Biocontrol agents Pseudomonas brassicacearum DF41 & Pseudomonas chlororaphis PA23: Investigation of fungal suppression and defense against Caenorhabditis elegans
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Date
2015-04-22
Authors
Nandi, Munmun
Journal Title
Journal ISSN
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Publisher
PLoS One
Abstract
The success of biocontrol bacteria is often restrained due to their low persistence in the rhizosphere and fluctuations in expression of antagonistic compounds. In the first part of this thesis the ability of the biocontrol agents (BCAs) Pseudomonas brassicacearum DF41 and Pseudomonas chlororaphis PA23 to resist grazing by the bacterivorous nematode Caenorhabditis elegans was investigated. We found that both BCAs are capable of killing the nematodes through exposure to toxic metabolites. We discovered that in addition to HCN, pyrrolnitrin (PRN) is a potent nematicide produced by PA23. Unique for a pseudomonad, DF41 was also found to kill the nematodes by forming biofilms on the nematode anterior, causing starvation. Biofilm formation was dependent upon the Gac two-component system and NaCl concentration of the media. Co-culturing these BCAs in the presence of nematodes increased expression of a number of genes associated with biocontrol. We observed elevated exoproduct formation, consistent with our gene expression analysis. The nematicidal activity exhibited by DF41 and PA23 towards C. elegans bodes well for their persistence in the environment.
In the second part of this thesis the role of hydrogen cyanide (HCN) and the anaerobic regulator ANR in PA23 biocontrol was explored. An hcn mutant was created and in vitro antifungal (AF) assays revealed the involvement of HCN in Sclerotinia sclerotiorum suppression. Addition of glycine promoted both AF activity and HCN production. In addition, HCN was found to be positively regulated by quorum sensing (QS). Besides a phz box, an anr box was identified in the hcnA promoter, suggesting a role for ANR in regulating hcnA. An anr mutant was generated and phenotypic characterization revealed that ANR is a key regulator governing PA23 secondary metabolite production. Through gene expression analysis, ANR was shown to positively regulate phzI/phzR and PhzR negatively regulate anr. Furthermore, expressing anr in trans partially complemented the QS-deficient phenotype with respect to several biocontrol genes and exoproducts. Overall, the global regulator ANR is vital for PA23-mediated biocontrol and a significant overlap exists between the QS and ANR regulons.
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Keywords
Bacterivorous, Biocontrol, Biofilm, Gene expression, Metabolites, Pseudomonas, Quorum sensing
Citation
Nandi, M., Selin, C., Brassinga, A.K.C., Belmonte, M.F., Fernando, W.G.D., Loewen, P.C. and de Kievit, T.R. 2015. Pyrrolnitrin and hydrogen cyanide production by Pseudomonas chlororaphis strain PA23 exhibits nematicidal and repellent activity against Caenorhabditis elegans. PloSOne/doi:10.1371/Journal.poine.0123184.