Immunization of Mice with Lactobacillus casei Expressing a Beta-Intimin Fragment Reduces Intestinal Colonization by Citrobacter rodentium

Abstract

Gastrointestinal infections are among the leading causes of childhood mortality around the world and are responsible for approximately 2 million deaths every year (12). Enteropathogenic Escherichia coli (EPEC) is one of the main causes of childhood diarrhea in developing countries and has been considered a worldwide pathogen (7,18,31,59). EPEC has been classified into typical or atypical according to the expression or lack of expression, respectively, of the bundle-forming pilus, encoded by the EAF plasmid (1, 50). Epidemiological studies suggest that the prevalence of atypical EPEC has increased in acute or persistent diarrhea cases in both children and adults (23,29,52,62). EPEC colonizes the intestinal mucosa, causing the characteristic attaching and effacing (A/E) lesions. The formation of the A/E lesion involves a type III secretion system (T3SS), encoded by the LEE pathogenicity island, which delivers a range of effector proteins into host cells. Using this strategy, the pathogen triggers actin polymerization, producing a pedestal-shaped structure to which the bacteria strongly attach through the interaction of the adhesin intimin with the translocated receptor Tir (27,38). Intimin is a 94-kDa protein inserted into the bacterial outer membranes from typical and atypical EPEC, enterohemorrhagic E. coli (EHEC), and the mouse pathogen Citrobacter rodentium. This adhesin is encoded by the eae gene and is essential for virulence (28,48). The N-terminal region is conserved among intimins, whereas the C-terminal portion is highly variable and has been used to classify intimin into 29 types that can be related to tropism for different intestinal tissues (26,35,49). Among them, the ␤ subtype is one of the intimins most frequently expressed by EPEC isolates (6,56,60). EPEC and EHEC infections can be prevented by breastfeeding (17,57), and antibodies against intimin are usually observed in the milk of women living in areas of endemicity (58,70). In addition, these antibodies are able to inhibit the adhesion of EPEC to epithelial cells in vitro (15,19). Several vaccination strategies using intimin have reduced colonization or mortality in animal models of infection with rabbit enteropathogenic E. coli (REPEC), EHEC, or C. rodentium (4,30,36,39), suggesting that intimin is a good candidate for the formulation of vaccines against these infections. A minimal intimin fragment able to bind to the receptor Tir is composed of the last 190 amino acids, but high-affinity binding was observed only with a larger fragment comprising the last 280 amino acid residues (Int 280 ) (8). In fact, subcutaneous immunization of mice with the Int 280 fragment from ␣-intimin reduced intestinal colonization by a C. rodentium strain expressing an intimin from the same subtype (30). However, analysis of the immunodominant regions inside Int 280 have shown that antibodies against this fragment were directed to 2 regions in the N terminus of Int 280 , more precisely, a

A Self-Assembling Whole-Cell Vaccine Antigen Presentation Platform

“…3I; see also Table S2 in the supplemental material), while IgG was not detected in the serum throughout the study period. These findings were consistent with a previous report that sublingual immunization with a live bacterial vector generated antigen-specific fecal IgA but not serum IgG (18). These results demonstrate, for the first time, that a protein antigen anchored to the biofilm matrix of a bacterial cell can be delivered sublingually to stimulate an antigenspecific mucosal immune response.…”

TACI deficiency enhances antibody avidity and clearance of an intestinal pathogen

“…While the neutralizing targets of anti-Citrobacter antibodies are incompletely known, Ferreira et al (38) and Ghaem-Maghami et al (35) showed that antibodies directed against intimin, a surface protein, protect. Accordingly, we tested the average avidity of IgG antibodies directed against intimin in TACI-deficient and WT mice.…”

The role of specific IgG and complement in combating aprimary mucosal infection of the gut epithelium

“…It was demonstrated that subcutaneous immunization with purified int cv induced concentration of ant-Int cv IgG1 ans IgG2a in sera, indicating a balanced Th1:Th2 profile [38]. In contrast, protection elicited by L. casei -Intcv vaccines was observed in conditions where no significant induction of anti-Intcv IgG in sera was detected [38]. These data implicate that other mechanisms are involved in the protection elicited by L. casei -Int cv oral vaccines as compared to subcutaneous immunization.…”