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Immunization with a Double-Mutant (R192G/L211A) of the Heat-Labile Enterotoxin of Escherichia coli Offers Partial Protection against Campylobacter jejuni in an Adult Mouse Intestinal Colonization Model.

Albert MJ, Haridas S, Ebenezer M, Raghupathy R, Khan I - PLoS ONE (2015)

Bottom Line: Unlike LT which is reactogenic, dmLT has been shown to be safe for human use.However, the differences were not statistically significant.As dmLT is safe for humans, it could be incorporated into a C. jejuni vaccine to enhance its efficacy.

View Article: PubMed Central - PubMed

Affiliation: Departments of Microbiology, Faculty of Medicine, Kuwait University, Jabriya, Kuwait.

ABSTRACT
We have previously shown that antibodies to cholera toxin (CT) reacted with the major outer membrane proteins (MOMPs) from Campylobacter jejuni strains on Western blot. Further, oral immunization with CT significantly protected against challenge with C. jejuni in an adult mouse colonization model of infection. CT and the heat-labile enterotoxin (LT) of enterotoxigenic Escherichia coli are structurally and functionally related. LT and its mutants including the double-mutant LT (R192G/L211A) (dmLT), are powerful mucosal adjuvants. Unlike LT which is reactogenic, dmLT has been shown to be safe for human use. In the current study, we determined whether rabbit anti-dmLT antibodies reacted with MOMPs from C. jejuni strains and whether immunization with dmLT would afford protection against C. jejuni. On Western blot, the MOMPs from C. jejuni 48 (Penner serotype O:19), C. jejuni 75 (O:3) and C. jejuni 111 (O:1,44) were probed with rabbit antibodies to dmLT or LT-E112K (a non-toxic LT mutant), which showed a lack of reaction. Adult BALB/c mice were orally immunized with dmLT and orally challenged with C. jejuni 48 or 111. Protection from colonization with the challenge bacteria was studied by enumerating Campylobacter colonies in feces daily for 9 days. Vaccination produced robust serum and stool antibody responses to dmLT and no antibody responses to C. jejuni MOMP. Vaccinated mice showed reduced colonization and excretion of both challenge strains compared to control mice. However, the differences were not statistically significant. The protective efficacy of the dmLT vaccine varied from 9.1% to 54.5%. The lack of cross-reaction between the MOMP and dmLT suggests that protection is not mediated by cross-reacting antibodies, but may be due to activation of innate immunity. As dmLT is safe for humans, it could be incorporated into a C. jejuni vaccine to enhance its efficacy.

No MeSH data available.


Related in: MedlinePlus

Western blots of enriched MOMPs from C. jejuni strains reacted with rabbit antiserum to the MOMP from C. jejuni strain 111 (A), with rabbit antibody to CT (Sigma) (B), and with rabbit antiserum to dmLT (C).SDS-12% PAGE-separated MOMPs were transferred to a nitrocellulose membrane and probed with the appropriate rabbit antiserum and then with anti-rabbit immunoglobulin. In panels A and B, lanes 1, 2 and 3 were loaded with MOMPs from C. jejuni 48, 75 and 111, respectively; lane 4 contains molecular weight markers. A prominent band of approximately 48 kDa corresponding to the MOMP is seen in all the three strains in panels A and B. The light band of approximately 74.1 kDa in panel B is a nonspecific band [9]. In panel C, lane 1 was loaded with dmLT, and 2 through 5, were the same as lanes 1 through 4 in panels A and B. There is a lack of reaction between the MOMPs and dmLT antibody in panel C. The different bands in lane 1 may represent different components of dmLT, namely B5, A, A1 and B1 (from highest to lowest molecular weight).
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pone.0142090.g001: Western blots of enriched MOMPs from C. jejuni strains reacted with rabbit antiserum to the MOMP from C. jejuni strain 111 (A), with rabbit antibody to CT (Sigma) (B), and with rabbit antiserum to dmLT (C).SDS-12% PAGE-separated MOMPs were transferred to a nitrocellulose membrane and probed with the appropriate rabbit antiserum and then with anti-rabbit immunoglobulin. In panels A and B, lanes 1, 2 and 3 were loaded with MOMPs from C. jejuni 48, 75 and 111, respectively; lane 4 contains molecular weight markers. A prominent band of approximately 48 kDa corresponding to the MOMP is seen in all the three strains in panels A and B. The light band of approximately 74.1 kDa in panel B is a nonspecific band [9]. In panel C, lane 1 was loaded with dmLT, and 2 through 5, were the same as lanes 1 through 4 in panels A and B. There is a lack of reaction between the MOMPs and dmLT antibody in panel C. The different bands in lane 1 may represent different components of dmLT, namely B5, A, A1 and B1 (from highest to lowest molecular weight).

Mentions: The homologous titer of rabbit antibodies to the MOMP of C. jejuni 111 in ELISA was 1 in 128,000. This antibody (at 1 in 10,000 dilution) reacted strongly on Western blot with all three strains (Fig 1A). This shows the sharing of MOMP antigens among the C. jejuni strains used for this study.


Immunization with a Double-Mutant (R192G/L211A) of the Heat-Labile Enterotoxin of Escherichia coli Offers Partial Protection against Campylobacter jejuni in an Adult Mouse Intestinal Colonization Model.

Albert MJ, Haridas S, Ebenezer M, Raghupathy R, Khan I - PLoS ONE (2015)

Western blots of enriched MOMPs from C. jejuni strains reacted with rabbit antiserum to the MOMP from C. jejuni strain 111 (A), with rabbit antibody to CT (Sigma) (B), and with rabbit antiserum to dmLT (C).SDS-12% PAGE-separated MOMPs were transferred to a nitrocellulose membrane and probed with the appropriate rabbit antiserum and then with anti-rabbit immunoglobulin. In panels A and B, lanes 1, 2 and 3 were loaded with MOMPs from C. jejuni 48, 75 and 111, respectively; lane 4 contains molecular weight markers. A prominent band of approximately 48 kDa corresponding to the MOMP is seen in all the three strains in panels A and B. The light band of approximately 74.1 kDa in panel B is a nonspecific band [9]. In panel C, lane 1 was loaded with dmLT, and 2 through 5, were the same as lanes 1 through 4 in panels A and B. There is a lack of reaction between the MOMPs and dmLT antibody in panel C. The different bands in lane 1 may represent different components of dmLT, namely B5, A, A1 and B1 (from highest to lowest molecular weight).
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Related In: Results  -  Collection

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pone.0142090.g001: Western blots of enriched MOMPs from C. jejuni strains reacted with rabbit antiserum to the MOMP from C. jejuni strain 111 (A), with rabbit antibody to CT (Sigma) (B), and with rabbit antiserum to dmLT (C).SDS-12% PAGE-separated MOMPs were transferred to a nitrocellulose membrane and probed with the appropriate rabbit antiserum and then with anti-rabbit immunoglobulin. In panels A and B, lanes 1, 2 and 3 were loaded with MOMPs from C. jejuni 48, 75 and 111, respectively; lane 4 contains molecular weight markers. A prominent band of approximately 48 kDa corresponding to the MOMP is seen in all the three strains in panels A and B. The light band of approximately 74.1 kDa in panel B is a nonspecific band [9]. In panel C, lane 1 was loaded with dmLT, and 2 through 5, were the same as lanes 1 through 4 in panels A and B. There is a lack of reaction between the MOMPs and dmLT antibody in panel C. The different bands in lane 1 may represent different components of dmLT, namely B5, A, A1 and B1 (from highest to lowest molecular weight).
Mentions: The homologous titer of rabbit antibodies to the MOMP of C. jejuni 111 in ELISA was 1 in 128,000. This antibody (at 1 in 10,000 dilution) reacted strongly on Western blot with all three strains (Fig 1A). This shows the sharing of MOMP antigens among the C. jejuni strains used for this study.

Bottom Line: Unlike LT which is reactogenic, dmLT has been shown to be safe for human use.However, the differences were not statistically significant.As dmLT is safe for humans, it could be incorporated into a C. jejuni vaccine to enhance its efficacy.

View Article: PubMed Central - PubMed

Affiliation: Departments of Microbiology, Faculty of Medicine, Kuwait University, Jabriya, Kuwait.

ABSTRACT
We have previously shown that antibodies to cholera toxin (CT) reacted with the major outer membrane proteins (MOMPs) from Campylobacter jejuni strains on Western blot. Further, oral immunization with CT significantly protected against challenge with C. jejuni in an adult mouse colonization model of infection. CT and the heat-labile enterotoxin (LT) of enterotoxigenic Escherichia coli are structurally and functionally related. LT and its mutants including the double-mutant LT (R192G/L211A) (dmLT), are powerful mucosal adjuvants. Unlike LT which is reactogenic, dmLT has been shown to be safe for human use. In the current study, we determined whether rabbit anti-dmLT antibodies reacted with MOMPs from C. jejuni strains and whether immunization with dmLT would afford protection against C. jejuni. On Western blot, the MOMPs from C. jejuni 48 (Penner serotype O:19), C. jejuni 75 (O:3) and C. jejuni 111 (O:1,44) were probed with rabbit antibodies to dmLT or LT-E112K (a non-toxic LT mutant), which showed a lack of reaction. Adult BALB/c mice were orally immunized with dmLT and orally challenged with C. jejuni 48 or 111. Protection from colonization with the challenge bacteria was studied by enumerating Campylobacter colonies in feces daily for 9 days. Vaccination produced robust serum and stool antibody responses to dmLT and no antibody responses to C. jejuni MOMP. Vaccinated mice showed reduced colonization and excretion of both challenge strains compared to control mice. However, the differences were not statistically significant. The protective efficacy of the dmLT vaccine varied from 9.1% to 54.5%. The lack of cross-reaction between the MOMP and dmLT suggests that protection is not mediated by cross-reacting antibodies, but may be due to activation of innate immunity. As dmLT is safe for humans, it could be incorporated into a C. jejuni vaccine to enhance its efficacy.

No MeSH data available.


Related in: MedlinePlus