@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/107496,
    title ="Draft Genome Sequence of the Free-Living, Iridescent Bacterium Tenacibaculum mesophilum Strain ECR",
    author = "Mickol, Rebecca L. and Louyakis, Artemis S.",
    journal = "Microbiology Resource Announcements",
    volume = "10",
    number = "1",
    pages = "Art. No. e01302-20",
    month = "January",
    year = "2021",
    doi = "10.1128/mra.01302-20",
    issn = "2576-098X",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20210114-164619982",
    note = "© 2021 Mickol et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license. \n\nReceived 13 November 2020. Accepted 16 December 2020. Published 7 January 2021. \n\nThis work was conducted as part of the 2016 and 2018 Microbial Diversity summer courses at Marine Biological Laboratory (Woods Hole, MA). \n\nFunding for R.L.M. was provided by the Helmsley Charitable Trust-Microbial Diversity, the Selman A. Waksman Endowed Scholarship in Microbial Diversity, and the Simons M.D. Scholarship. Funding for the Microbial Diversity course was provided by grants from the DOE, NASA, the NSF, Promega, and the Simons Foundation.\n\nData availability:\nThe GenBank accession number for this genome sequence is\nJAAQOW000000000 under NCBI BioProject PRJNA488075 (accession numbers SRR11313510, SRR11313511, and SRR11313512).",
    revision_no = "16",
    abstract = "Here, we report the genome sequence of Tenacibaculum mesophilum strain ECR, which was isolated from the river/ocean interface at Trunk River in Falmouth, Massachusetts. The isolation and sequencing were performed as part of the 2016 and 2018 Microbial Diversity courses at the Marine Biological Laboratory in Woods Hole, Massachusetts.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/106810,
    title ="Bidirectional redox cycling of phenazine-1-carboxylic acid by Citrobacter portucalensis MBL drives increased nitrate reduction",
    author = "Tsypin, Lev M. and Newman, Dianne K.",
    
    
    
    
    month = "November",
    year = "2020",
    
    
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20201124-104632686",
    note = "The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license. \n\nVersion 1 (November 24, 2020 - 08:09); this version posted November 25, 2020. \n\nWe would like to thank the members of the Newman lab, and especially Scott Saunders, Darcy McRose, Avi Flamholz, John Ciemniecki, Chelsey VanDrisse, and Justin Bois for their insight and helpful discussions throughout this work. We are grateful to Nathan Dalleska at the Environmental Analysis Center at Caltech for training LMT on the Dionex instrument and providing a facility for analytical chemistry. LMT was supported by the Rosen Endowment Fellowship at Caltech and the National Science Foundation Graduate Research Fellowship\n(DGE‐1745301). Additional support to DKN came from NIH (1R01AI127850-01A1 and 1R01HL152190-01) and ARO (W911NF-17-1-0024) grants.",
    revision_no = "20",
    abstract = "Phenazines are secreted metabolites that microbes use in diverse ways, from quorum sensing to antimicrobial warfare to energy conservation. Phenazines are able to contribute to these activities due to their redox activity. The physiological consequences of cellular phenazine reduction have been extensively studied, but the counterpart phenazine oxidation has been largely overlooked. Phenazine-1-carboxylic acid (PCA) is common in the environment and readily reduced by its producers. Here, we describe its anaerobic oxidation by Citrobacter portucalensis strain MBL, which was isolated from topsoil in Falmouth, MA, and which does not produce phenazines itself. This activity depends on the availability of a suitable terminal electron acceptor, specifically nitrate or fumarate. When C. portucalensis MBL is provided reduced PCA and either nitrate or fumarate, it continuously oxidizes the PCA. We compared this terminal electron acceptor-dependent PCA-oxidizing activity of C. portucalensis MBL to that of several other γ-proteobacteria with varying capacities to respire nitrate and/or fumarate. We found that PCA oxidation by these strains in a fumarate-or nitrate-dependent manner is decoupled from growth and correlated with their possession of the fumarate or periplasmic nitrate reductases, respectively. We infer that bacterial PCA oxidation is widespread and genetically determined. Notably, reduced PCA enhances the rate of nitrate reduction to nitrite by C. portucalensis MBL beyond the stoichiometric prediction, which we attribute to C. portucalensis MBL’s ability to also reduce oxidized PCA, thereby catalyzing a complete PCA redox cycle. This bidirectionality highlights the versatility of PCA as a biological redox agent.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/105842,
    title ="Keystone metabolites of crop rhizosphere microbiomes",
    author = "Dahlstrom, Kurt M. and McRose, Darcy L.",
    journal = "Current Biology",
    volume = "30",
    number = "19",
    pages = "R1131-R1137",
    month = "October",
    year = "2020",
    doi = "10.1016/j.cub.2020.08.005",
    issn = "0960-9822",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20201006-131015350",
    note = "© 2020 Elsevier Inc. \n\nAvailable online 5 October 2020. \n\nWe thank the following agencies and foundations for supporting our research: grants to D.K.N. from the ARO (W911NF-17-1-0024), NIH (1R01AI127850-01A1) and Caltech Resnick Sustainability Institute; the Life Sciences Research Foundation postdoctoral fellowship to K.M.D.; and the Caltech Biology & Biological Engineering Division and Simons Foundation Marine Microbial Ecology postdoctoral fellowships to D.L.M. We are grateful to A. Flamholz, D. Dar, L.S. Thomashow, L. Glass, C. Adams, and Z. Lonergan for insightful feedback on the manuscript. We also thank W.P. Falcon for helpful discussions about agriculture.",
    revision_no = "9",
    abstract = "The role of microbes in sustaining agricultural plant growth has great potential consequences for human prosperity. Yet we have an incomplete understanding of the basic function of rhizosphere microbial communities and how they may change under future stresses, let alone how these processes might be harnessed to sustain or improve crop yields. A reductionist approach may aid the generation and testing of hypotheses that can ultimately be translated to agricultural practices. With this in mind, we ask whether some rhizosphere microbial communities might be governed by ‘keystone metabolites’, envisioned here as microbially produced molecules that, through antibiotic and/or growth-promoting properties, may play an outsized role in shaping the development of the community spatiotemporally. To illustrate this point, we use the example of redox-active metabolites, and in particular phenazines, which are produced by many bacteria found in agricultural soils and have well-understood catalytic properties. Phenazines can act as potent antibiotics against a variety of cell types, yet they also can promote the acquisition of essential inorganic nutrients. In this essay, we suggest the ways these metabolites might affect microbial communities and ultimately agricultural productivity in two specific scenarios: firstly, in the biocontrol of beneficial and pathogenic fungi in increasingly arid crop soils and, secondly, through promotion of phosphorus bioavailability and sustainable fertilizer use. We conclude with specific proposals for future research.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/103785,
    title ="Global landscape of phenazine biosynthesis and biodegradation reveals species-specific colonization patterns in agricultural soils and crop microbiomes",
    author = "Dar, Daniel and Thomashow, Linda S.",
    journal = "eLife",
    volume = "2020",
    number = "9",
    pages = "Art. No. e59726",
    month = "September",
    year = "2020",
    doi = "10.7554/eLife.59726",
    issn = "2050-084X",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20200609-075446759",
    note = "© 2020 Dar et al. This article is distributed under the\nterms of the Creative Commons Attribution License, which\npermits unrestricted use and redistribution provided that the original author and source are credited. \n\nReceived: 05 June 2020; Accepted: 02 September 2020; Published: 15 September 2020. \n\nWe thank Jeff Dangl and Adam Deutschbauer for kindly providing us with the D. japonica UNC79MFTsu3.2 strain. We also thank Darcy McRose for help with LC-MS sample preparation and analysis, Megan Bergkessel and Kurt Dahlstrom for their help with mutant generation, Will DePas for assistance with HCR, as well as the rest of the Newman lab for fruitful discussions and comments. \n\nWe also thank Gil Sharon and Alon Philosof for critically reading the manuscript. We thank Mingming Yang with assistance in collecting, processing, and performing viable counts of phenazine-producing pseudomonads from Washington State University’s Lind Dryland Research Station. We also thank Bruce Sauer and Brian Fode for plot maintenance. Grants to DKN from the NIH (1R01AI127850-01A1) and ARO (W911NF-17-1-0024) supported this work. DD was supported by the Rothschild foundation, EMBO Long-Term, and the Helen Hay Whitney postdoctoral fellowships, as well as a Geobiology Postdoctoral Fellowship from the Division of Geological and Planetary Sciences, Caltech. \n\nThe funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication. \n\nData and software availability: The metagenomic DNA-sequencing data generated in this study were deposited in the Sequence Read Archive (SRA) under accession PRJNA634917. All public SRA samples analyzed in this study are indicated in Supplementary file 4. Code can be found at: https://github.com/daniedar/phenazines (Dar, 2020; copy archived at https://github.com/elifesciences-publications/phenazines). \n\nAuthor contributions: Daniel Dar, Conceptualization, Data curation, Formal analysis, Funding acquisition, Validation, Investigation, Visualization, Methodology, Writing - original draft, Writing - review and editing; Linda S Thomashow, David M Weller, Investigation, Writing - review and editing; Dianne K Newman, Conceptualization, Supervision, Funding acquisition, Writing - original draft, Writing - review and editing.",
    revision_no = "45",
    abstract = "Phenazines are natural bacterial antibiotics that can protect crops from disease. However, for most crops it is unknown which producers and specific phenazines are ecologically relevant, and whether phenazine biodegradation can counter their effects. To better understand their ecology, we developed and environmentally-validated a quantitative metagenomic approach to mine for phenazine biosynthesis and biodegradation genes, applying it to >800 soil and plant-associated shotgun-metagenomes. We discover novel producer-crop associations and demonstrate that phenazine biosynthesis is prevalent across habitats and preferentially enriched in rhizospheres, whereas biodegrading bacteria are rare. We validate an association between maize and Dyella japonica, a putative producer abundant in crop microbiomes. D. japonica upregulates phenazine biosynthesis during phosphate limitation and robustly colonizes maize seedling roots. This work provides a global picture of phenazines in natural environments and highlights plant-microbe associations of agricultural potential. Our metagenomic approach may be extended to other metabolites and functional traits in diverse ecosystems.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/100293,
    title ="Extracellular DNA Promotes Efficient Extracellular Electron Transfer by Pyocyanin in Pseudomonas aeruginosa Biofilms",
    author = "Saunders, Scott H. and Tse, Edmund C. M.",
    journal = "Cell",
    volume = "182",
    number = "4",
    pages = "919-932",
    month = "August",
    year = "2020",
    doi = "10.1016/j.cell.2020.07.006",
    issn = "0092-8674",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20191213-144717393",
    note = "© 2020 Elsevier Inc. \n\nReceived 12 December 2019, Revised 19 May 2020, Accepted 9 July 2020, Available online 6 August 2020. \n\nWe thank Jeanyoung Jo, Lars Dietrich, and Matthew Parsek for providing strains and the Biological Imaging Center, GPS Division Analytical Facility, and Beckman Institute Laser Resource Center at Caltech for supporting confocal imaging, SEM imaging, and time resolved spectroscopy, respectively. We also thank three anonymous reviewers for their constructive comments. This work was supported by NIH (1R01AI127850-01A1 to D.K.N. and GM126904 to J.K.B.); ARO (W911NF-17-1-0024 to D.K.N.); ONR (N0001418WX00436 to M.D.Y., S.A.T., and L.M.T.); and Rosen Bioengineering Center at Caltech (to S.H.S., D.K.N., and J.K.B.). E.C.M.T. was supported by a Croucher Foundation Research Fellowship. \n\nAuthor Contributions: Conceptualization, S.H.S., J.K.B., L.M.T., and D.K.N.; Methodology, S.H.S., E.C.M.T., M.D.Y., F.J.O., S.A.T., E.D.A.S., J.K.B., L.M.T., and D.K.N.; Formal Analysis, S.H.S. and L.M.T.; Investigation, S.H.S., E.C.M.T., M.D.Y., F.J.O., S.A.T., and E.D.A.S.; Resources, J.K.B., L.M.T., and D.K.N.; Writing – Original Draft, S.H.S. and D.K.N.; Writing – Review & Editing, S.H.S., E.C.M.T., M.D.Y., F.J.O., S.A.T., E.D.A.S., J.K.B., L.M.T., and D.K.N.; Visualization, S.H.S.; Supervision, J.K.B., L.M.T., and D.K.N.; Funding Acquisition, J.K.B., L.M.T., and D.K.N. \n\nThe authors declare no competing interests.",
    revision_no = "40",
    abstract = "Redox cycling of extracellular electron shuttles can enable the metabolic activity of subpopulations within multicellular bacterial biofilms that lack direct access to electron acceptors or donors. How these shuttles catalyze extracellular electron transfer (EET) within biofilms without being lost to the environment has been a long-standing question. Here, we show that phenazines mediate efficient EET through interactions with extracellular DNA (eDNA) in Pseudomonas aeruginosa biofilms. Retention of pyocyanin (PYO) and phenazine carboxamide in the biofilm matrix is facilitated by eDNA binding. In vitro, different phenazines can exchange electrons in the presence or absence of DNA and can participate directly in redox reactions through DNA. In vivo, biofilm eDNA can also support rapid electron transfer between redox active intercalators. Together, these results establish that PYO:eDNA interactions support an efficient redox cycle with rapid EET that is faster than the rate of PYO loss from the biofilm.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/104783,
    title ="Draft Genome Sequence of the Redox-Active Enteric Bacterium Citrobacter portucalensis Strain MBL",
    author = "Tsypin, Lev M. and Saunders, Scott H.",
    journal = "Microbiology Resource Announcements",
    volume = "9",
    number = "32",
    pages = "Art. No. e00695-20",
    month = "August",
    year = "2020",
    doi = "10.1128/mra.00695-20",
    issn = "2576-098X",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20200806-153947593",
    note = "© 2020 Tsypin et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license. \n\nReceived June 16, 2020. Accepted July 8, 2020. Published online August 6, 2020. \n\nFunding for the Microbial Diversity 2017 program at the MBL was provided by the Hibbit Endowed Education Fund, the Moshe Shilo Memorial Scholarship Fund, the Holger and Friederun Jannasch Scholarship, the U.S. DOE, NASA, the NSF, and the Simons Foundation. L.M.T. was supported by the Rosen Endowment Fellowship at Caltech and an NSF Graduate Research Fellowship (grant DGE‐1745301). Additional support came from NIH (1R01AI127850-01A1) and Army Research Office (W911NF-17-1-0024) grants to D.K.N.",
    revision_no = "26",
    abstract = "We grew a soil enrichment culture to identify organisms that anaerobically oxidize phenazine-1-carboxylic acid. A strain of Citrobacter portucalensis was isolated from this enrichment and sequenced by both Illumina and PacBio technologies. It has a genome with a length of 5.3\u2009Mb, a G+C content of 51.8%, and at least one plasmid.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/101377,
    title ="The potential for redox-active metabolites (RAMs) to enhance or unlock anaerobic survival metabolisms in aerobes",
    author = "Ciemniecki, John A. and Newman, Dianne K.",
    journal = "Journal of Bacteriology",
    volume = "202",
    number = "11",
    pages = "Art. No. e00797-19",
    month = "June",
    year = "2020",
    doi = "10.1128/jb.00797-19",
    issn = "0021-9193",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20200219-111358713",
    note = "© 2020 American Society for Microbiology. \n\nAccepted manuscript posted online 18 February 2020; Published 11 May 2020. \n\nWe thank Megan Bergkessel, Elena Perry, Lev Tsypin, David Basta, and Chelsey VanDrisse for constructive feedback on the manuscript. \n\nJ.A.C. is supported by an NIH Training Grant to Caltech’s BBE Division as well as by grants to D.K.N. from the NIH (1R01AI127850-01A1) and ARO (W911NF-17-1-0024).",
    revision_no = "23",
    abstract = "Classifying microorganisms as “obligate” aerobes has colloquially implied death without air, leading to the erroneous assumption that, without oxygen, they are unable to survive. However, over the past few decades, more than a few obligate aerobes have been found to possess anaerobic energy conservation strategies that sustain metabolic activity in the absence of growth or at very low growth rates. Similarly, studies emphasizing the aerobic prowess of certain facultative aerobes have sometimes led to underrecognition of their anaerobic capabilities. Yet an inescapable consequence of the affinity both obligate and facultative aerobes have for oxygen is that the metabolism of these organisms may drive this substrate to scarcity, making anoxic survival an essential skill. To illustrate this, we highlight the importance of anaerobic survival strategies for Pseudomonas aeruginosa and Streptomyces coelicolor, representative facultative and obligate aerobes, respectively. Included among these strategies, we describe a role for redox-active secondary metabolites (RAMs), such as phenazines made by P. aeruginosa, in enhancing substrate-level phosphorylation. Importantly, RAMs are made by diverse bacteria, often during stationary phase in the absence of oxygen, and can sustain anoxic survival. We present a hypothesis for how RAMs may enhance or even unlock energy conservation pathways that facilitate the anaerobic survival of both RAM producers and nonproducers.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/102971,
    title ="Upregulation of virulence genes promotes Vibrio cholerae biofilm hyperinfectivity",
    author = "Gallego-Hernandez, A. L. and DePas, W. H.",
    journal = "Proceedings of the National Academy of Sciences of the United States of America",
    volume = "117",
    number = "20",
    pages = "11010-11017",
    month = "May",
    year = "2020",
    doi = "10.1073/pnas.1916571117",
    issn = "0027-8424",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20200504-083728290",
    note = "© 2020 National Academy of Sciences. Published under the PNAS license. \n\nEdited by John J. Mekalanos, Harvard University, Boston, MA, and approved March 11, 2020 (received for review September 23, 2019). PNAS first published April 30, 2020. \n\nWe thank Benjamin Abrams, University of California, Santa Cruz (UCSC) Life Sciences Microscopy Center, for technical support during confocal imaging, which is supported by grant S10 OD023528 (to F.H.Y.) from the National Institutes of Health. We thank Ron Taylor and Karen Skorupski for the TcpA antibody. We thank Adam Alpine for his assistance with Fig. 2 A and B. We thank Molecular Technologies (Caltech) for assistance with HCR probe design. We would also like to acknowledge the members of the F.H.Y. laboratory and Karla J. F. Satchell for their useful input and contributions. This work was supported by NIH grants RO1 AI102584, RO1 AI114261, and RO1 AI055987 (to F.H.Y.); NIH grants 5R01HL117328-03 and 1R01AI127850-01A1 (to D.K.N.); and the DEPAS17F0 fellowship from the Cystic Fibrosis Foundation (to W.H.D.), as well as the European Research Council StG-716734, the Human Frontier Science Program CDA00084/2015-C, and the Deutsche Forschungsgemeinschaft SFB987 (to K.D.). A.L.G.-H. was supported by the University of California Institute for Mexico and the United States (UC MEXUS) and El Consejo Nacional de Ciencia y Tecnología (CONACYT) Postdoctoral Research fellowship. \n\nData Availability: RNA-seq data are available through NCBI GEO with series number GSE135887. \n\nAuthor contributions: A.L.G.-H., W.H.D., J.H.P., J.K.T., K.D., D.K.N., and F.H.Y. designed research; A.L.G.-H., W.H.D., J.H.P., and J.K.T. performed research; R.H. contributed new reagents/analytic tools; A.L.G.-H., W.H.D., J.H.P., J.K.T., R.H., H.J., K.D., S.B., D.K.N., and F.H.Y. analyzed data; and A.L.G.-H., W.H.D., J.H.P., J.K.T., K.D., D.K.N., and F.H.Y. wrote the paper. \n\nThe authors declare no competing interest. \n\nThis article is a PNAS Direct Submission. \n\nData deposition: RNA-seq data are available through National Center for Biotechnology Information (NCBI) Gene Expression Omnibus (GEO) (series number GSE135887). \n\nThis article contains supporting information online at https://www.pnas.org/lookup/suppl/doi:10.1073/pnas.1916571117/-/DCSupplemental.",
    revision_no = "24",
    abstract = "Vibrio cholerae remains a major global health threat, disproportionately impacting parts of the world without adequate infrastructure and sanitation resources. In aquatic environments, V. cholerae exists both as planktonic cells and as biofilms, which are held together by an extracellular matrix. V. cholerae biofilms have been shown to be hyperinfective, but the mechanism of hyperinfectivity is unclear. Here we show that biofilm-grown cells, irrespective of the surfaces on which they are formed, are able to markedly outcompete planktonic-grown cells in the infant mouse. Using an imaging technique designed to render intestinal tissue optically transparent and preserve the spatial integrity of infected intestines, we reveal and compare three-dimensional V. cholerae colonization patterns of planktonic-grown and biofilm-grown cells. Quantitative image analyses show that V. cholerae colonizes mainly the medial portion of the small intestine and that both the abundance and localization patterns of biofilm-grown cells differ from that of planktonic-grown cells. In vitro biofilm-grown cells activate expression of the virulence cascade, including the toxin coregulated pilus (TCP), and are able to acquire the cholera toxin-carrying CTXФ phage. Overall, virulence factor gene expression is also higher in vivo when infected with biofilm-grown cells, and modulation of their regulation is sufficient to cause the biofilm hyperinfectivity phenotype. Together, these results indicate that the altered biogeography of biofilm-grown cells and their enhanced production of virulence factors in the intestine underpin the biofilm hyperinfectivity phenotype.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/102703,
    title ="Bacterial defenses against a natural antibiotic promote collateral resilience to clinical antibiotics",
    author = "Meirelles, Lucas A. and Perry, Elena K.",
    
    
    
    
    month = "April",
    year = "2020",
    
    
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20200421-132753787",
    note = "The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license. \n\nPosted April 20, 2020. \n\nWe thank members of the Newman lab and Shashank Gandhi for constructive feedback throughout the project and on the manuscript. We also thank Steven Wilbert for assistance with image analysis, David Basta for providing the plasmid used for lptA deletion, and The Millard and Muriel Jacobs Genetics and Genomics Laboratory at Caltech and Igor Antoshechkin for support during library preparation and sequencing of the Tn-seq samples. Finally, we thank John LiPuma (CFF Burkholderia cepacia Research Laboratory and Repository at the University of Michigan) for providing clinical Burkholderia strains. Grants to D.K.N. from the NIH (1R01AI127850-01A1) and ARO (W911NF-17-1-0024) supported this work. E.K.P. was supported by a National Science Foundation Graduate Research Fellowship under Grant No. DGE-1745301. \n\nAuthor contributions: These authors contributed equally and are listed alphabetically: Lucas A. Meirelles and Elena K. Perry. Study conception: L.A.M., E.K.P., D.K.N. Study design: L.A.M., E.K.P., M.B., D.K.N. Tn-seq: L.A.M, M.B. Tolerance experiments: L.A.M. Resistance experiments: E.K.P. Manuscript preparation: E.K.P., L.A.M., M.B., D.K.N. Study supervision and funding: D.K.N. \n\nThe authors declare no competing interests. \n\nData and materials availability: Tn-seq data have been deposited at GEO under accession number GSE148769. Whole genome sequencing data for Δphz and ciprofloxacin-resistant mutants of P. aeruginosa and B. multivorans AU42096 have been deposited at NCBI under accession number PRJNA625945. All other data that support the findings of this study are available from the corresponding author upon reasonable request.",
    revision_no = "21",
    abstract = "As antibiotic-resistant infections become increasingly prevalent worldwide, understanding the factors that lead to antimicrobial treatment failure is essential to optimizing the use of existing drugs. Opportunistic human pathogens in particular typically exhibit high levels of intrinsic antibiotic resistance and tolerance1, leading to chronic infections that can be nearly impossible to eradicate2. We asked whether the recalcitrance of these organisms to antibiotic treatment could be driven in part by their evolutionary history as environmental microbes, which frequently produce or encounter natural antibiotics3,4. Using the opportunistic pathogen Pseudomonas aeruginosa as a model, we demonstrate that the self-produced natural antibiotic pyocyanin (PYO) activates bacterial defenses that confer collateral tolerance to certain synthetic antibiotics, including in a clinically-relevant growth medium. Non-PYO-producing opportunistic pathogens isolated from lung infections similarly display increased antibiotic tolerance when they are co-cultured with PYO-producing P. aeruginosa. Furthermore, we show that beyond promoting bacterial survival in the presence of antibiotics, PYO can increase the apparent rate of mutation to antibiotic resistance by up to two orders of magnitude. Our work thus suggests that bacterial production of natural antibiotics in infections could play an important role in modulating not only the immediate efficacy of clinical antibiotics, but also the rate at which antibiotic resistance arises in multispecies bacterial communities.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/101176,
    title ="Heat-shock proteases promote survival of Pseudomonas aeruginosa during growth arrest",
    author = "Basta, David W. and Angeles-Albores, David",
    journal = "Proceedings of the National Academy of Sciences of the United States of America",
    volume = "117",
    number = "8",
    pages = "4358-4367",
    month = "February",
    year = "2020",
    doi = "10.1073/pnas.1912082117",
    issn = "0027-8424",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20200206-163326208",
    note = "© 2020 The Author(s). Published under the PNAS license. \n\nEdited by Caroline S. Harwood, University of Washington, Seattle, WA, and approved January 8, 2020 (received for review July 14, 2019) PNAS first published February 6, 2020. \n\nWe thank members of the D.K.N. laboratory for thoughtful discussions and critical feedback on the manuscript, and Lisa Racki (The Scripps Research Institute) for the gift of pLREX97. This manuscript derives from a chapter in D.W.B.’s doctoral thesis from the California Institute of Technology. This work was supported by the Millard and Muriel Jacobs Genetics and Genomics Laboratory at the California Institute of Technology and by the NIH (1R01AI127850-01A1 and 1R21AI146987-01). \n\nAuthor contributions: D.W.B., D.A.-A., M.A.S., J.A.C., and D.K.N. designed research; D.W.B., D.A.-A., M.A.S., and J.A.C. performed research; D.W.B., D.A.-A., M.A.S., J.A.C., and D.K.N. analyzed data; and D.W.B. and D.K.N. wrote the paper. \n\nThe authors declare no competing interest. \n\nThis article is a PNAS Direct Submission. \n\nThis article contains supporting information online at https://www.pnas.org/lookup/suppl/doi:10.1073/pnas.1912082117/-/DCSupplemental.",
    revision_no = "30",
    abstract = "When nutrients in their environment are exhausted, bacterial cells become arrested for growth. During these periods, a primary challenge is maintaining cellular integrity with a reduced capacity for renewal or repair. Here, we show that the heat-shock protease FtsH is generally required for growth arrest survival of Pseudomonas aeruginosa, and that this requirement is independent of a role in regulating lipopolysaccharide synthesis, as has been suggested for Escherichia coli. We find that ftsH interacts with diverse genes during growth and overlaps functionally with the other heat-shock protease-encoding genes hslVU, lon, and clpXP to promote survival during growth arrest. Systematic deletion of the heat-shock protease-encoding genes reveals that the proteases function hierarchically during growth arrest, with FtsH and ClpXP having primary, nonredundant roles, and HslVU and Lon deploying a secondary response to aging stress. This hierarchy is partially conserved during growth at high temperature and alkaline pH, suggesting that heat, pH, and growth arrest effectively impose a similar type of proteostatic stress at the cellular level. In support of this inference, heat and growth arrest act synergistically to kill cells, and protein aggregation appears to occur more rapidly in protease mutants during growth arrest and correlates with the onset of cell death. Our findings suggest that protein aggregation is a major driver of aging and cell death during growth arrest, and that coordinated activity of the heat-shock response is required to ensure ongoing protein quality control in the absence of growth.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/91271,
    title ="Refinement of metabolite detection in cystic fibrosis sputum reveals heme correlates with lung function decline",
    author = "Glasser, Nathaniel R. and Hunter, Ryan C.",
    journal = "PLoS ONE",
    volume = "14",
    number = "12",
    pages = "Art. No. e0226578",
    month = "December",
    year = "2019",
    doi = "10.1371/journal.pone.0226578",
    issn = "1932-6203",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20181128-093526010",
    note = "© 2019 Glasser et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Received: August 26, 2019; Accepted: November 28, 2019; Published: December 18, 2019. The MWCFC Sputum Biomarker investigators included Frederick R Adler, Fadi Asfour, Barbara A Chatfield, Jessica A Francis, John R Hoidal, Judy L Jensen, Yanping Li, Theodore G Liou, Kristyn A Packer, Jane Vroom (University of Utah); Natalia Argel, Peggy Radford (Phoenix Children’s Hospital); Perry S Brown, Dixie Durham (St. Luke’s Cystic Fibrosis Center of Idaho); Cori L Daines, Osmara Molina (University of Arizona); Barbara Glover, Craig Nakamura, Ryan Yoshikawa (Cystic Fibrosis Center, Las Vegas); Theresa Heynekamp, Abby J Redway (University of New Mexico); Ruth Keogh (London School of Hygiene and Tropical Medicine); Carol M Kopecky, Scott D Sagel (Children's Hospital Colorado, University of Colorado School of Medicine); Noah Lechtzin (Johns Hopkins University School of Medicine); Jerimiah Lysinger, Shawna Sprandel (Montana Cystic Fibrosis Center, Billings Clinic); Katie R Poch, Jennifer L Taylor-Cousar (National Jewish Health); Alexandra L Quittner (Miami Children's Research Institute, Nicklaus Children's Hospital); John P Clancy (University of Cincinnati); J Stuart Elborn (Queen’s University, Belfast and Royal Brompton Hospital, London); Kenneth N Olivier (Laboratory of Chronic Airway Infection, Pulmonary Branch, National Heart Lung and Blood Institute, National Institutes of Health). All MWCFC Correspondence should be addressed to Theodore G. Liou (ted.liou@utah.edu). We thank Dr. Nathan Dalleska at the Environmental Analysis Center (Caltech) for analytical support. MWCFC work was supported by grants from the Cystic Fibrosis Foundation (LIOU13A0, LIOU14Y4), the National Center for Advancing Translational Science at the NIH (NCATS/NIH 8UL1TR000105 [formerly UL1RR025764]), the Ben B and Iris M Margolis Foundation of Utah and the Claudia Ruth Goodrich Stevens Endowment Fund. This work was also supported by grants to DKN from the NIH (5R01HL117328-03 and 1R01AI127850-01A1). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Data Availability: All relevant data are within the paper. Author Contributions: Conceptualization: Nathaniel R. Glasser, Dianne K. Newman. Data curation: Nathaniel R. Glasser, Ryan C. Hunter, Theodore G. Liou. Formal analysis: Nathaniel R. Glasser, Ryan C. Hunter, Theodore G. Liou, Dianne K. Newman. Funding acquisition: Theodore G. Liou, Dianne K. Newman. Investigation: Nathaniel R. Glasser, Theodore G. Liou, Dianne K. Newman. Methodology: Nathaniel R. Glasser. Project administration: Ryan C. Hunter, Theodore G. Liou, Dianne K. Newman. Supervision: Dianne K. Newman. Validation: Nathaniel R. Glasser. Writing – original draft: Nathaniel R. Glasser, Ryan C. Hunter, Theodore G. Liou, Dianne K. Newman. Writing – review & editing: Nathaniel R. Glasser, Ryan C. Hunter, Theodore G. Liou, Dianne K. Newman. The authors have declared that no competing interests exist.",
    revision_no = "25",
    abstract = "The bacterial growth environment within cystic fibrosis (CF) sputum is complex, dynamic, and shaped by both host and microbial processes. Characterization of the chemical parameters within sputum that stimulate the in vivo growth of airway pathogens (e.g. Pseudomonas aeruginosa) and their associated virulence factors may lead to improved CF treatment strategies. Motivated by conflicting reports of the prevalence and abundance of P. aeruginosa-derived metabolites known as phenazines within CF airway secretions, we sought to quantify these metabolites in sputum using quadrupole time-of-flight mass spectrometry. In contrast to our previous work, all phenazines tested (pyocyanin (PYO), phenazine-1-carboxylic acid (PCA), phenazine-1-carboxamide, and 1-hydroxyphenazine) were below detection limits of the instrument (0.1 μM). Instead, we identified a late-eluting compound that shared retention time and absorbance characteristics with PCA, yet generated mass spectra and a fragmentation pattern consistent with ferriprotoporphyrin IX, otherwise known as heme B. These data suggested that UV-vis chromatographic peaks previously attributed to PCA and PYO in sputum were mis-assigned. Indeed, retrospective analysis of raw data from our prior study found that the heme B peak closely matched the peaks assigned to PCA, indicating that the previous study likely uncovered a positive correlation between pulmonary function (percent predicted forced expiratory volume in 1 second, or ppFEV1) and heme B, not PCA or any other phenazine. To independently test this observation, we performed a new tandem mass-spectrometry analysis of 71 additional samples provided by the Mountain West CF Consortium Sputum Biomarker study and revealed a positive correlation (ρ = −0.47, p<0.001) between sputum heme concentrations and ppFEV1. Given that hemoptysis is strongly associated with airway inflammation, pulmonary exacerbations and impaired lung function, these new data suggest that heme B may be a useful biomarker of CF pathophysiology.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/95390,
    title ="Quantitative Visualization of Gene Expression in Mucoid and Nonmucoid Pseudomonas aeruginosa Aggregates Reveals Localized Peak Expression of Alginate in the Hypoxic Zone",
    author = "Jorth, Peter and Spero, Melanie A.",
    journal = "mBio",
    volume = "10",
    number = "6",
    pages = "Art. No. e02622-19",
    month = "December",
    year = "2019",
    doi = "10.1128/mbio.02622-19",
    issn = "2150-7511",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20190510-105529016",
    note = "© 2019 Jorth et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license. \n\nReceived 3 October 2019. Accepted 1 November 2019. Published 17 December 2019. \n\nWe thank Will DePas, Ruth Lee, Niles Pierce, Maayan Schwarzkopf, and the Programmable Molecular Technology Center at the Caltech Beckman Institute for technical assistance and advice. Confocal microscopy was performed in the Caltech Biological Imaging Facility at the Caltech Beckman Institute, which is supported by the Arnold and Mabel Beckman Foundation. Grants to D.K.N. from the Army Research Office (W911NF-17-1-0024) and National Institutes of Health (1R01AI127850-01A1 and 1R21AI146987-01) supported this research. P.J. was supported by postdoctoral fellowships from the Cystic Fibrosis Foundation (JORTH14F0 and JORTH17F5) and a grant from the National Institutes of Health (1K22AI127473-01A1). M.A.S. was supported by a gift from the Doren Family Foundation.",
    revision_no = "43",
    abstract = "It is well appreciated that oxygen- and other nutrient-limiting gradients characterize microenvironments within chronic infections that foster bacterial tolerance to treatment and the immune response. However, determining how bacteria respond to these microenvironments has been limited by a lack of tools to study bacterial functions at the relevant spatial scales in situ. Here, we report the application of the hybridization chain reaction (HCR) v3.0 to provide analog mRNA relative quantitation of Pseudomonas aeruginosa single cells as a step toward this end. To assess the potential for this method to be applied to bacterial populations, we visualized the expression of genes needed for the production of alginate (algD) and the dissimilatory nitrate reductase (narG) at single-cell resolution within laboratory-grown aggregates. After validating new HCR probes, we quantified algD and narG expression across microenvironmental gradients within both single aggregates and aggregate populations using the agar block biofilm assay (ABBA). For mucoid and nonmucoid ABBA populations, narG was expressed in hypoxic and anoxic regions, while alginate expression was restricted to the hypoxic zone (∼40 to 200\u2009μM O2). Within individual aggregates, surface-adjacent cells expressed alginate genes at higher levels than interior cells, revealing that alginate expression is not constitutive in mucoid P. aeruginosa but instead varies with oxygen availability. These results establish HCR v3.0 as a versatile and robust tool to resolve subtle differences in gene expression at spatial scales relevant to microbial assemblages. This advance has the potential to enable quantitative studies of microbial gene expression in diverse contexts, including pathogen activities during infections. IMPORTANCE: A goal for microbial ecophysiological research is to reveal microbial activities in natural environments, including sediments, soils, or infected human tissues. Here, we report the application of the hybridization chain reaction (HCR) v3.0 to quantitatively measure microbial gene expression in situ at single-cell resolution in bacterial aggregates. Using quantitative image analysis of thousands of Pseudomonas aeruginosa cells, we validated new P. aeruginosa HCR probes. Within in vitro P. aeruginosa aggregates, we found that bacteria just below the aggregate surface are the primary cells expressing genes that protect the population against antibiotics and the immune system. This observation suggests that therapies targeting bacteria growing with small amounts of oxygen may be most effective against these hard-to-treat infections. More generally, this proof-of-concept study demonstrates that HCR v3.0 has the potential to identify microbial activities in situ at small spatial scales in diverse contexts.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/98876,
    title ="Chlorate’s Potential as a Pro-Drug for Killing Antibiotic-Tolerant Pathogens in the Cystic Fibrosis Lung\n",
    author = "Spero, M. A. and Silveira, C. B.",
    journal = "Pediatric Pulmonology",
    volume = "54",
    number = "S2",
    pages = "S281",
    month = "October",
    year = "2019",
    
    issn = "8755-6863",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20190926-131636762",
    note = "© 2019 Wiley Periodicals, Inc. \n\nFirst published: 05 September 2019.",
    revision_no = "11",
    abstract = "Chronic lung infections are a major contributor to morbidity and\nmortality in cystic fibrosis (CF) patients. Perhaps the most notorious CF pathogen is Pseudomonas aeruginosa, which establishes decades-long lung infections despite aggressive antibiotic treatment. In part, drugs fail to clear P. aeruginosa lung infections because some pathogen populations exhibit antibiotic tolerance, a metabolic state that reduces a cell’s susceptibility to drugs. Antibiotic tolerance is associated with low metabolic activity, and P. aeruginosa growth is limited by oxygen availability in the largely hypoxic/anoxic CF sputum.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/91894,
    title ="Extended hopanoid loss reduces bacterial motility and surface attachment, and leads to heterogeneity in root nodule growth kinetics in a Bradyrhizobium-Aeschynomene symbiosis",
    author = "Belin, Brittany J. and Tookmanian, Elise T.",
    journal = "Molecular Plant-Microbe Interactions",
    volume = "32",
    number = "10",
    pages = "1415-1428",
    month = "October",
    year = "2019",
    doi = "10.1094/MPMI-04-19-0111-R",
    issn = "0894-0282",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20181218-110603835",
    note = "© 2019 The American Phytopathological Society. \n\nThis work was supported by grants from the HHMI (D.K.N.), NASA (NNX12AD93G, D.K.N.), Jane Coffin Childs Memorial Fund (B.J.B.), NIH (K99GM126141, B.J.B.), and Army Research Office (W911NF-18-1-0254, GW) and predoctoral 635 fellowships from NSF (E.T.) and the Ford Foundation (J.d.A.). We thank Dr. Eric Giraud for his generous gift of A. afraspera seeds and training on Aeschynomene symbioses and Drs. Hans-Martin Fischer and Raphael Ledermann for plasmids and technical advice for the genetic transformation of B. diazoefficiens. Dr. Nathan Dalleska of the Environmental Analysis Center at Caltech was instrumental in providing training and support for GC-MS analysis of acetylene reduction. We are grateful to Dr. Gargi Kulkarni and other members of the Newman lab, as well as Drs. Elliot Meyerowitz and Rob Phillips, for their collegiality and thoughtful discussions about this work. We are indebted to Ms. Shannon Park and Ms. Kristy Nguyen for providing the administrative assistance that allows us to focus on our research.",
    revision_no = "25",
    abstract = "Hopanoids are steroid-like bacterial lipids that enhance membrane rigidity and promote bacterial growth under diverse stresses. Roughly 10% of bacteria contain genes involved in hopanoid biosynthesis, and these genes are particularly conserved in plant-associated organisms. We previously found that the extended class of hopanoids (C35) in the nitrogen-fixing soil bacterium Bradyrhizobium diazoefficiens promotes its root nodule symbiosis with the tropical legume Aeschynomene afraspera. By quantitatively modeling root nodule development, we identify independent consequences of extended hopanoid loss in the initiation of root nodule formation and in the rate of root nodule maturation. In vitro studies demonstrate that extended hopanoids support B. diazoefficiens motility and surface attachment, which may correlate with stable root colonization in planta. Confocal microscopy of maturing root nodules reveals that root nodules infected with extended hopanoid-deficient B. diazoefficiens contain unusually low densities of bacterial symbionts, indicating that extended hopanoids are necessary for persistent, high levels of host infection.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/98873,
    title ="Nontuberculous Mycobacterial Aggregation is Regulated by C:N Balance",
    author = "DePas, W. and Bergkessel, M.",
    journal = "Pediatric Pulmonology",
    volume = "54",
    number = "S2",
    pages = "S310",
    month = "October",
    year = "2019",
    
    issn = "8755-6863",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20190926-103130771",
    note = "© 2019 Wiley Periodicals, Inc. \n\nFirst published: 05 September 2019. \n\nSupported by the Cystic Fibrosis Foundation and the National Institutes of Health.",
    revision_no = "9",
    abstract = "The current treatment regimen for nontuberculous mycobacteria (NTM) involves long courses of antibiotic cocktails that demonstrate limited efficacy and cause frequent and serious side effects. Mycobacterium abscessus, in particular, is difficult to treat, motivating studies to identify new therapeutic targets. Experiments using zebrafish and human cell culture lines have demonstrated that M. abscessus is more virulent when aggregated into cord-like biofilms, at least in part because of the decreased ability of phagocytes to efficiently engulf and kill corded M. abscessus cells.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/98789,
    title ="A contractile injection system stimulates tubeworm metamorphosis by translocating a proteinaceous effector",
    author = "Ericson, Charles F. and Eisenstein, Fabian",
    journal = "eLife",
    volume = "8",
    
    pages = "Art. No. e46845",
    month = "September",
    year = "2019",
    doi = "10.7554/elife.46845",
    issn = "2050-084X",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20190923-092135402",
    note = "© 2019 Ericson et al. This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited. \n\nReceived: 14 March 2019; Accepted: 05 August 2019; Published: 17 September 2019. \n\nWe thank Dr. Anca Segall and Dr. Manal Swairjo for reagents, technical support and constructive suggestions. We thank Ms. Amanda Alker and Ms. Nathalie Delherbe for their constructive suggestions. ScopeM at ETHZ and Ohad Medalia at the University of Zürich are acknowledged for instrument access. We thank Paula Picotti and Marco Faini for discussions of mass spectrometry experiments. This work was supported by the Harold and June Memorial Scholarship (CFE), Norma Sullivan Memorial Endowed Scholarship (CFE), Howard Hughes Medical Institute (DKN), NIH NIDCD (1R21DC013180-01A1, RWZ), Office of Naval Research (N00014-17-1-2677, NJS and SB), Office of Naval Research (N00014-16-1-2135, NJS), Office of Naval Research (N00014-14-1-0340, NJS and DKN), Alfred P Sloan Foundation, Sloan Research Fellowship (NJS), San Diego State University (NJS), European Research Council (679209, MP), Swiss National Science Foundation (31003A_179255, MP) and Gebert Rüf Foundation (MP). \n\nThe funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication. \n\nAuthor contributions: Charles F Ericson, Kyle E Malter, Formal analysis, Investigation, Visualization, Methodology, Writing—original draft, Writing—review and editing; Fabian Eisenstein, João M Medeiros, Formal analysis, Investigation, Visualization, Methodology, Writing—review and editing; Giselle S Cavalcanti, Investigation, Writing—review and editing; Robert W Zeller, Resources, Funding acquisition, Methodology, Writing—review and editing; Dianne K Newman, Conceptualization, Resources, Supervision, Funding acquisition, Validation, Project administration, Writing—review and editing; Martin Pilhofer, Conceptualization, Resources, Data curation, Formal analysis, Supervision, Funding acquisition, Investigation, Visualization, Methodology, Writing—original draft, Project administration, Writing—review and editing; Nicholas J Shikuma, Conceptualization, Resources, Supervision, Funding acquisition, Validation, Investigation, Visualization, Methodology, Writing—original draft, Project administration, Writing—review and editing. \n\nData availability: Subtomogram averages were deposited in the Electron Microscopy Data Bank (accession numbers EMD-4730 and EMD-4731).",
    revision_no = "14",
    abstract = "The swimming larvae of many marine animals identify a location on the sea floor to undergo metamorphosis based on the presence of specific bacteria. Although this microbe–animal interaction is critical for the life cycles of diverse marine animals, what types of biochemical cues from bacteria that induce metamorphosis has been a mystery. Metamorphosis of larvae of the tubeworm Hydroides elegans is induced by arrays of phage tail-like contractile injection systems, which are released by the bacterium Pseudoalteromonas luteoviolacea. Here we identify the novel effector protein Mif1. By cryo-electron tomography imaging and functional assays, we observe Mif1 as cargo inside the tube lumen of the contractile injection system and show that the mif1 gene is required for inducing metamorphosis. Purified Mif1 is sufficient for triggering metamorphosis when electroporated into tubeworm larvae. Our results indicate that the delivery of protein effectors by contractile injection systems may orchestrate microbe–animal interactions in diverse contexts.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/90464,
    title ="The dormancy-specific regulator, SutA, is intrinsically disordered and modulates transcription initiation in Pseudomonas aeruginosa",
    author = "Bergkessel, Megan and Babin, Brett M.",
    journal = "Molecular Microbiology",
    volume = "112",
    number = "3",
    pages = "992-1009",
    month = "September",
    year = "2019",
    doi = "10.1111/mmi.14337",
    issn = "0950-382X",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20181029-102517113",
    note = "© 2019 John Wiley & Sons Ltd. \n\nIssue Online: 10 September 2019; Version of Record online: 10 July 2019; Accepted manuscript online: 28 June 2019; Manuscript accepted: 25 June 2019. \n\nWe thank Ben Ramirez (University of Illinois at Chicago) for helping us with preliminary NMR studies of SutA, Jacqueline Barton (Caltech) for giving us access to her lab to perform experiments involving radioactivity, Nate Glasser for help with HPLC measurements to quantify SutA, Hsiau‐Wei (Jack) Lee and Aimee Marceau (University of California, Santa Cruz) for help with the NMR binding experiment using the Bruker AVIII HD 800 MHz NMR, Weidong Hu (City of Hope) for help with NMR experiments using the Bruker AV III 700 MHz spectrometer, and Julia Kardon and Niels Bradshaw (Brandeis University) and members of the Newman lab for feedback on the project at different stages. MB was supported by a postdoctoral fellowship from the Cystic Fibrosis Foundation (BERGKE16F0). Grants from the NIH (GM067153) to IA and grants from the NIH (R01HL117328 and 1R01AI127850‐01A1) to DKN supported this work. The Proteome Exploration Laboratory is supported by the Beckman Institute and NIH 1S10OD02001301. This work was also supported by the Institute for Collaborative Biotechnologies through grant W911NF‐09‐0001 from the U.S. Army Research Office. The content of the information does not necessarily reflect the position or the policy of the Government and no official endorsement should be inferred. \n\nThe authors have no conflicts of interest to disclose.",
    revision_no = "37",
    abstract = "Though most bacteria in nature are nutritionally limited and grow slowly, our understanding of core processes like transcription comes largely from studies in model organisms doubling rapidly. We previously identified a small protein of unknown function, SutA, in a screen of proteins synthesized in Pseudomonas aeruginosa during dormancy. SutA binds RNA polymerase (RNAP), causing widespread changes in gene expression, including upregulation of the ribosomal RNA genes. Here, using biochemical and structural methods, we examine how SutA interacts with RNAP and the functional consequences of these interactions. We show that SutA comprises a central α‐helix with unstructured N‐ and C‐terminal tails, and binds to the β1 domain of RNAP. It activates transcription from the rrn promoter by both the housekeeping sigma factor holoenzyme (Eσ^(70)) and the stress sigma factor holoenzyme (Eσ^S) in vitro, but has a greater impact on Eσ^S. In both cases, SutA appears to affect intermediates in the open complex formation and its N‐terminal tail is required for activation. The small magnitudes of in vitro effects are consistent with a role in maintaining activity required for homeostasis during dormancy. Our results add SutA to a growing list of transcription regulators that use their intrinsically disordered regions to remodel transcription complexes.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/95386,
    title ="Aggregation of nontuberculous mycobacteria is regulated by carbon:nitrogen balance",
    author = "DePas, William H. and Bergkessel, Megan",
    journal = "mBio",
    volume = "10",
    number = "4",
    pages = "Art. No. e01715-19",
    month = "July",
    year = "2019",
    doi = "10.1128/mBio.01715-19",
    issn = "2150-7511",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20190509-154449662",
    note = "© 2019 DePas et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license. \n\nReceived 28 June 2019; Accepted 22 July 2019; Published 13 August 2019. \n\nWe thank the Cystic Fibrosis Foundation (grants DEPAS17F0 to W.H.D. and BERGKE16F0 to M.B.) and the NIH (grant 1R01AI127850-01A1 to D.K.N.) for supporting our research. A portion of the imaging was performed in the Caltech Biological Imaging Facility, with the support of the Caltech Beckman Institute and the Arnold and Mabel Beckman Foundation. \n\nLindsay Caverly (CS Mott Children’s Hospital, University of Michigan, Ann Arbor, MI, USA) graciously provided M. abscessus clinical isolates (NTM0253a, NTM0253b, NTM0711a, and NTM0711b), William Jacobs (Albert Einstein College of Medicine, Bronx, NY, USA) provided WT M. smegmatis MC^2155, and William Bishai (Johns Hopkins University School of Medicine, Baltimore, MD, USA) provided plasmids pJV53 and pMH94. Igor Antoshechkin and the Millard and Muriel Jacobs Genetics and Genomics Laboratory at Caltech assisted with genome sequencing. Alex Sessions and Fenfang Wu (Caltech) helped with C/N measurements and analysis. Ion chromatography instrumentation used for this work is located in the Environmental Analysis Center at Caltech. We acknowledge Nathan F. Dalleska, Lev Tsypin, and Melanie Spero for ion chromatography method support. SEM was performed at the Caltech GPS Division Analytical Facility with the assistance of Chi Ma.",
    revision_no = "22",
    abstract = "Nontuberculous mycobacteria (NTM) are emerging opportunistic pathogens that colonize household water systems and cause chronic lung infections in susceptible patients. The ability of NTM to form surface-attached biofilms in the nonhost environment and corded aggregates in vivo is important to their ability to persist in both contexts. Underlying the development of these multicellular structures is the capacity of mycobacterial cells to adhere to one another. Unlike most other bacteria, NTM spontaneously and constitutively aggregate in vitro, hindering our ability to understand the transition between planktonic and aggregated cells. While culturing a model NTM, Mycobacterium smegmatis, in rich medium, we fortuitously discovered that planktonic cells accumulate after ∼3\u2009days of growth. By providing selective pressure for bacteria that disperse earlier, we isolated a strain with two mutations in the oligopeptide permease operon (opp). A mutant lacking the opp operon (Δopp) disperses earlier than wild type (WT) due to a defect in nutrient uptake. Experiments with WT M. smegmatis revealed that growth as aggregates is favored when carbon is replete, but under conditions of low available carbon relative to available nitrogen, M. smegmatis grows as planktonic cells. By adjusting carbon and nitrogen sources in defined medium, we tuned the cellular C/N ratio such that M. smegmatis grows either as aggregates or as planktonic cells. C/N-mediated aggregation regulation is widespread among NTM with the possible exception of rough-colony Mycobacterium abscessus isolates. Altogether, we show that NTM aggregation is a controlled process that is governed by the relative availability of carbon and nitrogen for metabolism.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/94806,
    title ="The transcription factors ActR and SoxR differentially affect the phenazine tolerance of Agrobacterium tumefaciens",
    author = "Perry, Elena K. and Newman, Dianne K.",
    journal = "Molecular Microbiology",
    volume = "112",
    number = "1",
    pages = "199-218",
    month = "July",
    year = "2019",
    doi = "10.1111/mmi.14263",
    issn = "0950-382X",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20190419-100142089",
    note = "© 2019 John Wiley & Sons. \n\nIssue Online: 09 July 2019; Version of Record online:\n03 May 2019; Accepted manuscript online: 19 April 2019;\nManuscript accepted: 13 April 2019.\n\nWe thank all members of the Newman lab for helpful advice, discussions and feedback on the manuscript, and Clay Fuqua for generously providing pNPTS138. We also thank the Marine Biological Laboratory Microbial Diversity Course, Class of 2017, for assistance with screening transposon mutants. This material is based upon work supported by the National Science Foundation Graduate Research Fellowship under Grant No. DGE‐1745301. This work was also supported by the Millard and Muriel Jacobs Genetics and Genomics Laboratory at the California Institute of Technology (Caltech), the Caltech Electron Paramagnetic Resonance Spectroscopy Facility and grants to DKN from the ARO (W911NF‐17‐1‐0024) and NIH 341 (1R01AI127850‐01A1).\n\nData availability statement:\nThe data that support the findings of this study are available from the corresponding author upon reasonable request.\n\nAuthor contributions:\nEKP and DKN conceived the study and designed the experiments. EKP performed the experiments, analyzed and interpreted data and wrote the manuscript. DKN contributed to data interpretation, obtained funding and edited the manuscript.",
    revision_no = "20",
    abstract = "Bacteria in soils encounter redox‐active compounds, such as phenazines, that can generate oxidative stress, but the mechanisms by which different species tolerate these compounds are not fully understood. Here, we identify two transcription factors, ActR and SoxR, that play contrasting yet complementary roles in the tolerance of the soil bacterium Agrobacterium tumefaciens to phenazines. We show that ActR promotes phenazine tolerance by proactively driving expression of a more energy‐efficient terminal oxidase at the expense of a less efficient alternative, which may affect the rate at which phenazines abstract electrons from the electron transport chain (ETC) and thereby generate reactive oxygen species. SoxR, on the other hand, responds to phenazines by inducing expression of several efflux pumps and redox‐related genes, including one of three copies of superoxide dismutase and five novel members of its regulon that could not be computationally predicted. Notably, loss of ActR is far more detrimental than loss of SoxR at low concentrations of phenazines, and also increases dependence on the otherwise functionally redundant SoxR‐regulated superoxide dismutase. Our results thus raise the intriguing possibility that the composition of an organism's ETC may be the driving factor in determining sensitivity or tolerance to redox‐active compounds.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/92544,
    title ="Draft Genome Sequence of the Iridescent Marine Bacterium Tenacibaculum discolor Strain IMLK18",
    author = "Kee, H. Lynn and Mikheyeva, Irina V.",
    journal = "Microbiology Resource Announcements",
    volume = "8",
    number = "5",
    pages = "Art. No. e01683-18",
    month = "January",
    year = "2019",
    doi = "10.1128/mra.01683-18",
    issn = "2576-098X",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20190131-104830799",
    note = "© 2019 Kee et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license. \n\nReceived 18 December 2018. Accepted 8 January 2019. Published 31 January 2019. \n\nThis isolation and culturing of T. discolor IMLK18 were conducted during the 2016 Microbial Diversity summer course at the Marine Biological Laboratory (Woods Hole, MA), and genomic sequencing was conducted during the 2018 Microbial Diversity summer course. We thank Patrick Degnan, Whitney England, Rachel Whitaker, George O’Toole, Callie Rogers, Jose de la Torre, Gabriela Kovacikova, Titus Brown, and Kyle Costa for their assistance and advice. The Promega Corporation donated the molecular reagents used in this project. \n\nH.L.K. was supported by funding from Stetson University and the Marine Biological Laboratory (Whitman Center Fellowship). Participation in and research activities during the summer program were supported by the Simons Foundation (grant 309981 to the Marine Biological Laboratory), the Helmsley Charitable Trust, the Waksman Foundation, Howard Hughes Medical Institute, the National Aeronautics and Space Administration (grant NNA13AA92A), the National Science Foundation (grant DEB-1822263), and the U.S. Department of Energy (grant DE-SC0016127). These funders had no role in the study design, data collection and interpretation, or the decision to submit the work for publication. \n\nData availability.The GenBank accession number for this genome sequence is RCVH00000000, and the SRA accession number for the Illumina sequencing run is SRS4170023.",
    revision_no = "11",
    abstract = "We report here the draft genome sequence of a strain of Tenacibaculum discolor (Bacteroidetes) that was isolated from the river-ocean interface at Trunk River in Falmouth, Massachusetts. The isolation and genomic sequencing were performed during the 2016 and 2018 Microbial Diversity summer programs at the Marine Biological Laboratory in Woods Hole, Massachusetts.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/90094,
    title ="Towards measuring growth rates of pathogens during infections by D_2O-labeling lipidomics",
    author = "Neubauer, Cajetan and Sessions, Alex L.",
    journal = "Rapid Communications in Mass Spectrometry",
    volume = "32",
    number = "24",
    pages = "2129-2140",
    month = "December",
    year = "2018",
    doi = "10.1002/rcm.8288",
    issn = "0951-4198",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20181003-105017642",
    note = "© 2018 John Wiley & Sons. \n\nVersion of Record online: 11 November 2018; Accepted manuscript online: 25 September 2018; Manuscript accepted: 18 September 2018; Manuscript revised: 10 August 2018; Manuscript received: 23 May 2018. \n\nWe thank all reviewers for comments. We are grateful to Drs. Fenfang Wu, Reto Wijker and Jesse Allen for technical advice on the use of instrumentation, and to Dr. Ajay Kasai for CF sputum collection. LC/MS data was collected at the Caltech Environmental Analysis Center (Pasadena) and with the assistance of Dr. Anastasia Kalli at Thermo Fisher Scientific (San Jose, CA, USA). This work was made possible in parts by grants from National Aeronautics and Space Administration (NNX12AD93G), the National Science Foundation (1224158), and the National Institutes of Health (R01HL117328). IRB was funded by CEMI (Caltech) and by the Leverhulme Trust.",
    revision_no = "22",
    abstract = "Rationale: Microbial growth rate is an important physiological parameter that is challenging to measure in situ, partly because microbes grow slowly in many environments. Recently, it has been demonstrated that generation times of S. aureus in cystic fibrosis (CF) infections can be determined by D_2O‐labeling of actively synthesized fatty acids. To improve species specificity and allow growth rate monitoring for a greater range of pathogens during the treatment of infections, it is desirable to accurately quantify trace incorporation of deuterium into phospholipids. \n\nMethods: Lipid extracts of D_2O‐treated E. coli cultures were measured on liquid chromatography/electrospray ionization mass spectrometry (LC/ESI‐MS) instruments equipped with time‐of‐flight (TOF) and orbitrap mass analyzers, and used for comparison with the analysis of fatty acids by isotope‐ratio gas chromatography (GC)/MS. We then developed an approach to enable tracking of lipid labeling, by following the transition from stationary into exponential growth in pure cultures. Lastly, we applied D_2O‐labeling lipidomics to clinical samples from CF patients with chronic lung infections. \n\nResults: Lipidomics facilitates deuterium quantification in lipids at levels that are useful for many labeling applications (>0.03 at% D). In the E. coli cultures, labeling dynamics of phospholipids depend largely on their acyl chains and between phospholipids we notice differences that are not obvious from absolute concentrations alone. For example, cyclopropyl‐containing lipids reflect the regulation of cyclopropane fatty acid synthase, which is predominantly expressed at the beginning of stationary phase. The deuterium incorporation into a lipid that is specific for S. aureus in CF sputum indicates an average generation time of the pathogen on the order of one cell doubling per day. \n\nConclusions: This study demonstrates how trace level measurement of stable isotopes in intact lipids can be used to quantify lipid metabolism in pure cultures and provides guidelines that enable growth rate measurements in microbiome samples after incubation with a low percentage of D_2O.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/89888,
    title ="Both toxic and beneficial effects of pyocyanin contribute to the lifecycle of Pseudomonas aeruginosa",
    author = "Meirelles, Lucas A. and Newman, Dianne K.",
    journal = "Molecular Microbiology",
    volume = "110",
    number = "6",
    pages = "995-1010",
    month = "December",
    year = "2018",
    doi = "10.1111/mmi.14132",
    issn = "0950-382X",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20180924-131756393",
    note = "© 2018 John Wiley & Sons Ltd. \n\nIssue Online: 05 December 2018; Version of Record online: 23 October 2018; Accepted manuscript online: 19 September 2018; Manuscript accepted: 14 September 2018. \n\nWe thank all the members of the Newman lab for experimental advice and feedback on the manuscript; Megan Bergkessel was particularly generous with her time and intellectual support. We also thank Scott Saunders for helping with pyocyanin quantification. Grants to D.K.N. from the ARO (W911NF‐17‐1‐0024) and NIH 341 (1R01AI127850‐01A1) supported this research.",
    revision_no = "19",
    abstract = "Pseudomonas aeruginosa, an opportunistic pathogen, produces redox‐active pigments called phenazines. Pyocyanin (PYO, the blue phenazine) plays an important role during biofilm development. Paradoxically, PYO auto‐poisoning can stimulate cell death and release of extracellular DNA (eDNA), yet PYO can also promote survival within biofilms when cells are oxidant‐limited. Here, we identify the environmental and physiological conditions in planktonic culture that promote PYO‐mediated cell death. We demonstrate that PYO auto‐poisoning is enhanced when cells are starved for carbon. In the presence of PYO, cells activate a set of genes involved in energy‐dependent defenses, including: (i) the oxidative stress response, (ii) RND efflux systems and (iii) iron‐sulfur cluster biogenesis factors. P. aeruginosa can avoid PYO poisoning when reduced carbon is available, but blockage of adenosine triphosphate (ATP) synthesis either through carbon limitation or direct inhibition of the F_0F_1‐ATP synthase triggers death and eDNA release. Finally, even though PYO is toxic to the majority of the population when cells are nutrient limited, a subset of cells is intrinsically PYO resistant. The effect of PYO on the producer population thus appears to be dynamic, playing dramatically different yet predictable roles throughout distinct stages of growth, helping rationalize its multifaceted contributions to biofilm development.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/90127,
    title ="Refining the Application of Microbial Lipids as Tracers of Staphylococcus aureus Growth Rates in Cystic Fibrosis Sputum",
    author = "Neubauer, Cajetan and Kasi, Ajay S.",
    journal = "Journal of Bacteriology",
    volume = "200",
    number = "24",
    pages = "Art. No. e00365-18",
    month = "December",
    year = "2018",
    doi = "10.1128/jb.00365-18",
    issn = "0021-9193",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20181004-081517527",
    note = "© 2018 American Society for Microbiology. \n\nReceived 14 June 2018; Accepted 19 September 2018; Accepted manuscript posted online 24 September 2018. \n\nWe thank Elise Cowley, Reto Wijker, and Fenfang Wu for helping with our study. We thank Dominique H. Limoli and George O'Toole for providing CFBE cells and Ram Balasubramanian for providing PMN and HL-60 cells. We thank Jennifer Dien Bard and Thomas G. Keens for their guidance with our study, the CHLA CF center team, and patients of the CHLA CF clinic for participating in this study. We thank the anonymous reviewers for their comments. \n\nThis work was funded by grants from the National Institutes of Health (R01HL117328).",
    revision_no = "31",
    abstract = "Chronic lung infections in cystic fibrosis (CF) could be treated more effectively if the effects of antimicrobials on pathogens in situ were known. Here, we compared changes in the microbial community composition and pathogen growth rates in longitudinal studies of seven pediatric CF patients undergoing intravenous antibiotic administration during pulmonary exacerbations. The microbial community composition was determined by counting rRNA with NanoString DNA analysis, and growth rates were obtained by incubating CF sputum with heavy water and tracing incorporation of deuterium into two branched-chain (“anteiso”) fatty acids (a-C_(15:0) and a-C_(17:0)) using gas chromatography-mass spectrometry (GC/MS). Prior to this study, both lipids were thought to be specific for Staphylococcaceae; hence, their isotopic enrichment was interpreted as a growth proxy for Staphylococcus aureus. Our experiments revealed, however, that Prevotella is also a relevant microbial producer of a-C_(17:0) fatty acid in some CF patients; thus, deuterium incorporation into these lipids is better interpreted as a more general pathogen growth rate proxy. Even accounting for a small nonmicrobial background source detected in some patient samples, a-C_(15:0) fatty acid still appears to be a relatively robust proxy for CF pathogens, revealing a median generation time of ∼1.5 days, similar to prior observations. Contrary to our expectation, pathogen growth rates remained relatively stable throughout exacerbation treatment. We suggest two straightforward “best practices” for application of stable-isotope probing to CF sputum metabolites: (i) parallel determination of microbial community composition in CF sputum using culture-independent tools and (ii) assessing background levels of the diagnostic metabolite.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/90892,
    title ="Extracellular Electron Transfer Transcends Microbe-Mineral Interactions",
    author = "Saunders, Scott H. and Newman, Dianne K.",
    journal = "Cell Host & Microbe",
    volume = "24",
    number = "5",
    pages = "611-613",
    month = "November",
    year = "2018",
    doi = "10.1016/j.chom.2018.10.018",
    issn = "1931-3128",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20181114-140802025",
    note = "© 2018 Elsevier Inc. \n\nWe thank Ken Nealson, Daniel Dar, and John Ciemniecki for constructive comments on the manuscript and the ARO (W911NF-17-1-0024) and NIH (1R01AI127850-01A1) for supporting our EET research.",
    revision_no = "12",
    abstract = "Extracellular electron transfer (EET) allows microbes to drive their metabolism through interactions with minerals or electrodes. In recent work, Light et al. (2018) discover a specialized EET pathway in Listeria monocytogenes with homologs in pathogens and gut commensals, suggesting that EET plays important roles in diverse environments.",
}
@article {CaltechAUTHORS_https://authors.library.caltech.edu/id/eprint/88863,
    title ="Scanning the isotopic structure of molecules by tandem mass spectrometry",
    author = "Neubauer, Cajetan and Sweredoski, Michael J.",
    journal = "International Journal of Mass Spectrometry",
    volume = "434",
    
    pages = "276-286",
    month = "November",
    year = "2018",
    doi = "10.1016/j.ijms.2018.08.001",
    issn = "1387-3806",
    
    url = "https://resolver.caltech.edu/CaltechAUTHORS:20180816-125733829",
    note = "© 2018 Elsevier B.V. \n\nReceived 10 May 2018, Revised 23 July 2018, Accepted 7 August 2018, Available online 16 August 2018.",
    revision_no = "12",
    abstract = "Biomolecules generally exist as mixtures of diverse isotopologues that differ in the number and sites of rare-isotope substitutions. The exact proportion of isotopologues of a biomolecule may depend on the molecular, cellular, organismal and environmental factors involved in its biosynthesis, localization and consumption. Molecular isotopic structure can thus be a valuable tool to elucidate biochemical mechanisms and for the reconstruction of physiological, ecological and climatic processes. However, little information about this record is accessible by conventional methods of stable isotope chemistry. Here, we report an easy to implement mass spectrometric method that quantifies intramolecular isotope distributions and is specifically designed for use on samples containing low, natural abundances of the rare isotopes. Its essential feature is the use of a narrow initial mass selection window to isolate ions that are heavier due to the presence of one or more isotopic substitutions. This pre-selection greatly increases the relative proportions of the various rare-isotope substituted isotopologues. The selected ions are then fragmented, and within seconds to minutes the isotopic pattern of the fragment peaks reveals information about the intramolecular distribution of isotopes. This approach requires ~0.1 to 10 nanomole of analyte, which is about five orders of magnitude less than NMR. We demonstrate the ability to measure the site-specific isotope ratios of metabolites by resolving the ^(13)C content in the amino acid methionine among multiple non-equivalent carbon sites. This technique enables rapid origin assignments for a wide range of organic molecules and can be used for new applications of molecular isotopic structure in medicine and environmental sciences.",
}