RNA stable isotope probing and high-throughput sequencing were used to characterize the active microbiomes of bacteria and fungi colonizing the roots and rhizosphere soil of oilseed rape to identify taxa assimilating plant-derived carbon following CO labeling. Root- and rhizosphere soil-associated communities of both bacteria and fungi differed from each other, and there were highly significant differences between their DNA- and RNA-based community profiles. ,, ,, ,, and were the most active bacterial phyla in the rhizosphere soil. were more active in roots. The most abundant bacterial genera were well represented in both the C- andC-RNA fractions, while the fungal taxa were more differentiated. ,, and were dominant in roots, whereas and () were dominant in rhizosphere soil. " Nitrososphaera" was enriched in C in rhizosphere soil. and were abundant in theC-RNA fraction of roots; was abundant in both roots and rhizosphere soil and heavilyC enriched. was dominant in rhizosphere soil and less abundant, but wasC enriched in roots. The patterns of colonization and C acquisition revealed in this study assist in identifying microbial taxa that may be superior competitors for plant-derived carbon in the rhizosphere of This microbiome study characterizes the active bacteria and fungi colonizing the roots and rhizosphere soil of using high-throughput sequencing and RNA-stable isotope probing. It identifies taxa assimilating plant-derived carbon following CO labeling and compares these with other less active groups not incorporating a plant assimilate. is an economically and globally important oilseed crop, cultivated for edible oil, biofuel production, and phytoextraction of heavy metals; however, it is susceptible to several diseases. The identification of the fungal and bacterial species successfully competing for plant-derived carbon, enabling them to colonize the roots and rhizosphere soil of this plant, should enable the identification of microorganisms that can be evaluated in more detailed functional studies and ultimately be used to improve plant health and productivity in sustainable agriculture.

Oilseed rape (Brassica napus) is one of the major oilseed crops in the world but is vulnerable to attack by many pathogens and insect pests. In addition to the host plant genotype, micro-organisms present in the rhizosphere and within plant tissues affect the susceptibility to plant pathogens. While rapid progress has been achieved concerning the concept of plant resistance genes, information on the role of the microbial community in plant protection is less apparent. We have studied the endophytic bacterial populations present in different tissues of oilseed rape and also analysed several cultivars (Express, Libraska, Maluka and Westar), which differ in their susceptibility to the wilt pathogen Verticillium longisporum. The population diversity was studied using agar plating assay, fatty acid methyl ester analysis and functional characterisation of isolated strains. Our work shows that already in the seeds there exists diversity in populations as well as in the total microbial load between two of the four tested cultivars. About 50% of the strains isolated from cultivars Express and Libraska showed moderate to strong direct inhibition of V. longisporum. The diversity of the endophytic flora isolated from oilseed rape and its implications in crop protection are discussed.

BackgroundImproved understanding of bacterial-fungal interactions in the rhizosphere should assist in the successful application of bacteria as biological control agents against fungal pathogens of plants, providing alternatives to chemicals in sustainable agriculture. Rhizoctonia solani is an important soil-associated fungal pathogen and its chemical treatment is not feasible or economic. The genomes of the plant-associated bacteria Serratia proteamaculans S4 and Serratia plymuthica AS13 have been sequenced, revealing genetic traits that may explain their diverse plant growth promoting activities and antagonistic interactions with R. solani. To understand the functional response of this pathogen to different bacteria and to elucidate whether the molecular mechanisms that the fungus exploits involve general stress or more specific responses, we performed a global transcriptome profiling of R. solani Rhs1AP anastomosis group 3 (AG-3) during interaction with the S4 and AS13 species of Serratia using RNA-seq.ResultsApproximately 104,504 million clean 75-100 bp paired-end reads were obtained from three libraries, each in triplicate (AG3-Control, AG3-S4 and AG3-AS13). Transcriptome analysis revealed that approximately 10 % of the fungal transcriptome was differentially expressed during challenge with Serratia. The numbers of S4- and AS13-specific differentially expressed genes (DEG) were 866 and 292 respectively, while there were 1035 common DEGs in the two treatment groups. Four hundred and sixty and 242 genes respectively had values of log2 fold-change > 3 and for further analyses this cut-off value was used. Functional classification of DEGs based on Gene Ontology enrichment analysis and on KEGG pathway annotations revealed a general shift in fungal gene expression in which genes related to xenobiotic degradation, toxin and antioxidant production, energy, carbohydrate and lipid metabolism and hyphal rearrangements were subjected to transcriptional regulation.ConclusionsThis RNA-seq profiling generated a novel dataset describing the functional response of the phytopathogen R. solani AG3 to the plant-associated Serratia bacteria S4 and AS13. Most genes were regulated in the same way in the presence of both bacterial isolates, but there were also some strain-specific responses. The findings in this study will be beneficial for further research on biological control and in depth exploration of bacterial-fungal interactions in the rhizosphere.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1758-z) contains supplementary material, which is available to authorized users.

The potential of bacteria that are adapted to the oilseed rape root environment for use in the biological control of Verticillium dahliae, Kleb was investigated in both controlled and non-sterile growth conditions. Bacterial strains dominated by the red-pigmented members of enterobacteriaceae were isolated from thoroughly washed and air-dried root segments of symptomless young rape plants. Other associated strains found either belonged to Alcaligenes sp., Stenotrophomonas spp. and Pseudomonas spp. (Pseudomonas acidovorans and Pseudomonas putida) or were unidenti®ed according to fatty acid methyl ester pro®le analysis. A total of 19 strains isolated in this study together with two previously studied strains, Serratia proteamaculans and Pseudomonas chlororaphis, were characterized on the basis of their interactions with V. dahliae and a number of functional characteristics. In line with earlier observations with root-colonizing fungi also from oilseed rape, all bacterial strains suppressed the pathogen not only directly and but also indirectly in in vitro assays. Mechanisms of suppression were apparently multifold among the strains, but production of hydrogen cyanide does not seem to be involved in indirect inhibition. The majority of the strains possessed the ability to produce cellulases, proteases and phosphatases and some even produced chitinases and induced hypersensitive responses, indicating their potential for nutrient acquisition as well as colonization capacity and active recognition by the plant cells. Investigations in non-sterile ®eld soil revealed that some strains protected rape plants from V. dahliae partly by delaying symptom development. None of the strains, however, was strongly deleterious to rape growth either in the presence or absence of the pathogen. Light microscopic observations of roots and results based on agar printing techniques revealed the potential of the studied strains to colonize or interfere with the pathogen colonization. This study provides some insight into the evolved relationship of bacterial residents with their host in terms of their potential importance in its ®tness. ZusammenfassungMerkmale von Raps isolierter Bakterien in bezug auf ihr Potential zur biologischen BekaÈ mpfung von Verticillium dahliae An den Lebensraum`Rapswurzel' angepaûte Bakterien wurden unter kontrollierten und unter unsterilen Bedingungen hinsichtlich ihres Potentials zur biologischen BekaÈ mpfung von Verticillium dahliae Kleb. gepruÈ ft. Von sorgsam gewaschenen und an der Luft getrockneten Wurzelsegmenten symptomloser junger Rapsp¯anzen wurden BakterienstaÈ mme isoliert, unter denen rot pigmentierte Enterobacteriaceae dominierten. Eine Analyse der FettsaÈ urenmethylesterpro®le ergab, daû die anderen assoziierten StaÈ mme entweder zu Alcaligenes sp., Stenotrophomonas spp. und Pseudomonas spp. (P. acidovorans und P. putida) gehoÈ rten oder aber mit diesem Verfahren nicht identi®ziert werden konnten. Im Rahmen dieser Untersuchung wurden insgesamt 19 StaÈ mme isoliert, die gemeinsam mit zwei zuvor unte...

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