Bacillus cereus MH778713 elicits tomato plant protection against Fusarium oxysporum.

AIM: The genus Fusarium comprises plant pathogenic species with agricultural relevance. Fusarium oxysporum causes tomato wilt disease with significant production losses. The use of agrochemicals to control the Fusarium wilt of tomato is not environmentally friendly. Bacillus species, as biocontrol agents, provide a safe and sustainable means to control Fusarium-induced plant diseases. In this study, the ability of Bacillus cereus MH778713, a strain isolated from root nodules of Prosopis laevigata, to protect tomato plants against Fusarium wilt was evaluated. METHODS AND RESULTS: Bacillus cereus MH778713 and its volatiles inhibited the radial growth of F. oxysporum and stimulated tomato seedling growth in in vitro and in vivo tests. When tomato plants growing in the greenhouse were inoculated with B. cereus MH778713, the percentage of wilted plants decreased from 96% to 12%, indicating an effective crop protection against Fusarium wilt. Among the metabolites produced by B. cereus MH778713, hentriacontane and 2,4-di-tert-butylphenol promoted tomato seedling growth and showed antifungal activity against the target pathogen. CONCLUSION: The inoculation of B. cereus MH778713 on tomato seedlings helped plants to manage Fusarium wilt, suggesting the potential of B. cereus MH778713 as a biocontrol agent. SIGNIFICANCE AND IMPACT OF THE STUDY: These results complement our previous studies on chromium tolerance and bioremediation traits of B. cereus MH778713 by highlighting the potential of this metal-resistant micro-organism to boost crop growth and disease resistance.

Diversity and phenotypic analyses of salt- and heat-tolerant wild bean Phaseolus filiformis rhizobia native of a sand beach in Baja California and description of Ensifer aridi sp. nov.

In northern Mexico, aridity, salinity and high temperatures limit areas that can be cultivated. To investigate the nature of nitrogen-fixing symbionts of Phaseolus filiformis, an adapted wild bean species native to this region, their phylogenies were inferred by MLSA. Most rhizobia recovered belong to the proposed new species Ensifer aridi. Phylogenetic analyses of nodC and nifH show that Mexican isolates carry symbiotic genes acquired through horizontal gene transfer that are divergent from those previously characterized among bean symbionts. These strains are salt tolerant, able to grow in alkaline conditions, high temperatures, and capable of utilizing a wide range of carbohydrates and organic acids as carbon sources for growth. This study improves the knowledge on diversity, geographic distribution and evolution of bean-nodulating rhizobia in Mexico and further enlarges the spectrum of microsymbiont with which Phaseolus species can interact with, including cultivated bean varieties, notably under stressed environments. Here, the species Ensifer aridi sp. nov. is proposed as strain type of the Moroccan isolate LMR001T (= LMG 31426T; = HAMBI 3707T) recovered from desert sand dune.

Genomic characterization of Ensifer aridi, a proposed new species of nitrogen-fixing rhizobium recovered from Asian, African and American deserts.

BACKGROUND: Nitrogen fixing bacteria isolated from hot arid areas in Asia, Africa and America but from diverse leguminous plants have been recently identified as belonging to a possible new species of Ensifer (Sinorhizobium). In this study, 6 strains belonging to this new clade were compared with Ensifer species at the genome-wide level. Their capacities to utilize various carbon sources and to establish a symbiotic interaction with several leguminous plants were examined. RESULTS: Draft genomes of selected strains isolated from Morocco (Merzouga desert), Mexico (Baja California) as well as from India (Thar desert) were produced. Genome based species delineation tools demonstrated that they belong to a new species of Ensifer. Comparison of its core genome with those of E. meliloti, E. medicae and E. fredii enabled the identification of a species conserved gene set. Predicted functions of associated proteins and pathway reconstruction revealed notably the presence of transport systems for octopine/nopaline and inositol phosphates. Phenotypic characterization of this new desert rhizobium species showed that it was capable to utilize malonate, to grow at 48 °C or under high pH while NaCl tolerance levels were comparable to other Ensifer species. Analysis of accessory genomes and plasmid profiling demonstrated the presence of large plasmids that varied in size from strain to strain. As symbiotic functions were found in the accessory genomes, the differences in symbiotic interactions between strains may be well related to the difference in plasmid content that could explain the different legumes with which they can develop the symbiosis. CONCLUSIONS: The genomic analysis performed here confirms that the selected rhizobial strains isolated from desert regions in three continents belong to a new species. As until now only recovered from such harsh environment, we propose to name it Ensifer aridi. The presented genomic data offers a good basis to explore adaptations and functionalities that enable them to adapt to alkalinity, low water potential, salt and high temperature stresses. Finally, given the original phylogeographic distribution and the different hosts with which it can develop a beneficial symbiotic interaction, Ensifer aridi may provide new biotechnological opportunities for degraded land restoration initiatives in the future.

Long-Chain Hydrocarbons (C21, C24, and C31) Released by Bacillus sp. MH778713 Break Dormancy of Mesquite Seeds Subjected to Chromium Stress.

Volatile organic compounds (VOCs) produced by rhizobacteria have been proven to stimulate plant growth during germination and seedling stages. However, the modulating effect of bacterial volatiles on the germination of seeds subjected to heavy metal stress is scarcely studied. In this work, the ability of volatiles released by Bacillus sp. MH778713 to induce seed dormancy breakage in Prosopis laevigata and Arabidopsis thaliana seeds were examined. The minimal inhibitory concentration of chromium (Cr) VI that prevents seed germination of P. laevigata and A. thaliana on water-Cr-agar plates was 2500 and 100 mg L-1, respectively. Remarkably, partitioned Petri-dish co-cultivation of Bacillus sp. MH778713 and plant seeds under Cr-stress showed the beneficial effect of volatiles emitted by Bacillus sp. MH778713, helping plant seeds to overcome Cr-stress. Among the metabolites emitted by Bacillus sp. MH778713, octadecane, heneicosane, 2,4-di-tert-butylphenol, hexadecane, eicosane, octacosane, and tetratriacontane were the most abundant. To confirm that these long-chain compounds produced by Bacillus sp. MH778713 could be responsible for the seed dormancy breakage, high pure organic compounds (2,4-di-tert-butylphenol, heneicosane, hentriacontane, and tetracosane) were used directly in germination assays of P. laevigata and A. thaliana seeds instead of volatiles emitted by Bacillus sp. MH778713. All organic compounds allowed Prosopis and Arabidopsis seeds to overcome Cr-toxicity and germinate. The results of this study provide new insight into the role of long-chain bacterial compounds produced by Bacillus sp. MH778713 as triggers of seed abiotic stress tolerance, surmounting chromium stress and stimulating seedling development.

Chromium Hyper-Tolerant Bacillus sp. MH778713 Assists Phytoremediation of Heavy Metals by Mesquite Trees (Prosopis laevigata).

Heavy metal accumulation in mesquite trees (Prosopis laevigata) growing in aluminum, titanium, chromium and zirconium-polluted soils of a semi-arid region in Mexico was investigated using wavelength dispersive X-ray fluorescence analysis. The results showed that P. laevigata trees can hyper accumulate up to 4100 mg/kg of Al, 14000 mg/kg of Fe, 1600 mg/kg of Ti, 2500 mg/kg of Zn, but not chromium, regarding high chromium concentrations found in soils (435 mg/kg). Since plant-associated microorganism can modulate phytoremediation efficiency, the biodiversity of P. laevigata associated bacteria was studied. Eighty-eight isolates from P. laevigata nodules were obtained; all isolates tolerated high concentrations of Al, Fe, Zn and Cr in vitro. The top-six chromium tolerant strains were identified by 16S rRNA sequence analysis as belonging to genus Bacillus. Bacillus sp. MH778713, close to Bacillus cereus group, showed to be the most resistant strain, tolerating up to 15000 mg/L Cr (VI) and 10000 mg/L of Al. Regarding the bioaccumulation traits, Bacillus sp. MH778713 accumulated up to 100 mg Cr(VI)/g of cells when it was exposed to 1474 mg/L of Cr VI. To assess Bacillus sp. MH778713 ability to assist Prosopis laevigata phytoremediation; twenty plants were inoculated or non-inoculated with Bacillus sp. MH778713 and grown in nitrogen-free Jensen's medium added with 0, 10 and 25 mg/L of Cr(VI). Only plants inoculated with Bacillus sp. grew in the presence of chromium showing the ability of this strain to assist chromium phytoremediation. P. laevigata and Bacillus spp. may be considered as good candidates for soil restoration of arid and semiarid sites contaminated with heavy metals.