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Differential expression of immunogenic proteins on virulent Mycobacterium tuberculosis clinical isolates.

Schierloh P, Klepp L, Vazquez C, Rocha RV, Blanco FC, Balboa L, López B, Ritacco V, Bigi F, Sasiain Mdel C - Biomed Res Int (2014)

Bottom Line: Recent studies have demonstrated that Mtb families found worldwide today differ in pathology, transmissibility, virulence, and development of immune response.We found that these proteins induced humoral immune responses in patients with drug-sensitive and drug-resistant tuberculosis with substantial cross-reactivity among the four proteins.Therefore, the present results describe new Mtb antigens with potential application as biomarkers of TB.

View Article: PubMed Central - PubMed

Affiliation: IMEX-CONICET, Academia Nacional de Medicina, Pacheco de Melo 3081, 1425 CABA, Argentina.

ABSTRACT
Molecular epidemiology has revealed that Mycobacterium tuberculosis (Mtb), formerly regarded as highly conserved species, displays a considerable degree of genetic variability that can influence the outcome of the disease as well as the innate and adaptive immune response. Recent studies have demonstrated that Mtb families found worldwide today differ in pathology, transmissibility, virulence, and development of immune response. By proteomic approaches seven proteins that were differentially expressed between a local clinical isolate from Latin-American-Mediterranean (LAM) and from Haarlem (H) lineages were identified. In order to analyze the immunogenic ability, recombinant Rv2241, Rv0009, Rv0407, and Rv2624c proteins were produced for testing specific antibody responses. We found that these proteins induced humoral immune responses in patients with drug-sensitive and drug-resistant tuberculosis with substantial cross-reactivity among the four proteins. Moreover, such reactivity was also correlated with anti-Mtb-cell surface IgM, but not with anti-ManLAM, anti-PPD, or anti-Mtb-surface IgG antibodies. Therefore, the present results describe new Mtb antigens with potential application as biomarkers of TB.

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Proteomics to identify differential proteins between LAM10406 and H12425 M. tuberculosis strains. (a) Equal amounts of proteins from culture supernatants of Mtb LAM10406 and H12425 were resolved on 1D 12% polyacrylamide gels, which were stained with colloidal Coomassie. The identified differential protein is indicated with the arrow. (b) Differential proteins identified by 2D gel electrophoresis. The first-dimension isoelectric focusing used a pH range of 4–7. The second-dimension SDS-PAGE was on 12% polyacrylamide gels, which were silver stained. Differential proteins present in the cell wall and cell-associated fractions of the Mtb LAM10406 and H12425 strains are indicated.
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fig1: Proteomics to identify differential proteins between LAM10406 and H12425 M. tuberculosis strains. (a) Equal amounts of proteins from culture supernatants of Mtb LAM10406 and H12425 were resolved on 1D 12% polyacrylamide gels, which were stained with colloidal Coomassie. The identified differential protein is indicated with the arrow. (b) Differential proteins identified by 2D gel electrophoresis. The first-dimension isoelectric focusing used a pH range of 4–7. The second-dimension SDS-PAGE was on 12% polyacrylamide gels, which were silver stained. Differential proteins present in the cell wall and cell-associated fractions of the Mtb LAM10406 and H12425 strains are indicated.

Mentions: The rationale behind searching for differentially expressed proteins between a strain belonging to LAM and a strain to Haarlem families was to find proteins that could explain the differential innate/adaptive immune responses developed by these two families [26–28]. To identify these proteins we compared the protein expression patterns of the H12425 and LAM10406 strains by 1D- and 2D-PAGE. In a first approach we prepared proteins from culture filtrates of both strains and resolved them by 1D electrophoresis gels, with subsequent colloidal Coomassie staining. In this case, we identified one differential band (Figure 1(a)) that was overexpressed in LAM10406. This band was cut from the gel and trypsin-digested to be identified by mass spectrometry (MS). Then, we analyzed the cell wall and cell associated protein fractions by silver-stained 2D electrophoresis gels. The comparison of two sets of 2D gels from each strain revealed four proteins present in the cell wall fraction of LAM10406 and absent in H12425, one protein from the same fraction present in H12425 and absent in LAM10406 and one protein only present in the cell associated fraction of LAM10406 (Figure 1(b)). All these proteins were identified by mass spectrometry and the results are summarized in Table 1. Although the functions of the proteins between both strains are diverse, most of them localize in the cell wall, where they may trigger a differential host immune response, which could impact the outcome/progression of the disease.


Differential expression of immunogenic proteins on virulent Mycobacterium tuberculosis clinical isolates.

Schierloh P, Klepp L, Vazquez C, Rocha RV, Blanco FC, Balboa L, López B, Ritacco V, Bigi F, Sasiain Mdel C - Biomed Res Int (2014)

Proteomics to identify differential proteins between LAM10406 and H12425 M. tuberculosis strains. (a) Equal amounts of proteins from culture supernatants of Mtb LAM10406 and H12425 were resolved on 1D 12% polyacrylamide gels, which were stained with colloidal Coomassie. The identified differential protein is indicated with the arrow. (b) Differential proteins identified by 2D gel electrophoresis. The first-dimension isoelectric focusing used a pH range of 4–7. The second-dimension SDS-PAGE was on 12% polyacrylamide gels, which were silver stained. Differential proteins present in the cell wall and cell-associated fractions of the Mtb LAM10406 and H12425 strains are indicated.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4109345&req=5

fig1: Proteomics to identify differential proteins between LAM10406 and H12425 M. tuberculosis strains. (a) Equal amounts of proteins from culture supernatants of Mtb LAM10406 and H12425 were resolved on 1D 12% polyacrylamide gels, which were stained with colloidal Coomassie. The identified differential protein is indicated with the arrow. (b) Differential proteins identified by 2D gel electrophoresis. The first-dimension isoelectric focusing used a pH range of 4–7. The second-dimension SDS-PAGE was on 12% polyacrylamide gels, which were silver stained. Differential proteins present in the cell wall and cell-associated fractions of the Mtb LAM10406 and H12425 strains are indicated.
Mentions: The rationale behind searching for differentially expressed proteins between a strain belonging to LAM and a strain to Haarlem families was to find proteins that could explain the differential innate/adaptive immune responses developed by these two families [26–28]. To identify these proteins we compared the protein expression patterns of the H12425 and LAM10406 strains by 1D- and 2D-PAGE. In a first approach we prepared proteins from culture filtrates of both strains and resolved them by 1D electrophoresis gels, with subsequent colloidal Coomassie staining. In this case, we identified one differential band (Figure 1(a)) that was overexpressed in LAM10406. This band was cut from the gel and trypsin-digested to be identified by mass spectrometry (MS). Then, we analyzed the cell wall and cell associated protein fractions by silver-stained 2D electrophoresis gels. The comparison of two sets of 2D gels from each strain revealed four proteins present in the cell wall fraction of LAM10406 and absent in H12425, one protein from the same fraction present in H12425 and absent in LAM10406 and one protein only present in the cell associated fraction of LAM10406 (Figure 1(b)). All these proteins were identified by mass spectrometry and the results are summarized in Table 1. Although the functions of the proteins between both strains are diverse, most of them localize in the cell wall, where they may trigger a differential host immune response, which could impact the outcome/progression of the disease.

Bottom Line: Recent studies have demonstrated that Mtb families found worldwide today differ in pathology, transmissibility, virulence, and development of immune response.We found that these proteins induced humoral immune responses in patients with drug-sensitive and drug-resistant tuberculosis with substantial cross-reactivity among the four proteins.Therefore, the present results describe new Mtb antigens with potential application as biomarkers of TB.

View Article: PubMed Central - PubMed

Affiliation: IMEX-CONICET, Academia Nacional de Medicina, Pacheco de Melo 3081, 1425 CABA, Argentina.

ABSTRACT
Molecular epidemiology has revealed that Mycobacterium tuberculosis (Mtb), formerly regarded as highly conserved species, displays a considerable degree of genetic variability that can influence the outcome of the disease as well as the innate and adaptive immune response. Recent studies have demonstrated that Mtb families found worldwide today differ in pathology, transmissibility, virulence, and development of immune response. By proteomic approaches seven proteins that were differentially expressed between a local clinical isolate from Latin-American-Mediterranean (LAM) and from Haarlem (H) lineages were identified. In order to analyze the immunogenic ability, recombinant Rv2241, Rv0009, Rv0407, and Rv2624c proteins were produced for testing specific antibody responses. We found that these proteins induced humoral immune responses in patients with drug-sensitive and drug-resistant tuberculosis with substantial cross-reactivity among the four proteins. Moreover, such reactivity was also correlated with anti-Mtb-cell surface IgM, but not with anti-ManLAM, anti-PPD, or anti-Mtb-surface IgG antibodies. Therefore, the present results describe new Mtb antigens with potential application as biomarkers of TB.

Show MeSH
Related in: MedlinePlus