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In Arabidopsis (Arabidopsis thaliana (L.) Heynh), exposure to volatile compounds (VCs) emitted by Penicillium aurantiogriseum promotes root hair (RH) proliferation and hyper-elongation through mechanisms involving ethylene, auxin and photosynthesis signaling. In addition, this treatment enhances the levels of the small signalling peptide RAPID ALKALINIZATION FACTOR 22 (RALF22). Here we used genetics to address the role of RALF22 in fungal VC-promoted RH growth and to identify the bioactive fungal VC. We found that RHs of ralf22 and feronia (fer-4) plants impaired in the expression of RALF22 and its receptor FERONIA, respectively, responded weakly to fungal VCs. Unlike in WT roots, fungal VC exposure did not enhance RALF22 transcript levels in roots of fer-4 and ethylene- and auxin- insensitive mutants. In ralf22 and fer-4 roots, this treatment did not enhance the levels of ERS2 transcripts encoding one member of the ethylene receptor family and those of some RH-related genes. RHs of ers2-1 and the rsl2rsl4 double mutants impaired in the expression of ERS2 and the ethylene- and auxin-responsive ROOT HAIR DEFECTIVE 6-LIKE 2 and 4 transcription factors, respectively, weakly responded to fungal VCs. Moreover, roots of plants defective in photosynthetic responsiveness to VCs exhibited weak RALF22 expression and RH growth responses to fungal VCs. VCs of ΔefeA strains of P. aurantiogriseum cultures impaired in ethylene synthesis weakly promoted RH proliferation and elongation in exposed plants. We conclude that RALF22 simultaneously functions as a transcriptionally regulated signaling molecule that participates in the ethylene, auxin and photosynthesis signaling-mediated RH growth response to fungal ethylene emissions and regulation of ethylene perception in RHs., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society of Plant Biologists. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Mediterranean high-mountain lakes are being increasingly affected by strong Saharan dust deposition events. However, the ecological impacts of these severe atmospheric episodes remain largely unknown. We examined the effects of a strong Saharan dust intrusion to the Iberian Peninsula in 2022 on the physicochemical parameters and prokaryotic communities in sediments of nine high-mountain lakes of Sierra Nevada (Spain) located above 2800 m.a.s.l and in different orientations (north vs. south). A previous year (2021), with lower Saharan dust deposition with respect to 2022, was used for interannual comparisons. The strong dust deposition to the high-mountain lakes resulted in a significant increase in sediment nutrient availability which was linked to changes in the composition of prokaryotic communities. Decreases in alpha diversity and changes in beta diversity of prokaryotic communities were mainly observed in lakes located in the south compared to the north orientation likely because the former was more affected by the atmospheric dust deposition episode. Dust intrusion to the high-mountain lakes resulted in significant changes in the relative abundance of specific genera involved in important nutrient cycling processes such as phosphate solubilization, nitrogen fixation, nitrification, and denitrification. Saharan dust deposition also increased predicted microbial functionality in all lakes. Our findings show that severe atmospheric dust inputs to remote high-mountain lakes of Sierra Nevada can have significant biogeochemical and biodiversity consequences through changes in nutrient availability and prokaryotic communities in sediments of these freshwater ecosystems. This information contributes to understanding how Mediterranean high-mountain lakes of Sierra Nevada face strong intrusions of Saharan dust and their ecological consequences., (© 2024. The Author(s).)

Bradyrhizobium diazoefficiens can live inside soybean root nodules and in free-living conditions. In both states, when oxygen levels decrease, cells adjust their protein pools by gene transcription modulation. PhaR is a transcription factor involved in polyhydroxyalkanoate (PHA) metabolism but also plays a role in the microaerobic network of this bacterium. To deeply uncover the function of PhaR, we applied a multipronged approach, including the expression profile of a phaR mutant at the transcriptional and protein levels under microaerobic conditions, and the identification of direct targets and of proteins associated with PHA granules. Our results confirmed a pleiotropic function of PhaR, affecting several phenotypes, in addition to PHA cycle control. These include growth deficiency, regulation of carbon and nitrogen allocation, and bacterial motility. Interestingly, PhaR may also modulate the microoxic-responsive regulatory network by activating the expression of fixK 2 and repressing nifA , both encoding two transcription factors relevant for microaerobic regulation. At the molecular level, two PhaR-binding motifs were predicted and direct control mediated by PhaR determined by protein-interaction assays revealed seven new direct targets for PhaR. Finally, among the proteins associated with PHA granules, we found PhaR, phasins, and other proteins, confirming a dual function of PhaR in microoxia.

The soybean endosymbiont Bradyrhizobium diazoefficiens harbours the complete denitrification pathway that is catalysed by a periplasmic nitrate reductase (Nap), a copper (Cu)-containing nitrite reductase (NirK), a c-type nitric oxide reductase (cNor), and a nitrous oxide reductase (Nos), encoded by the napEDABC, nirK, norCBQD, and nosRZDFYLX genes, respectively. Induction of denitrification genes requires low oxygen and nitric oxide, both signals integrated into a complex regulatory network comprised by two interconnected cascades, FixLJ-FixK2-NnrR and RegSR-NifA. Copper is a cofactor of NirK and Nos, but it has also a role in denitrification gene expression and protein synthesis. In fact, Cu limitation triggers a substantial down-regulation of nirK, norCBQD, and nosRZDFYLX gene expression under denitrifying conditions. Bradyrhizobium diazoefficiens genome possesses a gene predicted to encode a Cu-responsive repressor of the CsoR family, which is located adjacent to copA, a gene encoding a putative Cu+-ATPase transporter. To investigate the role of CsoR in the control of denitrification gene expression in response to Cu, a csoR deletion mutant was constructed in this work. Mutation of csoR did not affect the capacity of B. diazoefficiens to grow under denitrifying conditions. However, by using qRT-PCR analyses, we showed that nirK and norCBQD expression was much lower in the csoR mutant compared to wild-type levels under Cu-limiting denitrifying conditions. On the contrary, copA expression was significantly increased in the csoR mutant. The results obtained suggest that CsoR acts as a repressor of copA. Under Cu limitation, CsoR has also an indirect role in the expression of nirK and norCBQD genes., (© The Author(s) 2023. Published by Oxford University Press on behalf of FEMS.)

Current and continuing climate change in the Anthropocene epoch requires sustainable agricultural practices. Additionally, due to changing consumer preferences, organic approaches to cultivation are gaining popularity. The global market for organic grapes, grape products, and wine is growing. Biostimulant and biocontrol products are often applied in organic vineyards and can reduce the synthetic fertilizer, pesticide, and fungicide requirements of a vineyard. Plant growth promotion following application is also observed under a variety of challenging conditions associated with global warming. This paper reviews different groups of biostimulants and their effects on viticulture, including microorganisms, protein hydrolysates, humic acids, pyrogenic materials, and seaweed extracts. Of special interest are biostimulants with utility in protecting plants against the effects of climate change, including drought and heat stress. While many beneficial effects have been reported following the application of these materials, most studies lack a mechanistic explanation, and important parameters are often undefined (e.g., soil characteristics and nutrient availability). We recommend an increased study of the underlying mechanisms of these products to enable the selection of proper biostimulants, application methods, and dosage in viticulture. A detailed understanding of processes dictating beneficial effects in vineyards following application may allow for biostimulants with increased efficacy, uptake, and sustainability., Competing Interests: Author YA was employed by Chitose Laboratory Corp. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Jindo, Goron, Pizarro-Tobías, Sánchez-Monedero, Audette, Deolu-Ajayi, van der Werf, Goitom Teklu, Shenker, Pombo Sudré, Busato, Ochoa-Hueso, Nocentini, Rippen, Aroca, Mesa, Delgado and Tortosa.)

Nitrous oxide (N 2 O) is a powerful greenhouse gas that contributes to climate change. Denitrification is one of the largest sources of N 2 O in soils. The soybean endosymbiont Bradyrhizobium diazoefficiens is a model for rhizobial denitrification studies since, in addition to fixing N 2 , it has the ability to grow anaerobically under free-living conditions by reducing nitrate from the medium through the complete denitrification pathway. This bacterium contains a periplasmic nitrate reductase (Nap), a copper (Cu)-containing nitrite reductase (NirK), a c -type nitric oxide reductase (cNor), and a Cu-dependent nitrous oxide reductase (Nos) encoded by the napEDABC, nirK, norCBQD and nosRZDFYLX genes, respectively. In this work, an integrated study of the role of Cu in B. diazoefficiens denitrification has been performed. A notable reduction in nirK , nor, and nos gene expression observed under Cu limitation was correlated with a significant decrease in NirK, NorC and NosZ protein levels and activities. Meanwhile, nap expression was not affected by Cu, but a remarkable depletion in Nap activity was found, presumably due to an inhibitory effect of nitrite accumulated under Cu-limiting conditions. Interestingly, a post-transcriptional regulation by increasing Nap and NirK activities, as well as NorC and NosZ protein levels, was observed in response to high Cu. Our results demonstrate, for the first time, the role of Cu in transcriptional and post-transcriptional control of B. diazoefficiens denitrification. Thus, this study will contribute by proposing useful strategies for reducing N 2 O emissions from agricultural soils.

Two endophytic strains, coded MOVP5 T and MOPV6, were isolated from nodules of Phaseolus vulgaris plants grown on agricultural soil in Southeastern Spain, and were characterized through a polyphasic taxonomy approach. Their 16S rRNA gene sequences showed 99.3 and 99.4 %, 98.9 and 99.6 %, and 99.0 and 98.7% similarity to ' A. deltaense ' YIC 4121 T , A. radiobacter LGM 140 T , and A. pusense NRCPB10 T , respectively. Multilocus sequence analysis based on sequences of recA and atpD genes suggested that these two strains could represent a new Agrobacterium species with less than 96.5 % similarity to their closest relatives. PCR amplification of the telA gene, involved in synthesis of protelomerase, confirmed the affiliation of strains MOPV5 T and MOPV6 to the genus Agrobacterium . Whole genome average nucleotide identity and digital DNA-DNA hybridization average values were less than 95.1 and 66.7 %, respectively, with respect to its closest related species. Major fatty acids in strain MOPV5 T were C 18 : 1  ω7 c /C 18 : 1  ω6 c in summed feature 8, C 19 : 0 cyclo ω8 c , C 16 : 0 and C 16 : 0 3-OH. Colonies were small to medium, pearl-white coloured on YMA at 28 °C and growth was observed at 10-42 °C, pH 5.0-10.0 and with 0.0-0.5 % (w/v) NaCl. The DNA G+C content was 59.9 mol%. These two strains differ from all other genomovars of Agrobacterium found so far, including those that have not yet given a Latin name. The combined genotypic, phenotypic and chemotaxonomic data support the classification of strain MOPV5 T as representing a novel species of Agrobacterium , for which the name Agrobacterium leguminum sp. nov. is proposed. The type strain is MOPV5 T (=CECT 30096 T =LMG 31779 T ).

RNA-based high-throughput sequencing is a valuable tool in the discernment of the implication of metabolically active bacteria during composting. In this study, "alperujo" composting was used as microbial model for the elucidation of structure-function relationships with physicochemical transformation of the organic matter. DNA and RNA, subsequently retrotranscribed into cDNA, were isolated at the mesophilic, thermophilic and maturation phases. 16S rRNA gene was amplified by quantitative PCR (qPCR) and Illumina MiSeq platform to assess bacterial abundance and diversity, respectively. The results showed that the abundance of active bacteria assessed by qPCR was maximum at thermophilic phase, which confirm it as the most active stage of the process. Concerning diversity, Proteobacteria, Firmicutes, Bacteroidetes and Actinobacteria were the main phyla presented in composts. Concomitantly, three different behaviours were observed for bacterial dynamics: some genera decreased during the whole process meanwhile others proliferated only at thermophilic or maturation phase. Statistical correlation between physicochemical transformations of the organic matter and bacterial diversity revealed bacterial specialisation. This result indicated that specific groups of bacteria were only involved in the organic matter degradation during bio-oxidative phase or humification at maturation. Metabolic functions predictions confirmed that active bacteria were mainly involved in carbon (C) and nitrogen (N) cycles transformations, and pathogen reduction., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Legume-rhizobia symbiotic associations have beneficial effects on food security and nutrition, health and climate change. Hypoxia induced by flooding produces nitric oxide (NO) in nodules from soybean plants cultivated in nitrate-containing soils. As NO is a strong inhibitor of nitrogenase expression and activity, this negatively impacts symbiotic nitrogen fixation in soybean and limits crop production. In Bradyrhizobium diazoefficiens , denitrification is the main process involved in NO formation by soybean flooded nodules. In addition to denitrification, nitrate assimilation is another source of NO in free-living B. diazoefficiens cells and a single domain hemoglobin (Bjgb) has been shown to have a role in NO detoxification during nitrate-dependent growth. However, the involvement of Bjgb in protecting nitrogenase against NO in soybean nodules remains unclear. In this work, we have investigated the effect of inoculation of soybean plants with a bjgb mutant on biological nitrogen fixation. By analyzing the proportion of N in shoots derived from N 2 -fixation using the 15 N isotope dilution technique, we found that plants inoculated with the bjgb mutant strain had higher tolerance to flooding than those inoculated with the parental strain. Similarly, reduction of nitrogenase activity and nifH expression by flooding was less pronounced in bjgb than in WT nodules. These beneficial effects are probably due to the reduction of NO accumulation in bjgb flooded nodules compared to the wild-type nodules. This decrease is caused by an induction of expression and activity of the denitrifying NO reductase enzyme in bjgb bacteroids. As bjgb deficiency promotes NO-tolerance, the negative effect of NO on nitrogenase is partially prevented and thus demonstrates that inoculation of soybean plants with the B. diazoefficiens bjgb mutant confers protection of symbiotic nitrogen fixation during flooding., (Copyright © 2020 Salas, Tortosa, Hidalgo-García, Delgado, Bedmar, Richardson, Gates and Delgado.)

Metagenomic and transcriptomic techniques applied to composting could increase our understanding of the overall microbial ecology and could help us to optimise operational conditions which are directly related with economic interest. In this study, the fungal diversity and abundance of two-phase olive mill waste ("alperujo") composting was studied using Illumina MiSeq sequencing and quantitative PCR, respectively. The results showed an increase of the fungal diversity during the process, with Ascomycota being the predominant phylum. Penicillium was the main genera identified at the mesophilic and maturation phases, with Debaryomyces and Sarocladium at the thermophilic phase, respectively. The fungal abundance was increased during composting, which confirms their important role during thermophilic and maturation phases. Some Basidiomycota showed an increased during the process, which showed a positive correlation with the humification parameters. According to that, the genus Cystofilobasidium could be used as a potential fungal biomarker to assess alperujo compost maturation., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Microorganisms are the main contributing factor responsible for organic matter degradation during composting. In this research, the 454-pyrosequencing of the 16S rRNA gene was used to elucidate evolution of bacterial diversity during mesophilic, thermophilic and maturation composting stages of the two-phase olive mill waste ("alperujo"), the main by-product of the Spanish olive oil industry. Two similar piles were performance composting AL with sheep manure as bulking agent. Actinobacteria, Bacteriodetes, Firmicutes and Proteobacteria were the main phyla found in genomic libraries from each composting phase. Shannon and Chao1 biodiversity indices showed a clear difference between the mesophilic/thermophilic and maturation phases, which was mainly due to detection of new genera. PCA analysis of the relative number of sequences confirmed maturation affected bacterial population structure, and Pearson correlation coefficients between physicochemical composting parameters and relative number of genera sequences suggest that Planomicrobium and Ohtaekwangia could be considered as biomarkers for AL composting maturation., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Spatial and temporal variations related to hydric seasonality in abundance and diversity of denitrifier communities were examined in sediments taken from two sites differing in nitrate concentration along a stream Doñana National Park during a 3-year study. We found a positive relationship between the relative abundance of denitrifiers, determined as narG , napA , nirK , nirS and nosZ denitrification genes, and sediment nitrate content, with similar spatial and seasonal variations. However, we did not find association between denitrification activity and the community structure of denitrifiers. Because nosZ showed the strongest correlation with the content of nitrate in sediments, we used this gene as a molecular marker to construct eight genomic libraries. Analysis of these genomic libraries revealed that diversity of the nosZ -bearing communities was higher in the site with higher nitrate content. Regardless of nitrate concentration in the sediments, the Bradyrhizobiaceae and Rhodocyclaceae were the most abundant families. On the contrary, Rhizobiaceae was exclusively present in sediments with higher nitrate content. Results showed that differences in sediment nitrate concentration affect the composition and diversityof nosZ -bearing communities.

Constructed wetlands are used for biological treatment of wastewater from agricultural lands carrying pollutants such as nitrates. Nitrogen removal in wetlands occurs from direct assimilation by plants and through microbial nitrification and denitrification. We investigated the spatial distribution of N-cycling microbial communities and genes involved in nitrification and denitrification in constructed wetland sediments receiving irrigation water. We used quantitative real-time PCR (qPCR) to characterize microbial communities. Geostatistical variance analysis was used to relate them with vegetation cover and biogeochemical sediment properties. The spatial distribution of the N-cycling microbial communities of sediments was heterogeneous and complex. Total communities of bacteria and crenarchaea showed different spatial distributions. Analysis of autocorrelation patterns through semivariance indicated a tendency towards a patchy distribution over scales around 10 m for genes involved in the nitrification and denitrification processes. In contrast, biogeochemical sediment properties showed diverse spatial distributions. While almost no patchiness was found for pH and moisture, patchiness at scales between 8 and 10 m was detected for carbon, nitrate and ammonia. Denitrification variables showed spatial autocorrelation at scales comparable to genes. However, denitrifying enzyme activity and potential N(2)O production showed a common spatial pattern, different from that of the N(2)O/(N(2)O + N(2))., (© 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.)

The use of waste materials as organic amendments in soil remediation can affect metal solubility; this interaction will vary with the characteristics of the organic matter that is added to the soil. A pot experiment was carried out in a calcareous, metal-polluted soil, using Beta maritima L. as an indicator species for the treatment effects on metal solubility. The treatments were: fresh solid olive husk, a mature compost, their respective water extracts (as the most reactive and biodegradable fraction) and an unamended, control soil. The compost reduced metal availability and plant uptake, while fresh olive husk favoured Mn bioavailability and produced phytotoxicity. The water-soluble extract from fresh solid olive husk also provoked elevated Mn solubility in soil, but did not increase Mn uptake by plants. The application of water-soluble organic matter obtained from compost did not affect heavy metal solubility significantly. Therefore, composted olive husk seems to be the most-appropriate material for the development of bioremediation strategies., (Copyright © 2011 Elsevier B.V. All rights reserved.)