title: Nonribosomal peptides, key biocontrol components for Pseudomonas fluorescens In5, isolated from a Greenlandic suppressive soil.
abstract: Potatoes are cultivated in southwest Greenland without the use of pesticides and with limited crop rotation. Despite the fact that plant-pathogenic fungi are present, no severe-disease outbreaks have yet been observed. In this report, we document that a potato soil at Inneruulalik in southern Greenland is suppressive against Rhizoctonia solani Ag3 and uncover the suppressive antifungal mechanism of a highly potent biocontrol bacterium, Pseudomonas fluorescens In5, isolated from the suppressive potato soil. A combination of molecular genetics, genomics, and matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) imaging mass spectrometry (IMS) revealed an antifungal genomic island in P. fluorescens In5 encoding two nonribosomal peptides, nunamycin and nunapeptin, which are key components for the biocontrol activity by strain In5 in vitro and in soil microcosm experiments. Furthermore, complex microbial behaviors were highlighted. Whereas nunamycin was demonstrated to inhibit the mycelial growth of R. solani Ag3, but not that of Pythium aphanidermatum, nunapeptin instead inhibited P. aphanidermatum but not R. solani Ag3. Moreover, the synthesis of nunamycin by P. fluorescens In5 was inhibited in the presence of P. aphanidermatum. Further characterization of the two peptides revealed nunamycin to be a monochlorinated 9-amino-acid cyclic lipopeptide with similarity to members of the syringomycin group, whereas nunapeptin was a 22-amino-acid cyclic lipopeptide with similarity to corpeptin and syringopeptin.
  
  
  OBJECTIVE
  Crop rotation and systematic pest management are used to only a limited extent in Greenlandic potato farming. Nonetheless, although plant-pathogenic fungi are present in the soil, the farmers do not experience major plant disease outbreaks. Here, we show that a Greenlandic potato soil is suppressive against Rhizoctonia solani, and we unravel the key biocontrol components for Pseudomonas fluorescens In5, one of the potent biocontrol bacteria isolated from this Greenlandic suppressive soil. Using a combination of molecular genetics, genomics, and microbial imaging mass spectrometry, we show that two cyclic lipopeptides, nunamycin and nunapeptin, are important for the biocontrol activity of P. fluorescens In5 both in vitro and in microcosm assays. Furthermore, we demonstrate that the synthesis of nunamycin is repressed by the oomycete Pythium aphanidermatum. Overall, our report provides important insight into interkingdom interference between bacteria and fungi/oomycetes.
authors.affiliation: Department of Plant and Environmental Sciences, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Charlotte F
authors.initials: CF
authors.lastname: Michelsen
authors.forename: Jeramie
authors.initials: J
authors.lastname: Watrous
authors.affiliation: Department of Plant and Environmental Sciences, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Mikkel A
authors.initials: MA
authors.lastname: Glaring
authors.forename: Roland
authors.initials: R
authors.lastname: Kersten
authors.affiliation: Graduate School of Pharmaceutical Sciences, Kitasato University, Tokyo, Japan.
authors.forename: Nobuhiro
authors.initials: N
authors.lastname: Koyama
authors.affiliation: psg@plen.ku.dk pdorrestein@ucsd.edu.
authors.forename: Pieter C
authors.initials: PC
authors.lastname: Dorrestein
authors.affiliation: Department of Plant and Environmental Sciences, University of Copenhagen, Copenhagen, Denmark psg@plen.ku.dk pdorrestein@ucsd.edu.
authors.forename: Peter
authors.initials: P
authors.lastname: Stougaard
mesh_terms: D000935:Antifungal Agents / Q000378:metabolism*; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D005658:Fungi / Q000187:drug effects* / Q000254:growth & development; D044404:Genomic Islands; D023281:Genomics; D006116:Greenland; D008967:Molecular Biology; D046908:Peptide Biosynthesis, Nucleic Acid-Independent; D010455:Peptides / Q000378:metabolism*; D040901:Proteomics; D011551:Pseudomonas fluorescens / Q000302:isolation & purification* / Q000378:metabolism*; D017422:Sequence Analysis, DNA; D012988:Soil Microbiology; D011198:Solanum tuberosum / Q000254:growth & development; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D000935:Antifungal Agents; D004269:DNA, Bacterial; D010455:Peptides