Dissecting Disease-Suppressive Rhizosphere Microbiomes by Functional Amplicon Sequencing and 10× Metagenomics

Tracanna, Vittorio; Ossowicki, Adam; Petrus, Marloes L. C.; Overduin, Sam; Terlouw, Barbara R.; Lund, George; Robinson, Serina L.; Warris, Sven; Schijlen, Elio; Wezel, Gilles P. van; Raaijmakers, Jos M.; Garbeva, Paolina; Medema, Marnix H.

doi:10.1128/msystems.01116-20

Cited by 31 publications

(15 citation statements)

References 63 publications

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“…Pangenome analysis and metagenomics represent increasingly powerful routes to understanding the genetic determinants of biological activity in plant-associated microbes (Beskrovnaya et al, 2020;Biessy et al, 2019; V. J. Carrión et al, 2019;Melnyk et al, 2019;Mullins et al, 2019;Tracanna et al, 2021).…”

Section: Discussionmentioning

confidence: 99%

“…This population-level approach shows that the P. fluorescens population in a single field is highly complex, heterogeneous, and dynamic ( Figures 2 , 3 and 8 ), where the overall genotypic diversity is similar to the global diversity of P. fluorescens ( Garrido-Sanz et al, 2016 ). Pan-genome analysis and metagenomics represent increasingly powerful routes to understanding the genetic determinants of biological activity in plant-associated microbes ( Beskrovnaya et al, 2020 ; Biessy et al, 2019 ; Carrión et al, 2019 ; Melnyk et al, 2019 ; Mullins et al, 2019 ; Tracanna et al, 2021 ).…”

Section: Discussionmentioning

confidence: 99%

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Pan-genome analysis identifies intersecting roles for Pseudomonas specialized metabolites in potato pathogen inhibition

Pacheco-Moreno

Stefanato

Ford

et al. 2021

eLife

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Agricultural soil harbors a diverse microbiome that can form beneficial relationships with plants, including the inhibition of plant pathogens. Pseudomonas spp. are one of the most abundant bacterial genera in the soil and rhizosphere and play important roles in promoting plant health. However, the genetic determinants of this beneficial activity are only partially understood. Here, we genetically and phenotypically characterize the Pseudomonas fluorescens population in a commercial potato field, where we identify strong correlations between specialized metabolite biosynthesis and antagonism of the potato pathogens Streptomyces scabies and Phytophthora infestans. Genetic and chemical analyses identified hydrogen cyanide and cyclic lipopeptides as key specialized metabolites associated with S. scabies inhibition, which was supported by in planta biocontrol experiments. We show that a single potato field contains a hugely diverse and dynamic population of Pseudomonas bacteria, whose capacity to produce specialized metabolites is shaped both by plant colonization and defined environmental inputs.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Pan-genome analysis identifies intersecting roles for Pseudomonas specialized metabolites in potato pathogen inhibition

Pacheco-Moreno

Stefanato

Ford

et al. 2021

eLife

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“…This population level approach shows that the P. fluorescens population in a single field is highly complex, heterogeneous and dynamic (Figures 2, 3 and 8), where the overall genotypic diversity is similar to the global diversity of P. fluorescens (23). Pan-genome analysis and metagenomics represent increasingly powerful routes to understanding the genetic determinants of biological activity in plant-associated microbes (10,22,(92)(93)(94)(95).…”

Section: Discussionmentioning

confidence: 99%

Pan-genome analysis identifies intersecting roles for Pseudomonas specialized metabolites in potato pathogen inhibition

Stefanato

Trippel

Uszkoreit

et al. 2019

Preprint

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23Agricultural soil harbors a diverse microbiome that can form beneficial relationships with plants, 24 including the inhibition of plant pathogens. Pseudomonas are one of the most abundant 25 bacterial genera in the soil and rhizosphere and play important roles in promoting plant growth 26 and preventing disease. However, the genetic determinants of this beneficial activity are only 27 partially understood, especially in relation to specialized metabolite production. Here, we 28 genetically and phenotypically characterize the Pseudomonas fluorescens population in 29 commercial potato field soils and identify strong correlations between specialized metabolite 30 biosynthetic pathways and antagonism of the potato pathogens Streptomyces scabies and 31 Phytophthora infestans. Genetic and chemical analyses identified hydrogen cyanide and cyclic 32 lipopeptides as key specialized metabolites associated with S. scabies inhibition. We show that 33 a single potato field contains a hugely diverse and dynamic population of Pseudomonas 34 bacteria, whose capacity to produce specialized metabolites is shaped both by plant 35 colonization and defined environmental inputs. 36 2 37 CFC agar and incubated overnight at 28 °C before streaking to single colonies on King's B (KB) 595 agar plates (96). Six isolates were selected at random per soil sample and subjected to 596 phenotypic/genomic analysis. 597 598 Amplicon sequencing 599 Genomic DNA was isolated from 3 g of pooled soil samples using the FastDNA™ SPIN Kit for 600 soil (MP Biomedicals, UK) following the manufacturer's instructions. Genomic DNA 601 concentration and purity was determined by NanoDrop spectrophotometry as above. Microbial 602 16S rRNA genes were amplified from soil DNA samples with barcoded universal prokaryotic 603 23 primers (515F/R806) targeting the V4 region, and then subjected to Illumina® MiSeq 604 sequencing (600-cycle, 2x300 bp) at the DNA Sequencing Facility, Department of Biochemistry, 605

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“…As phylogenetic clustering of MacB amino acid sequences correlated well with the classification by family of LPs, it offers an accessory tool for the initial characterization of LP BGCs revealed by (meta)genomic sequencing. Next, we investigated the possibility of using macB for PCR-based detection of the presence of a LP BGC in Pseudomonas environmental isolates, as an alternative for methods targeting NRPS catalytic domains using degenerate primers, which have proven useful for detecting NRPS genes in Proteobacteria , including Pseudomonas , Firmicutes , Actinomycetes , and Cyanobacteria ( 28 – 32 ). Two sets of primers were designed based on the LP macB -like gene (which we propose to designate as pleB ) of known producers (i.e., chemically and/or genetically characterized strains) and putative producers identified by genome mining.…”

Section: Introductionmentioning

confidence: 99%

Transporter Gene-Mediated Typing for Detection and Genome Mining of Lipopeptide-Producing Pseudomonas

Girard

Geudens

Pauwels

et al. 2022

Appl Environ Microbiol

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Pseudomonas lipopeptides (LPs) are involved in diverse ecological functions and have biotechnological potential associated with their antimicrobial and/or anti-proliferative activities. They are synthesized by multi-modular non-ribosomal peptide synthetases which, together with transport and regulatory proteins, are encoded by large biosynthetic gene clusters (BGCs). These secondary metabolites are classified in distinct families based on sequence and length of the oligopeptide, and size of the macrocycle, if present. Phylogeny of PleB, the MacB-like transporter that is part of a dedicated ATP-dependent tripartite efflux system driving export of Pseudomonas LPs, revealed a strong correlation with LP chemical diversity. As each LP BGC carries its cognate pleB , PleB is suitable as a diagnostic sequence for genome mining, allowing assignment of the putative metabolite to a particular LP family. In addition, pleB proved a suitable target gene for an alternative PCR method to detect LP-producing Pseudomonas , not relying on amplification of catalytic domains of the biosynthetic enzymes. Combined with amplicon sequencing, this approach enabled typing of Pseudomonas strains as potential producers of a LP belonging to one of ten different families, underscoring its value for strain prioritization. This was validated by chemical characterization of known LPs from three different families secreted by novel producers isolated from the rice or maize rhizosphere, namely the type strains of Pseudomonas fulva (putisolvin), Pseudomonas zeae (tensin) and Pseudomonas xantholysinigenes (xantholysin). In addition, a new member of the Bananamide family, prosekin, was discovered in the type strain of Pseudomonas prosekii , an Antarctic isolate. Importance Pseudomonas are ubiquitous bacteria able to thrive in a wide range of ecological niches and lipopeptides often support their lifestyle but also their interaction with other micro- and macro-organisms. Therefore, the production of lipopeptides is widespread among Pseudomonas strains. Consequently, Pseudomonas lipopeptide research affects not only chemists and microbiologists but touches a much broader audience, including biochemists, ecologists and plant biologists. In this study we present a reliable transporter gene-guided approach for the detection and/or typing of Pseudomonas lipopeptide producers. Indeed, it allows to readily assess the lipopeptide diversity among sets of Pseudomonas isolates and differentiate strains likely to produce known lipopeptides from producers of potentially novel lipopeptides. This work provides a valuable tool that can also be integrated in a genome mining strategy and adapted for the typing of other specialized metabolites.

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Dissecting Disease-Suppressive Rhizosphere Microbiomes by Functional Amplicon Sequencing and 10× Metagenomics

Abstract: Soil-borne plant-pathogenic fungi continue to be a major threat to agriculture and horticulture. The genus Fusarium in particular is one of the most devastating groups of soilborne fungal pathogens for a wide range of crops.

Cited by 31 publications

References 63 publications

Pan-genome analysis identifies intersecting roles for Pseudomonas specialized metabolites in potato pathogen inhibition

Pan-genome analysis identifies intersecting roles for Pseudomonas specialized metabolites in potato pathogen inhibition

Pan-genome analysis identifies intersecting roles for Pseudomonas specialized metabolites in potato pathogen inhibition

Transporter Gene-Mediated Typing for Detection and Genome Mining of Lipopeptide-Producing Pseudomonas

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