{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,9]],"date-time":"2026-10-09T14:54:38Z","timestamp":1791557678596,"version":"4.3.4"},"reference-count":30,"publisher":"MDPI AG","issue":"19","license":[{"start":{"date-parts":[[2026,10,5]],"date-time":"2026-10-05T00:00:00Z","timestamp":1791158400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2026,10,5]],"date-time":"2026-10-05T00:00:00Z","timestamp":1791158400000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Science and Technology Project of Fujian Province","award":["2023J01763"],"award-info":[{"award-number":["2023J01763"]}]},{"name":"Science and Technology Project of Fujian Province","award":["2026J001772"],"award-info":[{"award-number":["2026J001772"]}]},{"name":"Xiamen Science and Technology Subsidy Project","award":["2024CXY0307"],"award-info":[{"award-number":["2024CXY0307"]}]},{"name":"Educational Office of Fujian Province","award":["[2023]20, FBJY20230312"],"award-info":[{"award-number":["[2023]20, FBJY20230312"]}]},{"award":["[2023]20, FBJY20230312"],"award-info":[{"award-number":["[2023]20, FBJY20230312"]}],"id":[{"id":"https:\/\/ror.org\/01d6wrk87","id-type":"ROR","asserted-by":"publisher"}]},{"name":"Open Research Fund Program of Fujian Provincial Key Laboratory of Marine Fishery Resources and Eco-environment","award":["JD202402"],"award-info":[{"award-number":["JD202402"]}]},{"name":"Fujian Provincial Department of Science and Technology","award":["[2023]15, 2023S2030"],"award-info":[{"award-number":["[2023]15, 2023S2030"]}],"id":[{"id":"https:\/\/ror.org\/00rgzpv08","id-type":"ROR","asserted-by":"crossref"}]},{"award":["[2023]15, 2023S2030"],"award-info":[{"award-number":["[2023]15, 2023S2030"]}],"id":[{"id":"https:\/\/ror.org\/00rgzpv08","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Animals"],"abstract":"<jats:p>Vibrio alginolyticus is a major aquaculture pathogen whose virulence is largely mediated by the type II secretion system (T2SS). However, the functions of key T2SS components, including gspJ, gspL, and gspM, remain unclear. Earlier research indicated that a Scutellaria baicalensis extract at 8 mg\/mL markedly suppressed the growth, motility, and biofilm-forming capacity of V. alginolyticus ND-01 (OD600 = 0.5) following 12 h of treatment, with the resulting stress substantially altering the expression of multiple genes, including gspJ, gspL, and gspM. The gspJ-RNAi, gspL-RNAi, and gspM-RNAi strains were constructed using RNAi. Biological functional assays demonstrated that stable silencing of gspJ, gspL, and gspM markedly decreased biofilm formation ability (84.01%, 24.99%, and 78.26%, respectively) and host cell adhesion (73.57%, 75.65%, and 79.61%, respectively); gspJ and gspM silencing reduced motility (54.82% and 57.42%, respectively), whereas gspL silencing enhanced motility by 38.49%, indicating their essential roles in virulence-associated phenotypes. Notably, in vivo infection of Epinephelus coioides revealed that gspL-RNAi significantly attenuated pathogenicity, as evidenced by reduced host mortality (50%) and decreased bacterial colonization in liver and spleen tissues, compared with the wild-type group. Consistently, Label-Free Proteomic Analysis of the gspL-RNAi strain identified 189 differentially expressed proteins in the gspL-silenced strain, with enrichment in two-component signaling, ABC transport systems, and amino acid metabolism. Network analysis further positioned gspL as a central regulatory hub linking T2SS activity to signal transduction, transport, and metabolic pathways. In addition, the expression of gspJ-RNAi, gspL-RNAi, and gspM-RNAi exhibited distinct responses to environmental stresses, including pH and temperature variation. Collectively, our findings highlight that gspJ, gspL, and gspM are critical regulators of virulence in V. alginolyticus, with gspL playing a central role in coordinating T2SS function with global regulatory networks. This study advances our understanding of the pathogenic mechanisms of V. alginolyticus and provides potential targets for the development of anti-virulence aquaculture strategies.<\/jats:p>","DOI":"10.3390\/ani16193121","type":"journal-article","created":{"date-parts":[[2026,10,5]],"date-time":"2026-10-05T08:57:18Z","timestamp":1791190638000},"page":"3121","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["T2SS Components GspJ, GspL, and GspM Regulate Virulence in Vibrio alginolyticus"],"prefix":"10.3390","volume":"16","author":[{"given":"Le","family":"Zhang","sequence":"first","affiliation":[{"name":"Fisheries College, Jimei University, Xiamen 361021, 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(2024). Assessment of Vibrionaceae prevalence in seafood from Qidong market and analysis of Vibrio parahaemolyticus strains. PLoS ONE, 19.","DOI":"10.1371\/journal.pone.0309304"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"4025","DOI":"10.1038\/s41467-023-39583-2","article-title":"Membrane translocation process revealed by in situ structures of type II secretion system secretins","volume":"14","author":"Yu","year":"2023","journal-title":"Nat. Commun."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"5437","DOI":"10.1038\/s41467-019-13301-3","article-title":"Core architecture of a bacterial type II secretion system","volume":"10","author":"Chernyatina","year":"2019","journal-title":"Nat. Commun."},{"key":"ref_5","first-page":"001198","article-title":"The long and short of it: Benchmarking viromics using Illumina, Nanopore and PacBio sequencing technologies","volume":"10","author":"Cook","year":"2024","journal-title":"Microb. Genom."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"1604","DOI":"10.1080\/22221751.2019.1687261","article-title":"Mechanisms underlying the virulence regulation of new Vibrio alginolyticus ncRNA Vvrr1 with a comparative proteomic analysis","volume":"8","author":"Zuo","year":"2019","journal-title":"Emerg. Microbes Infect."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"483","DOI":"10.1039\/D4MO00043A","article-title":"Discovery of candidate biomarkers from plasma-derived extracellular vesicles of patients with cirrhosis and hepatocellular carcinoma: An exploratory proteomic study","volume":"20","author":"Muriel","year":"2024","journal-title":"Mol. Omics"},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Stappenbeck, F., Wang, F., Sinha, S.K., Hui, S.T., Farahi, L., Mukhamedova, N., Fleetwood, A., Murphy, A.J., Sviridov, D., and Lusis, A.J. (2024). Anti-Inflammatory Oxysterol, Oxy210, Inhibits Atherosclerosis in Hyperlipidemic Mice and Inflammatory Responses of Vascular Cells. Cells, 13.","DOI":"10.3390\/cells13191632"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"665","DOI":"10.1016\/j.bbrc.2020.09.135","article-title":"Combination of metformin and RG7388 enhances inhibition of growth and induction of apoptosis of ovarian cancer cells through the PI3K\/AKT\/mTOR pathway","volume":"533","author":"Cui","year":"2020","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Diaz, F.C., Waldrup, B., Carranza, F.G., Manjarrez, S., and Velazquez-Villarreal, E. (2025). Artificial Intelligence-Enhanced Precision Medicine Reveals Prognostic Impact of TGF-Beta Pathway Alterations in FOLFOX-Treated Early-Onset Colorectal Cancer Among Disproportionately Affected Populations. Int. J. Mol. Sci., 26.","DOI":"10.1101\/2025.08.11.25333363"},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Guo, J., Wang, L., Song, K., Lu, K., Li, X., and Zhang, C. (2023). Physiological Response of Spotted Seabass (Lateolabrax maculatus) to Different Dietary Available Phosphorus Levels and Water Temperature: Changes in Growth, Lipid Metabolism, Antioxidant Status and Intestinal Microbiota. Antioxidants, 12.","DOI":"10.3390\/antiox12122128"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"100776","DOI":"10.1016\/j.animal.2023.100776","article-title":"The pigments in eggshell with different colour and the pigment regulatory gene expression in corresponding chicken\u2019s shell gland","volume":"17","author":"Wang","year":"2023","journal-title":"Animal"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"e202510595","DOI":"10.1002\/anie.202510595","article-title":"Control Over S(VI) Stereogenicity for the Asymmetric Synthesis of Sulfonimidoyl Derivatives by Isothiourea-Catalyzed Covalent Activation of Sulfur(VI) Atoms","volume":"64","author":"Li","year":"2025","journal-title":"Angew. Chem. Int. Ed. Engl."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"117076","DOI":"10.1016\/j.jpba.2025.117076","article-title":"Integrating proteomics, metabolomics, and network pharmacology to investigate the mechanism of Cordyceps sinensis in the treatment of COPD rats","volume":"266","author":"Zhou","year":"2025","journal-title":"J. Pharm. Biomed. Anal."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"111332","DOI":"10.1016\/j.fsi.2026.111332","article-title":"Influence of the two-component system PhoBR associated with phosphate-sensing capacity on the virulence mechanism of Vibrio alginolyticus","volume":"174","author":"Chen","year":"2026","journal-title":"Fish Shellfish Immunol."},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Yi, X., Xu, X., Qi, X., Chen, Y., Zhu, Z., Xu, G., Li, H., Kraco, E.-K., Shen, H., and Lin, M. (2022). Mechanisms Underlying the Virulence Regulation of Vibrio alginolyticus ND-01 pstS and pstB with a Transcriptomic Analysis. Microorganisms, 10.","DOI":"10.3390\/microorganisms10112093"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"111954","DOI":"10.1016\/j.mce.2023.111954","article-title":"Liraglutide alleviates myocardial ischemia-reperfusion injury in diabetic mice","volume":"572","author":"Fang","year":"2023","journal-title":"Mol. Cell. Endocrinol."},{"key":"ref_18","doi-asserted-by":"crossref","unstructured":"Zhang, Y., Zhang, A., Wang, L., Yang, T., Dong, B., Wang, Z., Bi, Y., Chen, G., and Chang, G. (2022). Metabolomics and Proteomics Characterizing Hepatic Reactions to Dietary Linseed Oil in Duck. Int. J. Mol. Sci., 23.","DOI":"10.3390\/ijms232415690"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"149986","DOI":"10.1016\/j.gene.2025.149986","article-title":"Ginsenoside Rg1 delays chronological aging in a yeast model via SSE1-Mediated mitophagy","volume":"982","author":"Yao","year":"2026","journal-title":"Gene"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"4732","DOI":"10.3168\/jds.2025-27418","article-title":"Alterations in the fat globule membrane proteome of cow, yak, buffalo, and camel milk in response to homogenization and heat treatment","volume":"109","author":"Shi","year":"2026","journal-title":"J. Dairy Sci."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"4093","DOI":"10.1128\/JB.01944-14","article-title":"The Type II Secretion System Delivers Matrix Proteins for Biofilm Formation by Vibrio cholerae","volume":"196","author":"Johnson","year":"2014","journal-title":"J. Bacteriol."},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Sikora, A.E. (2013). Proteins Secreted via the Type II Secretion System: Smart Strategies of Vibrio cholerae to Maintain Fitness in Different Ecological Niches. PLoS Pathog., 9.","DOI":"10.1371\/journal.ppat.1003126"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"2672","DOI":"10.1016\/j.cub.2025.04.072","article-title":"Activation of an antifungal pathway in Yersinia pseudotuberculosis by chitin-receptor-mediated fungal recognition","volume":"35","author":"Liu","year":"2025","journal-title":"Curr. Biol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"110847","DOI":"10.1016\/j.celrep.2022.110847","article-title":"Mitochondrial damage-associated molecular patterns trigger arginase-dependent lymphocyte immunoregulation","volume":"39","author":"Westhaver","year":"2022","journal-title":"Cell Rep."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"e0032825","DOI":"10.1128\/mra.00328-25","article-title":"Draft genomes of two Vibrio spp. isolated from the brown alga Sargassum ilicifolium in Singapore","volume":"14","author":"Luk","year":"2025","journal-title":"Microbiol. Resour. Announc."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"537","DOI":"10.1007\/s00253-024-13351-8","article-title":"TssL2 of T6SS2 is required for mobility, biofilm formation, wrinkly phenotype formation, and virulence of Vibrio parahaemolyticus SH112","volume":"108","author":"Bai","year":"2024","journal-title":"Appl. Microbiol. Biotechnol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"742604","DOI":"10.1016\/j.aquaculture.2025.742604","article-title":"Bile acid- and taurocholate-mediated virulence in AHPND-causing Vibrio parahaemolyticus is independent of the LuxO-OpaR quorum sensing cascade","volume":"606","author":"Kumar","year":"2025","journal-title":"Aquaculture"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"12","DOI":"10.1016\/j.drup.2018.03.002","article-title":"Fighting bacterial persistence: Current and emerging anti-persister strategies and therapeutics","volume":"38","author":"Defraine","year":"2018","journal-title":"Drug Resist. Updat."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"115945","DOI":"10.1016\/j.ejmech.2023.115945","article-title":"Novel 3-hydroxypyridin-4(1H)-One derivatives as ferroptosis inhibitors with iron-chelating and reactive oxygen species scavenging activities and therapeutic effect in cisplatin-induced cytotoxicity","volume":"263","author":"Zhang","year":"2024","journal-title":"Eur. J. Med. Chem."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"294","DOI":"10.1007\/s11274-024-04101-z","article-title":"Acetic and citric acids effect the type II secretion system and decrease the metabolic activities of salmon spoilage-related Rahnella aquatilis KM05","volume":"40","author":"Myszka","year":"2024","journal-title":"World J. Microbiol. Biotechnol."}],"container-title":["Animals"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-2615\/16\/19\/3121\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,10,9]],"date-time":"2026-10-09T14:43:51Z","timestamp":1791557031000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-2615\/16\/19\/3121"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,10,5]]},"references-count":30,"journal-issue":{"issue":"19","published-online":{"date-parts":[[2026,10]]}},"alternative-id":["ani16193121"],"URL":"https:\/\/doi.org\/10.3390\/ani16193121","relation":{},"ISSN":["2076-2615"],"issn-type":[{"value":"2076-2615","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,10,5]]}}}