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Characterization of the Burkholderia mallei tonB Mutant and Its Potential as a Backbone Strain for Vaccine Development.

Mott TM, Vijayakumar S, Sbrana E, Endsley JJ, Torres AG - PLoS Negl Trop Dis (2015)

Bottom Line: At 21 days post-immunization, TMM001-treated animals showed significantly higher levels of B. mallei-specific IgG1, IgG2a and IgM when compared to PBS-treated controls.At 48 h post-challenge, PBS-treated controls exhibited higher levels of serum inflammatory cytokines and more severe pathological damage to target organs compared to animals receiving TMM001.Although further work is needed to prevent chronic infection by TMM001 while maintaining immunogenicity, our attenuated strain demonstrates great potential as a backbone strain for future vaccine development against both glanders and melioidosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, Texas, United States of America.

ABSTRACT

Background: In this study, a Burkholderia mallei tonB mutant (TMM001) deficient in iron acquisition was constructed, characterized, and evaluated for its protective properties in acute inhalational infection models of murine glanders and melioidosis.

Methodology/principal findings: Compared to the wild-type, TMM001 exhibits slower growth kinetics, siderophore hyper-secretion and the inability to utilize heme-containing proteins as iron sources. A series of animal challenge studies showed an inverse correlation between the percentage of survival in BALB/c mice and iron-dependent TMM001 growth. Upon evaluation of TMM001 as a potential protective strain against infection, we found 100% survival following B. mallei CSM001 challenge of mice previously receiving 1.5 x 10(4) CFU of TMM001. At 21 days post-immunization, TMM001-treated animals showed significantly higher levels of B. mallei-specific IgG1, IgG2a and IgM when compared to PBS-treated controls. At 48 h post-challenge, PBS-treated controls exhibited higher levels of serum inflammatory cytokines and more severe pathological damage to target organs compared to animals receiving TMM001. In a cross-protection study of acute inhalational melioidosis with B. pseudomallei, TMM001-treated mice were significantly protected. While wild type was cleared in all B. mallei challenge studies, mice failed to clear TMM001.

Conclusions/significance: Although further work is needed to prevent chronic infection by TMM001 while maintaining immunogenicity, our attenuated strain demonstrates great potential as a backbone strain for future vaccine development against both glanders and melioidosis.

No MeSH data available.


Related in: MedlinePlus

Histopathology of TMM001- vs PBS-treated mice 48 h post challenge.H&E-stained lung (A, B) and spleen (C, D) of CSM001 (1.5 x 105 CFU) challenged mice previously immunized with PBS (A, C) or TMM001 (1.5 x 104 CFU) (B, D). Scale bar = 100μM. Histological scores were assigned for the liver (E), lungs (F), and spleen (G) tissue sections after microscopic examination. Mean ± SEM, representative of three animals, is plotted. Statistical significance was determined by the Mann-Whitney test. ★ p ≤ 0.05.
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pntd.0003863.g006: Histopathology of TMM001- vs PBS-treated mice 48 h post challenge.H&E-stained lung (A, B) and spleen (C, D) of CSM001 (1.5 x 105 CFU) challenged mice previously immunized with PBS (A, C) or TMM001 (1.5 x 104 CFU) (B, D). Scale bar = 100μM. Histological scores were assigned for the liver (E), lungs (F), and spleen (G) tissue sections after microscopic examination. Mean ± SEM, representative of three animals, is plotted. Statistical significance was determined by the Mann-Whitney test. ★ p ≤ 0.05.

Mentions: The mouse tissues (lungs, liver and spleen) from the TMM001 titration study (n = 3 per treatment) at 0 h and 48 h post-challenge were processed for histology. Representative images of the lungs, liver and spleen from PBS- and TMM001 (1.5 x 104 CFU)-immunized mice are presented in S6 Fig. At 0 h, the lungs, livers and spleens of PBS-treated mice were unremarkable, presenting as normal healthy tissue with normal architecture (S6 Fig, panels A-C). BALB/c mice immunized with TMM001 presented with mild-to-moderate changes in pathology: perivascular and peribronchial inflammatory infiltrates in the lung sections (S6 Fig, panel D), hepatitis with multifocal necrosis and scattered abscesses in the liver sections (S6 Fig, panel E), and necrosis of follicles and accumulation of neutrophils in spleen sections (S6 Fig, panel F). At 48 h post challenge with CSM001 (1.4 x 104 CFU), PBS-treated mice showed moderate-to-severe pathological changes, such as abscesses and multifocal inflammatory infiltrates in the lungs (S6 Fig, panel G and Fig 6A), areas of hepatocellular necrosis, occasional abscesses with necrotic cores and areas of focal necrosis in the liver (S6 Fig, panel H), and congestion of the red pulp, proliferation of large foamy macrophages (inset of Fig 6, panel C) and necrosis affecting the mantle zone (S6 Fig, panel I and Fig 6, panel C). Similarly, TMM001-immunized mice showed moderate-to-severe changes in pathology, but with a few differences. In the lungs, large, multifocal inflammatory infiltrates, as well as abscesses, were present with focal consolidation observed as well (S6 Fig, panel J and Fig 6, panel B). The liver presented with hepatitis and multiple foci of hepatocellular necrosis (S6 Fig, panel K), and large granulomas were formed in the spleen (S6 Fig, panel L and Fig 6, panel D). Histopathology scores showed significant differences due to treatment over time in the lungs (★★★★ p ≤ 0.0001), liver (★★★★ p ≤ 0.0001) and spleen (★★ p ≤ 0.001). When comparing the differences in treatment at 0 h and 48 h, the lungs (★ p = 0.05), liver (★ p = 0.05) and spleen (★ p = 0.05) showed a robust trend toward significance (Fig 6, panels E-G). Overall, TMM001-immunization alone does cause some histopathology as evident by the histopathology at 0 h. That being said, PBS-immunized animals exuded more extensive pathology 48 h after CSM001 challenge compared to TMM001-immunized animals.


Characterization of the Burkholderia mallei tonB Mutant and Its Potential as a Backbone Strain for Vaccine Development.

Mott TM, Vijayakumar S, Sbrana E, Endsley JJ, Torres AG - PLoS Negl Trop Dis (2015)

Histopathology of TMM001- vs PBS-treated mice 48 h post challenge.H&E-stained lung (A, B) and spleen (C, D) of CSM001 (1.5 x 105 CFU) challenged mice previously immunized with PBS (A, C) or TMM001 (1.5 x 104 CFU) (B, D). Scale bar = 100μM. Histological scores were assigned for the liver (E), lungs (F), and spleen (G) tissue sections after microscopic examination. Mean ± SEM, representative of three animals, is plotted. Statistical significance was determined by the Mann-Whitney test. ★ p ≤ 0.05.
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Related In: Results  -  Collection

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pntd.0003863.g006: Histopathology of TMM001- vs PBS-treated mice 48 h post challenge.H&E-stained lung (A, B) and spleen (C, D) of CSM001 (1.5 x 105 CFU) challenged mice previously immunized with PBS (A, C) or TMM001 (1.5 x 104 CFU) (B, D). Scale bar = 100μM. Histological scores were assigned for the liver (E), lungs (F), and spleen (G) tissue sections after microscopic examination. Mean ± SEM, representative of three animals, is plotted. Statistical significance was determined by the Mann-Whitney test. ★ p ≤ 0.05.
Mentions: The mouse tissues (lungs, liver and spleen) from the TMM001 titration study (n = 3 per treatment) at 0 h and 48 h post-challenge were processed for histology. Representative images of the lungs, liver and spleen from PBS- and TMM001 (1.5 x 104 CFU)-immunized mice are presented in S6 Fig. At 0 h, the lungs, livers and spleens of PBS-treated mice were unremarkable, presenting as normal healthy tissue with normal architecture (S6 Fig, panels A-C). BALB/c mice immunized with TMM001 presented with mild-to-moderate changes in pathology: perivascular and peribronchial inflammatory infiltrates in the lung sections (S6 Fig, panel D), hepatitis with multifocal necrosis and scattered abscesses in the liver sections (S6 Fig, panel E), and necrosis of follicles and accumulation of neutrophils in spleen sections (S6 Fig, panel F). At 48 h post challenge with CSM001 (1.4 x 104 CFU), PBS-treated mice showed moderate-to-severe pathological changes, such as abscesses and multifocal inflammatory infiltrates in the lungs (S6 Fig, panel G and Fig 6A), areas of hepatocellular necrosis, occasional abscesses with necrotic cores and areas of focal necrosis in the liver (S6 Fig, panel H), and congestion of the red pulp, proliferation of large foamy macrophages (inset of Fig 6, panel C) and necrosis affecting the mantle zone (S6 Fig, panel I and Fig 6, panel C). Similarly, TMM001-immunized mice showed moderate-to-severe changes in pathology, but with a few differences. In the lungs, large, multifocal inflammatory infiltrates, as well as abscesses, were present with focal consolidation observed as well (S6 Fig, panel J and Fig 6, panel B). The liver presented with hepatitis and multiple foci of hepatocellular necrosis (S6 Fig, panel K), and large granulomas were formed in the spleen (S6 Fig, panel L and Fig 6, panel D). Histopathology scores showed significant differences due to treatment over time in the lungs (★★★★ p ≤ 0.0001), liver (★★★★ p ≤ 0.0001) and spleen (★★ p ≤ 0.001). When comparing the differences in treatment at 0 h and 48 h, the lungs (★ p = 0.05), liver (★ p = 0.05) and spleen (★ p = 0.05) showed a robust trend toward significance (Fig 6, panels E-G). Overall, TMM001-immunization alone does cause some histopathology as evident by the histopathology at 0 h. That being said, PBS-immunized animals exuded more extensive pathology 48 h after CSM001 challenge compared to TMM001-immunized animals.

Bottom Line: At 21 days post-immunization, TMM001-treated animals showed significantly higher levels of B. mallei-specific IgG1, IgG2a and IgM when compared to PBS-treated controls.At 48 h post-challenge, PBS-treated controls exhibited higher levels of serum inflammatory cytokines and more severe pathological damage to target organs compared to animals receiving TMM001.Although further work is needed to prevent chronic infection by TMM001 while maintaining immunogenicity, our attenuated strain demonstrates great potential as a backbone strain for future vaccine development against both glanders and melioidosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, Texas, United States of America.

ABSTRACT

Background: In this study, a Burkholderia mallei tonB mutant (TMM001) deficient in iron acquisition was constructed, characterized, and evaluated for its protective properties in acute inhalational infection models of murine glanders and melioidosis.

Methodology/principal findings: Compared to the wild-type, TMM001 exhibits slower growth kinetics, siderophore hyper-secretion and the inability to utilize heme-containing proteins as iron sources. A series of animal challenge studies showed an inverse correlation between the percentage of survival in BALB/c mice and iron-dependent TMM001 growth. Upon evaluation of TMM001 as a potential protective strain against infection, we found 100% survival following B. mallei CSM001 challenge of mice previously receiving 1.5 x 10(4) CFU of TMM001. At 21 days post-immunization, TMM001-treated animals showed significantly higher levels of B. mallei-specific IgG1, IgG2a and IgM when compared to PBS-treated controls. At 48 h post-challenge, PBS-treated controls exhibited higher levels of serum inflammatory cytokines and more severe pathological damage to target organs compared to animals receiving TMM001. In a cross-protection study of acute inhalational melioidosis with B. pseudomallei, TMM001-treated mice were significantly protected. While wild type was cleared in all B. mallei challenge studies, mice failed to clear TMM001.

Conclusions/significance: Although further work is needed to prevent chronic infection by TMM001 while maintaining immunogenicity, our attenuated strain demonstrates great potential as a backbone strain for future vaccine development against both glanders and melioidosis.

No MeSH data available.


Related in: MedlinePlus