Prideapplication/xmlftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD073462/Spectronaut.sneftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD073462/ETBF.d.zipftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD073462/NTBF-BFT.d.zipprimaryOK200ygoo@wustl.eduByoung-Kyu ChoMass SpectrometryData-independent acquisitionCftrBacteroides fragilisOuter membrane vesiclesColon organoidsTransepithelial resistancehttps://www.ebi.ac.uk/pride/archive/projects/PXD073462Isolated OMVs were purified by trichloroacetic acid precipitation. After pelleting and washing with ice-cold acetone, the resulting protein pellet was resuspended in 8 M urea and 0.4 M ammonium bicarbonate, reduced with 4 mM dithiothreitol, and alkylated with 18 mM iodoacetamide. The solution was then diluted to <2 M urea, and 1 µg of trypsin was added for overnight digestion at 37°C. The resulting peptides were desalted using C18 solid-phase extraction spin columns, and eluates were dried under vacuum using a SpeedVac concentrator. Peptides were analyzed by LC–MS/MS using a nanoElute2 coupled to a timsTOF Pro2 mass spectrometer (Bruker Daltonics). Samples were separated on a C18 column (150 µm × 25 cm, 1.5 µm; Bruker). The mobile phases consisted of 0.1% formic acid (FA) in water (mobile phase A) and 0.1% FA in acetonitrile (mobile phase B). Peptides were separated using a 54-minute linear gradient from 3% to 35% mobile phase B, with a total run time of 60 minutes at a constant flow rate of 600 nL/min. The timsTOF Pro2 was operated in dia-PASEF mode. Spectra were acquired across an m/z range of 300-1200, using 28 mass windows per cycle. The inverse reduced ion mobility (1/K₀) range was set to 0.65–1.35 V·s/cm². The trapped ion mobility spectrometry (TIMS) analyzer operated at a 100% duty cycle with a ramp time of 75 ms, yielding an overall cycle time of approximately 1.20 seconds. Collision energy was linearly ramped from 59 eV at 1/K₀ = 1.6 to 20 eV at 1/K₀ = 0.6.PrideNot availabledia-PASEF data files were analyzed using directDIA™ workflow embedded in Spectronaut 18 (Biognosys AG) against the Bacteroides fragilis database. Search parameters included trypsin digestion with cleavage after K or R, allowance for up to two missed cleavages, carbamidomethylation of cysteine (static modification), and variable modifications including oxidized methionine and Protein N-terminal acetylation. Peptide spectrum matches (PSMs), peptides, and protein groups were identified using a false discovery rate (FDR) threshold of 0.01. For quantification, all fragment ions passing the FDR threshold were used, and protein abundances were calculated as the area under the curve of extracted ion chromatograms within the defined peak boundaries for each targeted ion.ProteomicsYoung Ah GooMass Spectrometry Technology Access Center, McDonnell Genome Institute, Washington University School of MedicinePARTIALBacteroides FragilisNot availablebyoung-kyu@wustl.eduWashington University in St. LouisUnited StatesfalseOuter Membrane Vesicles Secreted by Bacteroides fragilis Inhibit CFTR Chloride Secretion by Human Colon OrganoidsThe goals of this study were to develop a model to study host pathogen interactions in primary human colon organoids and to test the hypothesis that Bacteroides fragilis toxin (BFT-2) secreted in outer membrane vesicles (OMVs) modulates mucosal immunity and CFTR Cl- secretion. Since Bacteroides species often resides in mucus, OMVs are likely to represent a mechanism of communication between Bacteroides and the host. Two strains of Bacteroides were studied, Enterotoxigenic Bacteroides fragilis (ETBF), which produces BFT-2, and the non-toxigenic Bacteroides fragilis strain NCTC 9343 (NTBF) that does not produce BFT-2. We also utilized two additional strains of Bacteroides fragilis: one in which bft-2 was knocked out (ETBF Δbft), and one that was engineered to contain bft-2 (NTBF+bft). We report that most Bacteroides fragilis OMVs reduced CFTR Cl- secretion but had no effect on tight junction or cell adhesion proteins, transepithelial resistance (TER) or cytokine secretion by primary human colon organoids. We conclude that OMVs secreted by Bacteroides can be an important mechanism of host pathogen interactions in the colon by reducing CFTR Cl- secretion.2026-05-212026-01-23PXD073462NEWT:6945NEWT:184922NEWT:3555NEWT:2NEWT:157546NEWT:35554NEWT:38942NEWT:307972NEWT:32046NEWT:544496NEWT:2042546NEWT:45351NEWT:43179NEWT:4513NEWT:5722NEWT:55153NCBITaxon:10407NEWT:1736309NEWT:309800NEWT:1211601NEWT:876138NEWT:3654NEWT:237561NEWT:5833NEWT:6928NEWT:10036NEWT:36745NEWT:1351NEWT:1438992NEWT:2649997NEWT:272563NEWT:224326NCBITaxon:79857NEWT:1096976NEWT:95648NEWT:3885NEWT:3888NEWT:1589NEWT:135622NCBITaxon:4896NEWT:6915NEWT:3649NEWT:101510NEWT:3880NEWT:272559NEWT:3641NEWT:383379NEWT:466585NEWT:10029NEWT:913645NEWT:1000589NEWT:85963NEWT:85962NEWT:317447NEWT:7955NEWT:7959NEWT:2261NEWT:31156NEWT:398580NEWT:4565NEWT:1264690NEWT:515619NEWT:192875NEWT:34305NEWT:59729NCBITaxon:183674NEWT:224308NEWT:84645NEWT:626528NEWT:139927NEWT:4558NEWT:209285NEWT:5888NEWT:1283NEWT:931281NEWT:4550NEWT:1000561NEWT:197NEWT:1390363NEWT:288705NCBITaxon:79824NEWT:4787NCBITaxon:4563NEWT:5755NEWT:44689NEWT:3218NEWT:5759NEWT:1736231NEWT:1270NEWT:374990NEWT:2242NEWT:4784NEWT:11320NEWT:360106NEWT:286NEWT:391619NEWT:287NEWT:10117NEWT:10239NEWT:10116NEWT:1280NEWT:1735272NEWT:83334NEWT:83332NEWT:44685NEWT:317513NEWT:1148NEWT:580240NEWT:294128NEWT:11676NEWT:55571NEWT:100226NEWT:4530NEWT:4896NEWT:75058NEWT:13616NEWT:1390NEWT:1094343NEWT:296543NEWT:1773NEWT:1895NEWT:1182590NEWT:3712NEWT:105023NEWT:935293NEWT:64152NEWT:4924NEWT:749200NEWT:375146NEWT:990346NEWT:145953NEWT:257309NEWT:100816NEWT:263NEWT:230741NEWT:52283NEWT:284812NCBITaxon:1313NEWT:43330NEWT:1603293NEWT:408169NEWT:44544NEWT:4911NEWT:645463NEWT:3702NEWT:129249NEWT:243277NEWT:990119NEWT:408172NEWT:408170NEWT:493760NEWT:260710NEWT:257313NEWT:400772NEWT:3708NEWT:128161NEWT:332648NEWT:106592NEWT:536231NEWT:460519NEWT:1187947NEWT:1432138NEWT:10312NEWT:1424507NCBITaxon:1773NEWT:9598NEWT:8030NEWT:1639NEWT:188229NEWT:3818NEWT:480NEWT:4909NEWT:67767NEWT:432359NEWT:46835NEWT:1182263NEWT:2711NEWT:376686NEWT:95486NEWT:9103NEWT:29159NEWT:253NEWT:10306NCBITaxon:2759NEWT:1233435NEWT:93061NEWT:8022NEWT:145943NCBITaxon:4932NEWT:595536NEWT:240906NEWT:593117NEWT:89920NEWT:3635NEWT:5811NEWT:235443NEWT:272623NEWT:272624NEWT:411483NEWT:884019NEWT:198215NEWT:411490NEWT:983964NEWT:169963NEWT:32644NEWT:225117NEWT:499175NEWT:109779NEWT:476272NEWT:3747NEWT:195051NEWT:367830NEWT:1255228NEWT:178616NEWT:410289NEWT:373153NEWT:352472NEWT:357NEWT:360094NEWT:470NEWT:1313NEWT:411469NEWT:84023NEWT:559292NEWT:39491NCBITaxon:5811NEWT:411464NEWT:411460NEWT:2014887NEWT:2762NEWT:1174673NEWT:562NEWT:411470NEWT:33952NEWT:2094720NCBITaxon:2697049NEWT:571256NEWT:28038NEWT:1663NEWT:1423NEWT:4932NEWT:3603NEWT:2759NEWT:3847NEWT:327159NEWT:178876NEWT:327160NEWT:573NEWT:9031NEWT:7091NEWT:108931NEWT:241368NEWT:42528NEWT:190802NEWT:9778NEWT:150475NEWT:303NEWT:9417NEWT:7111NEWT:347515NEWT:1216979NEWT:5180NEWT:256737NEWT:9541NEWT:115104NEWT:1121114NEWT:663NEWT:1081927NEWT:1238993NEWT:67825NEWT:185579NEWT:941442NEWT:220668NEWT:13076NEWT:1249668NEWT:7108NEWT:317NEWT:7227NEWT:7469NEWT:885318NEWT:9402NEWT:415540NEWT:550NEWT:675060NEWT:4081NEWT:334542NEWT:554NEWT:98334NEWT:426428NEWT:7574NEWT:1715256NEWT:7215NEWT:575412NEWT:29204NEWT:2172103NEWT:507601NEWT:643680NCBITaxon:6157NEWT:746360NEWT:6239NEWT:470150NEWT:216257NEWT:102169NEWT:9986NEWT:4054NEWT:73239NEWT:226186NEWT:1268063NEWT:8782NEWT:1263854NEWT:435590NEWT:1902NEWT:160488NEWT:28104NEWT:1908NEWT:13164NEWT:216129NCBITaxon:2NEWT:985076NEWT:1215323NEWT:52641NEWT:7038NEWT:6192NEWT:28532NCBITaxon:38727NEWT:353152NEWT:2829NEWT:366581NEWT:216599NEWT:216595NEWT:1194669NEWT:243230NEWT:8355NEWT:9685NEWT:7029NEWT:1080772NEWT:8479NEWT:1283300NEWT:6183NEWT:6063NEWT:630NEWT:334747NEWT:61235NEWT:15368NEWT:6289NEWT:436486NEWT:6287NEWT:300641NEWT:727NEWT:9796NEWT:725NEWT:170187NEWT:469008NEWT:260707NEWT:256318NCBITaxon:6191NEWT:1836NEWT:185431NEWT:29760NEWT:260704NEWT:703612NEWT:260705NEWT:80863NEWT:2697049NEWT:105231NEWT:1216981NEWT:4097NCBITaxon:6073NEWT:884204NEWT:6279NEWT:1123869NEWT:9544NEWT:7370NEWT:83906NEWT:607699NEWT:6282NEWT:208964NEWT:1134506NEWT:575584NEWT:38783NEWT:8727NEWT:4006NEWT:8726NEWT:6426NEWT:6669NEWT:10090NEWT:4120NEWT:51515NEWT:5693NEWT:8724NEWT:51511NEWT:92867NEWT:8723NEWT:5334NEWT:37334NEWT:382352NCBITaxon:10359NEWT:242619NEWT:632957NEWT:34073NEWT:373995NEWT:5689NEWT:544404NEWT:9925NEWT:8839NEWT:4232NEWT:2758385NEWT:37349NEWT:4113NEWT:837NEWT:11298NEWT:171101NEWT:714NEWT:36015NEWT:421932NEWT:196627NEWT:5691NEWT:48479NEWT:627025NEWT:1097677NEWT:61674NEWT:1117957NEWT:9913NEWT:8709NEWT:4100NEWT:1076NEWT:6763NEWT:803NEWT:29722NEWT:380394NEWT:1692259NEWT:180066NEWT:135588NEWT:1843183NEWT:58002NEWT:326423NEWT:36111NEWT:4577NEWT:1416333NEWT:5664NEWT:2157NEWT:1678078NEWT:749906NEWT:418985NEWT:146479NEWT:749907NEWT:1911079NEWT:69373NEWT:264203NEWT:1480154NEWT:1274414NEWT:27606NEWT:59202NEWT:9975NEWT:9612NEWT:38865NEWT:51953NEWT:3197NEWT:9615NEWT:10299NEWT:860688NEWT:112273NEWT:36329NEWT:1147787NCBITaxon:3044782NEWT:1440772NEWT:72407NEWT:1355477NEWT:349741NEWT:9605NEWT:9606NEWT:90371NEWT:157295NEWT:641501NEWT:7668NEWT:915099NEWT:74940NEWT:9721NEWT:137221NEWT:9600NEWT:1450511NEWT:1143193NEWT:411901NEWT:9838NEWT:105884NCBITaxon:9615NEWT:9839NEWT:58334NEWT:1193501NEWT:3055NEWT:58331NEWT:6326NEWT:6689NEWT:5476NEWT:299767NEWT:1274432NEWT:1274426NEWT:70448NEWT:9825NEWT:1274423NEWT:393765NEWT:698936NEWT:39946NEWT:9823NEWT:9940NEWT:95602NEWT:8732NEWT:521001NEWT:39947NEWT:224911NEWT:1274420NEWT:5784580000-0003-3370-6985