Molecular Characterization of Putative Virulence Determinants in Burkholderia pseudomallei
The Gram-negative saprophyte Burkholderia pseudomallei is the causative agent of melioidosis, an infectious disease which is endemic in Southeast Asia and northern Australia. This bacterium possesses many virulence factors which are thought to contribute to its survival and pathogenicity. Using a vi...
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| Language: | English |
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Wiley
2014-01-01
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| Series: | The Scientific World Journal |
| Online Access: | http://dx.doi.org/10.1155/2014/590803 |
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| author | Suat Moi Puah S. D. Puthucheary Jin Town Wang Yi Jiun Pan Kek Heng Chua |
| author_facet | Suat Moi Puah S. D. Puthucheary Jin Town Wang Yi Jiun Pan Kek Heng Chua |
| author_sort | Suat Moi Puah |
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| description | The Gram-negative saprophyte Burkholderia pseudomallei is the causative agent of melioidosis, an infectious disease which is endemic in Southeast Asia and northern Australia. This bacterium possesses many virulence factors which are thought to contribute to its survival and pathogenicity. Using a virulent clinical isolate of B. pseudomallei and an attenuated strain of the same B. pseudomallei isolate, 6 genes BPSL2033, BP1026B_I2784, BP1026B_I2780, BURPS1106A_A0094, BURPS1106A_1131, and BURPS1710A_1419 were identified earlier by PCR-based subtractive hybridization. These genes were extensively characterized at the molecular level, together with an additional gene BPSL3147 that had been identified by other investigators. Through a reverse genetic approach, single-gene knockout mutants were successfully constructed by using site-specific insertion mutagenesis and were confirmed by PCR. BPSL2033::Km and BURPS1710A_1419::Km mutants showed reduced rates of survival inside macrophage RAW 264.7 cells and also low levels of virulence in the nematode infection model. BPSL2033::Km demonstrated weak statistical significance (P=0.049) at 8 hours after infection in macrophage infection study but this was not seen in BURPS1710A_1419::Km. Nevertheless, complemented strains of both genes were able to partially restore the gene defects in both in vitro and in vivo studies, thus suggesting that they individually play a minor role in the virulence of B. pseudomallei. |
| format | Article |
| id | doaj-art-213dfe0c6fe84b76b3b961dff6918761 |
| institution | Kabale University |
| issn | 2356-6140 1537-744X |
| language | English |
| publishDate | 2014-01-01 |
| publisher | Wiley |
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| series | The Scientific World Journal |
| spelling | doaj-art-213dfe0c6fe84b76b3b961dff69187612025-02-03T01:26:09ZengWileyThe Scientific World Journal2356-61401537-744X2014-01-01201410.1155/2014/590803590803Molecular Characterization of Putative Virulence Determinants in Burkholderia pseudomalleiSuat Moi Puah0S. D. Puthucheary1Jin Town Wang2Yi Jiun Pan3Kek Heng Chua4Department of Biomedical Science, Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur, MalaysiaDepartment of Medical Education, Research and Evaluation, Duke-NUS Graduate Medical School Singapore, 8 College Road, 169857, SingaporeDepartment of Microbiology, National Taiwan University College of Medicine, Section 1, Jen-Ai Road, Taipei 10051, TaiwanDepartment of Microbiology, National Taiwan University College of Medicine, Section 1, Jen-Ai Road, Taipei 10051, TaiwanDepartment of Biomedical Science, Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur, MalaysiaThe Gram-negative saprophyte Burkholderia pseudomallei is the causative agent of melioidosis, an infectious disease which is endemic in Southeast Asia and northern Australia. This bacterium possesses many virulence factors which are thought to contribute to its survival and pathogenicity. Using a virulent clinical isolate of B. pseudomallei and an attenuated strain of the same B. pseudomallei isolate, 6 genes BPSL2033, BP1026B_I2784, BP1026B_I2780, BURPS1106A_A0094, BURPS1106A_1131, and BURPS1710A_1419 were identified earlier by PCR-based subtractive hybridization. These genes were extensively characterized at the molecular level, together with an additional gene BPSL3147 that had been identified by other investigators. Through a reverse genetic approach, single-gene knockout mutants were successfully constructed by using site-specific insertion mutagenesis and were confirmed by PCR. BPSL2033::Km and BURPS1710A_1419::Km mutants showed reduced rates of survival inside macrophage RAW 264.7 cells and also low levels of virulence in the nematode infection model. BPSL2033::Km demonstrated weak statistical significance (P=0.049) at 8 hours after infection in macrophage infection study but this was not seen in BURPS1710A_1419::Km. Nevertheless, complemented strains of both genes were able to partially restore the gene defects in both in vitro and in vivo studies, thus suggesting that they individually play a minor role in the virulence of B. pseudomallei.http://dx.doi.org/10.1155/2014/590803 |
| spellingShingle | Suat Moi Puah S. D. Puthucheary Jin Town Wang Yi Jiun Pan Kek Heng Chua Molecular Characterization of Putative Virulence Determinants in Burkholderia pseudomallei The Scientific World Journal |
| title | Molecular Characterization of Putative Virulence Determinants in Burkholderia pseudomallei |
| title_full | Molecular Characterization of Putative Virulence Determinants in Burkholderia pseudomallei |
| title_fullStr | Molecular Characterization of Putative Virulence Determinants in Burkholderia pseudomallei |
| title_full_unstemmed | Molecular Characterization of Putative Virulence Determinants in Burkholderia pseudomallei |
| title_short | Molecular Characterization of Putative Virulence Determinants in Burkholderia pseudomallei |
| title_sort | molecular characterization of putative virulence determinants in burkholderia pseudomallei |
| url | http://dx.doi.org/10.1155/2014/590803 |
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