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Rhizobial inoculants are sold either as rhizobia within a liquid matrix; or as rhizobia adhered to granules composed of peat prill or finely ground peat moss. During the production of peat-based inoculants, immediately after mixing the rhizobia culture with partially dry sterile peat, the inoculant is stored for a period of 4-5 weeks, inducing a series of changes that results in an increased capability of the rhizobia to survive in the seeds. The number of viable rhizobia on preinoculated seeds at the point of sale, however, is often a limiting factor, as is the inefficiency of the inoculant bacteria to compete with the local rhizobia for the host colonization. In the present work, we used STM-seq for the genomewide screening of Ensifer meliloti mutants affected in the survival during the maturation of peat-based inoculant formulations. Through this approach, we identified hundreds of genes that proved to be relevant to this process. These results also provide a base knowledge that could be used to more completely understand the survival mechanisms used by rhizobia during the maturation of peat-based inoculants, as well as for the design of new inoculant formulations.HighlightsRhizobial inoculants provide an ecological means of nitrogen fertilization compatible with the implementation of sustainable agricultural practices. Their successful usage, however, suffers from two main limitations: the low number of viable rhizobia on preinoculated seeds at the point of sale, and the inefficiency to compete with the local rhizobia for host colonization. Here, we used a high-throughput mutant-screening technology, STM-seq, to uncover which rhizobial genes are involved in the rhizobial survival during the preparation and storage of peat-based inoculant formulations. Our findings provide useful information about the stresses faced by rhizobia during peat-inoculant maturation and storage, which could assist both for the selection of better rhizobial strains, and for the improvement of the inoculant formulations.Graphical Abstract

The acidity of soils significantly reduces the productivity of legumes mainly because of the detrimental effects of hydrogen ions on the legume plants, leading to the establishment of an inefficient symbiosis and poor biological nitrogen fixation. We recently reported the analysis of the fully sequenced genome of Rhizobium favelukesii LPU83, an alfalfa-nodulating rhizobium with a remarkable ability to grow, nodulate and compete in acidic conditions. To gain more insight into the genetic mechanisms leading to acid tolerance in R. favelukesii LPU83, we constructed a transposon mutant library and screened for mutants displaying a more acid-sensitive phenotype than the parental strain. We identified mutant Tn833 carrying a single-transposon insertion within LPU83_2531, an uncharacterized short ORF located immediately upstream from ubiF homolog. This gene encodes a protein with an enzymatic activity involved in the biosynthesis of ubiquinone. As the transposon was inserted near the 3′ end of LPU83_2531 and these genes are cotranscribed as a part of the same operon, we hypothesized that the phenotype in Tn833 is most likely due to a polar effect on ubiF transcription. We found that a mutant in ubiF was impaired to grow at low pH and other abiotic stresses including 5 mM ascorbate and 0.500 mM Zn(2+). Although the ubiF mutant retained the ability to nodulate alfalfa and Phaseolus vulgaris, it was unable to compete with the R. favelukesii LPU83 wild-type strain for nodulation in Medicago sativa and P. vulgaris, suggesting that ubiF is important for competitiveness. Here, we report for the first time an ubiF homolog being essential for nodulation competitiveness and tolerance to specific stresses in rhizobia. GRAPHICAL ABSTRACT: [Image: see text]

Numerous therapies have been implemented to treat intermittent exotropia, including pharmacological penalization, minus lenses, orthoptics and visual training, prisms and surgery. We have been impr...

The biology and biochemistry of plasmid transfer in soil bacteria is currently under active investigation because of its central role in prokaryote adaptation and evolution. In this work, we examined the conjugal properties of the cryptic plasmids present in a collection of the N(2)-fixing legume-symbiont Sinorhizobium meliloti. The study was performed on 65 S. meliloti isolates recovered from 25 humic soils of Argentina, which were grouped into 22 plasmid-profile types [i.e. plasmid operational taxonomic units (OTUs)]. The cumulative Shannon index calculated for the observed plasmid profiles showed a clear saturation plateau, thus indicating an adequate representation of the S. meliloti plasmid-profile types in the isolates studied. The results show that isolates of nearly 14% of the plasmid OTUs hosted transmissible plasmids and that isolates of 29% of the plasmid OTUs were able to retransfer the previously characterized mobilizable-cryptic plasmid pSmeLPU88b to a third recipient strain. It is noteworthy that isolates belonging to 14% of the plasmid OTUs proved to be refractory to the entrance of the model plasmid pSmeLPU88b, suggesting either the presence of surface exclusion phenomena or the occurrence of restriction incompatibility with the incoming replicon. Incompatibility for replication between resident plasmids and plasmid pSmeLPU88b was observed in c. 20% of the OTUs. The results reported here reveal a widespread compatibility among the conjugal functions of the cryptic plasmids in S. meliloti, and this fact, together with the observed high proportion of existing donor genotypes, points to the extrachromosomal compartment of the species as being an extremely active plasmid mobilome.