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In bacteria, although the significance of Ca\n                      <jats:sup>2+<\/jats:sup>\n                      regulation is recognized, it is commonly attributed to its role as a first messenger, while the mechanistic basis for its capacity as a second messenger remains elusive. Previously, we have shown that\n                      <jats:italic toggle=\"yes\">Pseudomonas aeruginosa,<\/jats:italic>\n                      an opportunistic pathogen causing life-threatening infections in humans, possesses the key prerequisites for intracellular Ca\n                      <jats:sup>2+<\/jats:sup>\n                      (Ca\n                      <jats:sup>2+<\/jats:sup>\n                      <jats:sub>in<\/jats:sub>\n                      ) signaling, including tightly controlled Ca\n                      <jats:sup>2+<\/jats:sup>\n                      <jats:sub>in<\/jats:sub>\n                      homeostasis, transient changes in Ca\n                      <jats:sup>2+<\/jats:sup>\n                      <jats:sub>in<\/jats:sub>\n                      levels in response to exogenous Ca\n                      <jats:sup>2+<\/jats:sup>\n                      , and Ca\n                      <jats:sup>2+<\/jats:sup>\n                      -regulated gene expression. In addition, we and others identified several key components of the Ca\n                      <jats:sup>2+<\/jats:sup>\n                      signaling network\n                      <jats:italic toggle=\"yes\">,<\/jats:italic>\n                      including Ca\n                      <jats:sup>2+<\/jats:sup>\n                      sensors, transporters, and regulators. The remaining question is whether the levels of Ca\n                      <jats:sup>2+<\/jats:sup>\n                      <jats:sub>in<\/jats:sub>\n                      alter in response to any extracellular stimuli other than Ca\n                      <jats:sup>2+<\/jats:sup>\n                      . To determine this, we have used a cytoplasmic Ca\n                      <jats:sup>2+<\/jats:sup>\n                      probe, R-GECO1, that produces a Ca\n                      <jats:sup>2+<\/jats:sup>\n                      -dependent fluorescence and tested a variety of extracellular stimuli, including ions, monosaccharides, amino acids, nucleotides, quorum-sensing signaling molecules, reactive oxygen species, and antibiotics. We observed changes in Ca\n                      <jats:sup>2+<\/jats:sup>\n                      <jats:sub>in<\/jats:sub>\n                      levels in response to several molecules, particularly glucose, asparagine, ATP, ADP, AMP, HOCl, and cycloserine. The results provide the first experimental evidence that, like eukaryotes,\n                      <jats:italic toggle=\"yes\">P. aeruginosa<\/jats:italic>\n                      responds to select, chemically diverse host-related stimuli by altering its Ca\n                      <jats:sup>2+<\/jats:sup>\n                      <jats:sub>in<\/jats:sub>\n                      levels, supporting a potential signaling role of Ca\u00b2\n                      <jats:sup>+<\/jats:sup>\n                      <jats:sub>in<\/jats:sub>\n                      in bacteria.\n                    <\/jats:p>\n                    <jats:sec>\n                      <jats:title>IMPORTANCE<\/jats:title>\n                      <jats:p>\n                        Cells use signaling to communicate and respond to their environment. Although the role of calcium ions (Ca\u00b2\n                        <jats:sup>+<\/jats:sup>\n                        ) as a signaling molecule, regulating multiple essential processes, has been well established in eukaryotes, Ca\u00b2\n                        <jats:sup>+<\/jats:sup>\n                        signaling in bacteria remains understudied. Previous research has indicated the role of Ca\u00b2\n                        <jats:sup>+<\/jats:sup>\n                        in bacterial regulation as a first messenger; however, its role as a second messenger is not yet recognized. Here, we show, for the first time, that\n                        <jats:italic toggle=\"yes\">P. aeruginosa<\/jats:italic>\n                        , an important human pathogen, alters its intracellular Ca\u00b2\n                        <jats:sup>+<\/jats:sup>\n                        levels in response to various external stimuli commonly associated with the host environment. These findings provide the mechanistic basis for\n                        <jats:italic toggle=\"yes\">P. aeruginosa<\/jats:italic>\n                        to use Ca\u00b2\n                        <jats:sup>+<\/jats:sup>\n                        as a second messenger. Understanding these Ca\n                        <jats:sup>2+<\/jats:sup>\n                        responses provides novel insights into Ca\n                        <jats:sup>2+<\/jats:sup>\n                        -mediated adaptation of invading pathogens to the host environment and may inform the development of new antimicrobial strategies.\n                      <\/jats:p>\n                    <\/jats:sec>\n                  <\/jats:sec>","DOI":"10.1128\/aem.01359-26","type":"journal-article","created":{"date-parts":[[2026,8,28]],"date-time":"2026-08-28T13:00:38Z","timestamp":1787922038000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":0,"title":["Host-derived stimuli alter intracellular calcium levels in\n                    <i>Pseudomonas aeruginosa,<\/i>\n                    revealing its potential role as a second messenger"],"prefix":"10.1128","volume":"92","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-2365-4704","authenticated-orcid":false,"given":"Aya","family":"Kubo","sequence":"first","affiliation":[{"id":[{"id":"https:\/\/ror.org\/01g9vbr38","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Microbiology and Molecular Genetics, Oklahoma State University","place":["Stillwater, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0002-7566-6803","authenticated-orcid":true,"given":"Reygan E.","family":"Braga","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/01g9vbr38","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Microbiology and Molecular Genetics, Oklahoma State University","place":["Stillwater, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dulandika","family":"Malalage","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/01g9vbr38","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Microbiology and Molecular Genetics, Oklahoma State University","place":["Stillwater, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Maha M.","family":"Achour","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/01g9vbr38","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Microbiology and Molecular Genetics, Oklahoma State University","place":["Stillwater, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0495-3867","authenticated-orcid":true,"given":"Marianna A.","family":"Patrauchan","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/01g9vbr38","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Microbiology and Molecular Genetics, Oklahoma State University","place":["Stillwater, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"235","reference":[{"key":"e_1_3_2_2_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2007.11.028"},{"key":"e_1_3_2_3_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.jinorgbio.2008.09.006"},{"key":"e_1_3_2_4_2","doi-asserted-by":"publisher","DOI":"10.1038\/nrm1155"},{"key":"e_1_3_2_5_2","doi-asserted-by":"publisher","DOI":"10.3390\/molecules25092148"},{"key":"e_1_3_2_6_2","doi-asserted-by":"publisher","unstructured":"Gilabert JA. 2012. 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