Salmonella enterica Serovar Enteritidis Ghosts Carrying the Escherichia coli Heat-Labile Enterotoxin B Subunit Are Capable of Inducing Enhanced Protective Immune Responses

ABSTRACTTheEscherichia coliheat-labile enterotoxin B subunit (LTB) is a potent vaccine adjuvant.Salmonella entericaserovar Enteritidis ghosts carrying LTB (S. Enteritidis-LTB ghosts) were genetically constructed using a novel plasmid, pJHL187-LTB, designed for the coexpression of the LTB and E lysis proteins.S. Enteritidis-LTB ghosts were characterized using scanning electron microscopy to visualize their transmembrane tunnel structures. The expression of LTB inS. Enteritidis-LTB ghost preparations was confirmed by immunoblot and enzyme-linked immunosorbent assays. The parenteral adjuvant activity of LTB was demonstrated by immunizing chickens with eitherS. Enteritidis-LTB ghosts orS. Enteritidis ghosts. Chickens were intramuscularly primed at 5 weeks of age and subsequently boosted at 8 weeks of age. In total, 60 chickens were equally divided into three groups (n= 20 for each): group A, nonvaccinated control; group B, immunized withS. Enteritidis-LTB ghosts; and group C, immunized withS. Enteritidis ghosts. Compared with the nonimmunized chickens (group A), the immunized chickens (groups B and C) exhibited increased titers of plasma IgG and intestinal secretory IgA antibodies. The CD3+CD4+subpopulation of T cells was also significantly increased in both immunized groups. Among the immunized chickens, those in group B exhibited significantly increased titers of specific plasma IgG and intestinal secretory IgA (sIgA) antibodies compared with those in group C, indicating the immunomodulatory effects of the LTB adjuvant. Furthermore, both immunized groups exhibited decreased bacterial loads in their feces and internal organs. These results indicate that parenteral immunization withS. Enteritidis-LTB ghosts can stimulate superior induction of systemic and mucosal immune responses compared to immunization withS. Enteritidis ghosts alone, thus conferring efficient protection against salmonellosis.

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Adjuvant effect of Escherichia coli heat labile enterotoxin B subunit against internal egg contamination in domestic fowl immunised with a live Salmonella enterica serovar Enteritidis vaccine

The Veterinary Journal ◽

10.1016/j.tvjl.2013.03.024 ◽

2013 ◽

Vol 197 (3) ◽

pp. 861-867 ◽

Cited By ~ 7

Author(s):

Rahul M. Nandre ◽

Chetan V. Jawale ◽

John Hwa Lee

Keyword(s):

Escherichia Coli ◽

Salmonella Enterica ◽

Domestic Fowl ◽

Salmonella Enterica Serovar Enteritidis ◽

Adjuvant Effect ◽

B Subunit ◽

Heat Labile ◽

Enterotoxin B

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Comparison of the immune responses induced by oral immunization of mice with Lactobacillus casei-expressing porcine parvovirus VP2 and VP2 fused to Escherichia coli heat-labile enterotoxin B subunit protein

Comparative Immunology Microbiology and Infectious Diseases ◽

10.1016/j.cimid.2010.02.004 ◽

2011 ◽

Vol 34 (1) ◽

pp. 73-81 ◽

Cited By ~ 29

Author(s):

Diqiu Liu ◽

Xiangqing Wang ◽

Junwei Ge ◽

Songmei Liu ◽

Yijing Li

Keyword(s):

Escherichia Coli ◽

Immune Responses ◽

Lactobacillus Casei ◽

Oral Immunization ◽

Porcine Parvovirus ◽

Subunit Protein ◽

B Subunit ◽

Heat Labile ◽

Enterotoxin B

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The B subunits of cholera and Escherichia coli heat-labile toxins enhance the immune responses in mice orally immunised with a recombinant live P-fimbrial vaccine for avian pathogenic E. coli

Acta Veterinaria Hungarica ◽

10.1556/avet.2014.006 ◽

2014 ◽

Vol 62 (3) ◽

pp. 293-303

Author(s):

In-Gyeong Oh ◽

Chetan Jawale ◽

John Lee

Keyword(s):

Escherichia Coli ◽

Immune Responses ◽

Cholera Toxin B Subunit ◽

Adjuvant Effect ◽

Toxin B ◽

E Coli ◽

Cholera Toxin B ◽

B Subunit ◽

Heat Labile ◽

Enterotoxin B

This study aimed to investigate the adjuvant effect of recombinant attenuatedSalmonellaexpressing cholera toxin B subunit (CTB) andEscherichia coliheat-labile enterotoxin B subunit (LTB) for the P-fimbriae subunit-based vaccine of avian pathogenicE. coli(APEC) in a murine model. The PapA-specific sIgA and IgG responses were significantly enhanced after immunisation with theSalmonella-PapA vaccine in the presence of CTB or LTB. The group immunised with theSalmonella-LTB strain promoted Th1-type immunity, whereas that immunised with theSalmonella-CTB strain produced Th2-type immunity. We concluded that bothSalmonella-CTB and -LTB strains can enhance the immune response to PapA, and that the LTB strain may be a more effective adjuvant for APEC vaccination, which requires higher Th1-type immunity for protection. Thus, our findings provide evidence that immunisation with an adjuvant, LTB, is one of the strategies of developing effective vaccines against P-fimbriated APEC.

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Enhanced protective immune responses against Salmonella Enteritidis infection by Salmonella secreting an Escherichia coli heat-labile enterotoxin B subunit protein

Comparative Immunology Microbiology and Infectious Diseases ◽

10.1016/j.cimid.2013.06.002 ◽

2013 ◽

Vol 36 (5) ◽

pp. 537-548 ◽

Cited By ~ 15

Author(s):

Rahul M. Nandre ◽

Chetan V. Jawale ◽

John Hwa Lee

Keyword(s):

Escherichia Coli ◽

Immune Responses ◽

Salmonella Enteritidis ◽

Subunit Protein ◽

B Subunit ◽

Heat Labile ◽

Enterotoxin B

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The A Subunit of Escherichia coli Heat-Labile Enterotoxin Functions as a Mucosal Adjuvant and Promotes IgG2a, IgA, and Th17 Responses to Vaccine Antigens

Infection and Immunity ◽

10.1128/iai.00181-12 ◽

2012 ◽

Vol 80 (7) ◽

pp. 2426-2435 ◽

Cited By ~ 50

Author(s):

Elizabeth B. Norton ◽

Louise B. Lawson ◽

Zaid Mahdi ◽

Lucy C. Freytag ◽

John D. Clements

Keyword(s):

Escherichia Coli ◽

Immune Responses ◽

Fluid Secretion ◽

Proteolytic Degradation ◽

Mucosal Adjuvant ◽

Content Type ◽

B Subunit ◽

Heat Labile ◽

A Subunit ◽

Camp Induction

ABSTRACTEnterotoxigenicEscherichia coli(ETEC) produces both heat-labile (LT) and heat-stable (ST) enterotoxins and is a major cause of diarrhea in infants in developing countries and in travelers to those regions. In addition to inducing fluid secretion, LT is a powerful mucosal adjuvant capable of promoting immune responses to coadministered antigens. In this study, we examined purified A subunit to further understand the toxicity and adjuvanticity of LT. Purified A subunit was enzymatically active but sensitive to proteolytic degradation and unable to bind gangliosides, and even in the presence of admixed B subunit, it displayed low cyclic AMP (cAMP) induction and no enterotoxicity. Thus, the AB5 structure plays a key role in protecting the A subunit from proteolytic degradation and in delivering the enzymatic signals required for secretion. In contrast, the A subunit alone was capable of activating dendritic cells and enhanced immune responses to multiple antigens following intranasal immunization; therefore, unlike toxicity, LT adjuvanticity is not dependent on the AB5 holotoxin structure or the presence of the B subunit. However, immune responses were maximal when signals were received from both subunits either in an AB5 structure or with A and B admixed. Furthermore, the quality of the immune response (i.e., IgG1/IgG2 balance and mucosal IgA and IL-17 secretion) was determined by the presence of an A subunit, revealing for the first time induction of Th17 responses with the A subunit alone. These results have important implications for understanding ETEC pathogenesis, unraveling immunologic responses induced by LT-based adjuvants, and developing new mucosal vaccines.

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Functional Pentameric Formation via Coexpression of the Escherichia coli Heat-Labile Enterotoxin B Subunit and Its Fusion Protein Subunit with a Neutralizing Epitope of ApxIIA Exotoxin Improves the Mucosal Immunogenicity and Protection against Challenge by Actinobacillus pleuropneumoniae

Clinical and Vaccine Immunology ◽

10.1128/cvi.05230-11 ◽

2011 ◽

Vol 18 (12) ◽

pp. 2168-2177 ◽

Cited By ~ 7

Author(s):

Jung-Mi Kim ◽

Seung-Moon Park ◽

Jung-Ae Kim ◽

Jin-Ah Park ◽

Min-Hee Yi ◽

...

Keyword(s):

Escherichia Coli ◽

Fusion Protein ◽

Actinobacillus Pleuropneumoniae ◽

Enzyme Linked Immunosorbent Assay ◽

Recombinant Yeast ◽

Content Type ◽

Fusion Construct ◽

B Subunit ◽

Heat Labile ◽

Enterotoxin B

ABSTRACTA coexpression strategy inSaccharomyces cerevisiaeusing episomal and integrative vectors for theEscherichia coliheat-labile enterotoxin B subunit (LTB) and a fusion protein of an ApxIIA toxin epitope produced byActinobacillus pleuropneumoniaecoupled to LTB, respectively, was adapted for the hetero-oligomerization of LTB and the LTB fusion construct. Enzyme-linked immunosorbent assay (ELISA) with GM1 ganglioside indicated that the LTB fusion construct, along with LTB, was oligomerized to make the functional heteropentameric form, which can bind to receptors on the mucosal epithelium. The antigen-specific antibody titer of mice orally administered antigen was increased when using recombinant yeast coexpressing the pentameric form instead of recombinant yeast expressing either the LTB fusion form or antigen alone. Better protection against challenge infection withA. pleuropneumoniaewas also observed for coexpression in recombinant yeast compared with others. The present study clearly indicated that the coexpression strategy enabled the LTB fusion construct to participate in the pentameric formation, resulting in an improved induction of systemic and mucosal immune responses.

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Attenuated Salmonella Gallinarum secreting an Escherichia coli heat-labile enterotoxin B subunit protein as an adjuvant for oral vaccination against fowl typhoid

Veterinary Immunology and Immunopathology ◽

10.1016/j.vetimm.2012.09.006 ◽

2012 ◽

Vol 150 (3-4) ◽

pp. 149-160 ◽

Cited By ~ 9

Author(s):

Byung Woo Jeon ◽

Chetan V. Jawale ◽

Seung Hwan Kim ◽

John Hwa Lee

Keyword(s):

Escherichia Coli ◽

Oral Vaccination ◽

Salmonella Gallinarum ◽

Subunit Protein ◽

B Subunit ◽

Heat Labile ◽

Fowl Typhoid ◽

Enterotoxin B

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Tissue culture and expression of Escherichia coli heat-labile enterotoxin B subunit in transgenic Peperomia pellucida

Protein Expression and Purification ◽

10.1016/j.pep.2010.02.010 ◽

2010 ◽

Vol 72 (1) ◽

pp. 82-86 ◽

Cited By ~ 11

Author(s):

Nguyen Hoang Loc ◽

Nguyen Hoang Bach ◽

Tae-Geum Kim ◽

Moon-Sik Yang

Keyword(s):

Escherichia Coli ◽

Tissue Culture ◽

B Subunit ◽

Heat Labile ◽

Enterotoxin B

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Escherichia coli Heat-Labile Enterotoxin B Subunit Is a More Potent Mucosal Adjuvant than Its Closely Related Homologue, the B Subunit of Cholera Toxin

Infection and Immunity ◽

10.1128/iai.69.5.3476-3482.2001 ◽

2001 ◽

Vol 69 (5) ◽

pp. 3476-3482 ◽

Cited By ~ 57

Author(s):

Douglas G. Millar ◽

Timothy R. Hirst ◽

Denis P. Snider

Keyword(s):

Escherichia Coli ◽

Cholera Toxin ◽

Lymphocyte Proliferation ◽

Vaccine Adjuvants ◽

B Subunit ◽

Heat Labile ◽

Antibody Titers ◽

Enterotoxin B ◽

Draining Lymph Nodes ◽

Stimulation Of

ABSTRACT Although cholera toxin (Ctx) and Escherichia coliheat-labile enterotoxin (Etx) are known to be potent mucosal adjuvants, it remains controversial whether the adjuvanticity of the holotoxins extends to their nontoxic, receptor-binding B subunits. Here, we have systematically evaluated the comparative adjuvant properties of highly purified recombinant EtxB and CtxB. EtxB was found to be a more potent adjuvant than CtxB, stimulating responses to hen egg lysozyme when the two were coadministered to mice intranasally, as assessed by enhanced serum and secretory antibody titers as well as by stimulation of lymphocyte proliferation in spleen and draining lymph nodes. These results indicate that, although structurally very similar, EtxB and CtxB have strikingly different immunostimulatory properties and should not be considered equivalent as prospective vaccine adjuvants.

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