{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,5,27]],"date-time":"2026-05-27T12:54:10Z","timestamp":1779886450183,"version":"3.53.1"},"reference-count":38,"publisher":"AIP Publishing","issue":"20","funder":[{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["R01 AI113219"],"award-info":[{"award-number":["R01 AI113219"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["pubs.aip.org"],"crossmark-restriction":true},"short-container-title":[],"published-print":{"date-parts":[[2021,5,28]]},"abstract":"<jats:p>Pseudomonas aeruginosa is an opportunistic human pathogen implicated in both acute and chronic diseases, which resists antibiotic treatment, in part by forming physical and chemical barriers such as biofilms. Here, we explore the use of confocal Raman imaging to characterize the three-dimensional (3D) spatial distribution of alkyl quinolones (AQs) in P. aeruginosa biofilms by reconstructing depth profiles from hyperspectral Raman data. AQs are important to quorum sensing (QS), virulence, and other actions of P. aeruginosa. Three-dimensional distributions of three different AQs (PQS, HQNO, and HHQ) were observed to have a significant depth, suggesting 3D anisotropic shapes\u2014sheet-like rectangular solids for HQNO and extended cylinders for PQS. Similar to observations from 2D imaging studies, spectral features characteristic of AQs (HQNO or PQS) and the amide I vibration from peptide-containing species were found to correlate with the PQS cylinders typically located at the tips of the HQNO rectangular solids. In the QS-deficient mutant lasIrhlI, a small globular component was observed, whose highly localized nature and similarity in size to a P. aeruginosa cell suggest that the feature arises from HHQ localized in the vicinity of the cell from which it was secreted. The difference in the shapes and sizes of the aggregates of the three AQs in wild-type and mutant P. aeruginosa is likely related to the difference in the cellular response to growth conditions, environmental stress, metabolic levels, or other structural and biochemical variations inside biofilms. This study provides a new route to characterizing the 3D structure of biofilms and shows the potential of confocal Raman imaging to elucidate the nature of heterogeneous biofilms in all three spatial dimensions. These capabilities should be applicable as a tool in studies of infectious diseases.<\/jats:p>","DOI":"10.1063\/5.0052785","type":"journal-article","created":{"date-parts":[[2021,5,25]],"date-time":"2021-05-25T09:58:44Z","timestamp":1621936724000},"update-policy":"https:\/\/doi.org\/10.1063\/aip-crossmark-policy-page","source":"Crossref","is-referenced-by-count":8,"title":["Depth distributions of signaling molecules in <i>Pseudomonas aeruginosa<\/i> biofilms mapped by confocal Raman microscopy"],"prefix":"10.1063","volume":"154","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-5854-1972","authenticated-orcid":false,"given":"Tianyuan","family":"Cao","sequence":"first","affiliation":[{"name":"Department of Chemistry and Biochemistry, University of Notre Dame 1 , Notre Dame, Indiana 46556, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3757-2902","authenticated-orcid":false,"given":"Abigail A.","family":"Weaver","sequence":"additional","affiliation":[{"name":"Department of Civil and Environmental Engineering and Earth Sciences, University of Notre Dame 2 , Notre Dame, Indiana 46556, USA"},{"name":"Institute for Precision Health, University of Notre Dame 3 , Notre Dame, Indiana 46556, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2191-6723","authenticated-orcid":false,"given":"Seol","family":"Baek","sequence":"additional","affiliation":[{"name":"Department of Chemistry and Biochemistry, University of Notre Dame 1 , Notre Dame, Indiana 46556, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2194-5607","authenticated-orcid":false,"given":"Jin","family":"Jia","sequence":"additional","affiliation":[{"name":"Department of Chemistry and Biochemistry, University of Notre Dame 1 , Notre Dame, Indiana 46556, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9509-2187","authenticated-orcid":false,"given":"Joshua D.","family":"Shrout","sequence":"additional","affiliation":[{"name":"Department of Civil and Environmental Engineering and Earth Sciences, University of Notre Dame 2 , Notre Dame, Indiana 46556, USA"},{"name":"Institute for Precision Health, University of Notre Dame 3 , Notre Dame, Indiana 46556, USA"},{"name":"Eck Institute for Global Health, University of Notre Dame 4 , Notre Dame, Indiana 46556, USA"},{"name":"Department of Biological Sciences, University of Notre Dame 5 , Notre Dame, Indiana 46556, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9052-0349","authenticated-orcid":false,"given":"Paul W.","family":"Bohn","sequence":"additional","affiliation":[{"name":"Department of Chemistry and Biochemistry, University of Notre Dame 1 , Notre Dame, Indiana 46556, USA"},{"name":"Institute for Precision Health, University of Notre Dame 3 , Notre Dame, Indiana 46556, USA"},{"name":"Department of Chemical and Biomolecular Engineering, University of Notre Dame 6 , Notre Dame, Indiana 46556, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"317","published-online":{"date-parts":[[2021,5,25]]},"reference":[{"key":"2023062610432342800_c1","doi-asserted-by":"publisher","first-page":"759348","DOI":"10.1155\/2015\/759348","article-title":"The formation of biofilms by Pseudomonas aeruginosa: A review of the natural and synthetic compounds interfering with control mechanisms","volume":"2015","journal-title":"Biomed. Res. Int."},{"key":"2023062610432342800_c2","doi-asserted-by":"publisher","first-page":"e1000354","DOI":"10.1371\/journal.ppat.1000354","article-title":"Assembly and development of the Pseudomonas aeruginosa biofilm matrix","volume":"5","year":"2009","journal-title":"PLoS Pathog."},{"key":"2023062610432342800_c3","doi-asserted-by":"publisher","first-page":"1318","DOI":"10.1126\/science.284.5418.1318","article-title":"Bacterial biofilms: A common cause of persistent infections","volume":"284","year":"1999","journal-title":"Science"},{"key":"2023062610432342800_c4","doi-asserted-by":"publisher","first-page":"95","DOI":"10.1038\/nrmicro821","article-title":"Bacterial biofilms: From the natural environment to infectious diseases","volume":"2","year":"2004","journal-title":"Nat. Rev. Microbiol."},{"key":"2023062610432342800_c5","doi-asserted-by":"publisher","first-page":"484","DOI":"10.1111\/j.1365-2672.1992.tb05009.x","article-title":"Effect of polysaccharide interactions on antibiotic susceptibility of Pseudomonas aeruginosa","volume":"73","year":"1992","journal-title":"J. Appl. Bacteriol."},{"key":"2023062610432342800_c6","doi-asserted-by":"publisher","first-page":"3492","DOI":"10.1128\/jb.00119-09","article-title":"Pseudomonas aeruginosa small-colony variants have adaptations that likely promote persistence in the cystic fibrosis lung","volume":"191","year":"2009","journal-title":"J. Bacteriol."},{"key":"2023062610432342800_c7","doi-asserted-by":"publisher","first-page":"261","DOI":"10.1016\/j.femsre.2003.09.004","article-title":"Quorum sensing and swarming migration in bacteria","volume":"28","year":"2004","journal-title":"FEMS Microbiol. Rev."},{"key":"2023062610432342800_c8","doi-asserted-by":"publisher","first-page":"165","DOI":"10.1146\/annurev.micro.55.1.165","article-title":"Quorum sensing in bacteria","volume":"55","year":"2001","journal-title":"Annu. Rev. Microbiol."},{"key":"2023062610432342800_c9","doi-asserted-by":"publisher","first-page":"1127","DOI":"10.1126\/science.8493556","article-title":"Expression of Pseudomonas aeruginosa virulence genes requires cell-to-cell communication","volume":"260","year":"1993","journal-title":"Science"},{"key":"2023062610432342800_c10","doi-asserted-by":"publisher","first-page":"295","DOI":"10.1126\/science.280.5361.295","article-title":"The involvement of cell-to-cell signals in the development of a bacterial biofilm","volume":"280","year":"1998","journal-title":"Science"},{"key":"2023062610432342800_c11","doi-asserted-by":"publisher","first-page":"2519","DOI":"10.3390\/s120302519","article-title":"Quorum sensing and bacterial social interactions in biofilms","volume":"12","year":"2012","journal-title":"Sensors"},{"key":"2023062610432342800_c12","doi-asserted-by":"publisher","first-page":"586","DOI":"10.1016\/j.tim.2012.09.004","article-title":"Quorum sensing and the confusion about diffusion","volume":"20","year":"2012","journal-title":"Trends Microbiol."},{"key":"2023062610432342800_c13","doi-asserted-by":"publisher","first-page":"339","DOI":"10.1038\/nchembio.1225","article-title":"A cell-cell communication signal integrates quorum sensing and stress response","volume":"9","year":"2013","journal-title":"Nat. Chem. Biol."},{"key":"2023062610432342800_c14","doi-asserted-by":"publisher","first-page":"26","DOI":"10.1007\/s13238-014-0100-x","article-title":"The hierarchy quorum sensing network in Pseudomonas aeruginosa","volume":"6","year":"2015","journal-title":"Protein Cell"},{"key":"2023062610432342800_c15","doi-asserted-by":"publisher","first-page":"6770","DOI":"10.1128\/jb.05929-11","article-title":"2-heptyl-4-quinolone, a precursor of the Pseudomonas quinolone signal molecule, modulates swarming motility in Pseudomonas aeruginosa","volume":"193","year":"2011","journal-title":"J. Bacteriol."},{"key":"2023062610432342800_c16","doi-asserted-by":"publisher","first-page":"413","DOI":"10.1111\/j.1574-6941.2011.01121.x","article-title":"The Pseudomonas quinolone signal (PQS), and its precursor HHQ, modulate interspecies and interkingdom behaviour","volume":"77","year":"2011","journal-title":"FEMS Microbiol. Ecol."},{"key":"2023062610432342800_c17","doi-asserted-by":"publisher","first-page":"e00873","DOI":"10.1128\/mbio.00873-17","article-title":"Pseudomonas aeruginosa alters Staphylococcus aureus sensitivity to vancomycin in a biofilm model of cystic fibrosis infection","volume":"8","year":"2017","journal-title":"mBio"},{"key":"2023062610432342800_c18","doi-asserted-by":"publisher","first-page":"6544","DOI":"10.1039\/c5an01149c","article-title":"Multimodal chemical imaging of molecular messengers in emerging Pseudomonas aeruginosa bacterial communities","volume":"140","year":"2015","journal-title":"Analyst"},{"key":"2023062610432342800_c19","doi-asserted-by":"publisher","first-page":"e00426","DOI":"10.1128\/msphere.00426-20","article-title":"Spatiotemporal distribution of Pseudomonas aeruginosa alkyl quinolones under metabolic and competitive stress","volume":"5","year":"2020","journal-title":"mSphere"},{"key":"2023062610432342800_c20","doi-asserted-by":"publisher","first-page":"9544","DOI":"10.1074\/jbc.ra118.002605","article-title":"Spatially dependent alkyl quinolone signaling responses to antibiotics in Pseudomonas aeruginosa swarms","volume":"293","year":"2018","journal-title":"J. Biol. Chem."},{"key":"2023062610432342800_c21","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1177\/1178636118817738","article-title":"Surface growing communities of Pseudomonas aeruginosa exhibit distinct alkyl quinolone signatures","volume":"11","year":"2018","journal-title":"Microbiol. Insights"},{"key":"2023062610432342800_c22","doi-asserted-by":"publisher","first-page":"6558","DOI":"10.1128\/jb.173.20.6558-6567.1991","article-title":"Optical sectioning of microbial biofilms","volume":"173","year":"1991","journal-title":"J. Bacteriol."},{"key":"2023062610432342800_c23","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1099\/jmm.0.015420-0","article-title":"Visualization of adherent micro-organisms using different techniques","volume":"59","year":"2010","journal-title":"J. Med. Microbiol."},{"key":"2023062610432342800_c24","doi-asserted-by":"publisher","first-page":"1359","DOI":"10.1099\/jmm.0.011213-0","article-title":"Bacterial colonization of enamel in situ investigated using fluorescence in situ hybridization","volume":"58","year":"2009","journal-title":"J. Med. Microbiol."},{"key":"2023062610432342800_c25","doi-asserted-by":"publisher","first-page":"1924","DOI":"10.1039\/c3an36416j","article-title":"Correlated imaging\u2014A grand challenge in chemical analysis","volume":"138","year":"2013","journal-title":"Analyst"},{"key":"2023062610432342800_c26","doi-asserted-by":"publisher","first-page":"875","DOI":"10.1039\/b602376m","article-title":"Metabolic fingerprinting in disease diagnosis: Biomedical applications of infrared and Raman spectroscopy","volume":"131","year":"2006","journal-title":"Analyst"},{"key":"2023062610432342800_c27","doi-asserted-by":"publisher","first-page":"4578","DOI":"10.1039\/c4an00826j","article-title":"Correlated mass spectrometry imaging and confocal Raman microscopy for studies of three-dimensional cell culture sections","volume":"139","year":"2014","journal-title":"Analyst"},{"key":"2023062610432342800_c28","doi-asserted-by":"publisher","first-page":"2312","DOI":"10.1039\/c8an02031k","article-title":"A three-dimensional principal component analysis approach for exploratory analysis of hyperspectral data: Identification of ovarian cancer samples based on Raman microspectroscopy imaging of blood plasma","volume":"144","year":"2019","journal-title":"Analyst"},{"key":"2023062610432342800_c29","doi-asserted-by":"publisher","first-page":"98","DOI":"10.1021\/ac3019119","article-title":"Quantitative vibrational imaging by hyperspectral stimulated Raman scattering microscopy and multivariate curve resolution analysis","volume":"85","year":"2013","journal-title":"Anal. Chem."},{"key":"2023062610432342800_c30","doi-asserted-by":"publisher","first-page":"7790","DOI":"10.1021\/acs.analchem.9b01359","article-title":"Confocal Raman microscopy investigation of self-assembly of hybrid phospholipid bilayers within individual porous silica chromatographic particles","volume":"91","year":"2019","journal-title":"Anal. Chem."},{"key":"2023062610432342800_c31","doi-asserted-by":"publisher","first-page":"1049","DOI":"10.1021\/acs.analchem.8b04390","article-title":"Single layer graphene for estimation of axial spatial resolution in confocal Raman microscopy depth profiling","volume":"91","year":"2019","journal-title":"Anal. Chem."},{"key":"2023062610432342800_c32","doi-asserted-by":"publisher","first-page":"707","DOI":"10.1021\/acs.analchem.9b02593","article-title":"Spatiotemporal organization of biofilm matrix revealed by confocal Raman mapping integrated with non-negative matrix factorization analysis","volume":"92","year":"2020","journal-title":"Anal. Chem."},{"key":"2023062610432342800_c33","doi-asserted-by":"publisher","first-page":"158","DOI":"10.1080\/05704928.2018.1558232","article-title":"Bacteria induced pH changes in tissue-engineered human skin detected non-invasively using Raman confocal spectroscopy","volume":"55","year":"2019","journal-title":"Appl. Spectrosc. Rev."},{"key":"2023062610432342800_c34","doi-asserted-by":"publisher","first-page":"365","DOI":"10.1007\/s10924-016-0925-x","article-title":"Biosynthesis, microstructural characterisations and investigation of in-vitro mutagenic and eco-toxicological response of a novel microbial exopolysaccharide based biopolymer","volume":"26","year":"2017","journal-title":"J. Polym. Environ."},{"key":"2023062610432342800_c35","doi-asserted-by":"publisher","first-page":"273","DOI":"10.1046\/j.1365-2818.1997.2620818.x","article-title":"Axial resolution of confocal Raman microscopes: Gaussian beam theory and practice","volume":"188","year":"1997","journal-title":"J. Microsc."},{"key":"2023062610432342800_c36","doi-asserted-by":"publisher","first-page":"2512","DOI":"10.1039\/c0an00371a","article-title":"Confocal Raman microscopy: Common errors and artefacts","volume":"135","year":"2010","journal-title":"Analyst"},{"key":"2023062610432342800_c37","doi-asserted-by":"publisher","first-page":"993004","DOI":"10.1117\/12.2236695","article-title":"Label-free molecular imaging of bacterial communities of the opportunistic pathogen Pseudomonas aeruginosa","volume":"9930","year":"2016","journal-title":"Proc. SPIE"},{"key":"2023062610432342800_c38","doi-asserted-by":"publisher","first-page":"1254","DOI":"10.1038\/nprot.2007.158","article-title":"Biosensor-based assays for PQS, HHQ and related 2-alkyl-4-quinolone quorum sensing signal molecules","volume":"2","year":"2007","journal-title":"Nat. Protoc."}],"container-title":["The Journal of Chemical Physics"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/pubs.aip.org\/aip\/jcp\/article-pdf\/doi\/10.1063\/5.0052785\/14004290\/204201_1_online.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/pubs.aip.org\/aip\/jcp\/article-pdf\/doi\/10.1063\/5.0052785\/14004290\/204201_1_online.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,6,26]],"date-time":"2023-06-26T15:24:29Z","timestamp":1687793069000},"score":1,"resource":{"primary":{"URL":"https:\/\/pubs.aip.org\/jcp\/article\/154\/20\/204201\/200064\/Depth-distributions-of-signaling-molecules-in"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,5,25]]},"references-count":38,"journal-issue":{"issue":"20","published-print":{"date-parts":[[2021,5,28]]}},"URL":"https:\/\/doi.org\/10.1063\/5.0052785","relation":{},"ISSN":["0021-9606","1089-7690"],"issn-type":[{"value":"0021-9606","type":"print"},{"value":"1089-7690","type":"electronic"}],"subject":[],"published-other":{"date-parts":[[2021,5,28]]},"published":{"date-parts":[[2021,5,25]]},"article-number":"204201"}}