Humoral and Cellular Responses to a conserved bacterial antigen: effects on microbiota

Commensal microorganisms like Escherichia coli can also behave as pathobionts. A key virulence mechanism in this bacterium is the overexpression of the AcrAB-TolC efflux pump; TolC, located in the outer membrane, is immunogenic and displays antibody-recognizable epitopes. Although recombinant TolC provides protection against multidrug-resistant E. coli, its mucosal immune effects remain poorly understood. This project evaluates humoral and cellular responses in the intestinal mucosa (colonic lamina propria (LP), mesenteric lymph node (mLN), and Peyer’s patches (PP)) and systemically, following intraperitoneal immunization of C57BL/6 mice with TolC+Alum. Naturally, anti-TolC IgM is detected in peritoneal lavage across all age groups, with higher levels in adult spleens. Anti-TolC IgG rises in neonatal lavage (p < 0.0001), and IgA increases in 4- and 8-week-old mice. Seven days after a single dose of TolC+Alum, mLN showed a significant increase in the frequency of IgM⁺ plasmablasts (p < 0.01), IgG⁺ plasmablasts (p < 0.05), and IgM⁺ plasma cells (p < 0.01). A reduction in IgG⁺ memory B cells was also observed (p < 0.01). In Peyer’s patches, there was an increase in the relative frequency of IgG⁺ plasmablasts (p < 0.05), IgG⁺ plasma cells (p < 0.05), IgG⁺ memory B cells (p < 0.05), and IgA⁺ memory B cells (p < 0.01). In the colonic lamina propria, a decrease in the frequency of IgG⁺ memory B cells was observed (p < 0.05). After three doses (42 dpi), a sustained increase in serum anti-TolC IgG was observed, with no changes in total or specific IgM/IgA. In mucosa, CD4⁺ and Th17 increased in PP, while lamina propria showed reductions in CD45⁺, CD4⁺, Th17, and IgM⁺ plasmablasts. Together, these results confirm TolC as a highly immunogenic antigen that modulates both systemic and mucosal immunity, including shifts in memory B cells and antibody-producing cells, with a tendency toward immune homeostasis after repeated exposure.