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Targeting the c-di-GMP network may be a promising anti-biofilm approach, but most strategies studied so far aimed at eliminating surface-attached biofilms, while in vivo <jats:italic>P. aeruginosa<\/jats:italic> biofilms often occur as suspended aggregates. Here, the expression profile of c-di-GMP metabolism-related genes was analysed among 32\u2009<jats:italic>P. aeruginosa<\/jats:italic> strains grown as aggregates in synthetic cystic fibrosis sputum. The diguanylate cyclase SiaD proved essential for auto-aggregation under in vivo-like conditions. Virtual screening predicted a high binding affinity of echinacoside towards the active site of SiaD. Echinacoside reduced c-di-GMP levels and aggregate sizes and potentiated tobramycin activity against aggregates in &gt;80% of strains tested. This synergism was also observed in <jats:italic>P. aeruginosa<\/jats:italic>-infected 3-D alveolar epithelial cells and murine lungs, demonstrating echinacoside\u2019s potential as an adjunctive therapy for recalcitrant <jats:italic>P. aeruginosa<\/jats:italic> infections.<\/jats:p>","DOI":"10.1038\/s41522-025-00673-2","type":"journal-article","created":{"date-parts":[[2025,3,7]],"date-time":"2025-03-07T11:21:58Z","timestamp":1741346518000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":17,"title":["Echinacoside reduces intracellular c-di-GMP levels and potentiates tobramycin activity against Pseudomonas aeruginosa biofilm aggregates"],"prefix":"10.1038","volume":"11","author":[{"given":"Yu-Ming","family":"Cai","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Feng","family":"Hong","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Amber","family":"De Craemer","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jacob George","family":"Malone","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Aur\u00e9lie","family":"Crabb\u00e9","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tom","family":"Coenye","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2025,3,7]]},"reference":[{"key":"673_CR1","doi-asserted-by":"publisher","first-page":"608","DOI":"10.1038\/s41579-022-00767-0","volume":"20","author":"K Sauer","year":"2022","unstructured":"Sauer, K. et al. The biofilm life cycle: expanding the conceptual model of biofilm formation. Nat. Rev. Microbiol. 20, 608\u2013620 (2022).","journal-title":"Nat. Rev. Microbiol."},{"key":"673_CR2","doi-asserted-by":"publisher","first-page":"100028","DOI":"10.1016\/j.bioflm.2020.100028","volume":"2","author":"KP Rumbaugh","year":"2020","unstructured":"Rumbaugh, K. P. How well are we translating biofilm research from bench-side to bedside? Biofilm 2, 100028 (2020).","journal-title":"Biofilm"},{"key":"673_CR3","doi-asserted-by":"publisher","first-page":"fuad050","DOI":"10.1093\/femsre\/fuad050","volume":"47","author":"M Lichtenberg","year":"2023","unstructured":"Lichtenberg, M., Coenye, T., Parsek, M. R., Bjarnsholt, T. & Jakobsen, T. H. What\u2019s in a name? Characteristics of clinical biofilms. FEMS Microbiol. Rev. 47, fuad050 (2023).","journal-title":"FEMS Microbiol. 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Biofilm-related infections: bridging the gap between clinical management and fundamental aspects of recalcitrance toward antibiotics. Microbiol. Mol. Biol. Rev. 78, 510\u2013543 (2014).","journal-title":"Microbiol. Mol. Biol. Rev."},{"key":"673_CR7","doi-asserted-by":"publisher","DOI":"10.1128\/aac.00535-24","volume":"15","author":"V Gigante","year":"2024","unstructured":"Gigante, V. et al. Multi-year analysis of the global preclinical antibacterial pipeline: trends and gaps. Antimicrob. Agents Chemother. 15, e0053524 (2024).","journal-title":"Antimicrob. Agents Chemother."},{"key":"673_CR8","unstructured":"Theuretzbacher, U. Evaluating the innovative potential of the global antibacterial pipeline. Clin. Microbiol. Infect. 0, S1198-743X(23)00490-1 (2023)."},{"key":"673_CR9","doi-asserted-by":"publisher","first-page":"753","DOI":"10.1007\/s12275-017-7274-x","volume":"55","author":"XH Li","year":"2017","unstructured":"Li, X. H. & Lee, J. H. 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