Search

8 páginas, 4 tablas, 3 figuras, A bacterial strain designated as RZME10T was isolated from a Zea mays L. root collected in Spain. Results of analysis of the 16S rRNA gene sequence showed that this strain belongs to the genus Agrobacterium with Agrobacterium larrymoorei ATCC 51759T being the most closely related species with 99.9 % sequence similarity. The similarity values of the rpoB, recA, gyrB, atpD and glnII genes between strain RZME10T and A. larrymoorei ATCC 51759T were 93.5, 90.0, 88.7, 87.9 and 90.1 %, respectively. The estimated average nucleotide identity using blast and digital DNA–DNA hybridization values between these two strains were 80.4 and 30.2 %, respectively. The major fatty acids of strain RZME10T are those from summed feature 8 (C18 : 1 ω6c/C18 : 1 ω7c) and C16 : 0. Pathogenicity tests on tomato and carrot roots showed that strain RZME10T was not able to induce plant tumours. Based on the results of genomic, chemotaxonomic and phenotypic analyses, we propose that strain RZME10T represents a novel species named Agrobacterium cavarae sp. nov. (type strain RZME10T=CECT 9795T=LMG 31257T)., This work was supported by an MINECO (Spanish Central Government) grant AGL2013-48098-P to E.V. E.M. was granted a postdoctoral contract associated with this project. J.D.F.F. was recipient of a predoctoral fellowship from the University of Salamanca.

A bacterial strain designated CRZM18RT was isolated from a root of Zea mays in Spain. The analysis of the 16S rRNA gene sequence showed that this strain belongs to the genus Rhizobium , with Rhizobium cellulosilyticum ALA10B2T and Rhizobium yantingense H66T being the most closely related species with 98.3 and 97.9 % sequence similarity, respectively. The analysis of the concatenated recA and atpD genes showed that strain CRZM18RT forms a cluster with these species and also with Rhizobium smilacinae PTYR-5T, but the recA and atpD genes of strain CRZM18RT were phylogenetically distant, with identities lower than 90 and 96 %, respectively. DNA–DNA hybridization analysis showed mean relatedness of 43, 22 and 38 % with respect to R. cellulosilyticum ALA10B2T, R. yantingense LMG 28229T and R. smilacinae LMG 27604T. Phenotypic characteristics also differed from those of the most closely related species of the genus Rhizobium . The major fatty acids were those from summed feature 8 (C18 : 1ω6c/C18 : 1ω7c) and C16 : 0. Based on the genotypic, chemotaxonomic and phenotypic data obtained in this study, we propose to classify strain CRZM18RT in a novel species named Rhizobium zeae sp. nov. (type strain CRZM18RT=LMG 29735T=CECT 9169T).

Polyhydroxyalkanoic acids (PHAs) are natural polyesters that can be used to produce bioplastics which are biodegradable. Numerous microorganisms accumulate PHAs as energy reserves. Combinations of different PHAs monomers lead to the production of bioplastics with very different properties. In the present work, we show the capability of strains belonging to various phylogenetic lineages within the genus Mesorhizobium, isolated from Lotus corniculatus nodules, to produce different PHA monomers. Among our strains, we found the production of 3-hydroxybutyrate, 3-hydroxyvalerate, 3-hydroxydodecanoate, and 3-hydroxyhexadecanoate. Most of the PHA-positive strains were phylogenetically related to the species M. jarvisii. However, our findings suggest that the ability to produce different monomers forming PHAs is strain-dependent.

A bacterial strain designated AMTAE16 was isolated from a root of wheat in Spain. Phylogenetic analyses based on 16S rRNA gene sequences placed the isolate in the genus Paenibacilluswith its closest relative being Paenibacillus daejeonensis AP-20 with 99.0 % 16S rRNA gene sequence similarity. DNA–DNA hybridization studies showed a mean of 30 % DNADNA relatedness between strain AMTAE16 and the type strain of P. daejeonensis. The isolate was a Gram-stainvariable, motile and sporulating rod. Catalase and oxidase activities were positive. Gelatin and starch were hydrolysed but not casein. Growth was supported by many carbohydrates and organic acids as carbon source. MK-7 was the only menaquinone detected and anteiso-C, C and iso-C were the major fatty acids. The major polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, two unidentified aminophospholipids, four unidentified phospholipids and two unidentified lipids. meso- Diaminopimelic acid was detected in the peptidoglycan. The DNA G+C content was 55.4 mol%. Phylogenetic, chemotaxonomic and phenotypic analyses showed that strain AMTAE16represents a novel species of the genus Paenibacillus, for which the name Paenibacillus hispanicus sp. nov. is proposed. The type strain is AMTAE16(=LMG 29501=CECT 9124)., This work was supported by MINECO (Spanish Central Government) Grants AGL2013-48098-P and AGL2013-40758-R. EM is granted by a postdoctoral contract of this project.

The infection of legume plants by rhizobia is tightly regulated to ensure accurate bacterial penetration, infection, and development of functionally efficient nitrogen-fixing root nodules. Rhizobial Nod factors (NF) have key roles in the elicitation of nodulation signaling. Infection of white clover roots also involves the tightly regulated specific breakdown of the noncrystalline apex of cell walls in growing root hairs, which is mediated by Rhizobium leguminosarum bv. trifolii cellulase CelC2. Here, we have analyzed the impact of this endoglucanase on symbiotic signaling in the model legume Medicago truncatula. Ensifer meliloti constitutively expressing celC gene exhibited delayed nodulation and elicited aberrant ineffective nodules, hampering plant growth in the absence of nitrogen. Cotreatment of roots with NF and CelC2 altered Ca spiking in root hairs and induction of the early nodulin gene ENOD11. Our data suggest that CelC2 alters early signaling between partners in the rhizobia-legume interaction., Ministerio de Economía y Competitividad Grant/Award Number: AGL2015-70510-R

Aims: To study multiple bacterial colonization In vitro, several limitations are obvious. One of these limitations is the plant autofluorescence generally between green and red fluorescence depending on the plant sections. The most important limitation is the bacterial fluorescence labelling, compromised by different kind of variables. Here we report the use of a secure stain, Calcofluor White, in rhizobial and other kind of beneficial bacteria colonization studies. Study Design: Root colonization assays were designed to confirm the stability of Calcofluor White stain (Sigma) in root cell walls. Place and Duration of Study: Every assay developed in this method article was performed using the technical resources at the Department of Microbiology and Genetics in the University of Salamanca (Spain) in 2013. Methodology: We have labelled rhizobia with two different kinds of fluorescent protein genes (gfp Method Article Flores-Felix et al.; JABB, 2(1): 65-70, 2015; Article no.JABB.2015.009 66 and rfp). We have co-inoculated Lactuca sativa and Daucus carota seedlings with two rhizobia: GFP-tagged Rhizobium sp. PEPV16 and RFP-tagged Mesorhizobium sp. CSLC01. Colonization assays were perfomed in several days post-inoculation, staining inoculated lettuce and carrot roots with Calcofluor White stain (Sigma). Samples were monitorized for several days, using a fluorescence microscope (NIKON Eclipse 80i). Results: Bacterial attachment to plant tissues is observed by fluorescence microscopy after their labelling with fluorescent proteins. Our results show how Calcofluor White staining for plant tissues improves bacterial visualization in contrast with tissues stained with propidium iodide, a carcinogenic agent that cannot be used when bacteria are tagged with red fluorescent proteins such as RFP or mCherry. Conclusion: Calcofluor white is a non-carcinogenic and low toxic compound that has been classically used to stain fungi and plant tissues for different uses. Due to its low wavelength, calcofluor white may be used in combination with several fluorophores. In the present work we showed that this compound is a reliable alternative to propidium iodide for plant tissues staining in multiple rhizobial/bacterial colonization studies.

5 pages, 1 figure, and 2 statistics tables, A bacterial strain designated RTAE36T was isolated from wheat roots in northern Spain. Phylogenetic analyses based on 16S rRNA gene sequence placed the isolate into the genus Paenibacillus with its closest relative being Paenibacillus borealis DSM 13188T with 97.7% sequence similarity. Cells of the isolate were facultatively anaerobic, Gram-stain-positive, motile and sporulating rods. Catalase and oxidase were positive. Gelatin, casein and starch were not hydrolysed. Growth was supported by many carbohydrates and organic acids as carbon sources. MK-7 was the only menaquinone detected, and anteiso-C15 : 0, C16 : 0, iso-C14 : 0 and iso-C16 : 0 were the major fatty acids. The polar lipids profile consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, one unidentified aminolipid, two unidentified phospholipids, three unidentified phosphoaminolipids, one unidentified glycolipid and one unidentified lipid. meso-Diaminopimelic acid was detected in the cell-wall peptidoglycan. Strains RTAE36T and P. borealis DSM 13188 had an mean DNA–DNA relatedness of 39% and differed in several phenotypic and chemotaxonomic characteristics, confirming that strain RTAE36 should be considered as a representative of a novel species of the genus Paenibacillus, for which the name Paenibacillus tritici sp. nov. is proposed. The type strain is RTAE36 (=LMG 29502=CECT 9125)., This work was supported by MINECO (Spanish Central Government) grants AGL2013-48098-P and AGL2013-40758-R to E. V. and M. F. P., respectively. E. M. is supported by a postdoctoral contract associated with the project AGL2013-48098-P. J. D. F. F. is supported by a Fellowship of Salamanca University.

In this study, three strains belonging to the genus Mesorhizobium , CSLC115NT, CSLC19N and CSLC37N, isolated from Lotus corniculatus nodules in Spain, were characterized. Their 16S rRNA gene sequences were closely related to those of Mesorhizobium metallidurans STM 2683T, Mesorhizobium tianshanense A-1BST, Mesorhizobium tarimense CCBAU 83306T, Mesorhizobium gobiense CCBAU 83330T and Mesorhizobium caraganae CCBAU 11299T with similarity values higher than 99.7 %. The analysis of concatenated recA and glnII genes showed that the most closely related type strains were M. metallidurans STM 2683T, M. tianshanense A-1BST and M. tarimense CCBAU 83306T with 96, 95 and 94 % similarity values in the recA gene and 95, 94 and 94 % in the glnII gene, respectively. M. metallidurans LMG 24485T, M. tianshanense USDA 3592T and M. tarimense LMG 24338T showed means of 44, 41 and 42 % DNA–DNA relatedness, respectively, with respect to strain CSLC115NT. The major fatty acids were those from summed feature 8 (C18 : 1ω7c/C18 : 1ω6c), C16 : 0 and C18 : 1ω7c 11-methyl. The results of phenotypic characterization support that the L. corniculatus nodulating strains analysed in this work belong to a novel species of the genus Mesorhizobium for which the name Mesorhizobium helmanticense sp. nov. is proposed, and the type strain is CSLC115NT (= LMG 29734T=CECT 9168T).

A bacterial strain designated GTAE24 was isolated from a root of wheat growing in soil from the Canary Islands, Spain. Phylogenetic analyses based on 16S rRNA gene sequences placed the isolate in the genus Brevundimonas with Brevundimonas abyssalis TAR-001 as its closest relative at 99.4% similarity. DNA–DNA hybridization studies showed an average of 38% relatedness between strain GTAE24 and the type strain of B. abyssalis. Cells were Gram-stain-negative and motile by polar flagella. The strain was positive for oxidase and weakly positive for catalase. Gelatin, starch and casein were not hydrolysed. Growth was supported by many carbohydrates and organic acids as carbon source. Ubiquinone Q-10 was the predominant isoprenoid quinone and C03C97c/Cω6c (summed feature 8) and C were the major fatty acids. The major polar lipids were phosphatidylglycerol, 1,2-di-O-acyl-3-O-[D-glucopyranosyl-(1,4)-a-D-glucopyranuronosyl] glycerol, 1,2- diacyl-3-O-[6¢-phosphatidyl-a-D-glucopyranosyl] glycerol, 1,2-di-O-acyl-3-O-a-D-glucopyranosyl glycerol, and 1,2-di-O-acyl-3- O-a-D-glucopyranuronosyl glycerol. The DNA G+C content was 63.9 mol%. Phylogenetic, chemotaxonomic and phenotypic analyses showed that strain GTAE24 should be considered as representing a novel species of the genus Brevundimonas, for which the name Brevundimonas canariensis sp. nov. is proposed. The type strain is GTAE24 (=LMG 29500=CECT 9126)., This work was supported by MINECO (Spanish Central Government) Grants AGL2013-48098 P to E. V. and AGL2013-40758 R to M. F. P. E. M. was funded by a postdoctoral contract associated with the project AGL2013-48098 P. L. C. was funded by a postdoctoral contract from Newcastle University.

6 pages, 1 figure, and 2 statistics tables, A bacterial strain designated PM10T was isolated from root nodules of Periandra mediterranea in Brazil. Phylogenetic analyses based on 16S rRNA gene sequences placed the isolate in the genus Paenibacillus with its closest relatives being Paenibacillus vulneris CCUG 53270T and Paenibacillus yunnanensis YN2T with 95.6 and 95.9 % 16S rRNA gene sequence similarity, respectively. The isolate was a Gram-stain-variable, motile, sporulating rod that was catalase-negative and oxidase-positive. Caseinase was positive, amylase was weakly positive and gelatinase was negative. Growth was supported by many carbohydrates and organic acids as carbon sources. MK-7 was the only menaquinone detected and anteiso-C15 : 0 was the major fatty acid. Major polar lipids were diphosphatidylglycerol, phosphatidylmonomethylethanolamine, phosphatidylethanolamine, phosphatidylglycerol and two unidentified lipids. meso-Diaminopimelic acid was detected in the peptidoglycan. The DNA G+C content was 52.9 mol%. Phylogenetic, chemotaxonomic and phenotypic analyses showed that strain PM10T should be considered representative of a novel species of the genus Paenibacillus, for which the name Paenibacillus periandrae sp. nov. is proposed. The type strain is PM10T (=LMG 28691 =CECT 8827T)., CAPES (CoordenaÇao de AperfeiÇoamento de Pessoal de Nível Superior) for scholarships and financial support, IEF (Instituto Estadual de Floresta) for logistical support in Serra do Rola MoÇa Park.

Several molecular chaperones are known to be involved in bacteria stress response. To investigate the role of chaperone ClpB in rhizobia stress tolerance as well as in the rhizobia-plant symbiosis process, the clpB gene from a chickpea microsymbiont, strain Mesorhizobium ciceri LMS-1, was identified and a knockout mutant was obtained. The ClpB knockout mutant was tested to several abiotic stresses, showing that it was unable to grow after a heat shock and it was more sensitive to acid shock than the wild-type strain. A plant-growth assay performed to evaluate the symbiotic performance of the clpB mutant showed a higher proportion of ineffective root nodules obtained with the mutant than with the wild-type strain. Nodulation kinetics analysis showed a 6- to 8-day delay in nodule appearance in plants inoculated with the ΔclpB mutant. Analysis of nodC gene expression showed lower levels of transcript in the ΔclpB mutant strain. Analysis of histological sections of nodules formed by the clpB mutant showed that most of the nodules presented a low number of bacteroids. No differences in the root infection abilities of green fluorescent protein–tagged clpB mutant and wild-type strains were detected. To our knowledge, this is the first study that presents evidence of the involvement of the chaperone ClpB from rhizobia in the symbiotic nodulation process.

We isolated a strain coded Esc2AmT during a study focused on the microbial diversity of adult specimens of the bark beetle Hylesinus fraxini. Its 16S rRNA gene sequence had 99.4 % similarity with respect to its closest relative, Pseudomonas rhizosphaerae IH5T. The analysis of partial sequences of the housekeeping genes rpoB, rpoD and gyrB confirmed that strain Esc2AmT formed a cluster with P. rhizosphaerae IH5T clearly separated from the remaining species of the genus Pseudomonas. Strain Esc2AmT had polar flagella and could grow at temperatures from 4 °C to 30 °C. The respiratory quinone was Q9 and the main fatty acids were C16 : 0, C18 : 1ω7c and/or C18 : 1ω6c in summed feature 8 and C16 : 1ω7c and/or C16 : 1ω6c in summed feature 3. DNA–DNA hybridization results showed 51 % relatedness with respect to P. rhizosphaerae IH5T. Oxidase, catalase and urease-positive, the arginine dihydrolase system was present but nitrate reduction and β-galactosidase production were negative. Aesculin hydrolysis was positive. Based on the results from the genotypic, phenotypic and chemotaxonomic analyses, we propose the classification of strain Esc2AmT as representing a novel species of the genus Pseudomonas, for which we propose the name Pseudomonas coleopterorum sp. nov. The type strain is Esc2AmT ( = LMG 28558T = CECT 8695T).