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In vitro adhesion properties of Shiga toxin-producing Escherichia coli isolated from cattle, food, and humans.

Pradel N, Etienne-Mesmin L, Thévenot J, Cordonnier C, Blanquet-Diot S, Livrelli V - Front Microbiol (2015)

Bottom Line: We analyzed the ability of 256 STEC strains belonging to seropathotype A (the most virulent O157:H7 serotype) to seropathotype E (not involved in human disease) to adhere to HEp-2, HCT-8, and T84 cell lines.Of the 256 STEC tested most (82%) were non-adherent in our assays.The adhesion levels were globally low and were not related to pathogenicity, although the highest levels were associated to O26:H11 and O103:H2 strains of seropathotype B (associated with HUS but less commonly than serotype O157:H7), possessing both the eae and toxB genes.

View Article: PubMed Central - PubMed

Affiliation: Centre de Recherche en Nutrition Humaine Auvergne, M2iSH, 'Microbes, Intestin, Inflammation et Susceptibilité de l'Hôte' UMR INSERM/Université d'Auvergne U1071 USC-INRA 2018, Clermont Université - Université d'Auvergne Clermont-Ferrand, France.

ABSTRACT
Shiga toxin-producing Escherichia coli (STEC) are able to cause serious illnesses ranging from diarrhea to hemorrhagic colitis and hemolytic-uremic syndrome (HUS). These bacteria colonize the digestive tract of humans and produce Shiga-toxins, which are considered to be essential for virulence and are crucial in lethal infection. Colon colonization is supposed to be a determinant step in the development of the infection, but the virulence traits that mediate this step are unclear. We analyzed the ability of 256 STEC strains belonging to seropathotype A (the most virulent O157:H7 serotype) to seropathotype E (not involved in human disease) to adhere to HEp-2, HCT-8, and T84 cell lines. Of the 256 STEC tested most (82%) were non-adherent in our assays. The adhesion levels were globally low and were not related to pathogenicity, although the highest levels were associated to O26:H11 and O103:H2 strains of seropathotype B (associated with HUS but less commonly than serotype O157:H7), possessing both the eae and toxB genes.

No MeSH data available.


Related in: MedlinePlus

Determination of experimental conditions for bacterial adherence of Escherichia coli strains to human colonic carcinoma T84 cells. Cells were infected with bacteria grown for 3 h (exponential growth phase) or 18 h (stationary growth phase), in Luria Bertani Broth (LB) or in Cell Culture Medium (CCM), at 37°C before infection. The cells were then washed, treated with Triton X-100, and the recovered adherent bacteria were diluted and plated for colony forming unit counting. Results are expressed as percentage of adherent bacteria, relative to adherent bacteria obtained after a 3 h growth in LB taken as 100%. EDL933: EHEC O157:H7; CH014: non-O157 STEC; C600: K-12 E. coli; E2348/69: EPEC. All assays were performed independently at least three times. Results are means ± SEM of bacteria adhering to T84 cells.
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Figure 1: Determination of experimental conditions for bacterial adherence of Escherichia coli strains to human colonic carcinoma T84 cells. Cells were infected with bacteria grown for 3 h (exponential growth phase) or 18 h (stationary growth phase), in Luria Bertani Broth (LB) or in Cell Culture Medium (CCM), at 37°C before infection. The cells were then washed, treated with Triton X-100, and the recovered adherent bacteria were diluted and plated for colony forming unit counting. Results are expressed as percentage of adherent bacteria, relative to adherent bacteria obtained after a 3 h growth in LB taken as 100%. EDL933: EHEC O157:H7; CH014: non-O157 STEC; C600: K-12 E. coli; E2348/69: EPEC. All assays were performed independently at least three times. Results are means ± SEM of bacteria adhering to T84 cells.

Mentions: Different bacterial growth phases (exponential growth phase vs. late-log phase), and types of bacterial culture media (LB vs. CCM) were tested in preliminary infection experiments on the T84 human colonic carcinoma cell line. These assays were performed with two virulent STEC strains (O157:H7 strain EDL933 and O91:H21 strain CH014), the EPEC reference strain (E2348/69) as a positive control, and the E. coli strain K-12 C600 as a negative control (Figure 1). The best adhesion levels were obtained with strains grown for 3 h in CCM. For further quantitative experiments, we thus chose to incubate the bacteria for 3 h at 37°C in CCM before infection. Quantitative experiments, performed on 17 STEC strains isolated from human cases, indicated that the number of bacteria adhering to the T84 cells was between 2.4 × 103 and 4.4 × 105 CFU/well (Figure 2A). The number reached 2 × 106 CFU/well for the EPEC reference strain, indicating that STEC adhered to very low levels compared to the EPEC strain, and just above the level of the negative control C600. Adherence patterns were investigated after staining with Giemsa and by SEM observations. The STEC strains adhered sparsely, whereas the EPEC formed micro-colonies (Figure 2B).


In vitro adhesion properties of Shiga toxin-producing Escherichia coli isolated from cattle, food, and humans.

Pradel N, Etienne-Mesmin L, Thévenot J, Cordonnier C, Blanquet-Diot S, Livrelli V - Front Microbiol (2015)

Determination of experimental conditions for bacterial adherence of Escherichia coli strains to human colonic carcinoma T84 cells. Cells were infected with bacteria grown for 3 h (exponential growth phase) or 18 h (stationary growth phase), in Luria Bertani Broth (LB) or in Cell Culture Medium (CCM), at 37°C before infection. The cells were then washed, treated with Triton X-100, and the recovered adherent bacteria were diluted and plated for colony forming unit counting. Results are expressed as percentage of adherent bacteria, relative to adherent bacteria obtained after a 3 h growth in LB taken as 100%. EDL933: EHEC O157:H7; CH014: non-O157 STEC; C600: K-12 E. coli; E2348/69: EPEC. All assays were performed independently at least three times. Results are means ± SEM of bacteria adhering to T84 cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4343011&req=5

Figure 1: Determination of experimental conditions for bacterial adherence of Escherichia coli strains to human colonic carcinoma T84 cells. Cells were infected with bacteria grown for 3 h (exponential growth phase) or 18 h (stationary growth phase), in Luria Bertani Broth (LB) or in Cell Culture Medium (CCM), at 37°C before infection. The cells were then washed, treated with Triton X-100, and the recovered adherent bacteria were diluted and plated for colony forming unit counting. Results are expressed as percentage of adherent bacteria, relative to adherent bacteria obtained after a 3 h growth in LB taken as 100%. EDL933: EHEC O157:H7; CH014: non-O157 STEC; C600: K-12 E. coli; E2348/69: EPEC. All assays were performed independently at least three times. Results are means ± SEM of bacteria adhering to T84 cells.
Mentions: Different bacterial growth phases (exponential growth phase vs. late-log phase), and types of bacterial culture media (LB vs. CCM) were tested in preliminary infection experiments on the T84 human colonic carcinoma cell line. These assays were performed with two virulent STEC strains (O157:H7 strain EDL933 and O91:H21 strain CH014), the EPEC reference strain (E2348/69) as a positive control, and the E. coli strain K-12 C600 as a negative control (Figure 1). The best adhesion levels were obtained with strains grown for 3 h in CCM. For further quantitative experiments, we thus chose to incubate the bacteria for 3 h at 37°C in CCM before infection. Quantitative experiments, performed on 17 STEC strains isolated from human cases, indicated that the number of bacteria adhering to the T84 cells was between 2.4 × 103 and 4.4 × 105 CFU/well (Figure 2A). The number reached 2 × 106 CFU/well for the EPEC reference strain, indicating that STEC adhered to very low levels compared to the EPEC strain, and just above the level of the negative control C600. Adherence patterns were investigated after staining with Giemsa and by SEM observations. The STEC strains adhered sparsely, whereas the EPEC formed micro-colonies (Figure 2B).

Bottom Line: We analyzed the ability of 256 STEC strains belonging to seropathotype A (the most virulent O157:H7 serotype) to seropathotype E (not involved in human disease) to adhere to HEp-2, HCT-8, and T84 cell lines.Of the 256 STEC tested most (82%) were non-adherent in our assays.The adhesion levels were globally low and were not related to pathogenicity, although the highest levels were associated to O26:H11 and O103:H2 strains of seropathotype B (associated with HUS but less commonly than serotype O157:H7), possessing both the eae and toxB genes.

View Article: PubMed Central - PubMed

Affiliation: Centre de Recherche en Nutrition Humaine Auvergne, M2iSH, 'Microbes, Intestin, Inflammation et Susceptibilité de l'Hôte' UMR INSERM/Université d'Auvergne U1071 USC-INRA 2018, Clermont Université - Université d'Auvergne Clermont-Ferrand, France.

ABSTRACT
Shiga toxin-producing Escherichia coli (STEC) are able to cause serious illnesses ranging from diarrhea to hemorrhagic colitis and hemolytic-uremic syndrome (HUS). These bacteria colonize the digestive tract of humans and produce Shiga-toxins, which are considered to be essential for virulence and are crucial in lethal infection. Colon colonization is supposed to be a determinant step in the development of the infection, but the virulence traits that mediate this step are unclear. We analyzed the ability of 256 STEC strains belonging to seropathotype A (the most virulent O157:H7 serotype) to seropathotype E (not involved in human disease) to adhere to HEp-2, HCT-8, and T84 cell lines. Of the 256 STEC tested most (82%) were non-adherent in our assays. The adhesion levels were globally low and were not related to pathogenicity, although the highest levels were associated to O26:H11 and O103:H2 strains of seropathotype B (associated with HUS but less commonly than serotype O157:H7), possessing both the eae and toxB genes.

No MeSH data available.


Related in: MedlinePlus