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Mapping of the denitrification pathway in Burkholderia thailandensis by genome-wide mutant profiling


Vitale, Alessandra; Paszti, Sarah; Takahashi, Kohei; Toyofuku, Masanori; Pessi, Gabriella; Eberl, Leo (2020). Mapping of the denitrification pathway in Burkholderia thailandensis by genome-wide mutant profiling. Journal of Bacteriology, 202(23):00304-20.

Abstract

Burkholderia thailandensis is a soil saprophyte that is closely related to the pathogen Burkholderia pseudomallei, the etiological agent of melioidosis in humans. The environmental niches and infection sites occupied by these bacteria are thought to contain only limited concentrations of oxygen, where they can generate energy via denitrification. However, knowledge of the underlying molecular basis of the denitrification pathway in these bacteria is scarce. In this study, we employed a transposon sequencing (Tn-Seq) approach to identify genes conferring a fitness benefit for anaerobic growth of B. thailandensis. Of the 180 determinants identified, several genes were shown to be required for growth under denitrifying conditions: the nitrate reductase operon narIJHGK2K1, the aniA gene encoding a previously unknown nitrite reductase, and the petABC genes encoding a cytochrome bc1, as well as three novel regulators that control denitrification. Our Tn-Seq data allowed us to reconstruct the entire denitrification pathway of B. thailandensis and shed light on its regulation. Analyses of growth behaviors combined with measurements of denitrification metabolites of various mutants revealed that nitrate reduction provides sufficient energy for anaerobic growth, an important finding in light of the fact that some pathogenic Burkholderia species can use nitrate as a terminal electron acceptor but are unable to complete denitrification. Finally, we demonstrated that a nitrous oxide reductase mutant is not affected for anaerobic growth but is defective in biofilm formation and accumulates N2O, which may play a role in the dispersal of B. thailandensis biofilms.

Abstract

Burkholderia thailandensis is a soil saprophyte that is closely related to the pathogen Burkholderia pseudomallei, the etiological agent of melioidosis in humans. The environmental niches and infection sites occupied by these bacteria are thought to contain only limited concentrations of oxygen, where they can generate energy via denitrification. However, knowledge of the underlying molecular basis of the denitrification pathway in these bacteria is scarce. In this study, we employed a transposon sequencing (Tn-Seq) approach to identify genes conferring a fitness benefit for anaerobic growth of B. thailandensis. Of the 180 determinants identified, several genes were shown to be required for growth under denitrifying conditions: the nitrate reductase operon narIJHGK2K1, the aniA gene encoding a previously unknown nitrite reductase, and the petABC genes encoding a cytochrome bc1, as well as three novel regulators that control denitrification. Our Tn-Seq data allowed us to reconstruct the entire denitrification pathway of B. thailandensis and shed light on its regulation. Analyses of growth behaviors combined with measurements of denitrification metabolites of various mutants revealed that nitrate reduction provides sufficient energy for anaerobic growth, an important finding in light of the fact that some pathogenic Burkholderia species can use nitrate as a terminal electron acceptor but are unable to complete denitrification. Finally, we demonstrated that a nitrous oxide reductase mutant is not affected for anaerobic growth but is defective in biofilm formation and accumulates N2O, which may play a role in the dispersal of B. thailandensis biofilms.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:07 Faculty of Science > Department of Plant and Microbial Biology
07 Faculty of Science > Zurich-Basel Plant Science Center
Dewey Decimal Classification:580 Plants (Botany)
Scopus Subject Areas:Life Sciences > Microbiology
Life Sciences > Molecular Biology
Uncontrolled Keywords:Molecular Biology, Microbiology
Language:English
Date:8 September 2020
Deposited On:20 Jan 2021 11:55
Last Modified:26 Jan 2024 02:39
Publisher:American Society for Microbiology
ISSN:0021-9193
OA Status:Closed
Free access at:Publisher DOI. An embargo period may apply.
Publisher DOI:https://doi.org/10.1128/jb.00304-20
Project Information:
  • : FunderSNSF
  • : Grant ID31003A_153374
  • : Project TitleUnraveling the molecular basis of the nitrogen-fixing symbiosis between legumes and Ã�-rhizobia: a functional analysis of the Phaseolus-Burkholderia association
  • : FunderSNSF
  • : Grant ID31003A_143773
  • : Project TitleAnalysis of the quorum sensing circuitry of Burkholderia cenocepacia H111: The role of the Burkholderia diffusible signal factor (BDSF)