Jos M. Raaijmakers

Deciphering the rhizosphere microbiome for disease-suppressive bacteria.

A common plant pathogen induces the growth of disease-suppressive microbes in local soil communities. Disease-suppressive soils are exceptional ecosystems in which crop plants suffer less from specific soil-borne pathogens than expected owing to the activities of other soil microorganisms. For most disease-suppressive soils, the microbes and mechanisms involved in pathogen control are unknown. By coupling PhyloChip-based metagenomics of the rhizosphere microbiome with culture-dependent functional analyses, we identified key bacterial taxa and genes involved in suppression of a fungal root pathogen. More than 33,000 bacterial and archaeal species were detected, with Proteobacteria, Firmicutes, and Actinobacteria consistently associated with disease suppression. Members of the γ-Proteobacteria were shown to have disease-suppressive activity governed by nonribosomal peptide synthetases. Our data indicate that upon attack by a fungal root pathogen, plants can exploit microbial consortia from soil for protection against infections.

Going back to the roots: the microbial ecology of the rhizosphere

The rhizosphere is the interface between plant roots and soil where interactions among a myriad of microorganisms and invertebrates affect biogeochemical cycling, plant growth and tolerance to biotic and abiotic stress. The rhizosphere is intriguingly complex and dynamic, and understanding its ecology and evolution is key to enhancing plant productivity and ecosystem functioning. Novel insights into key factors and evolutionary processes shaping the rhizosphere microbiome will greatly benefit from integrating reductionist and systems-based approaches in both agricultural and natural ecosystems. Here, we discuss recent developments in rhizosphere research in relation to assessing the contribution of the micro- and macroflora to sustainable agriculture, nature conservation, the development of bio-energy crops and the mitigation of climate change.

The rhizosphere: a playground and battlefield for soilborne pathogens and beneficial microorganisms

The rhizosphere is a hot spot of microbial interactions as exudates released by plant roots are a main food source for microorganisms and a driving force of their population density and activities. The rhizosphere harbors many organisms that have a neutral effect on the plant, but also attracts organisms that exert deleterious or beneficial effects on the plant. Microorganisms that adversely affect plant growth and health are the pathogenic fungi, oomycetes, bacteria and nematodes. Most of the soilborne pathogens are adapted to grow and survive in the bulk soil, but the rhizosphere is the playground and infection court where the pathogen establishes a parasitic relationship with the plant. The rhizosphere is also a battlefield where the complex rhizosphere community, both microflora and microfauna, interact with pathogens and influence the outcome of pathogen infection. A wide range of microorganisms are beneficial to the plant and include nitrogen-fixing bacteria, endo- and ectomycorrhizal fungi, and plant growth-promoting bacteria and fungi. This review focuses on the population dynamics and activity of soilborne pathogens and beneficial microorganisms. Specific attention is given to mechanisms involved in the tripartite interactions between beneficial microorganisms, pathogens and the plant. We also discuss how agricultural practices affect pathogen and antagonist populations and how these practices can be adopted to promote plant growth and health.

Antibiotic production by bacterial biocontrol agents

Interest in biological control of plant pathogens has been stimulated in recent years by trends in agriculture towards greater sustainability and public concern about the use of hazardous pesticides. There is now unequivocal evidence that antibiotics play a key role in the suppression of various soilborne plant pathogens by antagonistic microorganisms. The significance of antibiotics in biocontrol, and more generally in microbial interactions, often has been questioned because of the indirect nature of the supporting evidence and the perceived constraints to antibiotic production in rhizosphere environments. Reporter gene systems and bio-analytical techniques have clearly demonstrated that antibiotics are produced in the spermosphere and rhizosphere of a variety of host plants. Several abiotic factors such as oxygen, temperature, specific carbon and nitrogen sources, and microelements have been identified to influence antibiotic production by bacteria biocontrol agents. Among the biotic factors that may play a determinative role in antibiotic production are the plant host, the pathogen, the indigenous microflora, and the cell density of the producing strain. This review presents recent advances in our understanding of antibiotic production by bacterial biocontrol agents and their role in microbial interactions.

The rhizosphere microbiome: significance of plant beneficial, plant pathogenic, and human pathogenic microorganisms

Microbial communities play a pivotal role in the functioning of plants by influencing their physiology and development. While many members of the rhizosphere microbiome are beneficial to plant growth, also plant pathogenic microorganisms colonize the rhizosphere striving to break through the protective microbial shield and to overcome the innate plant defense mechanisms in order to cause disease. A third group of microorganisms that can be found in the rhizosphere are the true and opportunistic human pathogenic bacteria, which can be carried on or in plant tissue and may cause disease when introduced into debilitated humans. Although the importance of the rhizosphere microbiome for plant growth has been widely recognized, for the vast majority of rhizosphere microorganisms no knowledge exists. To enhance plant growth and health, it is essential to know which microorganism is present in the rhizosphere microbiome and what they are doing. Here, we review the main functions of rhizosphere microorganisms and how they impact on health and disease. We discuss the mechanisms involved in the multitrophic interactions and chemical dialogues that occur in the rhizosphere. Finally, we highlight several strategies to redirect or reshape the rhizosphere microbiome in favor of microorganisms that are beneficial to plant growth and health.

Natural functions of lipopeptides from Bacillus and Pseudomonas: more than surfactants and antibiotics

Lipopeptides constitute a structurally diverse group of metabolites produced by various bacterial and fungal genera. In the past decades, research on lipopeptides has been fueled by their antimicrobial, antitumour, immunosuppressant and surfactant activities. However, the natural functions of lipopeptides in the lifestyles of the producing microorganisms have received considerably less attention. The substantial structural diversity of lipopeptides suggests that these metabolites have different natural roles, some of which may be unique to the biology of the producing organism. This review gives a detailed overview of the versatile functions of lipopeptides in the biology of Pseudomonas and Bacillus species, and highlights their role in competitive interactions with coexisting organisms, including bacteria, fungi, oomycetes, protozoa, nematodes and plants. Their functions in cell motility, leading to colonization of novel habitats, and in the formation and development of highly structured biofilms are discussed in detail. Finally, this review provides an update on lipopeptide detection and discovery as well as on novel regulatory mechanisms and genes involved in lipopeptide biosynthesis in these two bacterial genera.

Mass spectral molecular networking of living microbial colonies

Integrating the governing chemistry with the genomics and phenotypes of microbial colonies has been a “holy grail” in microbiology. This work describes a highly sensitive, broadly applicable, and cost-effective approach that allows metabolic profiling of live microbial colonies directly from a Petri dish without any sample preparation. Nanospray desorption electrospray ionization mass spectrometry (MS), combined with alignment of MS data and molecular networking, enabled monitoring of metabolite production from live microbial colonies from diverse bacterial genera, including Bacillus subtilis, Streptomyces coelicolor, Mycobacterium smegmatis, and Pseudomonas aeruginosa. This work demonstrates that, by using these tools to visualize small molecular changes within bacterial interactions, insights can be gained into bacterial developmental processes as a result of the improved organization of MS/MS data. To validate this experimental platform, metabolic profiling was performed on Pseudomonas sp. SH-C52, which protects sugar beet plants from infections by specific soil-borne fungi [R. Mendes et al. (2011) Science 332:1097–1100]. The antifungal effect of strain SH-C52 was attributed to thanamycin, a predicted lipopeptide encoded by a nonribosomal peptide synthetase gene cluster. Our technology, in combination with our recently developed peptidogenomics strategy, enabled the detection and partial characterization of thanamycin and showed that it is a monochlorinated lipopeptide that belongs to the syringomycin family of antifungal agents. In conclusion, the platform presented here provides a significant advancement in our ability to understand the spatiotemporal dynamics of metabolite production in live microbial colonies and communities.

NATURAL PLANT PROTECTION BY 2, 4- DIACETYLPHLOROGLUCINOL PRODUCING PSEUDOMONAS SPP. IN TAKE ALL DECLINE SOILS

Take-all decline (TAD) is a natural biological control of the wheat root disease “take-all” that develops in response to the disease during extended monoculture of wheat. The research to date on TAD has been mostly descriptive and no particular occurrence is yet fully understood. We demonstrate that root-associated fluorescent Pseudomonas spp. producing the antibiotic 2,4-diacetylphloroglucinol (Phl) are key components of the natural biological control that operates in TAD soils in Washington State (U.S.A.). Phl-producing Pseudomonas spp. were present on roots of wheat grown in TAD soils at or above the threshold population density required for significant suppression of take-all of wheat. The specific suppression that operates in TAD soils was lost when Phl-producing fluorescent Pseudomonas spp. were eliminated, and conducive soils gained suppressiveness to take-all when Phl-producing Pseudomonas strains were introduced via mixing in small amounts of TAD soil. Introduction of selected Phl-producing strai...

Comparative Genomics of Plant-Associated Pseudomonas spp.: Insights into Diversity and Inheritance of Traits Involved in Multitrophic Interactions

We provide here a comparative genome analysis of ten strains within the Pseudomonas fluorescens group including seven new genomic sequences. These strains exhibit a diverse spectrum of traits involved in biological control and other multitrophic interactions with plants, microbes, and insects. Multilocus sequence analysis placed the strains in three sub-clades, which was reinforced by high levels of synteny, size of core genomes, and relatedness of orthologous genes between strains within a sub-clade. The heterogeneity of the P. fluorescens group was reflected in the large size of its pan-genome, which makes up approximately 54% of the pan-genome of the genus as a whole, and a core genome representing only 45–52% of the genome of any individual strain. We discovered genes for traits that were not known previously in the strains, including genes for the biosynthesis of the siderophores achromobactin and pseudomonine and the antibiotic 2-hexyl-5-propyl-alkylresorcinol; novel bacteriocins; type II, III, and VI secretion systems; and insect toxins. Certain gene clusters, such as those for two type III secretion systems, are present only in specific sub-clades, suggesting vertical inheritance. Almost all of the genes associated with multitrophic interactions map to genomic regions present in only a subset of the strains or unique to a specific strain. To explore the evolutionary origin of these genes, we mapped their distributions relative to the locations of mobile genetic elements and repetitive extragenic palindromic (REP) elements in each genome. The mobile genetic elements and many strain-specific genes fall into regions devoid of REP elements (i.e., REP deserts) and regions displaying atypical tri-nucleotide composition, possibly indicating relatively recent acquisition of these loci. Collectively, the results of this study highlight the enormous heterogeneity of the P. fluorescens group and the importance of the variable genome in tailoring individual strains to their specific lifestyles and functional repertoire.

Cyclic lipopeptide production by plant-associated Pseudomonas spp : Diversity, activity, biosynthesis, and regulation

Cyclic lipopeptides (CLPs) are versatile molecules produced by a variety of bacterial genera, including plant-associated Pseudomonas spp. CLPs are composed of a fatty acid tail linked to a short oligopeptide, which is cyclized to form a lactone ring between two amino acids in the peptide chain. CLPs are very diverse both structurally and in terms of their biological activity. The structural diversity is due to differences in the length and composition of the fatty acid tail and to variations in the number, type, and configuration of the amino acids in the peptide moiety. CLPs have received considerable attention for their antimicrobial, cytotoxic, and surfactant properties. For plant-pathogenic Pseudomonas spp., CLPs constitute important virulence factors, and pore formation, followed by cell lysis, is their main mode of action. For the antagonistic Pseudomonas sp., CLPs play a key role in antimicrobial activity, motility, and biofilm formation. CLPs are produced via nonribosomal synthesis on large, multifunctional peptide synthetases. Both the structural organization of the CLP synthetic templates and the presence of specific domains and signature sequences within peptide synthetase genes will be described for both pathogenic and antagonistic Pseudomonas spp. Finally, the role of various genes and regulatory mechanisms in CLP production by Pseudomonas spp., including two-component regulation and quorum sensing, will be discussed in detail.