Shifts in the bacterial rhizospheric communities of canola plants in response to seed inoculation with a beneficial endophyte

NIEVA, AMIRA SUSANA; ROSSI FRANCO RUBÉN; SOLMI, LEANDRO; RUÍZ, OSCAR ADOLFO; GARRIZ, ANDRES; ROMERO FERNANDO MATÍAS

Simposio; Young Microbiologists Symposium; 2020

Many endophyticbacteria promote plant growth and protect them against pathogens. We haveisolated and characterized a Pseudomonas viridiflava endophytic strain that colonizesthe apoplast of canola leaves. This isolate showed antagonism against differentphytopathogens, such as Xanthomonascampestris, in vitro and in planta. The mechanisms proposedto be involved include the production of antimicrobial compounds as well as theability to induce defence mechanisms mediated by defense hormones in the host.The aim of this work was to analyse the modifications in the abundance ofrhizospheric bacterial communities in canola plants in response to P. viridiflava inoculation. Total DNAwas purified from rhizospheric soil from seed and mock-inoculated plants. Forbacterial communities, 16S gene libraries were constructed with the universalprimers 341F - 805R and sequenced thought the Illumina MiSeq technology.Pair-end sequences were joined, aligned and clustered into OperationalTaxonomic Units (OTUs) using the open reference strategy. Taxonomic identitywas assigned by the Greengenesdatabase. Singletons, chloroplasts, and mitochondria were filtered out of theanalysis. The OTUs table was subsampled and transformed for the furtherdiversity analysis. First, we measured different indexes regarding communities?diversity, such as richness, Simpson and Shannon for alpha diversity andBray-Curtis or Jaccard for beta diversity, we found no differences on everyindex analysed between control and inoculated samples. Taxonomic composition ofboth communities was similar. The dominant phylum of both, control andinoculated communities were Proteobacteria (33-41%), Actinobacteria (16-25%)and Acidobacteria (10-13%). The evaluation of abundances at phylum level showedthat Choloroflexi, Firmicutes and Actinobacteria were increased in therhizhosphere of P. viridiflava-inoculatedplants. While, the phylum Spirochaetes was diminished in the inoculatedsamples. When we further analysed in depth the phylum Firmicutes we observedthat the order Bacillales was increased in inoculated samples, while the Clostridialeswas reduced. In the case of Actinobacteria, the class Actinobacteria was theone showing changes, particularly the orders Micrococcales and Actinomycetaleswere increased in response to endophyte inoculation. Other phylum broadly involvedin relation with plants is Proteobacteria, which include several plantpathogens, such as X. campestris. Inthis way, we observed that the order Xanthomonadales was decreased in responseto the inoculation with P. viridiflava.On the other hand, we measured by qPCR the abundance of different genesinvolved in nitrogen metabolism, such as nitrogenase (nifH) and nitrous oxidereductase (nosZ), production of DAPG (phlD) and phosphate solubilisation, suchas alkaline phosphatase (phoD). Thus, we observed an increase in abundance ofthe genes nosZ and phoD upon inoculation with the beneficial endophyte, whilethe other genes analysed did not change between treatments. Altogether, ourresults show that inoculation with the beneficial endophyte P. viridiflava did not affect diversityon bacterial communities associated with canola rhizosphere. However, itmodifies the composition of specific taxons that could be involved in thegrowth and defence of the plants host such as Bacillales or Actinomycetales.Moreover, endophyte inoculation induces the accumulation of genes involved innitrogen metabolism and phosphate solubilisation. In conclusion, besides thedirect effect of endophyte inoculation, it could have an indirect effect onplant health and growth by recruiting different microorganisms with beneficialcharacteristics.