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Immunization with an autotransporter protein of Orientia tsutsugamushi provides protective immunity against scrub typhus.

Ha NY, Sharma P, Kim G, Kim Y, Min CK, Choi MS, Kim IS, Cho NH - PLoS Negl Trop Dis (2015)

Bottom Line: Despite the wide range of preventative approaches that have been attempted in the past 70 years, all have failed to develop an effective prophylactic vaccine.Currently, the selection of the proper antigens is one of the critical barriers to generating cross-protective immunity against antigenically-variable strains of O. tsutsugamushi.Our findings demonstrate that ScaA functions as a bacterial adhesion factor, and anti-ScaA antibody significantly neutralizes bacterial infection of host cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Seoul National University College of Medicine, Seoul, Republic of Korea; Department of Biomedical Sciences, Seoul National University College of Medicine, Seoul, Republic of Korea.

ABSTRACT

Background: Scrub typhus is an acute febrile disease caused by Orientia tsutsugamushi infection. Recently, the rapid increase of scrub typhus incidence in several countries within the endemic region has become a serious public health issue. Despite the wide range of preventative approaches that have been attempted in the past 70 years, all have failed to develop an effective prophylactic vaccine. Currently, the selection of the proper antigens is one of the critical barriers to generating cross-protective immunity against antigenically-variable strains of O. tsutsugamushi.

Methodology/principal findings: We examined the potential role of ScaA protein, an autotransporter protein of O. tsutsugamushi, in bacterial pathogenesis and evaluated the protective attributes of ScaA immunization in lethal O. tsutsugamushi infection in mice. Our findings demonstrate that ScaA functions as a bacterial adhesion factor, and anti-ScaA antibody significantly neutralizes bacterial infection of host cells. In addition, immunization with ScaA not only provides protective immunity against lethal challenges with the homologous strain, but also confers significant protection against heterologous strains when combined with TSA56, a major outer membrane protein of O. tsutsugamushi.

Conclusions/significance: Immunization of ScaA proteins provides protective immunity in mice when challenged with the homologous strain and significantly enhanced protective immunity against infection with heterologous strains. To our knowledge, this is the most promising result of scrub typhus vaccination trials against infection of heterologous strains in mouse models thus far.

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Related in: MedlinePlus

Expression of ScaA by O. tsutsugamushi.(A) Immunoblot analysis of whole proteins from L929 cells infected with O. tsutsugamushi proteins by using anti-ScaA serum (right panel). Anti-ScaA serum detected a protein with a molecular mass of approximately 150 kDa. Immunoblotting using anti-TAS56 was performed as a control (left panel). (B) Immunofluorescence confocal microscopy using preimmune serum or anti-ScaA serum (α-ScaA) showed ScaA in the O. tsutsugamushi-infected L929 cells. The left panels show bacteria stained with the pooled sera of scrub typhus patients (α-OT). Magnified images are shown in the lower panels (inset boxes). Scale bars, 5 μm.
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pntd.0003585.g001: Expression of ScaA by O. tsutsugamushi.(A) Immunoblot analysis of whole proteins from L929 cells infected with O. tsutsugamushi proteins by using anti-ScaA serum (right panel). Anti-ScaA serum detected a protein with a molecular mass of approximately 150 kDa. Immunoblotting using anti-TAS56 was performed as a control (left panel). (B) Immunofluorescence confocal microscopy using preimmune serum or anti-ScaA serum (α-ScaA) showed ScaA in the O. tsutsugamushi-infected L929 cells. The left panels show bacteria stained with the pooled sera of scrub typhus patients (α-OT). Magnified images are shown in the lower panels (inset boxes). Scale bars, 5 μm.

Mentions: In order to examine whether the ScaA is expressed in O. tsutsugamushi, we generated a polyclonal anti-ScaA antiserum by immunizing mice with purified ScaA passenger domain (amino acids 30 to 1000). The specificity of this antiserum was confirmed by ELISA and immunoblot analysis using the recombinant Sca proteins (S1 Fig). To identify endogenous ScaA protein in O. tsutsugamushi, the anti-ScaA serum was reacted with the cell lysates of O. tsutsugamushi-infected cells. Immunoblot analysis showed that a ~ 150 kDa protein was recognized by the anti-ScaA serum in infected cells but not in uninfected control (Fig. 1A). The full-length ScaA protein was predicted to have a mass of 156 kDa. Anti-ScaA serum was also weakly reacted with a few bands lower than ~ 150 kDa, suggesting a cross-reactive antigens or fragmented ScaA protein in infected cells. The TSA56 protein, a major outer membrane protein of O. tsutsugamushi, was used as a positive control [32]. To further confirm the specificity of the anti-ScaA antiserum for O. tsutsugamushi, intracellular bacteria were stained using the pooled sera of scrub typhus patients together with anti-ScaA serum or preimmune mouse serum. As shown in Fig. 1B, anti-ScaA serum readily detected the bacteria within the host cells, whereas the preimmune serum did not. In addition, we found that the ScaA proteins were located on the periphery of bacterial cells (Fig. 1B. lower panels, inset boxes). Taken together, these results confirm that the scaA gene is actively translated in O. tsutsugamushi within eukaryotic host cells and that the protein might be expressed on the outer membrane of the bacteria.


Immunization with an autotransporter protein of Orientia tsutsugamushi provides protective immunity against scrub typhus.

Ha NY, Sharma P, Kim G, Kim Y, Min CK, Choi MS, Kim IS, Cho NH - PLoS Negl Trop Dis (2015)

Expression of ScaA by O. tsutsugamushi.(A) Immunoblot analysis of whole proteins from L929 cells infected with O. tsutsugamushi proteins by using anti-ScaA serum (right panel). Anti-ScaA serum detected a protein with a molecular mass of approximately 150 kDa. Immunoblotting using anti-TAS56 was performed as a control (left panel). (B) Immunofluorescence confocal microscopy using preimmune serum or anti-ScaA serum (α-ScaA) showed ScaA in the O. tsutsugamushi-infected L929 cells. The left panels show bacteria stained with the pooled sera of scrub typhus patients (α-OT). Magnified images are shown in the lower panels (inset boxes). Scale bars, 5 μm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4359152&req=5

pntd.0003585.g001: Expression of ScaA by O. tsutsugamushi.(A) Immunoblot analysis of whole proteins from L929 cells infected with O. tsutsugamushi proteins by using anti-ScaA serum (right panel). Anti-ScaA serum detected a protein with a molecular mass of approximately 150 kDa. Immunoblotting using anti-TAS56 was performed as a control (left panel). (B) Immunofluorescence confocal microscopy using preimmune serum or anti-ScaA serum (α-ScaA) showed ScaA in the O. tsutsugamushi-infected L929 cells. The left panels show bacteria stained with the pooled sera of scrub typhus patients (α-OT). Magnified images are shown in the lower panels (inset boxes). Scale bars, 5 μm.
Mentions: In order to examine whether the ScaA is expressed in O. tsutsugamushi, we generated a polyclonal anti-ScaA antiserum by immunizing mice with purified ScaA passenger domain (amino acids 30 to 1000). The specificity of this antiserum was confirmed by ELISA and immunoblot analysis using the recombinant Sca proteins (S1 Fig). To identify endogenous ScaA protein in O. tsutsugamushi, the anti-ScaA serum was reacted with the cell lysates of O. tsutsugamushi-infected cells. Immunoblot analysis showed that a ~ 150 kDa protein was recognized by the anti-ScaA serum in infected cells but not in uninfected control (Fig. 1A). The full-length ScaA protein was predicted to have a mass of 156 kDa. Anti-ScaA serum was also weakly reacted with a few bands lower than ~ 150 kDa, suggesting a cross-reactive antigens or fragmented ScaA protein in infected cells. The TSA56 protein, a major outer membrane protein of O. tsutsugamushi, was used as a positive control [32]. To further confirm the specificity of the anti-ScaA antiserum for O. tsutsugamushi, intracellular bacteria were stained using the pooled sera of scrub typhus patients together with anti-ScaA serum or preimmune mouse serum. As shown in Fig. 1B, anti-ScaA serum readily detected the bacteria within the host cells, whereas the preimmune serum did not. In addition, we found that the ScaA proteins were located on the periphery of bacterial cells (Fig. 1B. lower panels, inset boxes). Taken together, these results confirm that the scaA gene is actively translated in O. tsutsugamushi within eukaryotic host cells and that the protein might be expressed on the outer membrane of the bacteria.

Bottom Line: Despite the wide range of preventative approaches that have been attempted in the past 70 years, all have failed to develop an effective prophylactic vaccine.Currently, the selection of the proper antigens is one of the critical barriers to generating cross-protective immunity against antigenically-variable strains of O. tsutsugamushi.Our findings demonstrate that ScaA functions as a bacterial adhesion factor, and anti-ScaA antibody significantly neutralizes bacterial infection of host cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Seoul National University College of Medicine, Seoul, Republic of Korea; Department of Biomedical Sciences, Seoul National University College of Medicine, Seoul, Republic of Korea.

ABSTRACT

Background: Scrub typhus is an acute febrile disease caused by Orientia tsutsugamushi infection. Recently, the rapid increase of scrub typhus incidence in several countries within the endemic region has become a serious public health issue. Despite the wide range of preventative approaches that have been attempted in the past 70 years, all have failed to develop an effective prophylactic vaccine. Currently, the selection of the proper antigens is one of the critical barriers to generating cross-protective immunity against antigenically-variable strains of O. tsutsugamushi.

Methodology/principal findings: We examined the potential role of ScaA protein, an autotransporter protein of O. tsutsugamushi, in bacterial pathogenesis and evaluated the protective attributes of ScaA immunization in lethal O. tsutsugamushi infection in mice. Our findings demonstrate that ScaA functions as a bacterial adhesion factor, and anti-ScaA antibody significantly neutralizes bacterial infection of host cells. In addition, immunization with ScaA not only provides protective immunity against lethal challenges with the homologous strain, but also confers significant protection against heterologous strains when combined with TSA56, a major outer membrane protein of O. tsutsugamushi.

Conclusions/significance: Immunization of ScaA proteins provides protective immunity in mice when challenged with the homologous strain and significantly enhanced protective immunity against infection with heterologous strains. To our knowledge, this is the most promising result of scrub typhus vaccination trials against infection of heterologous strains in mouse models thus far.

Show MeSH
Related in: MedlinePlus