{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T16:54:45Z","timestamp":1785430485952,"version":"3.56.0"},"reference-count":34,"publisher":"MDPI AG","issue":"6","license":[{"start":{"date-parts":[[2026,6,9]],"date-time":"2026-06-09T00:00:00Z","timestamp":1780963200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"National Science Center of Poland","award":["2017\/27\/B\/NZ6\/01544"],"award-info":[{"award-number":["2017\/27\/B\/NZ6\/01544"]}]},{"award":["2017\/27\/B\/NZ6\/01544"],"award-info":[{"award-number":["2017\/27\/B\/NZ6\/01544"]}],"id":[{"id":"https:\/\/ror.org\/03ha2q922","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Pathogens"],"abstract":"<jats:p>Legionella pneumophila is widely recognized as the principal organism responsible for Legionnaires\u2019 disease. Modifications of surface-exposed lipopolysaccharide in L. pneumophila are key determinants of bacterial adaptation to host cells. Using a mutant strain deficient in N-methylation of legionaminic acid and fluorescence-based biophysical analyses, it was demonstrated that N-methyl groups linked to the acetimidoylamino group of legionaminic acid modulate the physicochemical properties of the bacterial cell surface and influence the interaction of L. pneumophila with macrophages. Loss of N-methyl groups reduced the efficiency of bacterial interaction with THP-1-derived macrophages and impaired intracellular proliferation. In addition, the presence of N-methyl groups in legionaminic acid contributes to increased TNF-\u03b1 production in THP-1 macrophages stimulated with L. pneumophila sg1 LPS, without affecting IL-6 induction, suggesting that N-methylation of legionaminic acid may skew the early pro-inflammatory response toward TNF-\u03b1-dominated signaling during L. pneumophila infection.<\/jats:p>","DOI":"10.3390\/pathogens15060615","type":"journal-article","created":{"date-parts":[[2026,6,10]],"date-time":"2026-06-10T09:25:17Z","timestamp":1781083517000},"page":"615","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["N-Methylation of Legionaminic Acid in Legionella pneumophila Serogroup 1 LPS Affects Cell Surface Properties, Intracellular Growth, and Cytokine Response"],"prefix":"10.3390","volume":"15","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-2827-1596","authenticated-orcid":false,"given":"Bo\u017cena","family":"Kowalczyk","sequence":"first","affiliation":[{"name":"Department of Genetics and Microbiology, Institute of Biological Sciences, Faculty of Biology and Biotechnology, Maria Curie-Sk\u0142odowska University, Akademicka 19 St., 20-033 Lublin, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Agnieszka","family":"Szuster-Ciesielska","sequence":"additional","affiliation":[{"name":"Department of Virology and Immunology, Institute of Biological Sciences, Faculty of Biology and Biotechnology, Maria Curie-Sk\u0142odowska University, Akademicka 19 St., 20-033 Lublin, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1156-8059","authenticated-orcid":false,"given":"Rafal","family":"Luchowski","sequence":"additional","affiliation":[{"name":"Department of Biophysics, Institute of Physics, Faculty of Mathematics, Physics and Computer Science, Maria Curie-Sk\u0142odowska University, Radziszewskiego 10 St., 20-031 Lublin, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8245-3913","authenticated-orcid":false,"given":"Wies\u0142aw I.","family":"Gruszecki","sequence":"additional","affiliation":[{"name":"Department of Biophysics, Institute of Physics, Faculty of Mathematics, Physics and Computer Science, Maria Curie-Sk\u0142odowska University, Radziszewskiego 10 St., 20-031 Lublin, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0386-3544","authenticated-orcid":false,"given":"Piotr","family":"Koper","sequence":"additional","affiliation":[{"name":"Department of Genetics and Microbiology, Institute of Biological Sciences, Faculty of Biology and Biotechnology, Maria Curie-Sk\u0142odowska University, Akademicka 19 St., 20-033 Lublin, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6622-5550","authenticated-orcid":false,"given":"Markus","family":"Petzold","sequence":"additional","affiliation":[{"name":"Institute of Medical Microbiology and Virology, University Hospital Carl Gustav Carus, University of Technology Dresden, 01069 Dresden, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4687-7126","authenticated-orcid":false,"given":"Jacek","family":"Tarasiuk","sequence":"additional","affiliation":[{"name":"Department of Genetics and Microbiology, Institute of Biological Sciences, Faculty of Biology and Biotechnology, Maria Curie-Sk\u0142odowska University, Akademicka 19 St., 20-033 Lublin, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sofia","family":"Barigelli","sequence":"additional","affiliation":[{"name":"Laboratory of Applied and Environmental Microbiology, Department of Chemistry, Biology and Biotechnology, University of Perugia, 06122 Perugia, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4983-1967","authenticated-orcid":false,"given":"Ermanno","family":"Federici","sequence":"additional","affiliation":[{"name":"Laboratory of Applied and Environmental Microbiology, Department of Chemistry, Biology and Biotechnology, University of Perugia, 06122 Perugia, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8540-3084","authenticated-orcid":false,"given":"Marta","family":"Palusi\u0144ska-Szysz","sequence":"additional","affiliation":[{"name":"Department of Genetics and Microbiology, Institute of Biological Sciences, Faculty of Biology and Biotechnology, Maria Curie-Sk\u0142odowska University, Akademicka 19 St., 20-033 Lublin, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,6,9]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"376","DOI":"10.1016\/S0140-6736(15)60078-2","article-title":"Legionnaires\u2019 disease","volume":"387","author":"Cunha","year":"2016","journal-title":"Lancet"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"101031","DOI":"10.1016\/j.lanmic.2024.101031","article-title":"Global surge of Legionnaires\u2019 disease in 2024: Urgent call for heightened awareness and preparedness","volume":"6","author":"Pareek","year":"2024","journal-title":"Lancet Microbe"},{"key":"ref_3","first-page":"30566","article-title":"European Legionnaires\u2019 Disease Surveillance Network. Legionnaires\u2019 disease in Europe 2011\u20132015","volume":"22","year":"2017","journal-title":"Eurosurveillance"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"439","DOI":"10.1146\/annurev-pathmechdis-012419-032742","article-title":"Legionnaires\u2019 disease: State of the art knowledge of pathogenesis mechanisms of Legionella","volume":"15","author":"Mondino","year":"2020","journal-title":"Annu. Rev. Pathol."},{"key":"ref_5","doi-asserted-by":"crossref","unstructured":"Kowalczyk, B., Chmiel, E., and Palusinska-Szysz, M. (2021). The role of lipids in Legionella-host interaction. Int. J. Mol. Sci., 22.","DOI":"10.3390\/ijms22031487"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1111\/j.1574-6968.1992.tb05430.x","article-title":"Identification of 27-oxo-octacosanoic acid and heptacosane-1,27-dioic acid in Legionella pneumophila","volume":"76","author":"Moll","year":"1992","journal-title":"FEMS Microbiol. Lett."},{"key":"ref_7","first-page":"113","article-title":"The lipopolysaccharide of Legionella pneumophila (strain Philadelphia 1): Chemical structure and biological significance","volume":"392","author":"Knirel","year":"1995","journal-title":"Prog. Clin. Biol. Res."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"91","DOI":"10.1016\/S0008-6215(97)00210-3","article-title":"Identification of an \u03b1-d-Manp-(1\u21928)-Kdo disaccharide in the inner core region and the structure of the complete core region of the Legionella pneumophila serogroup 1 lipopolysaccharide","volume":"304","author":"Moll","year":"1997","journal-title":"Carbohydr. Res."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"223","DOI":"10.1016\/0008-6215(96)00194-2","article-title":"Structural study of a highly O-acetylated core of Legionella pneumophila serogroup 1 lipopolysaccharide","volume":"293","author":"Knirel","year":"1996","journal-title":"Carbohydr. Res."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"239","DOI":"10.1111\/j.1432-1033.1994.tb18734.x","article-title":"The structure of the O-specific chain of Legionella pneumophila serogroup 1 lipopolysaccharide","volume":"221","author":"Knirel","year":"1994","journal-title":"Eur. J. Biochem."},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Petzold, M., Th\u00fcrmer, A., Menzel, S., Mouton, J.W., Heuner, K., and L\u00fcck, C. (2013). A structural comparison of lipopolysaccharide biosynthesis loci of Legionella pneumophila serogroup 1 strains. BMC Microbiol., 13.","DOI":"10.1186\/1471-2180-13-198"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1555","DOI":"10.1101\/gr.209536.116","article-title":"Multiple major disease-associated clones of Legionella pneumophila have emerged recently and independently","volume":"26","author":"David","year":"2016","journal-title":"Genome Res."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"7165","DOI":"10.1038\/s41467-021-27478-z","article-title":"Population analysis of Legionella pneumophila reveals a basis for resistance to complement-mediated killing","volume":"12","author":"Wee","year":"2021","journal-title":"Nat. Commun."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"eadx9722","DOI":"10.1126\/sciadv.adx9722","article-title":"The discovery of a Legionella phage explains a key determinant of human disease","volume":"11","author":"Nicholson","year":"2025","journal-title":"Sci. Adv."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"1975","DOI":"10.1111\/j.1365-2672.2007.03434.x","article-title":"Random mutagenesis of Legionella pneumophila reveals genes associated with lipopolysaccharide synthesis and recognition by typing monoclonal antibodies","volume":"103","author":"Wagner","year":"2007","journal-title":"J. Appl. Microbiol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"90","DOI":"10.1111\/j.1365-2958.2005.04539.x","article-title":"IcmS-dependent translocation of SdeA into macrophages by the Legionella pneumophila type IV secretion system","volume":"56","author":"Bardill","year":"2005","journal-title":"Mol. Microbiol."},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Daigneault, M., Preston, J.A., Marriott, H.M., Whyte, M.K.B., and Dockrell, D.H. (2010). The identification of markers of macrophage differentiation in PMA-stimulated THP-1 cells and monocyte-derived macrophages. PLoS ONE, 5.","DOI":"10.1371\/journal.pone.0008668"},{"key":"ref_18","first-page":"83","article-title":"Bacterial lipopolysaccharide. Extraction with phenol-water and further applications of the procedure","volume":"5","author":"Westphal","year":"1965","journal-title":"Methods Carbohydr. Chem."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"603","DOI":"10.1111\/j.1432-1033.1975.tb04172.x","article-title":"Electrodialysis of lipopolysaccharides and their conversion to uniform salt forms","volume":"54","author":"Galanos","year":"1975","journal-title":"Eur. J. Biochem."},{"key":"ref_20","unstructured":"Lakowicz, J.R. (2010). Energy Transfer. Principles of Fluorescence Spectroscopy, Springer."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"159720","DOI":"10.1016\/j.bbalip.2026.159720","article-title":"Substituents of the polysaccharide region of LPS in Legionella pneumophila sg1 modulate interactions with host cells","volume":"1871","author":"Kowalczyk","year":"2026","journal-title":"Biochim. Biophys. Acta Mol. Cell Biol. Lipids"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"116244","DOI":"10.1016\/j.bios.2024.116244","article-title":"Bacterial detection based on F\u00f6rster resonance energy transfer","volume":"255","author":"Zhang","year":"2024","journal-title":"Biosens. Bioelectron."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"495","DOI":"10.1078\/1438-4221-00307","article-title":"Analysing the action of bacterial toxins in living cells with fluorescence resonance energy transfer (FRET)","volume":"293","author":"Majoul","year":"2004","journal-title":"Int. J. Med. Microbiol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"4151","DOI":"10.1128\/IAI.66.9.4151-4157.1998","article-title":"Low endotoxic potential of Legionella pneumophila lipopolysaccharide due to failure of interaction with the monocyte lipopolysaccharide receptor CD14","volume":"66","author":"Neumeister","year":"1998","journal-title":"Infect. Immun."},{"key":"ref_25","doi-asserted-by":"crossref","unstructured":"Kowalczyk, B., Petzold, M., Kaczynski, Z., Szuster-Ciesielska, A., Luchowski, R., Gruszecki, W.I., Fuchs, B., Galuska, C.E., Choma, A., and Tarasiuk, J. (2023). Lipopolysaccharide of Legionella pneumophila serogroup 1 facilitates interaction with host cells. Int. J. Mol. Sci., 24.","DOI":"10.3390\/ijms241914602"},{"key":"ref_26","doi-asserted-by":"crossref","unstructured":"Palusinska-Szysz, M., Luchowski, R., Gruszecki, W.I., Choma, A., Szuster-Ciesielska, A., L\u00fcck, C., Petzold, M., Sroka-Bartnicka, A., and Kowalczyk, B. (2019). The role of Legionella pneumophila serogroup 1 lipopolysaccharide in host-pathogen interaction. Front. Microbiol., 10.","DOI":"10.3389\/fmicb.2019.02890"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"560","DOI":"10.1046\/j.0014-2956.2001.02683.x","article-title":"N-methylation in polylegionaminic acid is associated with the phase-variable epitope of Legionella pneumophila serogroup 1 lipopolysaccharide. Identification of 5-(N,N-di-methylacetimidoyl) amino and 5-acetimidoyl(N-methyl)amino-7-acetamido-3,5,7,9-tetradeoxynon-2-ulosonic acid in the O-chain polysaccharide","volume":"269","author":"Kooistra","year":"2002","journal-title":"Eur. J. Biochem."},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"Shevchuk, O., J\u00e4ger, J., and Steinert, M. (2011). Virulence properties of the Legionella pneumophila cell envelope. Front. Microbiol., 2.","DOI":"10.3389\/fmicb.2011.00074"},{"key":"ref_29","doi-asserted-by":"crossref","unstructured":"Grigoryeva, L.S., and Cianciotto, N.P. (2021). Human macrophages utilize a wide range of pathogen recognition receptors to recognize Legionella pneumophila, including Toll-Like Receptor 4 engaging Legionella lipopolysaccharide and the Toll-like Receptor 3 nucleic-acid sensor. PLoS Pathog., 7.","DOI":"10.1371\/journal.ppat.1009781"},{"key":"ref_30","doi-asserted-by":"crossref","unstructured":"Cochet, F., and Peri, F. (2017). The role of carbohydrates in the lipopolysaccharide (LPS)\/Toll-Like Receptor 4 (TLR4) signalling. Int. J. Mol. Sci., 18.","DOI":"10.3390\/ijms18112318"},{"key":"ref_31","doi-asserted-by":"crossref","unstructured":"Mazgaeen, L., and Gurung, P. (2020). Recent Advances in Lipopolysaccharide Recognition Systems. Int. J. Mol. Sci., 21.","DOI":"10.3390\/ijms21020379"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"2395","DOI":"10.1016\/j.bpj.2014.04.016","article-title":"Lipopolysaccharide-induced dynamic lipid membrane reorganization: Tubules, perforations, and stacks","volume":"106","author":"Adams","year":"2014","journal-title":"Biophys. J."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"142","DOI":"10.1186\/s12964-025-02149-4","article-title":"An examination of the LPS-TLR4 immune response through the analysis of molecular structures and protein\u2013protein interactions","volume":"23","author":"Yao","year":"2025","journal-title":"Cell Commun. Signal."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"22","DOI":"10.1097\/SHK.0000000000000470","article-title":"The role of lipopolysaccharide structure in monocyte activation and cytokine secretion","volume":"45","author":"Plevin","year":"2016","journal-title":"Shock"}],"container-title":["Pathogens"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-0817\/15\/6\/615\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,6,10]],"date-time":"2026-06-10T10:05:45Z","timestamp":1781085945000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-0817\/15\/6\/615"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,6,9]]},"references-count":34,"journal-issue":{"issue":"6","published-online":{"date-parts":[[2026,6]]}},"alternative-id":["pathogens15060615"],"URL":"https:\/\/doi.org\/10.3390\/pathogens15060615","relation":{},"ISSN":["2076-0817"],"issn-type":[{"value":"2076-0817","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,6,9]]}}}