aureus</i> is one of the most common pathogens isolated from the lungs of people with cystic fibrosis (CF), but little is known about its ability to colonize this niche. We performed a transposon-sequencing (Tn-seq) screen to identify genes necessary for <i>S. aureus</i> growth in media prepared from <i>ex vivo</i> CF sputum. We identified 19 genes that were required for growth in all sputum media tested and dozens more that were required for growth in at least one sputum medium. Depleted mutants of interest included insertions in many genes important for surviving metal starvation, as well as the primary regulator of cysteine metabolism, <i>cymR</i>. To investigate the mechanisms by which these genes contribute to <i>S. aureus</i> growth in sputum, we quantified low-molecular-weight thiols, nutrient transition metals, and the host metal-sequestration protein calprotectin in sputum from 11 individuals with CF. In all samples, the abundance of calprotectin exceeded nutrient metal concentration, explaining the <i>S. aureus</i> requirement for metal-starvation genes. Furthermore, all samples contain potentially toxic quantities of cysteine and sufficient glutathione to satisfy the organic sulfur requirements of <i>S. aureus</i>. Deletion of the cysteine importer genes <i>tcyA</i> and <i>tcyP</i> in the ∆<i>cymR</i> background restored growth to wild-type levels in CF sputum, suggesting that the mechanism by which <i>cymR</i> is required for growth in sputum is to prevent uncontrolled import of cysteine or cystine from this environment. Overall, this work demonstrates that calprotectin and cysteine limit <i>S. aureus</i> growth in CF sputum.IMPORTANCE<i>Staphylococcus aureus</i> is a major cause of lung infections in people with cystic fibrosis (CF). This work identifies genes required for <i>S. aureus</i> growth in this niche, which represent potential targets for anti-Staphylococcal treatments. We show that genes involved in surviving metal starvation are required for growth in CF sputum. We also found that the primary regulator of cysteine metabolism, CymR, plays a critical role in preventing cysteine intoxication during growth in CF sputum. To support these models, we analyzed sputum from 11 individuals with CF to determine concentrations of calprotectin, nutrient metals, and low-molecular-weight thiols, which have not previously been quantified together in the same samples.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0037425"},"PeriodicalIF":5.1,"publicationDate":"2025-04-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143764271","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}