Introducing THOR, a model microbiome for genetic dissection of community behavior

The quest to manipulate microbiomes has intensified, but many microbial communities have proven recalcitrant to sustained change. Developing model communities amenable to genetic dissection will underpin successful strategies for shaping microbiomes by advancing understanding of community interactions. We developed a model community with representatives from three dominant rhizosphere taxa: the Firmicutes, Proteobacteria, and Bacteroidetes. We chose Bacillus cereus as a model rhizosphere Firmicute and characterized twenty other candidates, including hitchhikers that co-isolated with B. cereus from the rhizosphere. Pairwise analysis produced a hierarchical interstrain-competition network. We chose two hitchhikers - Pseudomonas koreensis from the top tier of the competition network and Flavobacterium johnsoniae from the bottom of the network to represent the Proteobacteria and Bacteroidetes, respectively. The model community has several emergent properties - induction of dendritic expansion of B. cereus colonies by either of the other members and production of more robust biofilms by the three members together than individually. Moreover, P. koreensis produces a novel family of alkaloid antibiotics that inhibit growth of F. johnsoniae, and production is inhibited by B. cereus. We designate this community THOR, because the members are the hitchhikers of the rhizosphere. The genetic, genomic, and biochemical tools available for dissection of THOR provide the means to achieve a new level of understanding of microbial community behavior.

The Pseudomonas aeruginosa T6SS-VgrG1b spike is topped by a PAAR protein eliciting DNA damage to bacterial competitors

The type VI secretion system (T6SS) is a supramolecular complex involved in the delivery of potent toxins during bacterial competition. Pseudomonas aeruginosa possesses three T6SS gene clusters and several hcp and vgrG gene islands, the latter encoding the spike at the T6SS tip. The vgrG1b cluster encompasses seven genes whose organization and sequences are highly conserved in P. aeruginosa genomes, except for two genes that we called tse7 and tsi7. We show that Tse7 is a Tox-GHH2 domain nuclease which is distinct from other T6SS nucleases identified thus far. Expression of this toxin induces the SOS response, causes growth arrest and ultimately results in DNA degradation. The cytotoxic domain of Tse7 lies at its C terminus, while the N terminus is a predicted PAAR domain. We find that Tse7 sits on the tip of the VgrG1b spike and that specific residues at the PAAR–VgrG1b interface are essential for VgrG1b-dependent delivery of Tse7 into bacterial prey. We also show that the delivery of Tse7 is dependent on the H1-T6SS cluster, and injection of the nuclease into bacterial competitors is deployed for interbacterial competition. Tsi7, the cognate immunity protein, protects the producer from the deleterious effect of Tse7 through a direct protein–protein interaction so specific that toxin/immunity pairs are effective only if they originate from the same P. aeruginosa isolate. Overall, our study highlights the diversity of T6SS effectors, the exquisite fitting of toxins on the tip of the T6SS, and the specificity in Tsi7-dependent protection, suggesting a role in interstrain competition.

Competition for nodule occupancy of introduced Bradyrhizobium japonicum strain SMGS1 in French soils already containing Bradyrhizobium japonicum strain G49

The competitiveness of Bradyrhizobium japonicum strains G49 and SMGS1 was first studied in the greenhouse in sterilized sand, with or without added soil. Strain SMGS1 was more competitive than strain G49 with soybean (Glycine max L.) cultivar Labrador but the two strains showed equivalent competitiveness with cultivar Kingsoy. When soil was added, nodule occupancy of strain G49 was only 22% with this cultivar. In field experiments, conducted over 2 years in soils already containing strain G49 (1.5 × 103 to 4.0 × 104 cells/g of soil), nodule occupancy of inoculated strain SMGS1 ranged from 20 to 90%. Nodule occupancy was 3–22% higher when inoculation was done by peat seed coating or with liquid inoculation in the row than with peat-coated clay microgranulars. Nodule occupancy was also dependent on the physiological state of the inoculated cells. When an inoculum stored at 28 °C for 1 year was used at the same viable cell rate, nodule occupancy of strain SMGS1 was 4–20% lower than with a recently made inoculum. Pot experiments with soil from field experiments carried out in the 1st year showed that the inoculated strain continued forming nodules without further inoculation, with a recovery rate equivalent to that of field experiment in the previous year.Key words: Bradyrhizobium japonicum, interstrain competition, inoculation technology, ELISA, field trials.

Environmental effects on competition for nodule occupancy between introduced and indigenous rhizobia and among introduced strains

Understanding the impact of environmental variables on interstrain competition is important to ensure the successful use of rhizobial inoculant. In eight inoculation trials conducted at five diverse sites on Maui, Hawaii, equal numbers of three serologically distinct strains of effective, homologous rhizobia in a peat-based inoculant were applied to seeds of soybean, bush bean, cowpea, lima bean, peanut, leucaena, clover, and tinga pea. We studied the influence of environmental variables on interstrain competition between applied and indigenous rhizobia and among the three strains comprising the inoculum. Although temperature and soil fertility were correlated with nodule occupancy by inoculant strains in a few cases, the most significant environmental variable controlling their competitive success was the size of the indigenous rhizobial population. Nodule occupancy was best described (r2 = 0.51, p < 0.001) by the equation y = 97.88 − 15.03(log10(x + 1)), where y is percent nodule occupancy by inoculant rhizobia and x is the most probable number of indigenous rhizobia per gram soil. For each legume, one of the three inoculant strains was a poor competitor across sites. Competition between the other two strains varied between sites, but was infrequently related to environmental variables. Results indicated that competitive strains could be selected that perform well across a range of environments. Key words: competition, rhizobial ecology, inoculation response, competitiveness index.