Alexandra Díez-Méndez

Rhizobium cellulosilyticum as a co-inoculant enhances Phaseolus vulgaris grain yield under greenhouse conditions

The Rhizobium-legume symbiosis is a complex partnership with many factors, with initial bacterial colonization of the plant root surface and primary infection as key early stages. Two molecules are strongly involved in these processes: the structural carbohydrate cellulose and the enzyme cellulase, which breaks down the former and allows rhizobia to infect the roots. Here, we report the effect on common bean (Phaseolus vulgaris L.) after co-inoculation of the non-nodulating, cellulase-overproducing strain Rhizobium cellulosilyticum ALA10B2T and the P. vulgaris-nodulating R. leguminosarum strain TPV08. In order to elucidate the effect of combined inoculation with both strains, we designed greenhouse assays, including single inoculation with strain TPV08, co-inoculation with both strains and an uninoculated treatment in non-sterile peat. Chemical fertilizers were not added. Chlorophyll content in the leaves was measured after the flowering stage by spectrophotometry and was considered to be indicative of the nutrient status of the plants. Nodule formation was observed on roots of the inoculated plants, while no nodulation was observed on roots of the uninoculated plants. The results indicate a synergistic effect between the two Rhizobium strains. Co-inoculated plants exhibited significant increases in seed yield and nitrogen content in comparison with the uninoculated control plants and with plants inoculated with a single strain. It is suggested that co-inoculation with strain ALA10B2T greatly increased the efficiency of N fixation by strain TPV08.

On the bright side of a forest pest-the metabolic potential of bark beetles' bacterial associates

Bark beetles reproduce and overwinter under the bark of trees, and are associated with bacteria that may influence the fitness of their hosts. As regard the aim of this study was to test the metabolic potential of bacterial strains, isolated from the bark beetle species Cryphalus piceae, Ips typographus and Pityophthorus pityophthorus and collected in the Czech Republic from fir, spruce and pine trees, respectively, to degrade plant cell compounds. The bacterial strains were identified as belonging to the genera Curtobacterium, Erwinia, Pantoea, Pseudomonas, Rahnella, Staphylococcus, and Yersinia. Several activities related to the degradation of lignocellulosic materials, such as cellulose, xylan and starch, were found. Moreover, the genomes of three of these strains were sequenced and analyzed, and the presence of the enzymatic machinery required for biomass hydrolysis was discovered. This finding supports the idea that bacteria aid in the provision of nutrients to the beetle from the hydrolysis of tree compounds, results that are relevant for studying the ecological implication of bacterial strains in the bark beetle life cycle. In addition, the activities found in association with the bacterial strains could be useful in biotechnological processes, such as the production of biofuels from biomass, colorant degradation, in the textile industry and for wastewater treatments. Furthermore, the gene sequences of the lignocellulolytic enzymes found within the genomes serve as a basis for future studies regarding the potential application of these bacteria, and their metabolic machinery, in processes such as biomass hydrolysis and bioremediation.

Bacterial Probiotics: A Truly Green Revolution

Throughout history, the evolution and progress of all human civilizations have been closely linked to the evolution and development of agriculture, since this is the basis of food production to sustain population and ensure social stability.

Bacillus terrae sp. nov. isolated from Cistus ladanifer rhizosphere soil.

A bacterial strain designated RA9T was isolated from a root of Cistus ladanifer in Spain. Phylogenetic analyses based on 16S rRNA gene sequences placed the isolate into the genus Bacillus with its closest relatives being Bacillus fortis R-6514T and Bacillus fordii R-7190T with 98.2 % similarity in both cases. DNA-DNA hybridization studies showed mean relatedness values of 29 and 30 %, respectively, between strain RA9T and the type strains of B. fortis and B. fordii. Cells of the isolate were Gram-stain-positive, motile, sporulating rods. Catalase and oxidase were positive. Gelatin, starch and casein were not hydrolysed. Menaquinone MK-7 was the only menaquinone detected and iso-C15 : 0 and anteiso-C15 : 0 were the major fatty acids. The polar lipid profile consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, one unidentified aminophospholipid, one unidentified phospholipid, one unidentifed glycolipid and one unidentified lipid. meso-Diaminopimelic acid was detected in the peptidoglycan. The DNA G+C content was 43.1 mol%. Phylogenetic, chemotaxonomic and phenotypic analyses showed that strain RA9T should be considered as representing a novel species of the genus Bacillus, for which the name Bacillus terrae sp. nov. is proposed. The type strain is RA9T (=LMG 29736T=CECT 9170T).

Improvement of saffron production using Curtobacterium herbarum as a bioinoculant under greenhouse conditions

Plant Growth Promoting Rhizobacteria (PGPR) are natural soil bacteria which establish a beneficial relationship with their host. This microbiota community exists in the rhizosphere and inside plant tissues and stimulates plant growth by a variety of direct or indirect mechanisms. These bacterial plant promoters are frequently present in different environments, and are associated with many plant species, both wild and agricultural. Saffron is the dried stigmas of Crocus sativus (L.) and is the most expensive spice in the world. Remarkably, saffron cultivation and collection is carried out by hand and does not involve the use of machines. Additionally, 150 flowers are needed to produce one gram of dried stigmas. Hence, a slight increase in the size of the saffron filaments per plant would result in a significant increase in the production of this spice. In this study, we report the improved production of saffron using Curtobacterium herbarum Cs10, isolated from Crocus seronitus subs clusii, as a bioinoculant. The bacterial strain was selected owing to its multifunctional ability to produce siderophores, solubilize phosphate and to produce plant growth hormones like IAA. Furthermore, the isolate was tested on saffron producing plants under greenhouse conditions. The results indicate that Curtobacterium herbarum Cs10 improves the number of flowers and significantly enhances the length of the saffron filaments and overall saffron production compared to the control treated plants.

Discovery of Phloeophagus Beetles as a Source of Pseudomonas Strains That Produce Potentially New Bioactive Substances and Description of Pseudomonas bohemica sp. nov.

Antimicrobial resistance is a worldwide problem that threatens the effectiveness of treatments for microbial infection. Consequently, it is essential to study unexplored niches that can serve for the isolation of new microbial strains able to produce antimicrobial compounds to develop new drugs. Bark beetles live in phloem of host trees and establish symbioses with microorganisms that provide them with nutrients. In addition, some of their associated bacteria play a role in the beetle protection by producing substances that inhibit antagonists. In this study the capacity of several bacterial strains, isolated from the bark beetles Ips acuminatus, Pityophthorus pityographus Cryphalus piceae, and Pityogenes bidentatus, to produce antimicrobial compounds was analyzed. Several isolates exhibited the capacity to inhibit Gram-positive and Gram-negative bacteria, as well as fungi. The genome sequence analysis of three Pseudomonas isolates predicted the presence of several gene clusters implicated in the production of already described antimicrobials and moreover, the low similarity of some of these clusters with those previously described, suggests that they encode new undescribed substances, which may be useful for developing new antimicrobial agents. Moreover, these bacteria appear to have genetic machinery for producing antitumoral and antiviral substances. Finally, the strain IA19T showed to represent a new species of the genus Pseudomonas. The 16S rRNA gene sequence analysis showed that its most closely related species include Pseudomonas lutea, Pseudomonas graminis, Pseudomonas abietaniphila and Pseudomonas alkylphenolica, with 98.6, 98.5 98.4, and 98.4% identity, respectively. MLSA of the housekeeping genes gyrB, rpoB, and rpoD confirmed that strain IA19T clearly separates from its closest related species. Average nucleotide identity between strains IA19T and P. abietaniphila ATCC 700689T, P. graminis DSM 11363T, P. alkylphenolica KL28T and P. lutea DSM 17257T were 85.3, 80.2, 79.0, and 72.1%, respectively. Growth occurs at 4-37°C and pH 6.5-8. Optimal growth occurs at 28°C, pH 7–8 and up to 2.5% NaCl. Respiratory ubiquinones are Q9 (97%) and Q8 (3%). C16:0 and in summed feature 3 are the main fatty acids. Based on genotypic, phenotypic and chemotaxonomic characteristics, the description of Pseudomonas bohemica sp. nov. has been proposed. The type strain is IA19T (=CECT 9403T = LMG 30182T).

Rhizobium Symbiotic Enzyme Cellulase CelC2: Properties and Applications

Abstract Rhizobium leguminosarum bv. trifolii CelC2 cellulase is a 1,4-β- d -endoglucanase involved in the hydrolysis and biosynthesis of rhizobial cellulose. These special features make this enzyme essential in the plant infection process and important in root surface colonization and biofilm formation. A loss-of-function of the celC gene causes an increase in the production of external cellulose microfibrils. However, a gain-of-function derivative leads to an enhancement of rhizobial competitiveness, increasing the number of rhizobial cells within the plant. In this chapter, we review the properties and prospective applications of CelC2 cellulase, which reveals itself as a symbiotic enzyme with a high biotechnological potential.