title: Evaluation of recombinant leukocidin domain of VvhA exotoxin of Vibrio vulnificus as an effective toxoid in mouse model.
abstract: Vibrio vulnificus hemolysin A (VvhA) is a pore forming toxin and plays an important role in the pathogenesis. The hemolytic and cytotytic property of VvhA toxin is associated with N-terminal leukocidin domain which triggers apoptotic signaling cascade in epithelial cells. The present study was undertaken to assess the protective efficacy of recombinant VvhA leukocidin domain (rL/VvhA) against VvhA toxin challenge using in vitro and in vivo assays. The rL/VvhA protein was found to be non-toxic with no significant hemolytic or cytotoxic effects. Intraperitoneal (I.P.) immunization of BALB/c mice with rL/VvhA protein elicited significantly higher specific serum antibody titer with mixed Th1/Th2 mediated immune responses. HeLa cell monolayer supplemented with anti-rL/VvhA antibodies were effectively protected (viability 86.69%) against lethal 5 LD50 toxin challenge. An effective in vitro proliferation of lymphocyte was observed upon re-stimulation of rL/VvhA primed splenocytes with formalin inactivated VvhA toxin (fVvhA). Co-expression of Th1/Th2 polarized cytokines (IFN-γ, IL-12 and IL-4), were seen in the cell culture supernatant. In contrast to sham immunized mice, rL/VvhA immunized mice demonstrated significant protection (90% survival) against native toxin challenge in vitro and in vivo infection models. These results suggested leukocidin domain of the VvhA toxin as protective immunogen for possible protection against V. vulnificus VvhA.
authors.affiliation: Department of Microbiology, Defence Food Research Laboratory, Mysore 570011, India.
authors.forename: Guttinakere Krishnappa
authors.initials: GK
authors.lastname: Lohith
authors.affiliation: Department of Microbiology, Defence Food Research Laboratory, Mysore 570011, India.
authors.forename: Joseph Jeyabalaji
authors.initials: JJ
authors.lastname: Kingston
authors.affiliation: Department of Microbiology, Defence Food Research Laboratory, Mysore 570011, India.
authors.forename: Amit Kumar
authors.initials: AK
authors.lastname: Singh
authors.affiliation: Department of Microbiology, Defence Food Research Laboratory, Mysore 570011, India.
authors.forename: Harishchandra Sripathy
authors.initials: HS
authors.lastname: Murali
authors.affiliation: Department of Microbiology, Defence Food Research Laboratory, Mysore 570011, India. Electronic address: drharshvardhanbatra@gmail.com.
authors.forename: Harsh Vardhan
authors.initials: HV
authors.lastname: Batra
mesh_terms: D000818:Animals; D000907:Antibodies, Bacterial / Q000276:immunology; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D003001:Cloning, Molecular; D016207:Cytokines / Q000378:metabolism; D005260:Female; D015870:Gene Expression; D007114:Immunization; D007956:Leukocidins / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D008213:Lymphocyte Activation / Q000276:immunology; D008214:Lymphocytes / Q000276:immunology / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D009500:Neutralization Tests; D054730:Protein Interaction Domains and Motifs / Q000235:genetics / Q000276:immunology*; D011994:Recombinant Proteins; D013154:Spleen / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D014121:Toxoids / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D014735:Vibrio Infections / Q000276:immunology / Q000401:mortality / Q000517:prevention & control; D041261:Vibrio vulnificus / Q000235:genetics / Q000276:immunology*
chemical_list: D000907:Antibodies, Bacterial; D001426:Bacterial Proteins; D016207:Cytokines; D007956:Leukocidins; D011994:Recombinant Proteins; D014121:Toxoids; C470630:VvhA protein, Vibrio vulnificus
keywords: Leukocidin domain; Neutralizing antibodies; Recombinant protein; Vibrio vulnificus; VvhA toxin