Oral administration of mice Conventional BALB/c mice, 3 to 6 weeks of age were purchased from INRA animal care facilities (Jouy-en-Josas, Cytoskeletal Signaling inhibitor France), acclimatized for 1 week before immunization under standard animal husbandry conditions in the animal facility (Unité d’Expérimentation Animale, Jouy-en-Josas, France). Mice (n = 8) were intragastrically administered

with 1×109 (CFU) of strains, LL, LL-BLG or LLmInlA-BLG on 3 consecutive days using a gavage tube feeding. On the fourth day, the small intestine was collected for subsequent BLG quantification in isolated IECs. Intestinal epithelial cells isolation Mice were euthanized, and their small intestines were removed, rinsed with complete DMEM medium (containing 2 mM Thiazovivin nmr L-glutamine and 10% fetal calf serum).

The length of intestine was opened and submerged in buffer A (in mM: 120 NaCl, 4.7 KCl, 2.4 KCl, 1.2 KH2PO4, 1.2 Na2HP04, 25 NaHCO3, 10 HEPES, 5 EDTA, 0.5 DTT, 0.25% BSA; at pH 7.4 warmed to 37°C) for 20 min with agitation at 240 rpm [44]. Cells were find more collected by centrifugation (415.73 g – 2000 rpm – for 5 min) at room temperature, washed once with PBS and lysed by sonication (3 times, 10 s). The cell lysate was centrifuged for 10 min at 3143.98 g (5500 rpm), then the supernatant was recovered and stored at -80°C. The EIA to detect BLG was performed as described above. Statistical analyses The results are expressed as mean ± standard error (SE) values. Statistical significance between the groups was calculated using the One Way ANOVA (and nonparametric) test, followed by the “Bonferroni” post-test. Values of p < 0.05 were considered significant. Acknowledgements The research leading to these results has received funding from the European Community's Seventh Framework

Programme (FP7/2007-2013) under grant agreement n°215553-2. Antibodies and reagents were kindly provided by Karine Adel Patient and Jean-Michel Wal (INRA, UR496, Unité d’Immuno-Allergie Alimentaire, F-78352 BCKDHB Jouy-en-Josas, France; CEA, Institut de Biologie et de Technologie de Saclay, iBiTeC-S, Laboratoire d’Etudes et de Recherches en Immunoanalyse, F-91191 Gif-sur-Yvette, France). pPL2mInLA was a kind gift of Dr. Schubert (Helmholtz Centre for Infection Research, Inhoffenstr. 7, D-38124 Braunschweig, Germany). References 1. Donnelly JJ, Liu MA, Ulmer JB: Antigen presentation and DNA vaccines. Am J Respir Crit Care Med 2000, 162:190–193. 2. Ledwith BJ, Manam S, Troilo PJ, Barnum AB, Pauley CJ, Griffiths TG, Harper LB, Schock HB, Zhang H, Faris JE, Way PA, Beare CM, Bagdon WJ, Nichols WW: Plasmid DNA vaccines: assay for integration into host genomic DNA. Dev Biol 2000, 104:33–43. 3.