Genome Analysis of Endobacterium cerealis, a Novel Genus and Species Isolated from Zea mays Roots in North Spain.

In the present work, we analyse the genomic and phenotypic characteristics of a strain named RZME27T isolated from roots of a Zea mays plant grown in Spain. The phylogenetic analyses of 16S rRNA gene and whole genome sequences showed that the strain RZME27T clustered with the type strains of Neorhizobium galegae and Pseudorhizobium pelagicum from the family Rhizobiaceae. This family encompasses several genera establishing symbiosis with legumes, but the genes involved in nodulation and nitrogen fixation are absent in its genome. Nevertheless, genes related to plant colonization, such as those involved in motility, chemotaxis, quorum sensing, exopolysaccharide biosynthesis and hydrolytic enzymes production were found. The comparative pangenomic analyses showed that 78 protein clusters present in the strain RZME27T were not found in the type strains of its closest relatives N. galegae and P. pelagicum. The calculated average nucleotide identity (ANI) values between the strain RZME27T and the type strains of N. galegae and P. pelagicum were 75.61% and 75.1%, respectively, similar or lower than those found for other genera from family Rhizobiaceae. Several phenotypic differences were also found, highlighting the absence of the fatty acid C19:0 cyclo ω8c and propionate assimilation. These results support the definition of a novel genus and species named Endobacterium cerealis gen. nov. sp. nov. whose type strain is RZME27T

Modeling of a future scenario of potential sea level rise and consequences to land use in the Cananéia-Iguape Estuarine-Lagoonal Complex (Brazil)

The objective of this study was to develop possible future land use scenarios for the Cananéia-Iguape Estuarine-Lagoonal Complex by the analysis of the land use evolution along 24 years (1986-2010), and to predict scenarios for 2025. In order to predict the scenario of the land use and occupation classes of 2025 a simulation was carried out by means of the Markov chain methods and calibration of the simulation model in the IDRISI Andes program. In the phase prior to this step it was utilized TM sensor scenes (Landsat 5) and supervised classification techniques to simulating the land use and occupation map of 2010 from land use and occupation maps from 1986 and 1999. Then the land use and occupation map of 2010 was utilized inside de Markov chain to simulate the map of 2025. By applying the techniques proposed by Titus and Narayanan, Pfeffer, Ramhstorf and the Intergovernmental Panel on Climate Change (IPCC) model, the vulnerability of the study area was calculated taking into consideration a potential sea level rise in 2025, 2050 and 2100. In the last step of the applied method, land use and occupation classes were assessed, which will possibly be affected by a highest tide event in 2025. It was observed that 80.92% the area flooded during highest tide events in 1999 was composed of dense arboreal vegetation and it is concluded that this expansion of the dense arboreal vegetation area will also occur towards the coastal zone in 2025. We suggest that the local factor for sea level rise in the study area has a strong geological component, so that, instead of a mere sea level rise, an opposite movement is taking place, which is the subsidence of the plain due to modern tectonism.The objective of this study was to develop possible future land use scenarios for the Cananéia-Iguape Estuarine-Lagoonal Complex by the analysis of the land use evolution along 24 years (1986-2010), and to predict scenarios for 2025. In order to predict the scenario of the land use and occupation classes of 2025 a simulation was carried out by means of the Markov chain methods and calibration of the simulation model in the IDRISI Andes program. In the phase prior to this step it was utilized TM sensor scenes (Landsat 5) and supervised classification techniques to simulating the land use and occupation map of 2010 from land use and occupation maps from 1986 and 1999. Then the land use and occupation map of 2010 was utilized inside de Markov chain to simulate the map of 2025. By applying the techniques proposed by Titus and Narayanan, Pfeffer, Ramhstorf and the Intergovernmental Panel on Climate Change (IPCC) model, the vulnerability of the study area was calculated taking into consideration a potential sea level rise in 2025, 2050 and 2100. In the last step of the applied method, land use and occupation classes were assessed, which will possibly be affected by a highest tide event in 2025. It was observed that 80.92% the area flooded during highest tide events in 1999 was composed of dense arboreal vegetation and it is concluded that this expansion of the dense arboreal vegetation area will also occur towards the coastal zone in 2025. We suggest that the local factor for sea level rise in the study area has a strong geological component, so that, instead of a mere sea level rise, an opposite movement is taking place, which is the subsidence of the plain due to modern tectonism

Agrobacterium cavarae sp. nov., isolated from maize (Zea mays L.) roots

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)

Selection of the root endophyte Pseudomonas brassicacearum CDVBN10 as plant growth promoter for Brassica napus L. crops

Rapeseed (Brassica napus L.) is an important crop worldwide, due to its multiple uses, such as a human food, animal feed and a bioenergetic crop. Traditionally, its cultivation is based on the use of chemical fertilizers, known to lead to several negative effects on human health and the environment. Plant growth-promoting bacteria may be used to reduce the need for chemical fertilizers, but efficient bacteria in controlled conditions frequently fail when applied to the fields. Bacterial endophytes, protected from the rhizospheric competitors and extreme environmental conditions, could overcome those problems and successfully promote the crops under field conditions. Here, we present a screening process among rapeseed bacterial endophytes to search for an efficient bacterial strain, which could be developed as an inoculant to biofertilize rapeseed crops. Based on in vitro, in planta, and in silico tests, we selected the strain Pseudomonas brassicacearum CDVBN10 as a promising candidate; this strain produces siderophores, solubilizes P, synthesizes cellulose and promotes plant height in 5 and 15 days-post-inoculation seedlings. The inoculation of strain CDVBN10 in a field trial with no addition of fertilizers showed significant improvements in pod numbers, pod dry weight and shoot dry weight. In addition, metagenome analysis of root endophytic bacterial communities of plants from this field trial indicated no alteration of the plant root bacterial microbiome; considering that the root microbiome plays an important role in plant fitness and development, we suggest this maintenance of the plant and its bacterial microbiome homeostasis as a positive

Arabinose and protocatechuate catabolism genes are important for growth of Rhizobium leguminosarum biovar viciae in the pea rhizosphere

Background and aims: To form nitrogen-fixing nodules on pea roots, Rhizobium leguminosarum biovar viciae must be competitive in the rhizosphere. Our aim was to identify genes important for rhizosphere fitness. Methods: Signature-tagged mutants were screened using microarrays to identify mutants reduced for growth in pea rhizospheres. Candidate mutants were assessed relative to controls for growth in minimal medium, growth in pea rhizospheres and for infection of peas in mixed inoculants. Mutated genes were identified by DNA sequencing and confirmed by transduction. Results: Of 5508 signature-tagged mutants, microarrays implicated 50 as having decreased rhizosphere fitness. Growth tests identified six mutants with rhizosphere-specific phenotypes. The mutation in one of the genes (araE) was in an arabinose catabolism operon and blocked growth on arabinose. The mutation in another gene (pcaM), encoding a predicted solute binding protein for protocatechuate and hydroxybenzoate uptake, decreased growth on protocatechuate. Both mutants were decreased for nodule infection competitiveness with mixed inoculants, but nodulated peas normally when inoculated alone. Other mutants with similar phenotypes had mutations predicted to affect secondary metabolism. Conclusions: Catabolism of arabinose and protocatechuate in the pea rhizosphere is important for competitiveness of R.l. viciae. Other genes predicted to be involved in secondary metabolism are also important

Spatiotemporal Characteristics of the Largest HIV-1 CRF02_AG Outbreak in Spain: Evidence for Onward Transmissions

Background and Aim: The circulating recombinant form 02_AG (CRF02_AG) is the predominant clade among the human immunodeficiency virus type-1 (HIV-1) non-Bs with a prevalence of 5.97% (95% Confidence Interval-CI: 5.41–6.57%) across Spain. Our aim was to estimate the levels of regional clustering for CRF02_AG and the spatiotemporal characteristics of the largest CRF02_AG subepidemic in Spain.Methods: We studied 396 CRF02_AG sequences obtained from HIV-1 diagnosed patients during 2000–2014 from 10 autonomous communities of Spain. Phylogenetic analysis was performed on the 391 CRF02_AG sequences along with all globally sampled CRF02_AG sequences (N = 3,302) as references. Phylodynamic and phylogeographic analysis was performed to the largest CRF02_AG monophyletic cluster by a Bayesian method in BEAST v1.8.0 and by reconstructing ancestral states using the criterion of parsimony in Mesquite v3.4, respectively.Results: The HIV-1 CRF02_AG prevalence differed across Spanish autonomous communities we sampled from (p < 0.001). Phylogenetic analysis revealed that 52.7% of the CRF02_AG sequences formed 56 monophyletic clusters, with a range of 2–79 sequences. The CRF02_AG regional dispersal differed across Spain (p = 0.003), as suggested by monophyletic clustering. For the largest monophyletic cluster (subepidemic) (N = 79), 49.4% of the clustered sequences originated from Madrid, while most sequences (51.9%) had been obtained from men having sex with men (MSM). Molecular clock analysis suggested that the origin (tMRCA) of the CRF02_AG subepidemic was in 2002 (median estimate; 95% Highest Posterior Density-HPD interval: 1999–2004). Additionally, we found significant clustering within the CRF02_AG subepidemic according to the ethnic origin.Conclusion: CRF02_AG has been introduced as a result of multiple introductions in Spain, following regional dispersal in several cases. We showed that CRF02_AG transmissions were mostly due to regional dispersal in Spain. The hot-spot for the largest CRF02_AG regional subepidemic in Spain was in Madrid associated with MSM transmission risk group. The existence of subepidemics suggest that several spillovers occurred from Madrid to other areas. CRF02_AG sequences from Hispanics were clustered in a separate subclade suggesting no linkage between the local and Hispanic subepidemics

Biotechnological applications of bacterial cellulases

Cellulases have numerous applications in several industries, including biofuel production, food and feed industry, brewing, pulp and paper, textile, laundry, and agriculture.Cellulose-degrading bacteria are widely spread in nature, being isolated from quite different environments. Cellulose degradation is the result of a synergic process between an endoglucanase, an exoglucanase and a,β-glucosidase. Bacterial endoglucanases degrade ß-1,4-glucan linkages of cellulose amorphous zones, meanwhile exoglucanases cleave the remaining oligosaccharide chains, originating cellobiose, which is hydrolyzed by ß-glucanases. Bacterial cellulases (EC 3.2.1.4) are comprised in fourteen Glycosil Hydrolase families. Several advantages, such as higher growth rates and genetic versatility, emphasize the suitability and advantages of bacterial cellulases over other sources for this group of enzymes. This review summarizes the main known cellulolytic bacteria and the best strategies to optimize their cellulase production, focusing on endoglucanases, as well as it reviews the main biotechnological applications of bacterial cellulases in several industries, medicine and agriculture

History and current taxonomic status of genus Agrobacterium

The genus Agrobacterium was created a century ago by Conn who included it in the family Rhizobiaceae together with the genus Rhizobium. Initially, the genus Agrobacterium contained the non-pathogenic species Agrobacterium radiobacter and the plant pathogenic species Agrobacterium tumefaciens and Agrobacterium rhizogenes. At the end of the past century two new pathogenic species, Agrobacterium rubi and Agrobacterium vitis, were added to the genus. Already in the present century these species plus Agrobacterium larrymoorei were reclassified into genus Rhizobium. This reclassification was controversial and for a time both genus names were used when new species were described. Few years ago, after a taxonomic revision based on genomic data, the old species A. rhizogenes was maintained in the genus Rhizobium, the old species A. vitis was transferred to the genus Allorhizobium and several Rhizobium species were transferred to the genus Agrobacterium, which currently contains 14 species including the old species A. radiobacter, A. tumefaciens, A. rubi and A. larrymoorei. Most of these species are able to produce tumours in different plants, nevertheless the genus Agrobacterium also encompasses non-pathogenic species, one species able to nodulate legumes and one human pathogenic species. Taking into account that the species affiliations to five Agrobacterium genomospecies have not been determined yet, an increase in the number of species within this genus is expected in the near future

Alcanivorax balearicus sp nov., isolated from Lake Martel

A bacterial strain designated MACL04(T) was isolated from Lake Martel, a subterraneous saline lake in Mallorca (Spain). The complete 16S rRNA gene sequence of this strain showed nearly 100 % similarity to that of Alcanivorax dieselolei B-5(T). Despite this high similarity, strain MACL04(T) showed phenotypic, chemotaxonomic and molecular differences with respect to A. dieselolei, indicating that strain MACL04(T) represents a separate species. Cells of strain MACL04(T) were motile by means of a single polar or subpolar flagellum and colonies formed on media containing 1 % (v/v) Tween 20 were opaque and mucoid, with blue-green iridescence. The generation time of strain MACL04(T) in this medium was approximately half that of A. diesefolei B-5(T) and strain MACL04(T) did not produce lipases after incubation for 5 days. Strain MACL04(T) did not require NaCl for growth and grew in the presence of up to 15 % (w/v) NaCl. The strain was able to use alkanes as a sole carbon source; however, glucose could also be used, albeit weakly, as a carbon source. Several amino acids and organic acids were used as carbon sources. Strain MACL04(T) produced acid in media containing pyruvate as the sole carbon source. The major fatty acids were C-19: 0 cyclo omega 8c and C-16: 0. The fatty acid C-16: 1 omega 8C, present in strain MACL04(T), was not detected in the recognized Alcanivorax species. The sequences of the large and short 16S-23S intergenic spacer T regions showed similarities of 97.2 and 98.8 % (ungapped) with respect to A. dieselolei B-5(T) Partial sequences of gyrB and alkb genes showed 94.0 % similarity between strain MACL04(T) and A. dieselolei B-5(T). The G + C content of strain MACL04(T) was 62.8 mol%. The data from this polyphasic study indicate that strain MACL04(T) represents a novel species of the genus Alcanivorax, for which the name Alcanivorax balearicus sp. nov. is proposed. The type strain is MACL04(T) (= LMG 22508(T) =CECT 5683(T))

Design and application of microbial biofertilizers

Soil beneficial microbes, especially plant growth promoting microorganisms (bacteria and fungi), are susceptible to be used in microbial-based bioformulations. These products, which can also be referred as biofertilizers, have been studied for several decades and their benefits for the environment and agro-systems versus chemically synthetized products are sufficiently demonstrated. Here, we describe the key stages that a biofertilizer product goes through in the bioformulation processes, from the lab to the field, with special emphasis in the strain(s) selection, viability, competence, and stability. Moreover, we focus on carrier materials, their nature, and their roles to help the active microorganism to survive and persist under stress conditions, such as drought, pathogens, and soil degradation. Finally, we reviewed the current state of legislation and bioformulation marketing in several world regions. Overall, with this chapter we intend to enumerate the steps in the formulation of a successful biofertilizer and to further discuss key stages and handicaps in the process of production, registration, and safe application in the fields.Peer reviewe