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Protective Enterotoxigenic Escherichia coli Antigens in a Murine Intranasal Challenge Model.

Kumar A, Hays M, Lim F, Foster LJ, Zhou M, Zhu G, Miesner T, Hardwidge PR - PLoS Negl Trop Dis (2015)

Bottom Line: Immunization with either Skp or MipA provided an intermediate degree of protection, 68 and 64%, respectively.Protection was significantly correlated with the induction of a secretory immunoglobulin A response.This study has identified several proteins that are conserved among heterologous ETEC strains and may thus potentially improve cross-protective efficacy if incorporated into future vaccine designs.

View Article: PubMed Central - PubMed

Affiliation: College of Veterinary Medicine, Kansas State University, Manhattan, Kansas, United States of America.

ABSTRACT
Enterotoxigenic Escherichia coli (ETEC) is an endemic health threat in underdeveloped nations. Despite the significant effort extended to vaccine trials using ETEC colonization factors, these approaches have generally not been especially effective in mediating cross-protective immunity. We used quantitative proteomics to identify 24 proteins that differed in abundance in membrane protein preparations derived from wild-type vs. a type II secretion system mutant of ETEC. We expressed and purified a subset of these proteins and identified nine antigens that generated significant immune responses in mice. Sera from mice immunized with either the MltA-interacting protein MipA, the periplasmic chaperone seventeen kilodalton protein, Skp, or a long-chain fatty acid outer membrane transporter, ETEC_2479, reduced the adherence of multiple ETEC strains differing in colonization factor expression to human intestinal epithelial cells. In intranasal challenge assays of mice, immunization with ETEC_2479 protected 88% of mice from an otherwise lethal challenge with ETEC H10407. Immunization with either Skp or MipA provided an intermediate degree of protection, 68 and 64%, respectively. Protection was significantly correlated with the induction of a secretory immunoglobulin A response. This study has identified several proteins that are conserved among heterologous ETEC strains and may thus potentially improve cross-protective efficacy if incorporated into future vaccine designs.

No MeSH data available.


Related in: MedlinePlus

Adherence assays.A. Adherence of ETEC H10407 to HCT-8 cells in the presence of antisera. The fold-change in ETEC H10407 adherence is plotted as a function of incubation with the indicated antisera, after normalization to control samples. Asterisks indicate the antisera showing significant protection against ETEC H10407 that were selected for further characterization, n = 3/group. B. Analysis of antisera specificity. Deletion mutants were constructed for the skp, mipA, and ETEC_2479 genes and then analyzed for their adherence to HCT-8 cells in the presence of the indicated antisera. Asterisks indicate significant reduction in bacterial adherence, n = 4-12/group. C. Heat map displaying the fold-change in adherence of the indicated ETEC strains to HCT-8 cells after their incubation with the indicated antisera. The scale ranges from 0.5 to a 10-fold decrease in adherence, as compared with control samples, n = 3/group. Red colors depict reduced ETEC adherence in the presence of antisera, while green colors depict increased adherence. The midpoint of the color gradient was set as a 2-fold reduction in adherence (black).
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pntd.0003924.g002: Adherence assays.A. Adherence of ETEC H10407 to HCT-8 cells in the presence of antisera. The fold-change in ETEC H10407 adherence is plotted as a function of incubation with the indicated antisera, after normalization to control samples. Asterisks indicate the antisera showing significant protection against ETEC H10407 that were selected for further characterization, n = 3/group. B. Analysis of antisera specificity. Deletion mutants were constructed for the skp, mipA, and ETEC_2479 genes and then analyzed for their adherence to HCT-8 cells in the presence of the indicated antisera. Asterisks indicate significant reduction in bacterial adherence, n = 4-12/group. C. Heat map displaying the fold-change in adherence of the indicated ETEC strains to HCT-8 cells after their incubation with the indicated antisera. The scale ranges from 0.5 to a 10-fold decrease in adherence, as compared with control samples, n = 3/group. Red colors depict reduced ETEC adherence in the presence of antisera, while green colors depict increased adherence. The midpoint of the color gradient was set as a 2-fold reduction in adherence (black).

Mentions: We quantified the extent to which the mouse antisera would subsequently protect against ETEC H10407 adherence to HCT-8 cells. Of the 9 antigens tested, 3 antigens (Skp, MipA, and ETEC_2479) showed a significant ability to inhibit H10407 adherence (Fig 2A). To determine the specificity of these antisera for their respective antigens, we deleted individually the genes encoding Skp, MipA, and ETEC_2479 and then re-evaluated the ability of the respective antisera to protect against the adherence of these ETEC mutants to HCT-8 cells. The antisera inhibited the WT strain and the 2 heterologous ETEC mutants, but no longer inhibited the adherence of the corresponding homologous ETEC mutant to as great an extent, suggesting that the phenotypes observed are largely attributable to antisera specificity (Fig 2B).


Protective Enterotoxigenic Escherichia coli Antigens in a Murine Intranasal Challenge Model.

Kumar A, Hays M, Lim F, Foster LJ, Zhou M, Zhu G, Miesner T, Hardwidge PR - PLoS Negl Trop Dis (2015)

Adherence assays.A. Adherence of ETEC H10407 to HCT-8 cells in the presence of antisera. The fold-change in ETEC H10407 adherence is plotted as a function of incubation with the indicated antisera, after normalization to control samples. Asterisks indicate the antisera showing significant protection against ETEC H10407 that were selected for further characterization, n = 3/group. B. Analysis of antisera specificity. Deletion mutants were constructed for the skp, mipA, and ETEC_2479 genes and then analyzed for their adherence to HCT-8 cells in the presence of the indicated antisera. Asterisks indicate significant reduction in bacterial adherence, n = 4-12/group. C. Heat map displaying the fold-change in adherence of the indicated ETEC strains to HCT-8 cells after their incubation with the indicated antisera. The scale ranges from 0.5 to a 10-fold decrease in adherence, as compared with control samples, n = 3/group. Red colors depict reduced ETEC adherence in the presence of antisera, while green colors depict increased adherence. The midpoint of the color gradient was set as a 2-fold reduction in adherence (black).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4526226&req=5

pntd.0003924.g002: Adherence assays.A. Adherence of ETEC H10407 to HCT-8 cells in the presence of antisera. The fold-change in ETEC H10407 adherence is plotted as a function of incubation with the indicated antisera, after normalization to control samples. Asterisks indicate the antisera showing significant protection against ETEC H10407 that were selected for further characterization, n = 3/group. B. Analysis of antisera specificity. Deletion mutants were constructed for the skp, mipA, and ETEC_2479 genes and then analyzed for their adherence to HCT-8 cells in the presence of the indicated antisera. Asterisks indicate significant reduction in bacterial adherence, n = 4-12/group. C. Heat map displaying the fold-change in adherence of the indicated ETEC strains to HCT-8 cells after their incubation with the indicated antisera. The scale ranges from 0.5 to a 10-fold decrease in adherence, as compared with control samples, n = 3/group. Red colors depict reduced ETEC adherence in the presence of antisera, while green colors depict increased adherence. The midpoint of the color gradient was set as a 2-fold reduction in adherence (black).
Mentions: We quantified the extent to which the mouse antisera would subsequently protect against ETEC H10407 adherence to HCT-8 cells. Of the 9 antigens tested, 3 antigens (Skp, MipA, and ETEC_2479) showed a significant ability to inhibit H10407 adherence (Fig 2A). To determine the specificity of these antisera for their respective antigens, we deleted individually the genes encoding Skp, MipA, and ETEC_2479 and then re-evaluated the ability of the respective antisera to protect against the adherence of these ETEC mutants to HCT-8 cells. The antisera inhibited the WT strain and the 2 heterologous ETEC mutants, but no longer inhibited the adherence of the corresponding homologous ETEC mutant to as great an extent, suggesting that the phenotypes observed are largely attributable to antisera specificity (Fig 2B).

Bottom Line: Immunization with either Skp or MipA provided an intermediate degree of protection, 68 and 64%, respectively.Protection was significantly correlated with the induction of a secretory immunoglobulin A response.This study has identified several proteins that are conserved among heterologous ETEC strains and may thus potentially improve cross-protective efficacy if incorporated into future vaccine designs.

View Article: PubMed Central - PubMed

Affiliation: College of Veterinary Medicine, Kansas State University, Manhattan, Kansas, United States of America.

ABSTRACT
Enterotoxigenic Escherichia coli (ETEC) is an endemic health threat in underdeveloped nations. Despite the significant effort extended to vaccine trials using ETEC colonization factors, these approaches have generally not been especially effective in mediating cross-protective immunity. We used quantitative proteomics to identify 24 proteins that differed in abundance in membrane protein preparations derived from wild-type vs. a type II secretion system mutant of ETEC. We expressed and purified a subset of these proteins and identified nine antigens that generated significant immune responses in mice. Sera from mice immunized with either the MltA-interacting protein MipA, the periplasmic chaperone seventeen kilodalton protein, Skp, or a long-chain fatty acid outer membrane transporter, ETEC_2479, reduced the adherence of multiple ETEC strains differing in colonization factor expression to human intestinal epithelial cells. In intranasal challenge assays of mice, immunization with ETEC_2479 protected 88% of mice from an otherwise lethal challenge with ETEC H10407. Immunization with either Skp or MipA provided an intermediate degree of protection, 68 and 64%, respectively. Protection was significantly correlated with the induction of a secretory immunoglobulin A response. This study has identified several proteins that are conserved among heterologous ETEC strains and may thus potentially improve cross-protective efficacy if incorporated into future vaccine designs.

No MeSH data available.


Related in: MedlinePlus