New flow cytometric method to quantify the inhibition of enteropathogenic <i>Escherichia coli</i> adhesion by anti‐adhesin antibodies

Boullier, Séverine; Tasca, Christian; Milon, Alain

doi:10.1002/cyto.a.10042

Cited by 10 publications

(6 citation statements)

References 23 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…This hypothesis was confirmed by our results ex vivo that showed a reduction in the level of adhesion of E22 bacteria to the ileum of suckling rabbits compared with that in weaned pups. It was also consistent with our results obtained by using a recently developed assay of inhibition of AF/R2-dependent adhesion of EPEC to HeLa cells (5). This assay is based on the ability of pathogenic E. coli O103 to adhere to HeLa cells in vitro via the AF/R2 adhesin (30).…”

Section: Discussionsupporting

confidence: 91%

“…pEGFP-E22 was cultured in Penassay broth (antibiotic medium 3; Difco) supplemented with 25 g/ml of carbenicillin, as described previously (5).…”

Section: Methodsmentioning

confidence: 99%

“…Incubation of the bacteria with the cells was realized in 500 l of MEM buffered with 5% FCS and 1% mannose, with a starting inoculum of 20 l (about 2 ϫ 10 7 bacteria per well). To test the inhibitory capacity of adhesion of the serum, milk, or lactoserum of the does, 20 l of strain pEGFP-E22 bacteria grown overnight (10 9 CFU/ml) in Penassay broth was preincubated with different amounts of serum, milk, or lactoserum for 1 h at room temperature, prior to incubation for 30 min with HeLa cells, as described previously (5). After 30 min of incubation of the cells with the bacteria at 37°C, the cells were washed three times with PBS.…”

Section: Methodsmentioning

confidence: 99%

“…We previously developed an in vitro test to quantify the capacity of rabbit sera to inhibit the AF/R2-dependent bacterial adhesion to HeLa cells (5). In order to assess whether the protective effect of doe's milk could be linked to an inhibiting adhesion effect in vitro, we adapted this test for milk and lactoserum samples.…”

Section: Vol 14 2007 Epec Colibacillosis Protection By Maternal Milmentioning

confidence: 99%

See 3 more Smart Citations

Maternal Milk Contains Antimicrobial Factors That Protect Young Rabbits from Enteropathogenic Escherichia coli Infection

Gallois

Gidenne

Tasca

et al. 2007

Clin Vaccine Immunol

Self Cite

View full text Add to dashboard Cite

Enteropathogenic Escherichia coli (EPEC) colibacillosis represents a major cause of lethal diarrhea in young children in developing countries. EPEC strains also infect numerous mammal species and represent a major economical problem in rabbit industry. Protection against this pathogen is a challenging goal both in humans and in other mammal species. Despite a good knowledge of the pathogenicity mechanisms of EPEC, the intrinsic and environmental factors that control the expression of EPEC virulence in mammals remain unknown. For instance, the exacerbated sensitivity of young mammals to EPEC infection is still unexplained. Our goal was to investigate if age or other factors, like milk consumption, could be determinants that trigger the disease. We used rabbits as an animal model to study the role of milk in the sensitivity to an EPEC infection. Weaned and suckling rabbits were orally inoculated with EPEC strain E22 (O103:H2:K؊) at 28 days of age, and the evolution of the disease was investigated in the two groups. In addition, in order to better characterize the interactions between milk and EPEC, we determined in vitro bacterial growth and the abilities of EPEC cells to adhere to epithelial cells in the presence of milk. Our results demonstrate a protective role of milk in vivo in association with in vitro antibacterial activity. These effects are independent of the presence of specific anti-EPEC antibodies.

show abstract

Section: Discussionsupporting

confidence: 91%

“…pEGFP-E22 was cultured in Penassay broth (antibiotic medium 3; Difco) supplemented with 25 g/ml of carbenicillin, as described previously (5).…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Vol 14 2007 Epec Colibacillosis Protection By Maternal Milmentioning

confidence: 99%

See 2 more Smart Citations

Maternal Milk Contains Antimicrobial Factors That Protect Young Rabbits from Enteropathogenic Escherichia coli Infection

Gallois

Gidenne

Tasca

et al. 2007

Clin Vaccine Immunol

Self Cite

View full text Add to dashboard Cite

show abstract

“…Flow cytometry was used, as previously described with modifications (2), to compare the numbers of HEp-2 cells to which green fluorescent protein (GFP)-expressing EPEC strains adhered following preincubation with normal serum or anti-HisInt 286. The flow cytometry was performed at the Uniformed Services University Biomedical Instrumentation Center.…”

mentioning

confidence: 99%

Antibody against the Carboxyl Terminus of Intimin α Reduces Enteropathogenic Escherichia coli Adherence to Tissue Culture Cells and Subsequent Induction of Actin Polymerization

et al. 2005

View full text Add to dashboard Cite

The C-terminal third of intimin binds to its translocated receptor (Tir) to promote attaching and effacing lesion formation during infection with enteropathogenic Escherichia coli (EPEC). We observed that the adherence of EPEC strains to HEp-2 cells was reduced and that actin polymerization was blocked by antibody raised against the C-terminal third of intimin ␣.Enteropathogenic Escherichia coli (EPEC) strains are pediatric pathogens associated with acute and protracted infantile diarrhea, particularly in developing countries. Outbreaks of disease among children and adults also occur sporadically worldwide (19). EPEC strains target the small intestinal mucosa where they adhere, efface the microvilli, and induce cytoskeletal changes and pedestal formation in the underlying enterocytes. These lesions, described in 1983 by Moon et al. as attaching and effacing (A/E) lesions, are the hallmark pathology of EPEC infection (18). The bacterial surface protein intimin is necessary for A/E lesion formation and for the full virulence of EPEC in human volunteer infections (3, 4, 11). Since the carboxyl-terminal 190 amino acids of intimin bind the translocated intimin receptor, Tir (1, 15), we reasoned that antibody directed against the binding domain of intimin might block lesion formation.Intimins expressed by various E. coli strains that produce A/E lesions in humans and animals have been divided into at least five distinct types based on the antigenic variability of the 280 C-terminal amino acids that comprise the eukaryotic binding domain. Previous studies in our laboratory showed that antibody directed against the binding domain of intimin ␥ effectively reduces binding of enterohemorrhagic E. coli (EHEC) O157:H7, but not EPEC strain E2348/69, to HEp-2 cells (5). Other investigators used rabbit polyclonal antibodies against a histidine-tagged 280-amino-acid portion of the C terminus of intimin ␣ to monitor intimin expression in the HEp-2 cell EPEC adherence model (12). Here we evaluated the capacity of antibody specific for the carboxyl-terminal binding domain of EPEC intimin ␣ to interfere with the binding of EPEC strains and to block actin polymerization in the HEp-2 cell adherence assay.

show abstract

Comparison of invasion of fibroblasts and macrophages by high- and low-virulence Leptospira strains: colonization of the host-cell nucleus and induction of necrosis by the virulent strain

Ojcius

Yan

2007

Arch Microbiol

View full text Add to dashboard Cite

The infection cycle of low- and high-virulence strains of Leptospira interrogans was compared in fibroblasts and macrophages. L. interrogans serovar Lai strain Lai was used as a representative high-virulence strain, while L. interrogans serovars Pomona strain Luo was used as a low-virulence strain. L. biflexa serovar Patoc strain Patoc I, a nonparasitic strain of Leptospira, was used as a control. Both the high- and low-virulence strains could adhere to fibroblasts and macrophages using one or both ends of the spirochete, which was followed by phagocytosis of both strains. Both strains adhered more strongly to macrophages than fibroblasts. However, the high-virulence strain could invade the host-cell nucleus, while the low-virulence strain remained in phagosomes. The L. biflexa strain neither adhered to nor invaded either cell type. Both of the L. interrogans strains also induced cell death (mostly necrosis) of macrophages, whether or not the spirochetes were viable, suggesting that leptospiral virulence is unrelated to macrophage death. However, the high-virulence strain induced mainly necrosis in fibroblasts, while the low-virulence strain induced more apoptosis. Thus, the main feature distinguishing the two L. interrogans strains is the ability of the high-virulence strain to invade the host-cell nucleus and induce pro-inflammatory necrosis in fibroblasts.

show abstract

New flow cytometric method to quantify the inhibition of enteropathogenic Escherichia coli adhesion by anti‐adhesin antibodies

Cited by 10 publications

References 23 publications

Maternal Milk Contains Antimicrobial Factors That Protect Young Rabbits from Enteropathogenic Escherichia coli Infection

Maternal Milk Contains Antimicrobial Factors That Protect Young Rabbits from Enteropathogenic Escherichia coli Infection

Antibody against the Carboxyl Terminus of Intimin α Reduces Enteropathogenic Escherichia coli Adherence to Tissue Culture Cells and Subsequent Induction of Actin Polymerization

Comparison of invasion of fibroblasts and macrophages by high- and low-virulence Leptospira strains: colonization of the host-cell nucleus and induction of necrosis by the virulent strain

Contact Info

Product

Resources

About