Plant growth-promoting rhizobacteria (PGPR) increase plant growth and crop productivity. The inoculation of plants with a bacterial mixture (consortium) apparently provides greater benefits to plant growth than inoculation with a single bacterial strain. In the present work, a bacterial consortium was formulated containing four compatible and desiccation-tolerant strains with potential as PGPR. The formulation had one moderately (Pseudomonas putida KT2440) and three highly desiccation-tolerant (Sphingomonas sp. OF178, Azospirillum brasilense Sp7 and Acinetobacter sp. EMM02) strains. The four bacterial strains were able to adhere to seeds and colonize the rhizosphere of plants when applied in both mono-inoculation and multi-inoculation treatments, showing that they can also coexist without antagonistic effects in association with plants. The effects of the bacterial consortium on the growth of blue maize were evaluated. Seeds inoculated with either individual bacterial strains or the bacterial consortium were subjected to two experimental conditions before sowing: normal hydration or desiccation. In general, inoculation with the bacterial consortium increased the shoot and root dry weight, plant height and plant diameter compared to the non-inoculated control or mono-inoculation treatments. The bacterial consortium formulated in this work had greater benefits for blue maize plants even when the inoculated seeds underwent desiccation stress before germination, making this formulation attractive for future field applications.

The potential of Pseudomonas putida KT2440 to act as a plant-growth promoter or as a bioremediator of toxic compounds can be affected by desiccation. In the present work, the bacterial survival ratio (BSR) in response to air desiccation was evaluated for P . putida KT2440 in the presence of different protectors. The BSR in the presence of nonreducing disaccharides, such as trehalose, was high after 15 days of desiccation stress (occurring at 30°C and 50% relative humidity), whereas in the absence of a protector the bacterial counts diminished to nondetectable numbers (ca 2.8 log CFU/mL). The LIVE/DEAD staining method showed that bacteria protected with trehalose maintained increased numbers of green cells after desiccation while cells without protection were all observed to be red. This indicated that nonprotected bacteria had compromised membrane integrity. However, when nonprotected bacteria subjected to 18 days of desiccation stress were rehydrated for a short time with maize root exudates or for 48 h with water (prolonged rehydration), the bacterial counts were as high as that observed for those not subjected to desiccation stress, suggesting that the cells entered the viable but nonculturable (VBNC) state under desiccation and that they returned to a culturable state after those means of rehydration. Interestingly an increase in the green color intensity of cells that returned to a culturable state was observed using LIVE/DEAD staining method, indicating an improvement in their membrane integrity. Cellular activity in the VBNC state was determined. A GFP-tagged P . putida strain expressing GFP constitutively was subjected to desiccation. After 12 days of desiccation, the GFP-tagged strain lost culturability, but it exhibited active GFP expression, which in turn made the cells green. Furthermore, the expression of 16S rRNA, rpoN (housekeeping), mutL , mutS (encoding proteins from the mismatch repair complex), and oprH (encoding an outer membrane protein) were examined by RT-PCR. All evaluated genes were expressed by both types of cells, culturable and nonculturable, indicating active molecular processes during the VBNC state.

It has been established that a decrease in the population of Gluconacetobacter diazotrophicus associated with sugarcane occurs after nitrogen fertilization. This fact could be due to a direct influence of NH(4)NO(3) on bacterial cells or to changes in plant physiology after fertilizer addition, affecting bacterial establishment. In this work, we observed that survival of G. diazotrophicus was directly influenced when 44.8mM of NH(4)NO(3) (640mgN/plant) was used for in vitro experiments. Furthermore, micropropagated sugarcane plantlets were inoculated with G. diazotrophicus and used for split root experiments, in which both ends of the system were fertilized with a basal level of NH(4)NO(3) (0.35mM; 10mgN/plant). Twenty days post inoculation (dpi) one half of the plants were fertilized with a high dose of NH(4)NO(3) (6.3mM; 180 mgN/plant) on one end of the system. This nitrogen level was lower than that directly affecting G. diazotrophicus cells; however, it caused a decrease in the bacterial population in comparison with control plants fertilized with basal nitrogen levels. The decrease in the population of G. diazotrophicus was higher in pots fertilized with a basal nitrogen level when compared with the corresponding end supplied with high levels of NH4NO3 (100dpi; 80 days post fertilization) of the same plant system. These observations suggest that the high nitrogen level added to the plants induce systemic physiological changes that affect the establishment of G. diazotrophicus.

Here we describe two methods for assessing the ability of one bacteria to inhibit the growth of another through the production of antimicrobial compounds. In the first assay, the double agar layer method, only the secreted substances but not the candidate antagonist bacteria interact with the sensitive microorganisms. In the second assay, the simultaneous inhibition method, both microorganisms are grown together allowing the competitive exclusion.

Introduction: Several plant-beneficial bacteria have the capability to promote the growth of plants through different mechanisms. The survival of such bacteria could be affected by environmental abiotic factors compromising their capabilities of phytostimulation. One of the limiting abiotic factors is low water availability. Materials and Methods: In extreme cases, bacterial cells can suffer desiccation, which triggers harmful effects on cells. Bacteria tolerant to desiccation have developed different strategies to cope with these conditions; however, the genes involved in these processes have not been sufficiently explored. Klebsiella variicola T29A is a beneficial bacterial strain that promotes the growth of corn plants and is highly tolerant to desiccation. In the present work, we investigated genes involved in desiccation tolerance. Results & Discussion: As a result, a library of 8974 mutants of this bacterial strain was generated by random mutagenesis with mini-Tn5 transposon, and mutants that lost the capability to tolerate desiccation were selected. We found 14 sensitive mutants; those with the lowest bacterial survival rate contained mini-Tn5 transposon inserted into genes encoding a protein domain related to BetR, putative secretion ATPase and dihydroorotase. The mutant in the betR gene had the lowest survival; therefore, the mutagenized gene was validated using specific amplification and sequencing. Conclusion: Trans complementation with the wild-type gene improved the survival of the mutant under desiccation conditions, showing that this gene is a determinant for the survival of K. variicola T29A under desiccation conditions.