Wheat genome architecture influences interactions with phytobeneficial microbial functional groups in the rhizosphere.

Wheat has undergone a complex evolutionary history, which led to allopolyploidization and the hexaploid bread wheat Triticum aestivum. However, the significance of wheat genomic architecture for beneficial plant-microbe interactions is poorly understood, especially from a functional standpoint. In this study, we tested the hypothesis that wheat genomic architecture was an overriding factor determining root recruitment of microorganisms with particular plant-beneficial traits. We chose five wheat species representing genomic profiles AA (Triticum urartu), BB {SS} (Aegilops speltoides), DD (Aegilops tauschii), AABB (Triticum dicoccon) and AABBDD (Triticum aestivum) and assessed by quantitative polymerase chain reaction their ability to interact with free-nitrogen fixers, 1-aminocyclopropane-1-carboxylate deaminase producers, 2,4-diacetylphloroglucinol producers and auxin producers via the phenylpyruvate decarboxylase pathway, in combination with Illumina MiSeq metabarcoding analysis of N fixers (and of the total bacterial community). We found that the abundance of the microbial functional groups could fluctuate according to wheat genomic profile, as did the total bacterial abundance. N fixer diversity and total bacterial diversity were also influenced significantly by wheat genomic profile. Often, rather similar results were obtained for genomes DD (Ae. tauschii) and AABBDD (T. aestivum), pointing for the first time that the D genome could be particularly important for wheat-bacteria interactions.

Genomic content of wheat has a higher influence than plant domestication status on the ability to interact with Pseudomonas plant growth-promoting rhizobacteria.

Plant evolutionary history has had profound effects on belowground traits, which is likely to have impacted the ability to interact with microorganisms, but consequences on root colonization and gene expression by plant growth-promoting rhizobacteria (PGPR) remain poorly understood. Here, we tested the hypothesis that wheat genomic content and domestication are key factors determining the capacity for PGPR interaction. Thus, 331 wheat representatives from eight Triticum or Aegilops species were inoculated under standardized conditions with the generalist PGPR Pseudomonas ogarae F113, using an autofluorescent reporter system for monitoring F113 colonization and expression of phl genes coding for the auxinic inducing signal 2,4-diacetylphloroglucinol. The interaction with P. ogarae F113 was influenced by ploidy level, presence of genomes AA, BB, DD, and domestication. While root colonization was higher for hexaploid and tetraploid species, and phl expression level higher for hexaploid wheat, the diploid Ae. tauschii displayed higher phl induction rate (i.e., expression:colonisation ratio) on roots. However, a better potential of interaction with F113 (i.e., under non-stress gnotobiotic conditions) did not translate, after seed inoculation, into better performance of wheat landraces in non-sterile soil under drought. Overall, results showed that domestication and especially plant genomic content modulate the PGPR interaction potential of wheats.

Ancient wheat varieties have a higher ability to interact with plant growth-promoting rhizobacteria.

Plant interactions with plant growth-promoting rhizobacteria (PGPR) are highly dependent on plant genotype. Modern plant breeding has largely sought to improve crop performance but with little focus on the optimization of plant × PGPR interactions. The interactions of the model PGPR strain Pseudomonas kilonensis F113 were therefore compared in 199 ancient and modern wheat genotypes. A reporter system, in which F113 colonization and expression of 2,4-diacetylphloroglucinol biosynthetic genes (phl) were measured on roots was used to quantify F113 × wheat interactions under gnotobiotic conditions. Thereafter, eight wheat accessions that differed in their ability to interact with F113 were inoculated with F113 and grown in greenhouse in the absence or presence of stress. F113 colonization was linked to improved stress tolerance. Moreover, F113 colonization and phl expression were higher overall on ancient genotypes than modern genotypes. F113 colonization improved wheat performance in the four genotypes that showed the highest level of phl expression compared with the four genotypes in which phl expression was lowest. Taken together, these data suggest that recent wheat breeding strategies have had a negative impact on the ability of the plants to interact with PGPR.

Anthropization level of Lascaux Cave microbiome shown by regional-scale comparisons of pristine and anthropized caves.

Limestone areas across the world develop karstic caves, which are populated by a wide range of macro- and microorganisms. Many of these caves display Paleolithic art or outstanding speleothems, and in the last century they have been subjected to anthropization due to touristic management and intense human frequentation. Despite their cultural importance and associated conservation issues, the impact of anthropization on cave biodiversity is not known. Here, we show that anthropization is associated with specific cave biota modifications. We compared diversity in four pristine caves, four anthropized show caves, and the iconic Lascaux Cave with even stronger anthropization. The predominant microbial higher taxa were the same in all caves, but the most anthropized cave (Lascaux) was unique as it differed from the eight others by a higher proportion of Bacteroidetes bacteria and the absence of Euryarchaeota and Woesearchaeota archaea. Anthropization resulted in lower diversity and altered community structure for bacteria and archaea on cave walls, especially in Lascaux, but with a more limited effect on microeukaryotes and arthropods. Our findings fill a key gap in our understanding of the response of karstic communities to anthropization, by revealing that tourism-related anthropization impacts on the prokaryotic microbiome rather than on eukaryotic residents, and that it shapes cave biota irrespective of cave natural features.

Rhizosphere ecology and phytoprotection in soils naturally suppressive to Thielaviopsis black root rot of tobacco.

Soil suppressiveness to disease is an intriguing emerging property in agroecosystems, with important implications because it enables significant protection of susceptible plants from soil-borne pathogens. Unlike many soils where disease suppressiveness requires crop monoculture to establish, certain soils are naturally suppressive to disease, and this type of specific disease suppressiveness is maintained despite crop rotation. Soils naturally suppressive to Thielaviopsis basicola-mediated black root rot of tobacco and other crops occur in Morens region (Switzerland) and have been studied for over 30 years. In Morens, vermiculite-rich suppressive soils formed on morainic deposits while illite-rich conducive soils developed on sandstone, but suppressiveness is of microbial origin. Antagonistic pseudomonads play a role in black root rot suppressiveness, including Pseudomonas protegens (formerly P. fluorescens) CHA0, a major model strain for research. However, other types of rhizobacterial taxa may differ in prevalence between suppressive and conducive soils, suggesting that the microbial basis of black root rot suppressiveness could be far more complex than solely a Pseudomonas property. This first review on black root rot suppressive soils covers early findings on these soils, the significance of recent results, and compares them with other types of suppressive soils in terms of rhizosphere ecology and plant protection mechanisms.

Root microbiome relates to plant host evolution in maize and other Poaceae.

Prokaryote-eukaryote interactions are primordial, but host selection of its bacterial community remains poorly understood. Because eukaryote evolution affects numerous traits shaping the ecology of their microbiome, we can expect that many evolutionary changes in the former will have the potential to impact on the composition of the latter. Consequently, the more phylogenetically distant the eukaryotic hosts, the more distinct their associated bacterial communities should be. We tested this with plants, by comparing the bacterial communities associated with maize genotypes or other Poaceae. 16S rRNA taxonomic microarray analysis showed that the genetic distance between rhizobacterial communities correlated significantly with the phylogenetic distance (derived from chloroplastic sequences) between Poaceae genotypes. This correlation was also significant when considering specific bacterial populations from all main bacterial divisions, instead of the whole rhizobacterial community. These results indicate that eukaryotic host's evolutionary history can be a significant factor shaping directly the assembly and composition of its associated bacterial compartment.

Frequent, independent transfers of a catabolic gene from bacteria to contrasted filamentous eukaryotes.

Even genetically distant prokaryotes can exchange genes between them, and these horizontal gene transfer events play a central role in adaptation and evolution. While this was long thought to be restricted to prokaryotes, certain eukaryotes have acquired genes of bacterial origin. However, gene acquisitions in eukaryotes are thought to be much less important in magnitude than in prokaryotes. Here, we describe the complex evolutionary history of a bacterial catabolic gene that has been transferred repeatedly from different bacterial phyla to stramenopiles and fungi. Indeed, phylogenomic analysis pointed to multiple acquisitions of the gene in these filamentous eukaryotes-as many as 15 different events for 65 microeukaryotes. Furthermore, once transferred, this gene acquired introns and was found expressed in mRNA databases for most recipients. Our results show that effective inter-domain transfers and subsequent adaptation of a prokaryotic gene in eukaryotic cells can happen at an unprecedented magnitude.

Stochastic and deterministic assembly processes of microbial communities in relation to natural attenuation of black stains in Lascaux Cave.

Community assembly processes are complex and understanding them represents a challenge in microbial ecology. Here, we used Lascaux Cave as a stable, confined environment to quantify the importance of stochastic vs deterministic processes during microbial community dynamics across the three domains of life in relation to an anthropogenic disturbance that had resulted in the side-by-side occurrence of a resistant community (unstained limestone), an impacted community (present in black stains), and a resilient community (attenuated stains). Metabarcoding data showed that the microbial communities of attenuated stains, black stains, and unstained surfaces differed, with attenuated stains being in an intermediate position. We found four scenarios to explain community response to disturbance in stable conditions for the three domains of life. Specifically, we proposed the existence of a fourth, not-documented yet scenario that concerns the always-rare microbial taxa, where stochastic processes predominate even after disturbance but are replaced by deterministic processes during post-disturbance recovery. This suggests a major role of always-rare taxa in resilience, perhaps because they might provide key functions required for ecosystem recovery.IMPORTANCEThe importance of stochastic vs deterministic processes in cave microbial ecology has been a neglected topic so far, and this work provided an opportunity to do so in a context related to the dynamics of black-stain alterations in Lascaux, a UNESCO Paleolithic cave. Of particular significance was the discovery of a novel scenario for always-rare microbial taxa in relation to disturbance, in which stochastic processes are replaced later by deterministic processes during post-disturbance recovery, i.e., during attenuation of black stains.

Natural plant disease suppressiveness in soils extends to insect pest control.

BACKGROUND: Since the 1980s, soils in a 22-km2 area near Lake Neuchâtel in Switzerland have been recognized for their innate ability to suppress the black root rot plant disease caused by the fungal pathogen Thielaviopsis basicola. However, the efficacy of natural disease suppressive soils against insect pests has not been studied. RESULTS: We demonstrate that natural soil suppressiveness also protects plants from the leaf-feeding pest insect Oulema melanopus. Plants grown in the most suppressive soil have a reduced stress response to Oulema feeding, reflected by dampened levels of herbivore defense-related phytohormones and benzoxazinoids. Enhanced salicylate levels in insect-free plants indicate defense-priming operating in this soil. The rhizosphere microbiome of suppressive soils contained a higher proportion of plant-beneficial bacteria, coinciding with their microbiome networks being highly tolerant to the destabilizing impact of insect exposure observed in the rhizosphere of plants grown in the conducive soils. We suggest that presence of plant-beneficial bacteria in the suppressive soils along with priming, conferred plant resistance to the insect pest, manifesting also in the onset of insect microbiome dysbiosis by the displacement of the insect endosymbionts. CONCLUSIONS: Our results show that an intricate soil-plant-insect feedback, relying on a stress tolerant microbiome network with the presence of plant-beneficial bacteria and plant priming, extends natural soil suppressiveness from soilborne diseases to insect pests. Video Abstract.

Effect of clay mineralogy on iron bioavailability and rhizosphere transcription of 2,4-diacetylphloroglucinol biosynthetic genes in biocontrol Pseudomonas protegens.

Pseudomonas strains producing 2,4-diacetylphloroglucinol (DAPG) can protect plants from soilborne phytopathogens and are considered the primary reason for suppressiveness of morainic Swiss soils to Thielaviopsis basicola-mediated black root-rot disease of tobacco, even though they also occur nearby in conducive sandstone soils. The underlying molecular mechanisms accounting for this discrepancy are not understood. In this study, we assessed the hypothesis that the presence of iron-rich vermiculite clay (dominant in suppressive soils) instead of illite (dominant in neighboring conducive soils) translates into higher levels of iron bioavailability and transcription of Pseudomonas DAPG synthetic genes in the tobacco rhizosphere. Rhizosphere monitoring of reporter gene systems pvd-inaZ and phlA-gfp in Pseudomonas protegens indicated that the level of iron bioavailability and the number of cells expressing phl genes (DAPG synthesis), respectively, were higher in vermiculitic than in illitic artificial soils. This was in accordance with the effect of iron on phlA-gfp expression in vitro and, indeed, iron addition to the illitic soil increased the number of cells expressing phlA-gfp. Similar findings were made in the presence of the pathogen T. basicola. Altogether, results substantiate the hypothesis that iron-releasing minerals may confer disease suppressiveness by modulating iron bioavailability in the rhizosphere and expression of biocontrol-relevant genes in antagonistic P. protegens.

Unexpected phytostimulatory behavior for Escherichia coli and Agrobacterium tumefaciens model strains.

Plant-beneficial effects of bacteria are often underestimated, especially for well-studied strains associated with pathogenicity or originating from other environments. We assessed the impact of seed inoculation with the emblematic bacterial models Agrobacterium tumefaciens C58 (plasmid-cured) or Escherichia coli K-12 on maize seedlings in nonsterile soil. Compared with the noninoculated control, root biomass (with A. tumefaciens or E. coli) and shoot biomass (with A. tumefaciens) were enhanced at 10 days for 'PR37Y15' but not 'DK315', as found with the phytostimulator Azospirillum brasilense UAP-154 (positive control). In roots as well as in shoots, Agrobacterium tumefaciens and E. coli triggered similar (in PR37Y15) or different (in DK315) changes in the high-performance liquid chromatography profiles of secondary metabolites (especially benzoxazinoids), distinct from those of Azospirillum brasilense UAP-154. Genome sequence analysis revealed homologs of nitrite reductase genes nirK and nirBD and siderophore synthesis genes for Agrobacterium tumefaciens, as well as homologs of nitrite reductase genes nirBD and phosphatase genes phoA and appA in E. coli, whose contribution to phytostimulation will require experimental assessment. In conclusion, the two emblematic bacterial models had a systemic impact on maize secondary metabolism and resulted in unexpected phytostimulation of seedlings in the Azospirillum sp.-responsive cultivar.

Comparison of prominent Azospirillum strains in Azospirillum-Pseudomonas-Glomus consortia for promotion of maize growth.

Azospirillum are prominent plant growth-promoting rhizobacteria (PGPR) extensively used as phytostimulatory crop inoculants, but only few studies are dealing with Azospirillum-containing mixed inocula involving more than two microorganisms. We compared here three prominent Azospirillum strains as part of three-component consortia including also the PGPR Pseudomonas fluorescens F113 and a mycorrhizal inoculant mix composed of three Glomus strains. Inoculant colonization of maize was assessed by quantitative PCR, transcription of auxin synthesis gene ipdC (involved in phytostimulation) in Azospirillum by RT-PCR, and effects on maize by secondary metabolic profiling and shoot biomass measurements. Results showed that phytostimulation by all the three-component consortia was comparable, despite contrasted survival of the Azospirillum strains and different secondary metabolic responses of maize to inoculation. Unexpectedly, the presence of Azospirillum in the inoculum resulted in lower phytostimulation in comparison with the Pseudomonas-Glomus two-component consortium, but this effect was transient. Azospirillum's ipdC gene was transcribed in all treatments, especially with three-component consortia, but not with all plants and samplings. Inoculation had no negative impact on the prevalence of mycorrhizal taxa in roots. In conclusion, this study brought new insights in the functioning of microbial consortia and showed that Azospirillum-Pseudomonas-Glomus three-component inoculants may be useful in environmental biotechnology for maize growth promotion.

Microscale dynamics of dark zone alterations in anthropized karstic cave shows abrupt microbial community switch.

Strong anthropization of karstic caves may result in formation of various wall alterations including dark zones, whose microbial community differs from that of non-altered surfaces nearby. Dark zones grow quickly and without gradual visual changes, leading to the hypothesis of a simple process rather than complex microbial successions, but this is counter-intuitive as underground microbial changes are typically slow and dark zones are microbiologically very distinct from unmarked surfaces. We tested this hypothesis in Paleolithic Lascaux Cave, across two years of microscale sampling. Indeed, Illumina MiSeq metabarcoding evidenced only three community stages for bacteria, fungi and all microeukaryotes together (i.e. unmarked surfaces, newly-formed dark zones and intermediate/old dark zones) and just two stages for archaea (unmarked surfaces vs dark zones), indicating abrupt community changes. The onset of dark zone formation coincided with the development of Ochroconis fungi, Bacteroidota and the bacterial genera Labrys, Nonomuraea and Sphingomonas, in parallel to Pseudomonas counter-selection. Modeling of community assembly processes highlighted that the dynamics of rare taxa in unmarked surfaces adjacent to dark zones and in newly-formed dark zones were governed in part by deterministic processes. This suggests that cooperative relationships between these taxa might be important to promote dark zone formation. Taken together, these findings indicate an abrupt community switch as these new alterations form on Lascaux cave walls.

Dark-zone alterations expand throughout Paleolithic Lascaux Cave despite spatial heterogeneity of the cave microbiome.

BACKGROUND: Cave anthropization related to rock art tourism can lead to cave microbiota imbalance and microbial alterations threatening Paleolithic artwork, but the underpinning microbial changes are poorly understood. Caves can be microbiologically heterogeneous and certain rock wall alterations may develop in different rooms despite probable spatial heterogeneity of the cave microbiome, suggesting that a same surface alteration might involve a subset of cosmopolitan taxa widespread in each cave room. We tested this hypothesis in Lascaux, by comparing recent alterations (dark zones) and nearby unmarked surfaces in nine locations within the cave. RESULTS: Illumina MiSeq metabarcoding of unmarked surfaces confirmed microbiome heterogeneity of the cave. Against this background, the microbial communities of unmarked and altered surfaces differed at each location. The use of a decision matrix showed that microbiota changes in relation to dark zone formation could differ according to location, but dark zones from different locations displayed microbial similarities. Thus, dark zones harbor bacterial and fungal taxa that are cosmopolitan at the scale of Lascaux, as well as dark zone-specific taxa present (i) at all locations in the cave (i.e. the six bacterial genera Microbacterium, Actinophytocola, Lactobacillus, Bosea, Neochlamydia and Tsukamurella) or (ii) only at particular locations within Lascaux. Scanning electron microscopy observations and most qPCR data evidenced microbial proliferation in dark zones. CONCLUSION: Findings point to the proliferation of different types of taxa in dark zones, i.e. Lascaux-cosmopolitan bacteria and fungi, dark zone-specific bacteria present at all locations, and dark zone-specific bacteria and fungi present at certain locations only. This probably explains why dark zones could form in various areas of the cave and suggests that the spread of these alterations might continue according to the area of distribution of key widespread taxa.

Microbial diversity in soils suppressive to Fusarium diseases.

Fusarium species are cosmopolitan soil phytopathogens from the division Ascomycota, which produce mycotoxins and cause significant economic losses of crop plants. However, soils suppressive to Fusarium diseases are known to occur, and recent knowledge on microbial diversity in these soils has shed new lights on phytoprotection effects. In this review, we synthesize current knowledge on soils suppressive to Fusarium diseases and the role of their rhizosphere microbiota in phytoprotection. This is an important issue, as disease does not develop significantly in suppressive soils even though pathogenic Fusarium and susceptible host plant are present, and weather conditions are suitable for disease. Soils suppressive to Fusarium diseases are documented in different regions of the world. They contain biocontrol microorganisms, which act by inducing plants' resistance to the pathogen, competing with or inhibiting the pathogen, or parasitizing the pathogen. In particular, some of the Bacillus, Pseudomonas, Paenibacillus and Streptomyces species are involved in plant protection from Fusarium diseases. Besides specific bacterial populations involved in disease suppression, next-generation sequencing and ecological networks have largely contributed to the understanding of microbial communities in soils suppressive or not to Fusarium diseases, revealing different microbial community patterns and differences for a notable number of taxa, according to the Fusarium pathosystem, the host plant and the origin of the soil. Agricultural practices can significantly influence soil suppressiveness to Fusarium diseases by influencing soil microbiota ecology. Research on microbial modes of action and diversity in suppressive soils should help guide the development of effective farming practices for Fusarium disease management in sustainable agriculture.

Symbiotic Variations among Wheat Genotypes and Detection of Quantitative Trait Loci for Molecular Interaction with Auxin-Producing Azospirillum PGPR.

Crop varieties differ in their ability to interact with Plant Growth-Promoting Rhizobacteria (PGPR), but the genetic basis for these differences is unknown. This issue was addressed with the PGPR Azospirillum baldaniorum Sp245, using 187 wheat accessions. We screened the accessions based on the seedling colonization by the PGPR and the expression of the phenylpyruvate decarboxylase gene ppdC (for synthesis of the auxin indole-3-acetic acid), using gusA fusions. Then, the effects of the PGPR on the selected accessions stimulating Sp245 (or not) were compared in soil under stress. Finally, a genome-wide association approach was implemented to identify the quantitative trait loci (QTL) associated with PGPR interaction. Overall, the ancient genotypes were more effective than the modern genotypes for Azospirillum root colonization and ppdC expression. In non-sterile soil, A. baldaniorum Sp245 improved wheat performance for three of the four PGPR-stimulating genotypes and none of the four non-PGPR-stimulating genotypes. The genome-wide association did not identify any region for root colonization but revealed 22 regions spread on 11 wheat chromosomes for ppdC expression and/or ppdC induction rate. This is the first QTL study focusing on molecular interaction with PGPR bacteria. The molecular markers identified provide the possibility to improve the capacity of modern wheat genotypes to interact with Sp245, as well as, potentially, other Azospirillum strains.

Two novel species isolated from wheat rhizospheres in Serbia: Pseudomonas serbica sp. nov. and Pseudomonas serboccidentalis sp. nov.

Pseudomonas strains IT-194P, IT-215P, IT-P366T and IT-P374T were isolated from the rhizospheres of wheat grown in soils sampled from different fields (some of them known to be disease-suppressive) located near Mionica, Serbia. Phylogenetic analysis of the 16S rRNA genes and of whole genome sequences showed that these strains belong to two potentially new species, one containing strains IT-P366T and IT-194P and clustering (whole genome analysis) next to P. umsongensis DSM16611T, and another species containing strains IT-P374T and IT-215P and clustering next to P. koreensis LMG21318T. Genome analysis confirmed the proposition of novel species, as ANI was below the threshold of 95% and dDDH below 70% for strains IT-P366T (compared with P. umsongensis DSM16611T) and IT-P374T (compared with P. koreensis LMG21318T). Unlike P. umsongensis DSM16611T, strains of P. serbica can grow on D-mannitol, but not on pectin, D-galacturonic acid, L-galactonic acid lactone and α-hydroxybutyric acid. In contrary to P. koreensis LMG21318T, strains of P. serboccidentalis can use sucrose, inosine and α-ketoglutaric acid (but not L-histidine) as carbon sources. Altogether, these results indicate the existence of two novel species for which we propose the names Pseudomonas serbica sp. nov., with the type strain IT-P366T (=CFBP 9060 T = LMG 32732 T = EML 1791 T) and Pseudomonas serboccidentalis sp. nov., with the type strain IT-P374T (=CFBP 9061 T = LMG 32734 T = EML 1792 T). Strains from this study presented a set of phytobeneficial functions modulating plant hormonal balance, plant nutrition and plant protection, suggesting a potential as Plant Growth-Promoting Rhizobacteria (PGPR).

Microbiome analysis in Lascaux Cave in relation to black stain alterations of rock surfaces and collembola.

Anthropization of Palaeolithic caves open for tourism may favour collembola invasion and result in the formation of black stains attributed to pigmented fungi. However, ecological processes underpinning black stain formation are not fully understood. Here, we tested the hypotheses that black stains from the Apse room of Lascaux Cave display a specific microbiota enriched in pigmented fungi, and that collembola thriving on the stains have the potential to consume and disseminate these black fungi. Metabarcoding showed that the microbiota of black stains and neighbouring unstained parts strongly differed, with in black stains a higher prevalence of Ochroconis and other pigmented fungi and the strong regression of Pseudomonas bacteria (whose isolates inhibited in vitro the growth of pigmented fungi). Isotopic analyses indicated that Folsomia candida collembola thriving on stains could feed on black stain in situ and assimilate the pigmented fungi they were fed with in vitro. They could carry these fungi and disseminate them when tested with complex black stains from Lascaux. This shows that black stain formation is linked to the development of pigmented fungi, which coincides with the elimination of antagonistic pseudomonads, and points towards a key role of F. candida collembola in the dynamics of pigmented fungi.

Is diversification history of maize influencing selection of soil bacteria by roots?

A wide range of plant lines has been propagated by farmers during crop selection and dissemination, but consequences of this crop diversification on plant-microbe interactions have been neglected. Our hypothesis was that crop evolutionary history shaped the way the resulting lines interact with soil bacteria in their rhizospheres. Here, the significance of maize diversification as a factor influencing selection of soil bacteria by seedling roots was assessed by comparing rhizobacterial community composition of inbred lines representing the five main genetic groups of maize, cultivated in a same European soil. Rhizobacterial community composition of 21-day-old seedlings was analysed using a 16S rRNA taxonomic microarray targeting 19 bacterial phyla. Rhizobacterial community composition of inbred lines depended on the maize genetic group. Differences were largely due to the prevalence of certain Betaproteobacteria and especially Burkholderia, as confirmed by quantitative PCR and cloning/sequencing. However, these differences in bacterial root colonization did not correlate with plant microsatellite genetic distances between maize genetic groups or individual lines. Therefore, the genetic structure of maize that arose during crop diversification (resulting in five main groups), but not the extent of maize diversification itself (as determined by maize genetic distances), was a significant factor shaping rhizobacterial community composition of seedlings.

Evolutionary history of synthesis pathway genes for phloroglucinol and cyanide antimicrobials in plant-associated fluorescent pseudomonads.

Plant-beneficial fluorescent Pseudomonas spp. play important ecological roles. Here, their evolutionary history was investigated by a multilocus approach targeting genes involved in synthesis of secondary antimicrobial metabolites implicated in biocontrol of phytopathogens. Some of these genes were proposed to be ancestral, and this was investigated using a worldwide collection of 30 plant-colonizing fluorescent pseudomonads, based on phylogenetic analysis of 14 loci involved in production of 2,4-diacetylphloroglucinol (phlACBDE, phlF, intergenic locus phlA/phlF), hydrogen cyanide (hcnABC, anr) or global regulation of secondary metabolism (gacA, gacS, rsmZ). The 10 housekeeping loci rrs, dsbA, gyrB, rpoD, fdxA, recA, rpoB, rpsL, rpsG, and fusA served as controls. Each strain was readily distinguished from the others when considering allelic combinations for these 14 biocontrol-relevant loci. Topology comparisons based on Shimodaira-Hasegawa tests showed extensive incongruence when comparing single-locus phylogenetic trees with one another, but less when comparing (after sequence concatenation) trees inferred for genes involved in 2,4-diacetylphloroglucinol synthesis, hydrogen cyanide synthesis, or secondary metabolism global regulation with trees for housekeeping genes. The 14 loci displayed linkage disequilibrium, as housekeeping loci did, and all 12 protein-coding loci were subjected to purifying selection except for one positively-selected site in HcnA. Overall, the evolutionary history of Pseudomonas genes involved in synthesis of secondary antimicrobial metabolites important for biocontrol functions is in fact similar to that of housekeeping genes, and results suggest that they are ancestral in pseudomonads producing hydrogen cyanide and 2,4-diacetylphloroglucinol.