Disassembly of F-actin cytoskeleton after interaction of Bacillus cereus with fully differentiated human intestinal Caco-2 cells

In the present study, the role of direct procaryote-eucaryote interactions in the virulence of Bacillus cereus was investigated. As a model of human enterocytes, differentiated Caco-2 cells were used. Infection of fully differentiated Caco-2 cells with B. cereus in the exponential phase of growth, in order to minimize the concentration of spores or sporulating microorganisms, shows that a strain-dependent cytopathic effect develops. Interestingly, addition of 3-h-old cultures of some strains resulted in complete detachment of the cultured cells after a 3-h infection whereas no such effect was found after a 3-h infection with 16-h-old cultures. Infection of enterocyte-like cells with B. cereus leads to disruption of the F-actin network and necrosis. Even though the effect of secreted factors cannot be ruled out, direct eucaryote-procaryote interaction seems to be necessary. In addition, we observed that some B. cereus strains were able to be internalized in Caco-2 cells. Our findings add a new insight into the mechanisms of virulence of B. cereus in the context of intestinal infection.Facultad de Ciencias ExactasCentro de Investigación y Desarrollo en Criotecnología de Alimento

Disassembly of F-actin cytoskeleton after interaction of Bacillus cereus with fully differentiated human intestinal Caco-2 cells

In the present study, the role of direct procaryote-eucaryote interactions in the virulence of Bacillus cereus was investigated. As a model of human enterocytes, differentiated Caco-2 cells were used. Infection of fully differentiated Caco-2 cells with B. cereus in the exponential phase of growth, in order to minimize the concentration of spores or sporulating microorganisms, shows that a strain-dependent cytopathic effect develops. Interestingly, addition of 3-h-old cultures of some strains resulted in complete detachment of the cultured cells after a 3-h infection whereas no such effect was found after a 3-h infection with 16-h-old cultures. Infection of enterocyte-like cells with B. cereus leads to disruption of the F-actin network and necrosis. Even though the effect of secreted factors cannot be ruled out, direct eucaryote-procaryote interaction seems to be necessary. In addition, we observed that some B. cereus strains were able to be internalized in Caco-2 cells. Our findings add a new insight into the mechanisms of virulence of B. cereus in the context of intestinal infection.Facultad de Ciencias ExactasCentro de Investigación y Desarrollo en Criotecnología de Alimento

Muc2 Protects against Lethal Infectious Colitis by Disassociating Pathogenic and Commensal Bacteria from the Colonic Mucosa

Despite recent advances in our understanding of the pathogenesis of attaching and effacing (A/E) Escherichia coli infections, the mechanisms by which the host defends against these microbes are unclear. The goal of this study was to determine the role of goblet cell-derived Muc2, the major intestinal secretory mucin and primary component of the mucus layer, in host protection against A/E pathogens. To assess the role of Muc2 during A/E bacterial infections, we inoculated Muc2 deficient (Muc2−/−) mice with Citrobacter rodentium, a murine A/E pathogen related to diarrheagenic A/E E. coli. Unlike wildtype (WT) mice, infected Muc2−/− mice exhibited rapid weight loss and suffered up to 90% mortality. Stool plating demonstrated 10–100 fold greater C. rodentium burdens in Muc2−/− vs. WT mice, most of which were found to be loosely adherent to the colonic mucosa. Histology of Muc2−/− mice revealed ulceration in the colon amid focal bacterial microcolonies. Metabolic labeling of secreted mucins in the large intestine demonstrated that mucin secretion was markedly increased in WT mice during infection compared to uninfected controls, suggesting that the host uses increased mucin release to flush pathogens from the mucosal surface. Muc2 also impacted host-commensal interactions during infection, as FISH analysis revealed C. rodentium microcolonies contained numerous commensal microbes, which was not observed in WT mice. Orally administered FITC-Dextran and FISH staining showed significantly worsened intestinal barrier disruption in Muc2−/− vs. WT mice, with overt pathogen and commensal translocation into the Muc2−/− colonic mucosa. Interestingly, commensal depletion enhanced C. rodentium colonization of Muc2−/− mice, although colonic pathology was not significantly altered. In conclusion, Muc2 production is critical for host protection during A/E bacterial infections, by limiting overall pathogen and commensal numbers associated with the colonic mucosal surface. Such actions limit tissue damage and translocation of pathogenic and commensal bacteria across the epithelium

Adherent Bacteria and Parasiticidal Secretion Products of Human Cervicovaginal Microbiota-Associated Lactobacillus gasseri Confer Non-Identical Cell Protection against Trichomonas vaginalis-Induced Cell Detachment

Trichomonas vaginalis, a protozoan parasite specific to the human genital tract, is one of the most common sexually transmitted pathogens. Its pathogenicity is strongly associated with its expression of a broad array of proteases triggering cytotoxic effects in host epithelial cells. Vaginal microbiota-associated Lactobacillus, including those of L. gasseri in particular, can counteract T. vaginalis pathogenesis, but the mechanisms involved have yet to be clarified. T. vaginalis strain G3 (Tv G3) cytotoxicity was assessed by examining cell morphology, cell detachment, and fluorescent labeling of the F-actin cytoskeleton and immunolabeling of vinculin-position focal adhesions (FAs) by confocal laser scanning electron microscopy on confluent cervicovaginal epithelial HeLa cell monolayers. The inhibitory effects of bacterial cells and secreted products of L. gasseri ATCC 9857 and KS 120.1 on the Tv G3 viability and parasite deleterious effects on HeLa cells were investigated. Pre-adhering L. gasseri cells delayed but did not inhibit Tv G3-induced cell detachment, F-actin cytoskeleton disorganization and the disappearance of vinculin-positive focal FAs. L. gasseri KS 120.1 secretion products had a rapid parasiticide activity by killing time- and concentration-dependent Tv G3 parasites after direct contact. By killing Tv G3 parasites already associated with the epithelial cells, secretion products have abolished parasite-induced cell detachment. Our findings suggest that vagina microbiota-associated L. gasseri creates a physical barrier and exerts pharmacological-type mechanisms to counteract the deleterious cytotoxic effects of T. vaginalis

The macrophage microtubule network acts as a key cellular controller of the intracellular fate of Leishmania infantum

International audienceThe parasitophorous vacuoles (PVs) that insulate Leishmania spp. in host macrophages are vacuolar compartments wherein promastigote forms differentiate into amastigote that are the replicative form of the parasite and are also more resistant to host responses. We revisited the biogenesis of tight-fitting PVs that insulate L. infantum in promastigote-infected macrophage-like RAW 264.7 cells by time-dependent confocal laser multidimensional imaging analysis. Pharmacological disassembly of the cellular microtubule network and silencing of the dynein gene led to an impaired interaction of L. infantum-containing phagosomes with late endosomes and lysosomes, resulting in the tight-fitting parasite-containing phagosomes never transforming into mature PVs. Analysis of the shape of the L. infantum parasite within PVs, showed that factors that impair promastigote-amastigote differentiation can also result in PVs whose maturation is arrested. These findings highlight the importance of the MT-dependent interaction of L. infantum-containing phagosomes with the host macrophage endolysosomal pathway to secure the intracellular fate of the parasite

Disassembly of F-Actin Cytoskeleton after Interaction of Bacillus cereus with Fully Differentiated Human Intestinal Caco-2 Cells

In the present study, the role of direct procaryote-eucaryote interactions in the virulence of Bacillus cereus was investigated. As a model of human enterocytes, differentiated Caco-2 cells were used. Infection of fully differentiated Caco-2 cells with B. cereus in the exponential phase of growth, in order to minimize the concentration of spores or sporulating microorganisms, shows that a strain-dependent cytopathic effect develops. Interestingly, addition of 3-h-old cultures of some strains resulted in complete detachment of the cultured cells after a 3-h infection whereas no such effect was found after a 3-h infection with 16-h-old cultures. Infection of enterocyte-like cells with B. cereus leads to disruption of the F-actin network and necrosis. Even though the effect of secreted factors cannot be ruled out, direct eucaryote-procaryote interaction seems to be necessary. In addition, we observed that some B. cereus strains were able to be internalized in Caco-2 cells. Our findings add a new insight into the mechanisms of virulence of B. cereus in the context of intestinal infection

Thermosensitive and Mucoadhesive Pluronic-Hydroxypropylmethylcellulose Hydrogel Containing the Mini-CD4 M48U1 Is a Promising Efficient Barrier against HIV Diffusion through Macaque Cervicovaginal Mucus

International audienc