Immunological characterization of a gidA mutant strain of Salmonella for potential use in a live-attenuated vaccine

BMC Microbiology, Nov 2012

Background Salmonella is often associated with gastrointestinal disease outbreaks in humans throughout the world due to the consumption of contaminated food. Our previous studies have shown that deletion of glucose-inhibited division gene (gidA) significantly attenuated Salmonella enterica serovar Typhimurium (STM) virulence in both in vitro and in vivo models of infection. Most importantly, immunization with the gidA mutant protected mice from a lethal dose challenge of wild-type STM. In this study, we further characterize the gidA mutant STM strain for potential use in a live-attenuated vaccine. Results The protective efficacy of immunization with the gidA mutant was evaluated by challenging immunized mice with a lethal dose of wild-type STM. Sera levels of IgG2a and IgG1, passive transfer of sera and cells, and cytokine profiling were performed to study the induction of humoral and cellular immune responses induced by immunization with the gidA mutant strain. Additionally, a lymphocyte proliferation assay was performed to gauge the splenocyte survival in response to treatment with STM cell lysate. Mice immunized with the gidA mutant strain were fully protected from a lethal dose challenge of wild-type STM. Naïve mice receiving either cells or sera from immunized mice were partially protected from a lethal dose challenge of wild-type STM. The lymphocyte proliferation assay displayed a significant response of splenocytes from immunized mice when compared to splenocytes from non-immunized control mice. Furthermore, the immunized mice displayed significantly higher levels of IgG1 and IgG2a with a marked increase in IgG1. Additionally, immunization with the gidA mutant strain evoked higher levels of IL-2, IFN-γ, and IL-10 cytokines in splenocytes induced with STM cell lysate. Conclusions Together, the results demonstrate that immunization with the gidA mutant strain protects mice by inducing humoral and cellular immune responses with the humoral immune response potentially being the main mechanism of protection.

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Immunological characterization of a gidA mutant strain of Salmonella for potential use in a live-attenuated vaccine

Daniel C Shippy 0 Amin A Fadl 0 0 Department of Animal Sciences, University of Wisconsin-Madison , 1675 Observatory Dr, Madison, WI 53706 , USA Background: Salmonella is often associated with gastrointestinal disease outbreaks in humans throughout the world due to the consumption of contaminated food. Our previous studies have shown that deletion of glucose-inhibited division gene (gidA) significantly attenuated Salmonella enterica serovar Typhimurium (STM) virulence in both in vitro and in vivo models of infection. Most importantly, immunization with the gidA mutant protected mice from a lethal dose challenge of wild-type STM. In this study, we further characterize the gidA mutant STM strain for potential use in a live-attenuated vaccine. Results: The protective efficacy of immunization with the gidA mutant was evaluated by challenging immunized mice with a lethal dose of wild-type STM. Sera levels of IgG2a and IgG1, passive transfer of sera and cells, and cytokine profiling were performed to study the induction of humoral and cellular immune responses induced by immunization with the gidA mutant strain. Additionally, a lymphocyte proliferation assay was performed to gauge the splenocyte survival in response to treatment with STM cell lysate. Mice immunized with the gidA mutant strain were fully protected from a lethal dose challenge of wild-type STM. Nave mice receiving either cells or sera from immunized mice were partially protected from a lethal dose challenge of wild-type STM. The lymphocyte proliferation assay displayed a significant response of splenocytes from immunized mice when compared to splenocytes from non-immunized control mice. Furthermore, the immunized mice displayed significantly higher levels of IgG1 and IgG2a with a marked increase in IgG1. Additionally, immunization with the gidA mutant strain evoked higher levels of IL-2, IFN-, and IL-10 cytokines in splenocytes induced with STM cell lysate. Conclusions: Together, the results demonstrate that immunization with the gidA mutant strain protects mice by inducing humoral and cellular immune responses with the humoral immune response potentially being the main mechanism of protection. - Background Salmonella is an enteric pathogen causing major public health problems throughout the world due to the consumption of contaminated food. Nontyphoidal Salmonella species, like Salmonella enterica serovar Typhimurium (STM), are the leading cause of hospitalization and death among the major foodborne pathogens [1]. Antibiotic resistance by Salmonella is dramatically increasing, so the development of an effective vaccine remains a global health priority [2,3]. Creating a safe and immunogenic vaccine strain is the biggest challenge in developing an effective liveattenuated Salmonella vaccine [4]. Several Salmonella vaccines, including whole-cell killed and live vaccines, have been developed with variable success [5,6]. These vaccines either required repeated administration or induced insufficient immune responses for long-lasting protection against lethal challenges with virulence Salmonella strains [7]. Many Salmonella vaccine strains carry deletion mutations affecting metabolic functions or virulence factors [8]. Several mutant strains of Salmonella have been investigated in the pursuit to develop optimal immune responses [9-11]. Our approach in constructing a live-attenuated Salmonella vaccine strain is to create a mutant defective in tRNA modification [12]. This strategy enables our vaccine strain to express multiple virulence factors at a significantly reduced level in order to obtain a safe and immunogenic vaccine candidate. Glucose-inhibited division (GidA) protein (also known as MnmG) was first described in Escherichia coli, where deletion of gidA resulted in a filamentous morphology when grown in a rich medium supplemented with glucose [13]. Further studies showed GidA is a flavin dinucleotide (FAD) binding enzyme involved in the fruiting body development of Myxococcus xanthus [14]. Furthermore, GidA has been shown to be a tRNA modification methylase in E. coli that forms a heterodimeric complex with MnmE (also known as TrmE) to catalyze the addition of a carboxymethylaminomethyl (cmnm) group at the 5 position of the wobble uridine (U34) of tRNAs [15-19]. Most importantly, deletion of gidA has been shown to attenuate the pathogenesis of some bacteria including Pseudomonas syringae, Aeromonas hydrophila, Streptococcus pyogenes, and Pseudomonas aeruginosa [20-23]. Our previous studies suggest a role for GidA in the regulation of Salmonella virulence and cell division [12,24]. In our initial study, the gidA mutant was attenuated in vitro and showed a significant decrease in ability to invade T84 intestinal epithelial cells as well as a significant decrease in ability to replicate and produce cytotoxic affects on macrophages. Furthermore, global transcriptional and proteomic profiling indicated a significant down-regulation in numerous ge (...truncated)


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Daniel C Shippy, Amin A Fadl. Immunological characterization of a gidA mutant strain of Salmonella for potential use in a live-attenuated vaccine, BMC Microbiology, 2012, pp. 286, 12, DOI: 10.1186/1471-2180-12-286