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Premise Methods Results Conclusions Legumes establish mutualistic interactions with pollinators and nitrogen (N)‐fixing bacteria that are critical for plant reproduction and ecosystem functioning. However, we know little about how N‐fixing bacteria and soil nutrient availability affect plant attractiveness to pollinators.In a two‐factorial greenhouse experiment to assess the impact of N‐fixing bacteria and soil types on floral traits and attractiveness to pollinators in Chamaecrista latistipula (Fabaceae), plants were inoculated with N‐fixing bacteria (NF+) or not (NF‐) and grown in N‐rich organic soil (+N organic soil) or N‐poor sand soil (‐N sand soil). We counted buds and flowers and measured plant size during the experiment. We also measured leaf, petal, and anther reflectance with a spectrophotometer and analyzed reflectance curves. Using the bee hexagon model, we estimated chromatic contrasts, a crucial visual cues for attracting bees that are nearby and more distant.NF+ plants in ‐N sand soil had a high floral display and color contrasts. On the other hand, NF‐ plants and/or plants in +N organic soil had severely reduced floral display and color contrasts, decreasing floral attractiveness to bee pollinators.Our findings indicate that the N‐fixing bacteria positively impact pollination, particularly when nutrients are limited. This study provides insights into the dynamics of plant–pollinator interactions and underscores the significant influence of root symbionts on key floral traits within tropical ecosystems. These results contribute to understanding the mechanisms governing mutualisms and their consequences for plant fitness and ecological dynamics. [ABSTRACT FROM AUTHOR]

Tuber magnatum is the most expensive truffle, but its large‐scale cultivation is still a challenge compared to other valuable Tuber species. T. magnatum mycelium has never been grown profitably until now, which has led to difficulties to studying it in vitro. This study describes beneficial interactions between T. magnatum mycelium and never before described bradyrhizobia, which allows the in vitro growth of T. magnatum mycelium. Three T. magnatum strains were co‐isolated on modified Woody Plant Medium (mWPM) with aerobic bacteria and characterised through microscopic observations. The difficulties of growing alone both partners, bacteria and T. magnatum mycelium, on mWPM demonstrated the reciprocal dependency. Three bacterial isolates for each T. magnatum strain were obtained and molecularly characterised by sequencing the 16S rRNA, glnII, recA and nifH genes. Phylogenetic analyses showed that all nine bacterial strains were distributed among five subclades included in a new monophyletic lineage belonging to the Bradyrhizobium genus within the Bradyrhizobium jicamae supergroup. The nifH genes were detected in all bacterial isolates, suggesting nitrogen‐fixing capacities. This is the first report of consistent T. magnatum mycelium growth in vitro conditions. It has important implications for the development of new technologies in white truffle cultivation and for further studies on T. magnatum biology and genetics. [ABSTRACT FROM AUTHOR]

Following the latest Permian mass extinction (LPME), abundant unusual sedimentary features and fabrics were widely distributed in Early Triassic carbonate platforms. As a unique type of carbonate grain, spheroids from the Lower Triassic are infrequently reported, yet differ significantly from giant ooids and oncoids. Griesbachian micritic spheroids are well preserved in the Lower Triassic Feixianguan Formation at the Baimiaozi (BMZ) outcrop in Beibei, Chongqing, southwestern China. These spheroids (diameter: 3.9–10.8 mm) were in a marl layer interbedded with massive oolites. They were deposited in the wave troughs of ripple marks comprising micritic clots, sparry calcite, clay minerals, organic matter and pyrite. A microbial spheroid origin is implied based on the coccoidal microbes (coccoid‐like microspherules and bacterial clump‐like microspherules), numerous pyrite framboids (indications suggest the presence of many sulphate‐reducing bacteria and the high iron content in seawater can promote nitrogen‐fixing cyanobacteria proliferation) and microbial‐derived microspherules. The dark‐coloured matrix between the spheroids primarily included microcrystalline calcite, clay minerals, organic matter and metazoan fossils. The spherical and ellipsoidal shapes of the spheroids suggested rapid accretion and lithification. As a unique carbonate depositional mode, Greisbachian spheroids may have been recorded for a palaeo‐ocean with dysoxic and calcium carbonate supersaturation shortly after the LPME. [ABSTRACT FROM AUTHOR]

An increase in the nitrogen (N) content in coarse woody debris (CWD) facilitates its decomposition, affecting the cycling of N and other nutrients in forest ecosystems. Bryophytes may increase the N content by transferring the N in bryophyte tissues to the underlying CWD. This study examined whether and how bryophytes increase N content in the underlying CWD using N‐stable isotope ratios (δ15N) as tracers for N sources. The N content and δ15N values in CWD with bryophytes were significantly higher than those in CWD without bryophytes. However, the δ15N values in CWD with bryophytes differed significantly from those in bryophytes on CWD, demonstrating that the N in bryophytes did not cause the increase in N content in CWD. The analyses using δ15N further indicated that the high N content in CWD with bryophytes may be attributed to increased N supply from wood‐decomposing fungi and N‐fixing bacteria. This increase in N content may result from the enhanced moisture content in CWD beneath bryophytes, which facilitates the activity of wood‐decomposing fungi and N‐fixing bacteria. Notably, the influence of bryophytes on the N content in CWD differed between bryophyte life forms: bryophytes that form dense mats increased the N content in CWD, whereas those with loose mats decreased the N content. This difference can be explained by the greater humidity experienced by CWD with bryophytes forming dense mats than that experienced by CWD with bryophytes forming loose mats. Given that the N content in CWD affects the decay processes, the results highlight the importance of biological interactions associated with bryophytes in forest ecosystems. [ABSTRACT FROM AUTHOR]

Plant root architecture is developmentally plastic in response to fluctuating nutrient levels in the soil. Part of this developmental plasticity is the formation of dedicated root cells and organs to host mutualistic symbionts. Structures like nitrogen‐fixing nodules serve as alternative nutrient acquisition strategies during starvation conditions. Some root systems can also form myconodules—globular root structures that can host mycorrhizal fungi. The myconodule association is different from the wide‐spread arbuscular mycorrhization. This range of symbiotic associations provides different degrees of compartmentalisation, from the cellular to organ scale, which allows the plant host to regulate the entry and extent of symbiotic interactions. In this review, we discuss the degrees of symbiont compartmentalisation by the plant host as a developmental strategy and speculate how spatial confinement mitigates risks associated with root symbiosis. Summary statement: In this review, we aim to synthesize a picture of how plants manage microorganism associations through a common theme of compartmentalising symbiotic microbes into specific cell‐types or organs. We discuss a range of examples of symbiosis, including 'myconodules'—nodule‐like organs that host endosymbiotic fungi and, in some species, are co‐colonized with nitrogen‐fixing bacteria. The context of growth and safety trade‐offs associated with microbe engagement may help explain both extant and extinct types of endosymbiotic nodules. [ABSTRACT FROM AUTHOR]

Summary: Symbiotic nitrogen fixation carried out by the interaction between legumes and rhizobia is the main source of nitrogen in natural ecosystems and in sustainable agriculture. For the symbiosis to be viable, nutrient exchange between the partners is essential. Transition metals are among the nutrients delivered to the nitrogen‐fixing bacteria within the legume root nodule cells. These elements are used as cofactors for many of the enzymes controlling nodule development and function, including nitrogenase, the only known enzyme able to convert N2 into NH3. In this review, we discuss the current knowledge on how iron, zinc, copper, and molybdenum reach the nodules, how they are delivered to nodule cells, and how they are transferred to nitrogen‐fixing bacteria within. [ABSTRACT FROM AUTHOR]

We report a rationally designed membrane‐intercalating conjugated oligoelectrolyte (COE), namely COE‐IC, which endows aerobic N2‐fixing bacteria Azotobacter vinelandii with a light‐harvesting ability that enables photosynthetic ammonia production. COE‐IC possesses an acceptor‐donor‐acceptor (A‐D‐A) type conjugated core, which promotes visible light absorption with a high molar extinction coefficient. Furthermore, COE‐IC spontaneously associates with A. vinelandii to form a biohybrid in which the COE is intercalated within the lipid bilayer membrane. In the presence of L‐ascorbate as a sacrificial electron donor, the resulting COE‐IC/A. vinelandii biohybrid showed a 2.4‐fold increase in light‐driven ammonia production, as compared to the control. Photoinduced enhancement of bacterial biomass and production of L‐amino acids is also observed. Introduction of isotopically enriched 15N2 atmosphere led to the enrichment of 15N‐containing intracellular metabolites, consistent with the products being generated from atmospheric N2. [ABSTRACT FROM AUTHOR]

A conductive polymer‐based photosynthetic biohybrid is constructed to enhance biological nitrogen fixation by increasing nitrogenase activity in the non‐photosynthetic bacterium Azotobacter Chroococcum (A. Chroococcum). The light‐harvesting cationic poly(fluorene‐alt‐phenylene) (PFP) electrostatically binds to the surface of the bacteria and possesses satisfactory conductivity to facilitate electron transfer to the bacterium, promoting the nitrogen fixation pathway through redox proteins on the surface of the bacteria when under illumination. Therefore, the nitrogenase activity, hydrogen, NH4+‐N and L‐amino acids production are increased by 260 %, 37 %, 44 %, and 47 %, respectively. The expression levels of nifD and nifK encoding molybdenum‐iron (MoFe) protein and relevant nitrogen‐fixing proteins are up‐regulated. These photoactive conductive polymer‐bacteria biohybrids provide a new method for improving the biological nitrogen fixation capability of non‐photosynthetic nitrogen‐fixing bacteria. [ABSTRACT FROM AUTHOR]

Recent studies have described active nitrogen fixation in high‐latitude waters, but the ecological controls on the occurrence or activity of nitrogen‐fixing organisms (diazotrophs) in such systems remain unknown. Turnover rates and top‐down controls are also general knowledge gaps for marine diazotrophs. We detected abundant UCYN‐A (endosymbiotic nitrogen‐fixing cyanobacteria) in the Gulf of Anadyr, western Bering Sea, which correlated with high dissolved iron to dissolved inorganic nitrogen ratios (Fe : DIN) due to riverine input. Growth and grazing mortality of UCYN‐A sublineages were almost balanced with higher biomass‐turnover rates compared to the whole phytoplankton community, indicating selective grazing of UCYN‐A in nitrogen‐depleted waters. Grazing rates on UCYN‐A1 (small cells) were higher than for UCYN‐A2/3/4 (large cells), consistent with the general size dependence of phytoplankton growth and grazing mortality. We found that Fe : DIN is a major determinant of UCYN‐A abundances in high‐latitude waters, where UCYN‐A could make substantial contributions to plankton food‐web cycling. [ABSTRACT FROM AUTHOR]

Root‐associated microbiomes play pivotal roles in rice plant nutrition and productivity. We studied how the Liaoxing 1 and Akitakomachi rice cultivars, which are characterized by high and low yields, respectively, modified the nutrient content, enzymatic activities in the rhizosphere, and bacterial community structure in the root endosphere, rhizosphere, and bulk soil with and without nitrogen application. Rice genotype and nitrogen application amount are both essential driving factors for changes in the nutrient content, enzymatic activities, and bacterial community. Bacterial community structure varied significantly across the three root‐associated compartments, and rhizosphere and bulk soil harbored significantly higher diversity and richness of bacteria than root endophytes. Bacterial communities in the root endosphere are mainly structured by rice cultivars. Nitrogen fertilization application influenced the compositions and functions of bacterial groups across the three compartments. Prolixibacteriaceae, as nitrogen‐fixing bacteria, were significantly enriched in the rhizosphere, and the relative abundance in the rhizosphere of the high‐yield cultivar was higher than that of the low‐yield cultivar. Rice genotype and nitrogen application amount are both essential driving factors for changes in nutrient content and enzymatic activities. In this study, it was found that the higher available nitrogen content and soil enzymatic activities in the rhizosphere of the high‐yield rice cultivar promoted rice nitrogen use, and they ultimately formed a positive feedback loop. Our results indicated that root‐associated bacterial communities combined with soil enzymatic activities strengthen the transformation of nitrogen nutrients, suggesting that the interactions could represent a mechanism for enhancing rice yield. [ABSTRACT FROM AUTHOR]

Evidence is accumulating that the soil microbiome—the community of microorganisms living in soils—has a major effect on plant traits and fitness. However, most work to date has taken place under controlled laboratory conditions and has not experimentally disentangled the effect of the soil microbiome on plant performance from the effects of key endosymbiotic constituents. As a result, it is difficult to extrapolate from existing data to understand the role of the soil microbiome in natural plant populations. To address this gap, we performed a field experiment using the black medick Medicago lupulina to test how the soil microbiome influences plant performance and colonization by two root endosymbionts (the mutualistic nitrogen‐fixing bacteria Ensifer spp. and the parasitic root‐knot nematode Meloidogyne hapla) under natural conditions. We inoculated all plants with nitrogen‐fixing bacteria and factorially manipulated the soil microbiome and nematode infection. We found that plants grown in microbe‐depleted soil exhibit greater mortality, but that among the survivors, there was no effect of the soil microbiome on plant performance (shoot biomass, root biomass, or shoot‐to‐root ratio). The soil microbiome also impacted parasitic nematode infection and affected colonization by mutualistic nitrogen‐fixing bacteria in a plant genotype‐dependent manner, increasing colonization in some plant genotypes and decreasing it in others. Our results demonstrate the soil microbiome has complex effects on plant–endosymbiont interactions and may be critical for survival under natural conditions. [ABSTRACT FROM AUTHOR]

Silicon (Si) uptake and deposition (silicification) in tissues is known to alleviate stresses and generally improve plant health. This is mostly studied in Si-high accumulators, such as grasses, with comparatively less known about its effects on other plant functional groups, such as legumes. There is speculation that Si may positively impact the symbiosis between legumes and the nitrogen-fixing bacteria (rhizobia) they associate with, but this is poorly understood. This study examined the effects of Si enrichment on legume species associated with rhizobia and the potential underlying mechanism of Si impacts. 2. We conducted a glasshouse experiment with lucerne Medicago sativa and barrel medic M. truncatula associated with a model rhizobial strain. Six genotypes (three per species) were either supplemented with Si (+Si) or untreated (-Si). We quantified 16 functional traits which could be classified as plant growth, physiology, elemental chemistry, nodule activity and nitrogen fixation. 3. The two legume species responded to Si distinctively. For example, Si supplementation increased shoot biomass by more than 10% in lucerne but growth was unaffected in barrel medic. Conversely, nitrogen-fixing enzyme (nitrogenase) activity was promoted by more than 85% in +Si barrel medic plants but not in lucerne. Moreover, Si supplementation of lucerne increased the concentrations of Si in leaves by more than 36% but not in root nodules. Increased foliar concentrations of Si in lucerne were positively associated with increased shoot and root biomass in Sequel and Trifecta genotypes, respectively. Conversely, Si supplementation of barrel medic increased the concentration of Si in root nodules by 29% but not that in foliar tissues. Nitrogenase activity and where silicification occurred, differed between genotypes in barrel medic; nitrogenase activity was correlated with concentrations of Si in root nodules rather than that in foliar tissues in one genotype (Sephi) but the reverse was true in another (Hannaford). 4. This study demonstrates that two closely related legume species can respond to Si in distinct ways, depending on plant genotype and symbiosis. These results present the overlooked function of Si in legume-rhizobia interactions, which could potentially enhance productivity of this important group of plants. [ABSTRACT FROM AUTHOR]

Nitrogen‐fixing bacteria (NFB) can reduce nitrogen at ambient pressure and temperature. In this study, we treated effluent from a paper mill in sequencing batch reactors (SBRs) and monitored the abundance and activity of NFB with a view to producing a sludge that could work as a biofertilizer. Four reactors were inoculated with activated sludge enriched with NFB and fed with a high C/N waste (100:0.5) from a paper mill. Though the reactors were able to reduce the organic load of the wastewater by up to 89%, they did not have any nitrogen‐fixing activity and showed a decrease in the putative number of NFB (quantified with qPCR). The most abundant species in the reactors treating high C/N paper mill wastewater was identified by Illumina MiSeq 16S rRNA gene amplicon sequencing as Methyloversatilis sp. (relative abundance of 4.4%). Nitrogen fixation was observed when the C/N ratio was increased by adding sucrose. We suspect that real‐world biological nitrogen fixation (BNF) will only occur where there is a C/N ratio ≤100:0.07. Consequently, operators should actively avoid adding or allowing nitrogen in the waste streams if they wish to valorize their sludge and reduce running costs. Practitioner points: Efficient biological wastewater treatment of low nitrogen paper mill effluent was achieved without nutrient supplementation.The sludge was still capable of fixing nitrogen although this process was not observed in the wastewater treatment system.This high C/N wastewater treatment technology could be used with effluents from cassava flour, olive oil, wine and dairy industries. [ABSTRACT FROM AUTHOR]

Nearly all organisms participate in multiple mutualisms, and complementarity within these complex interactions can result in synergistic fitness effects. However, it remains largely untested how multiple mutualisms impact eco‐evolutionary dynamics in interacting species. We tested how multiple microbial mutualists—N‐fixing bacteria and mycorrrhizal fungi—affected selection and heritability of traits in their shared host plant (Medicago truncatula), as well as fitness alignment between partners. Our results demonstrate for the first time that multiple mutualisms synergistically affect the selection and heritability of host traits and enhance fitness alignment between mutualists. Specifically, we found interaction with multiple microbial symbionts doubled the strength of natural selection on a plant architectural trait, resulted in 2‐ to 3‐fold higher heritability of plant reproductive success, and more than doubled fitness alignment between N‐fixing bacteria and plants. These findings show synergism generated by multiple mutualisms extends to key components of microevolutionary change, emphasising the importance of multiple mutualism effects on evolutionary trajectories. [ABSTRACT FROM AUTHOR]

In tropical forests, free‐living Biological nitrogen (N) fixation (BNF) in soil and litter tends to decrease when substrate N concentrations increase, whereas increasing phosphorus (P) and molybdenum (Mo) soil and litter concentrations have been shown to stimulate free‐living BNF rates. Yet, very few studies explored the effects of adding N, P, and Mo together in a single large‐scale fertilization experiment, which would teach us which of these elements constrain or limit BNF activities. At two distinct forest sites in French Guiana, we performed a 3‐year in situ nutrient addition study to explore the effects of N, P, and Mo additions on leaf litter and soil BNF. Additionally, we conducted a short‐term laboratory study with the same nutrient addition treatments (+N, +N+P, +P, +Mo, and +P+Mo). We found that N additions alone suppressed litter free‐living BNF in the field, but not in the short‐term laboratory study, while litter free‐living BNF remained unchanged in response to N+P additions. Additionally, we found that P and P+Mo additions stimulated BNF in leaf litter, both in the field and in the lab, while Mo alone yielded no changes. Soil BNF increased with P and P+Mo additions in only one of the field sites, while in the other site soil BNF increased with Mo and P+Mo additions. We concluded that increased substrate N concentrations suppress BNF. Moreover, both P and Mo have the potential to limit free‐living BNF in these tropical forests, but the balance between P versus Mo limitation is determined by site‐specific characteristics of nutrient supply and demand. Plain Language Summary: Nitrogen fixation by microorganisms is an important source of nitrogen for tropical forests. The controls over this process remain ambiguous but nutrient availability has been put forward as an important regulator. Especially nitrogen, phosphorus, and molybdenum have been shown to affect nitrogen fixation. In this experiment, we tested the effect of adding nitrogen, phosphorus, and molybdenum in different combinations to two mature tropical forest field sites over a period of 3 years. We found that phosphorus mainly causes nitrogen fixation to increase while nitrogen addition leads to a delayed decrease in nitrogen fixation. Molybdenum had a very site‐specific effect and only affected nitrogen fixation in one of the two sites. Key Points: Nitrogen additions cause delayed decreases in nitrogen fixationPhosphorus and molybdenum additions cause site‐specific increases in nitrogen fixationDifferences in soil and litter nutrient stoichiometry likely explain differences in responses to phosphorus and molybdenum fertilization [ABSTRACT FROM AUTHOR]

SUMMARY: Legumes and nitrogen‐fixing rhizobial bacteria establish root nodule symbiosis, which is orchestrated by several plant hormones. Exogenous addition of biologically active gibberellic acid (GA) is known to inhibit root nodule symbiosis. However, the precise role of GA has not been elucidated because of the trace amounts of these hormones in plants and the multiple functions of GAs. Here, we found that GA signaling acts as a key regulator in a long‐distance negative‐feedback system of root nodule symbiosis called autoregulation of nodulation (AON). GA biosynthesis is activated during nodule formation in and around the nodule vascular bundles, and bioactive GAs accumulate in the nodule. In addition, GA signaling induces expression of the symbiotic transcription factor NODULE INCEPTION (NIN) via a cis‐acting region on the NIN promoter. Mutants with deletions of this cis‐acting region have increased susceptibility to rhizobial infection and reduced GA‐induced CLE‐RS1 and CLE‐RS2 expression, suggesting that the inhibitory effect of GAs occurs through AON. This is supported by the GA‐insensitive phenotypes of an AON‐defective mutant of HYPERNODULATION ABERRANT ROOT FORMATION1 (HAR1) and a reciprocal grafting experiment. Thus, endogenous GAs induce NIN expression via its GA‐responsive cis‐acting region, and subsequently the GA‐induced NIN activates the AON system to regulate nodule formation. [ABSTRACT FROM AUTHOR]

The symbiosis between plants and nitrogen‐fixing bacteria is widespread among legumes and actinorhizal plants within the nitrogen‐fixing root nodule (NFN) clade. However, there are major differences, as well as similarities, in the symbioses between actinorhizal plants and Frankia and those of legumes and their associated rhizobia.This review provides an overview of NFN symbioses. We outline the evolution and biogeography of actinorhizal plants and legumes and compare and contrast their microsymbionts and symbiotic processes.Within the NFN clade, a far greater number of nodulated legumes exists, compared with actinorhizal plants, and legumes have a much wider biogeographical distribution. There are genetic and physiological differences between free‐living diazotrophic Frankia and the phylogenetically diverse rhizobia, most strains of which are unable to fix N2ex planta. Actinorhizal nodules are modified lateral roots with a central vascular system, whereas legume nodules are stem‐like organs with peripheral vascular systems. Most legumes contain their microsymbionts within symbiosomes, rather than the infection threads found in actinorhizal nodule cells. Legumes have greater control of their microsymbionts, and those within the Inverted Repeat Lacking Clade impose terminal differentiation on their bacteroids. Legumes also have effective processes for autoregulation of nodulation and downregulation of N2 fixation in response to high levels of soil N. These features of the legume‐rhizobia symbiosis have led to increased efficiencies in N2 fixation.Synthesis. We suggest that these characteristic features of the legume‐rhizobia symbiosis, specifically legumes' greater flexibility in the choice of microsymbiont partner and the evolution of increased efficiencies in N2 fixation, are factors that can explain why the majority of species within the Leguminosae have retained the ability to nodulate and how this has contributed to their evolutionary success. [ABSTRACT FROM AUTHOR]

Soil nitrogen (N)‐fixing bacteria community plays an important role in the N cycling process in soil, but there is still limited information about how the soil microbes that drive this process to respond to combined application of tillage and crop residue management under the double‐cropping rice (Oryza sativa L.) paddy field in southern of China. Therefore, the effects of 6‐years short‐term tillage treatment on soil N‐fixing bacteria community under the double‐cropping rice paddy field in southern China were studied by using the polymerase chain reaction‐denaturing gradient gel electrophoresis method. The field experiment included four tillage treatments: conventional tillage with crop residue incorporation (CT), rotary tillage with crop residue incorporation (RT), no‐tillage with crop residue retention (NT), rotary tillage with crop residue removed as control (RTO). The results showed that the diversity index and richness index of cbbLR and nifH genes with CT, RT, and NT treatments were increased, compared with RTO treatment. Compared with RTO treatment, the abundance of cbbLR gene with CT, RT, and NT treatments were increased by 6.54, 4.73, and 2.78 times, respectively. Meanwhile, the abundance of nifH gene with CT, RT, and NT treatments were 5.32, 3.71, and 2.45 times higher than that of RTO treatment. The results also indicated that soil autotrophic Azotobacter and nitrogenase activity with CT and RT treatments were significantly higher (p <.05) than that of RTO treatment. There was an obvious difference in characteristic of soil N‐fixing bacteria community between the application of crop residue and without crop residue input treatments. In summary, the results indicated that the abundance of N‐fixing bacteria community in the double‐cropping rice paddy field increased with conventional tillage and rotary tillage practice. [ABSTRACT FROM AUTHOR]

In the last decade, the known biogeography of nitrogen fixation in the ocean has been expanded to colder and nitrogen‐rich coastal environments. The symbiotic nitrogen‐fixing cyanobacteria group A (UCYN‐A) has been revealed as one of the most abundant and widespread nitrogen‐fixers, and includes several sublineages that live associated with genetically distinct but closely related prymnesiophyte hosts. The UCYN‐A1 sublineage is associated with an open ocean picoplanktonic prymnesiophyte, whereas UCYN‐A2 is associated with the coastal nanoplanktonic coccolithophore Braarudosphaera bigelowii, suggesting that different sublineages may be adapted to different environments. Here, we study the diversity of nifH genes present at the Santa Cruz Municipal Wharf in the Monterey Bay (MB), California, and report for the first time the presence of multiple UCYN‐A sublineages, unexpectedly dominated by the UCYN‐A2 sublineage. Sequence and quantitative PCR data over an 8‐year time‐series (2011–2018) showed a shift toward increasing UCYN‐A2 abundances after 2013, and a marked seasonality for this sublineage which was present during summer‐fall months, coinciding with the upwelling‐relaxation period in the MB. Increased abundances corresponded to positive temperature anomalies in MB, and we discuss the possibility of a benthic life stage of the associated coccolithophore host to explain the seasonal pattern. The dominance of UCYN‐A2 in coastal waters of the MB underscores the need to further explore the habitat preference of the different sublineages in order to provide additional support for the hypothesis that UCYN‐A1 and UCYN‐A2 sublineages are different ecotypes. [ABSTRACT FROM AUTHOR]

Many insects feed on a low‐nitrogen diet, and the origin of their nitrogen supply is poorly understood. It has been hypothesized that some insects rely on nitrogen‐fixing bacteria (diazotrophs) to supplement their diets. Nitrogen fixation by diazotrophs has been extensively studied and convincingly demonstrated in termites, while evidence for the occurrence and role of nitrogen fixation in the diet of other insects is less conclusive. Here, we summarize the methods to detect nitrogen fixation in insects and review the available evidence for its occurrence (focusing on insects other than termites). We distinguish between three aspects of nitrogen fixation investigations: (i) detecting the presence of potential diazotrophs; (ii) detecting the activity of the nitrogen‐fixing enzyme; and (iii) detecting the assimilation of fixed nitrogen into the insect tissues. We show that although evidence from investigations of the first aspect reveals ample opportunities for interactions with potential diazotrophs in a variety of insects, demonstrations of actual biological nitrogen fixation and the assimilation of fixed nitrogen are restricted to very few insect groups, including wood‐feeding beetles, fruit flies, leafcutter ants, and a wood wasp. We then discuss potential implications for the insect's fitness and for the ecosystem as a whole. We suggest that combining these multiple approaches is crucial for the study of nitrogen fixation in insects, and argue that further demonstrations are desperately needed in order to determine the relative importance of diazotrophs for insect diet and fitness, as well as to evaluate their overall impact on the ecosystem. [ABSTRACT FROM AUTHOR]

To date, the functional role of plant silicon has mostly been investigated in grasses (Poaceae). This potentially overlooks the importance of silicon in other plant functional groups such as legumes (Fabaceae). Legumes form a symbiotic relationship with nitrogen‐fixing bacteria (rhizobia) inside the root nodules for fixing atmospheric nitrogen. A small, but growing number of studies suggest that silicon promotes this symbiotic relationship.We consider how legumes may take up and deposit silicon relative to what is known about these processes in grasses. We synthesize information about how silicon affects legume growth and function in the context of environmental stresses and the legume–rhizobia symbiosis.The available literature indicates that silicon is broadly beneficial to legumes, alleviating the effects of stresses including metal toxicity, salinity, alkalinity and pathogens. Crucially, there is also evidence that silicon promotes the legume–rhizobia interaction including increased root nodulation, numbers of bacteroids and nitrogen fixation across several legume species.We propose a model for how silicon may benefit the legume–rhizobia interaction. We hypothesize that silicification in the tissues may reduce the high metabolic cost of carbon‐based compounds in cell wall construction, optimize solute transport and gas exchange in root nodules and/or promote protection against environmental stresses. We therefore propose a hypothetical framework to better understanding the impacts of silicon on legume–rhizobia relationships.We also suggest potential research priorities that would help us to better understand the functional role of silicon in nitrogen‐fixing legumes. These research priorities focus on characterizing how silicon affects the chemical dialogues between the host plant and its rhizobial partner, how silicon is deposited in legume roots and how resources are exchanged by the two. Given the growing importance of legumes at a global scale, silicon could play a vital role in improving legume health and productivity with manifold environmental benefits. A free Plain Language Summary can be found within the Supporting Information of this article. [ABSTRACT FROM AUTHOR]

l‐Tyrosine (Tyr) is an aromatic amino acid synthesized de novo in plants and microbes downstream of the shikimate pathway. In plants, Tyr and a Tyr pathway intermediate, 4‐hydroxyphenylpyruvate (HPP), are precursors to numerous specialized metabolites, which are crucial for plant and human health. Tyr is synthesized in the plastids by a TyrA family enzyme, arogenate dehydrogenase (ADH/TyrAa), which is feedback inhibited by Tyr. Additionally, many legumes possess prephenate dehydrogenases (PDH/TyrAp), which are insensitive to Tyr and localized to the cytosol. Yet the role of PDH enzymes in legumes is currently unknown. This study isolated and characterized Tnt1‐transposon mutants of MtPDH1 (pdh1) in Medicago truncatula to investigate PDH function. The pdh1 mutants lacked PDH transcript and PDH activity, and displayed little aberrant morphological phenotypes under standard growth conditions, providing genetic evidence that MtPDH1 is responsible for the PDH activity detected in M. truncatula. Though plant PDH enzymes and activity have been specifically found in legumes, nodule number and nitrogenase activity of pdh1 mutants were not significantly reduced compared with wild‐type (Wt) during symbiosis with nitrogen‐fixing bacteria. Although Tyr levels were not significantly different between Wt and mutants under standard conditions, when carbon flux was increased by shikimate precursor feeding, mutants accumulated significantly less Tyr than Wt. These data suggest that MtPDH1 is involved in Tyr biosynthesis when the shikimate pathway is stimulated and possibly linked to unidentified legume‐specific specialized metabolism. [ABSTRACT FROM AUTHOR]

Summary: The formation of nitrogen‐fixing nodules on legume hosts is a finely tuned process involving many components of both symbiotic partners. Production of the exopolysaccharide succinoglycan by the nitrogen‐fixing bacterium Sinorhizobium meliloti 1021 is needed for an effective symbiosis with Medicago spp., and the succinyl modification to this polysaccharide is critical. However, it is not known when succinoglycan intervenes in the symbiotic process, and it is not known whether the plant lysin‐motif receptor‐like kinase MtLYK10 intervenes in recognition of succinoglycan, as might be inferred from work on the Lotus japonicus MtLYK10 ortholog, LjEPR3. We studied the symbiotic infection phenotypes of S. meliloti mutants deficient in succinoglycan production or producing modified succinoglycan, in wild‐type Medicago truncatula plants and in Mtlyk10 mutant plants. On wild‐type plants, S. meliloti strains producing no succinoglycan or only unsuccinylated succinoglycan still induced nodule primordia and epidermal infections, but further progression of the symbiotic process was blocked. These S. meliloti mutants induced a more severe infection phenotype on Mtlyk10 mutant plants. Nodulation by succinoglycan‐defective strains was achieved by in trans rescue with a Nod factor‐deficient S. meliloti mutant. While the Nod factor‐deficient strain was always more abundant inside nodules, the succinoglycan‐deficient strain was more efficient than the strain producing only unsuccinylated succinoglycan. Together, these data show that succinylated succinoglycan is essential for infection thread formation in M. truncatula, and that MtLYK10 plays an important, but different role in this symbiotic process. These data also suggest that succinoglycan is more important than Nod factors for bacterial survival inside nodules. Significance Statement: This work should be of interest across the field of plant–microbe endosymbioses by providing insights into key determinants of both bacteria and plants for successful microbial host invasion. It is timely with much current interest in symbiotic roles of plant LysM‐RLK proteins and the evolutionary origins of nitrogen‐fixing endosymbiosis. MtLYK10 is a Medicago truncatula component that specifically intervenes in rhizobial infection, independently of succinoglycan recognition. [ABSTRACT FROM AUTHOR]

The availability of compatible mutualistic soil microbes could influence the invasion success of non‐native plant species. Specifically, there may be spatial variation in the distribution of compatible microbes, and species‐specific variation in plant host ability to associate with available microbes. Although either or both factors could promote or limit invasion, the scale over which most studies are conducted makes it difficult to examine these two possibilities simultaneously. However, this is critical to identifying a role of soil microbes in invasion.A series of recent research projects focused on interactions between Australian Acacia and nitrogen‐fixing bacteria (rhizobia) at multiple spatial scales, from the local to the inter‐continental, has allowed us to evaluate this question. Collectively, this research reveals that nodulation, performance and rhizobial community composition are all broadly similar across spatial scales and differentially invasive species.Synthesis and applications. We argue that current research provides convincing evidence that interactions with rhizobia do not determine invasion success in Acacia, but instead highlights key knowledge gaps that remain unfilled. Importantly, the ease with which non‐native Acacia species form mutualistic associations with rhizobia, regardless of invasive status, highlights the critical need to understand the impacts of all non‐native Acacia on native soil communities. [ABSTRACT FROM AUTHOR]

Increased stratification and mixed layer shoaling of the surface ocean resulting from warming can lead to exposure of marine dinitrogen (N2)‐fixing cyanobacteria to higher levels of inhibitory ultraviolet (UV) radiation. These same processes also reduce vertically advected supplies of the potentially limiting nutrient phosphorus (P) to N2 fixers. It is currently unknown how UV inhibition and P limitation interact to affect the biogeochemical cycles of nitrogen and carbon in these biogeochemically critical microbes. We investigated the responses of the important and widespread marine N2‐fixing cyanobacteria Crocosphaera (strain WH0005) and Trichodesmium (strains IMS 101 and GBR) to UV‐A and UV‐B under P‐replete and P‐limited conditions. Growth, N2 fixation, and carbon dioxide (CO2) fixation rates of Trichodesmium IMS 101 and Crocosphaera were negatively affected by UV exposure. This inhibition was greater for Trichodesmium IMS 101 than for Crocosphaera, which fixes N2 only during the night and so avoids direct UV damage. Negative effects of UV on both IMS 101 and Crocosphaera were less in P‐limited cultures than in P‐replete cultures. In contrast, no UV inhibition was observed in GBR, regardless of P availability. UV inhibition was related to different amounts of UV‐absorbing compounds produced by these isolates. Responses to UV radiation and P availability interactions were taxon‐specific, but our results indicated that in general, UV radiation effects on Trichodesmium and Crocosphaera range from negative to neutral. UV inhibition and its interactions with P limitation may thus have a substantial influence on the present day and future nitrogen and carbon cycles of the ocean. [ABSTRACT FROM AUTHOR]

2‐oxoglutarate (2‐OG) is a central metabolite that acts as a signaling molecule informing about the status of the carbon/nitrogen balance of the cell. In recent years, some transcriptional regulators and even two‐component systems have been described as 2‐OG sensors. In the nitrogen‐fixing cyanobacterium Anabaena sp. PCC 7120, two master regulators, NtcA and FurA, are deeply involved in the regulation of nitrogen metabolism. Both of them show a complex intertwined regulatory circuit to achieve a suitable regulation of nitrogen fixation. In this work, 2‐OG is found to bind FurA, modulating the specific binding of FurA to the ntcA promoter. This study provides evidence of a new additional control point in the complex network controlled by the NtcA and FurA proteins. [ABSTRACT FROM AUTHOR]

The current trend of increasing proportion of cultivation of transgenic Bt crops is pushing towards dramatic destabilization of the agroecosystem, thus raising severe concerns about the sustainability of transgenic Bt crops as an effective management tool for the control of target insect pests in the future. Rhizobacteria is the key biological regulator to ameliorate soil‐nitrogen utilization efficiency of crop plants, especially transgenic Bt crops. A laboratory study quantified the impacts of transgenic Bt maize (Line IE09S034 with Cry1Ie vs. non‐Bt maize cv. Xianyu335) inoculated with Azospirillum brasilense (AB) and Azotobacter chroococcum (AC) on the growth, development and food utilization of a target lepidopteran insect, Mythimna separata. The results showed that the inoculation of rhizobacteria significantly prolonged the larval lifespan and pupal duration, increased RCR and AD, reduced pupal weight, pupation rate, fecundity, RGR, ECD and ECI, and shortened adult longevity of M. separata fed on transgenic Bt maize, while exact opposite trends were found in these measured indexes of growth, development and food utilization for M. separata fed on non‐Bt maize inoculated with AB and AC compared with the buffer control in both years. Thus, the results clearly depicted that the inoculation of rhizobacteria had opposite influences on the growth, development and food utilization of M. separata fed on transgenic Bt maize. Presumably, rhizobacteria inoculation can be used to stimulate plant–soil‐nitrogen uptake and promote plant growth for transgenic Bt maize and non‐Bt maize, simultaneously increasing Bt toxin production and enhancing resistance efficiency against target lepidopteran pests for transgenic Bt maize. [ABSTRACT FROM AUTHOR]

Studies of plant resource‐use strategies along environmental gradients often assume that dry matter partitioning represents an individual's energy investment in foraging for above‐ versus below‐ground resources. However, ecosystem‐level studies of total below‐ground carbon allocation (TBCA) in forests do not support the equivalency of energy (carbon) and dry matter partitioning, in part because allocation of carbon to below‐ground pools and fluxes that are not accounted for by root biomass (e.g., mycorrhizal hyphae, rhizodeposition; root and soil respiration) can be substantial. Here, we apply this reasoning to individual plants in controlled environments and ask whether dry matter partitioning below‐ground (root mass fraction, RMF) accurately reflects TBCA in studies of optimal partitioning theory.We quantified the relationship between RMF and TBCA in individual plants, using 311 observations from 51 studies that simultaneously measured both allocation variables. Our analysis included tests of whether the RMF‐TBCA relationship depended on mutualist soil microbes, plant growth form, age and study methodology including isotopic pulse–chase duration.We found that RMF was a poor proxy for below‐ground energy investment. This disconnect of RMF and TBCA was driven in part by plants of low RMF (<0.4) exhibiting significantly higher rates of root and soil respiration per unit root mass than plants of high RMF. Root colonization by mutualist microbes, including arbuscular mycorrhizal fungi and nitrogen‐fixing bacteria, increased TBCA by 5%–7%, and TBCA was lower in grasses than other species by 9%–16%. These patterns were evident for relationships assessed both within and between species.We conclude that optimal partitioning studies of plants along environmental gradients are likely to underestimate plant energy allocation below‐ground if the C costs of root and soil respiration are ignored, especially under conditions favouring low RMF. Because energy rather than biomass better reflects how assimilated C supports fitness, this omission of respired C suggests existing studies misrepresent the significance of below‐ground processes to plant function. A free Plain Language Summary can be found within the Supporting Information of this article. [ABSTRACT FROM AUTHOR]

In Azospirillum brasilense, a gram‐negative nitrogen‐fixing bacterium, l‐arabinose is converted to α‐ketoglutarate through a nonphosphorylative metabolic pathway. In the first step in the pathway, l‐arabinose is oxidized to l‐arabino‐γ‐lactone by NAD(P)‐dependent l‐arabinose 1‐dehydrogenase (AraDH) belonging to the glucose‐fructose oxidoreductase/inositol dehydrogenase/rhizopine catabolism protein (Gfo/Idh/MocA) family. Here, we determined the crystal structures of apo‐ and NADP‐bound AraDH at 1.5 and 2.2 Å resolutions, respectively. A docking model of l‐arabinose and NADP‐bound AraDH and structure‐based mutational analyses suggest that Lys91 or Asp169 serves as a catalytic base and that Glu147, His153, and Asn173 are responsible for substrate recognition. In particular, Asn173 may play a role in the discrimination between l‐arabinose and d‐xylose, the C4 epimer of l‐arabinose. [ABSTRACT FROM AUTHOR]

Summary: Glycogen synthase kinase/SHAGGY‐like kinases (SKs) are a highly conserved family of signaling proteins that participate in many developmental, cell‐differentiation, and metabolic signaling pathways in plants and animals. Here, we investigate the involvement of SKs in legume nodulation, a process requiring the integration of multiple signaling pathways. We describe a group of SKs in the model legume Lotus japonicus (LSKs), two of which respond to inoculation with the symbiotic nitrogen‐fixing bacterium Mesorhizobium loti. RNAi knock‐down plants and an insertion mutant for one of these genes, LSK1, display increased nodulation. Ηairy‐root lines overexpressing LSK1 form only marginally fewer mature nodules compared with controls. The expression levels of genes involved in the autoregulation of nodulation (AON) mechanism are affected in LSK1 knock‐down plants at low nitrate levels, both at early and late stages of nodulation. At higher levels of nitrate, these same plants show the opposite expression pattern of AON‐related genes and lose the hypernodulation phenotype. Our findings reveal an additional role for the versatile SK gene family in integrating the signaling pathways governing legume nodulation, and pave the way for further study of their functions in legumes. Significance statement: SHAGGY‐like kinases are conserved eukaryotic signaling proteins involved in the control of developmental processes, the integration of cell division signals and in modulating metabolism. Their potential role in legume nodulation, where the integration of all these functions is required, has not yet been examined, however. We describe a family of six SHAGGY‐like kinases in the legume Lotus japonicus (LSKs) and provide evidence for a role of one family member, LSK1, in limiting the nodule number during colonization by symbiotic rhizobia. [ABSTRACT FROM AUTHOR]

Summary: Almost all plant species interact with one or more symbioses somewhere within their distribution range.Bringing together plant trait data and growth responses to symbioses spanning 552 plant species, we provide for the first time on a large scale (597 studies) a quantitative synthesis on plant performance differences between eight major types of symbiosis, including mycorrhizas, N‐fixing bacteria, fungal endophytes and ant–plant interactions.Frequency distributions of plant growth responses varied considerably between different types of symbiosis, in terms of both mean effect and 'risk', defined here as percentage of experiments reporting a negative effect of symbiosis on plants. Contrary to expectation, plant traits were poor predictors of growth response across and within all eight symbiotic associations. Our analysis showed no systematic additive effect when a host plant engaged in two functionally different symbioses.This synthesis suggests that plant species' ecological strategies have little effect in determining the influence of a symbiosis on host plant growth. Reliable quantification of differences in plant performance across symbioses will prove valuable for developing general hypotheses on how species become engaged in mutualisms without a guarantee of net returns. [ABSTRACT FROM AUTHOR]

A major goal in microbial ecology is to understand the factors that structure bacterial communities across space and time. For microbes that are plant symbionts, community assembly processes can lead to either a positive or negative relationship between plant size or age and soil microbe diversity. Here, we evaluated the extent to which such relationships exist within a single legume species (Acacia acuminata) and their naturally occurring symbiotic nitrogen‐fixing bacteria (rhizobia).We quantified the diversity of rhizobia that associate with A. acuminata trees of variable size spanning a large environmental gradient in southwest Australia (72 trees in 24 sites spread across ~300,000 km2), using metabarcoding. We modelled rhizobia diversity using 16S exact genetic variants, in a binomial multivariate statistical framework that controlled for climate and local soil characteristics.We identified two major phylogenetic clades of rhizobia that associate with A. acuminata. Soil sampled at the base of larger Acacia trees contained a higher richness of rhizobia genetic variants. Each major clade responds differently to environmental factors (climate and soil characteristics), but the positive association between tree size and rhizobia genetic diversity was mainly driven by responses from one of the two clades. Overall tree size explained more variation than any other factor, resulting in a ~3‐fold increase in total richness and clade diversity from the smallest to the largest trees.Synthesis. Previous studies have shown that plant host species is important in structuring microbial soil communities in the rhizosphere. Our results show that host size or age within a single plant species can also structure diversity of at least one group of soil microbes. A positive relationship between plant host size and rhizobia diversity suggests that hosts may modify the niche space of their surrounding soil (niche construction hypothesis) enabling a higher richness of microbial taxa. That different rhizobial groups responded differently to host size and other ecological factors suggests that rhizobia is not an ecologically uniform group, and that entirely neutral explanations for our results are unlikely. Host plants may be analogous to "islands," where larger plant hosts may accumulate diversity over time, through migration opportunities. Previous studies have shown that plant host species is important in structuring microbial soil communities in the rhizosphere. Our results show that host size or age within a single plant species can also structure diversity of at least one group of soil microbes. A positive relationship between plant host size and rhizobia diversity suggests that hosts may modify the niche space of their surrounding soil (niche construction hypothesis) enabling a higher richness of microbial taxa. That different rhizobial groups responded differently to host size and other ecological factors suggests that rhizobia is not an ecologically uniform group, and that entirely neutral explanations for our results are unlikely. Host plants may be analogous to "islands," where larger plant hosts may accumulate diversity over time, through migration opportunities. [ABSTRACT FROM AUTHOR]

Symbiotic nitrogen fixation in legume root nodules requires a steady supply of molybdenum for synthesis of the iron‐molybdenum cofactor of nitrogenase. This nutrient has to be provided by the host plant from the soil, crossing several symplastically disconnected compartments through molybdate transporters, including members of the MOT1 family. Medicago truncatula Molybdate Transporter (MtMOT) 1.2 is a Medicago truncatula MOT1 family member located in the endodermal cells in roots and nodules. Immunolocalization of a tagged MtMOT1.2 indicates that it is associated to the plasma membrane and to intracellular membrane systems, where it would be transporting molybdate towards the cytosol, as indicated in yeast transport assays. Loss‐of‐function mot1.2‐1 mutant showed reduced growth compared with wild‐type plants when nitrogen fixation was required but not when nitrogen was provided as nitrate. While no effect on molybdenum‐dependent nitrate reductase activity was observed, nitrogenase activity was severely affected, explaining the observed difference of growth depending on nitrogen source. This phenotype was the result of molybdate not reaching the nitrogen‐fixing nodules, since genetic complementation with a wild‐type MtMOT1.2 gene or molybdate‐fortification of the nutrient solution, both restored wild‐type levels of growth and nitrogenase activity. These results support a model in which MtMOT1.2 would mediate molybdate delivery by the vasculature into the nodules. Molybdenum is a key nutrient for symbiotic nitrogen fixation as an element participating in the iron‐molybdenum cofactor of nitrogenase. Here, we show that transporter MOT1.2 from model legume Medicago truncatula is responsible for directing this nutrient to nitrogen fixing nodules. [ABSTRACT FROM AUTHOR]

Summary: Dinitrogen (N2) fixing bacteria (diazotrophs) are an important source of new nitrogen in oligotrophic environments and represent stable members of the microbiome in tropical corals, while information on corals from temperate oligotrophic regions is lacking. Therefore, this study provides new insights into the diversity and activity of diazotrophs associated with the temperate coral Oculina patagonica from the Mediterranean Sea by combining metabarcoding sequencing of amplicons of both the 16S rRNA and nifH genes and 15N2 stable isotope tracer analysis to assess diazotroph‐derived nitrogen (DDN) assimilation by the coral. Results show that the diazotrophic community of O. patagonica is dominated by autotrophic bacteria (i.e. Cyanobacteria and Chlorobia). The majority of DDN was assimilated into the tissue and skeletal matrix, and DDN assimilation significantly increased in bleached corals. Thus, diazotrophs may constitute an additional nitrogen source for the coral host, when nutrient exchange with Symbiodinium is disrupted (e.g. bleaching) and external food supply is limited (e.g. oligotrophic summer season). Furthermore, we hypothesize that DDN can facilitate the fast proliferation of endolithic algae, which provide an alternative carbon source for bleached O. patagonica. Overall, O. patagonica could serve as a good model for investigating the importance of diazotrophs in coral recovery from bleaching. [ABSTRACT FROM AUTHOR]

Summary: The symbiotic unicellular cyanobacterium Candidatus Atelocyanobacterium thalassa (UCYN‐A) is one of the most abundant and widespread nitrogen (N2)‐fixing cyanobacteria in the ocean. Although it remains uncultivated, multiple sublineages have been detected based on partial nitrogenase (nifH) gene sequences, including the four most commonly detected sublineages UCYN‐A1, UCYN‐A2, UCYN‐A3 and UCYN‐A4. However, very little is known about UCYN‐A3 beyond the nifH sequences from nifH gene diversity surveys. In this study, single cell sorting, DNA sequencing, qPCR and CARD‐FISH assays revealed discrepancies involving the identification of sublineages, which led to new information on the diversity of the UCYN‐A symbiosis. 16S rRNA and nifH gene sequencing on single sorted cells allowed us to identify the 16S rRNA gene of the uncharacterized UCYN‐A3 sublineage. We designed new CARD‐FISH probes that allowed us to distinguish and observe UCYN‐A2 in a coastal location (SIO Pier; San Diego) and UCYN‐A3 in an open ocean location (Station ALOHA; Hawaii). Moreover, we reconstructed about 13% of the UCYN‐A3 genome from Tara Oceans metagenomic data. Finally, our findings unveil the UCYN‐A3 symbiosis in open ocean waters suggesting that the different UCYN‐A sublineages are distributed along different size fractions of the plankton defined by the cell‐size ranges of their prymnesiophyte hosts. [ABSTRACT FROM AUTHOR]

Contents Summary 1199 I. Introduction 1199 II. Selecting beneficial symbionts: one problem, many solutions 1200 III. Control and conflict over legume nodulation 1201 IV. Control and conflict over nodule growth and senescence 1204 V. Conclusion 1204 Acknowledgements 1205 References 1205 Summary: The legume–rhizobia association is a powerful model of the limits of host control over microbes. Legumes regulate the formation of root nodules that house nitrogen‐fixing rhizobia and adjust investment into nodule development and growth. However, the range of fitness outcomes in these traits reveals intense conflicts of interest between the partners. New work that we review and synthesize here shows that legumes have evolved varied mechanisms of control over symbionts, but that host control is often subverted by rhizobia. An outcome of this conflict is that both legumes and rhizobia have evolved numerous traits that can improve their own short‐term fitness in this interaction, but little evidence exists for any net improvement in the joint trait of nitrogen fixation. [ABSTRACT FROM AUTHOR]

Abstract: Grain legumes play a significant role in global food security. They have an advantage over cereals in that they can form symbiotic associations with nitrogen‐fixing bacteria, making them self‐sufficient in terms of nitrogen acquisition. In addition to this superior agronomic trait, grain legumes have excellent nutritional properties and are thus widely used as animal feed as well as in human nutrition. Current global trends towards increased legume consumption and availability of value‐added products, as well as legume production in developing countries require the provision of improved cultivars with better productivity and adaptability. Intensive efforts are thus underway to elaborate genomic resources and gain an improved knowledge base in a number of legume crops. There is also an emerging understanding of the beneficial interactions between legume‐associated organisms, particularly rhizobia and arbuscular mycorrhizal fungi, which result in improved nodulation and nutrient acquisition. The emerging focus on legume breeding for high sustainable yields as well as improved biotic and abiotic stress tolerance traits will serve to close the current gap between grain legume production and demand. With the support from policymakers, this increase in knowledge can be readily translated into increased crop production to meet the demands of an increasing global population. [ABSTRACT FROM AUTHOR]

Summary: The phototrophic alpha‐proteobacterium, Rhodopseudomonas palustris, is a model for studies of regulatory and physiological parameters that control the activity of nitrogenase. This enzyme produces the energy‐rich compound H2, in addition to converting N2 gas to NH3. Nitrogenase is an ATP‐requiring enzyme that uses large amounts of reducing power, but the electron transfer pathway to nitrogenase in R. palustris was incompletely known. Here, we show that the ferredoxin, Fer1, is the primary but not sole electron carrier protein encoded by R. palustris that serves as an electron donor to nitrogenase. A flavodoxin, FldA, is also an important electron donor, especially under iron limitation. We present a model where the electron bifurcating complex, FixABCX, can reduce both ferredoxin and flavodoxin to transfer electrons to nitrogenase, and we present bioinformatic evidence that FixABCX and Fer1 form a conserved electron transfer pathway to nitrogenase in nitrogen‐fixing proteobacteria. These results may be useful in the design of strategies to reroute electrons generated during metabolism of organic compounds to nitrogenase to achieve maximal activity. [ABSTRACT FROM AUTHOR]

Summary: Helicobacter pylori is anomalous among non nitrogen‐fixing bacteria in containing an incomplete NIF system for Fe–S cluster assembly comprising two essential proteins, NifS (cysteine desulfurase) and NifU (scaffold protein). Although nifU deletion strains cannot be obtained via the conventional gene replacement, a NifU‐depleted strain was constructed and shown to be more sensitive to oxidative stress compared to wild‐type (WT) strains. The hp1492 gene, encoding a putative Nfu‐type Fe–S cluster carrier protein, was disrupted in three different H. pylori strains, indicating that it is not essential. However, Δnfu strains have growth deficiency, are more sensitive to oxidative stress and are unable to colonize mouse stomachs. Moreover, Δnfu strains have lower aconitase activity but higher hydrogenase activity than the WT. Recombinant Nfu was found to bind either one [2Fe–2S] or [4Fe–4S] cluster/dimer, based on analytical, UV–visible absorption/CD and resonance Raman studies. A bacterial two‐hybrid system was used to ascertain interactions between Nfu, NifS, NifU and each of 36 putative Fe–S‐containing target proteins. Nfu, NifS and NifU were found to interact with 15, 6 and 29 putative Fe–S proteins respectively. The results indicate that Nfu, NifS and NifU play a major role in the biosynthesis and/or delivery of Fe–S clusters in H. pylori. [ABSTRACT FROM AUTHOR]

Due to their competitive advantage when inorganic nitrogen (N) sources are scarce, it is widely assumed that the abundance and N2‐fixation rate of N2‐fixing cyanobacteria (Nostocales) would increase in response to reduced N loading. If realized, this response would render efforts to improve water quality by N reduction ineffective. To assess this, we performed a microcosm experiment using water from an N limited lake, Langer See, in which phosphorus loading was held constant, while a gradient of decreasing N loading was simulated. Nostocales biovolume increased over time in all microcosms, regardless of the N addition rate, so that no difference in Nostocales biovolume developed between high and low N microcosms. In contrast, the biovolumes of other taxa (especially non‐fixing cyanobacteria) were lower at low N addition rates. N2‐fixation increased in low N microcosms, compensating for 36% of the difference in N addition by day 6. However, this compensation rate was achieved at Nostocales biovolumes far higher than those typical in the studied lake. At biovolumes typical for summer the compensation rate would only account for 7% of the omitted N. If these results were to hold over longer time scales, in shallow polymictic lakes like Langer See, reduced N loading may lower both in‐lake N concentrations and biovolumes of non‐fixing phytoplankton without significantly impacting Nostocales biovolume. [ABSTRACT FROM AUTHOR]

Abstract: Plants associated with symbiotic N‐fixing bacteria play important roles in early successional, riparian and semi‐dry ecosystems. These so‐called N‐fixing plants are widely used for reclamation of disturbed vegetation and improvement of soil fertility in agroforestry. Yet, available information about plants that are capable of establishing nodulation is fragmented and somewhat outdated. This article introduces the NodDB database of N‐fixing plants based on morphological and phylogenetic evidence (available at https://doi.org/10.15156/bio/587469) and discusses plant groups with conflicting reports and interpretation, such as certain legume clades and the Zygophyllaceae family. During angiosperm evolution, N‐fixing plants became common in the fabid rather than in the ‘nitrogen‐fixing’ clade. The global GBIF plant species distribution data indicated that N‐fixing plants tend to be relatively more diverse in savanna and semi‐desert biomes. The compiled and re‐interpreted information about N‐fixing plants enables accurate analyses of biogeography and community ecology of biological N fixation. [ABSTRACT FROM AUTHOR]

Summary: The recent discovery of completely nitrifying Nitrospira demands a re‐examination of nitrifying environments to evaluate their contribution to nitrogen cycling. To approach this challenge, tools are needed to detect and quantify comammox Nitrospira. We present primers for the simultaneous quantification and diversity assessement of both comammox Nitrospira clades. The primers cover a wide range of comammox diversity, spanning all available high quality sequences. We applied these primers to 12 groundwater‐fed rapid sand filters, and found comammox Nitrospira to be abundant in all filters. Clade B comammox comprise the majority (∼75%) of comammox abundance in all filters. Nitrosomonadaceae were present in all filters, although at low abundance (mean = 1.8%). Ordination suggests that temperature impacts the structure of nitrifying communities, and in particular that increasing temperature favours Nitrospira. The nitrogen content of the filter material, sulfate concentration and surface ammonium loading rates shape the structure of the comammox guild in the filters. This work provides an assay for simultaneous detection and diversity assessment of clades A and B comammox Nitrospira, expands our current knowledge of comammox Nitrospira diversity and demonstrates a key role for comammox Nitrospira in nitrification in groundwater‐fed biofilters. [ABSTRACT FROM AUTHOR]

The development of legume nitrogen-fixing nodules is regulated by reactive oxygen species (ROS) produced by symbionts. Several regulators from Rhizobium are involved in ROS sensing. In a previous study, we found that Sinorhizobium meliloti LsrB regulates lipopolysaccharide production and is associated with H2O2 accumulation in alfalfa ( Medicago sativa) nodules. However, its underlying regulatory mechanism remains unclear. Here, we report that the cysteine residues in LsrB are required for adaptation to oxidative stress, gene expression, alfalfa nodulation and nitrogen fixation. Moreover, LsrB directly activated the transcription of lrp3 and gshA (encoding γ-glutamylcysteine synthetase, responsible for glutathione synthesis) and this regulation required the cysteine (Cys) residues in the LsrB substrate-binding domain. The Cys residues could sense oxidative stress via the formation of intermolecular disulfide bonds, generating LsrB dimers and LsrB-DNA complexes. Among the Cys residues, C238 is a positive regulatory site for the induction of downstream genes, whereas C146 and C275 play negative roles in the process. The lsrB mutants with Cys-to-Ser substitutions displayed altered phenotypes in respect to their adaptation to oxidative stress, nodulation and nitrogen fixation-related plant growth. Our findings demonstrate that S. meliloti LsrB modulates alfalfa nodule development by directly regulating downstream gene expression via a post-translational strategy. [ABSTRACT FROM AUTHOR]

Sea buckthorn ( Hippophae rhamnoides L.) is a pioneer plant used for land reclamation and an appropriate material for studying the interactions of symbiotic microorganisms because of its nitrogen-fixing root nodules and mycorrhiza. We used high-throughput sequencing to reveal the diversities and community structures of rhizospheric fungi and their link with nitrogen-fixing Frankia harbored in sea buckthorn collected along an altitude gradient from the Qinghai Tibet Plateau to interior areas. We found that the fungal diversities and compositions varied between different sites. Ascomycota, Basidiomycota, and Zygomycota were the dominant phyla. The distribution of sea buckthorn rhizospheric fungi was driven by both environmental factors and the geographic distance. Among all examined soil characteristics, altitude, AP, and pH were found to have significant ( p < 0.05) effect on the rhizospheric fungal community. The rhizospheric fungal communities became more distinct as the distance increased. Moreover, co-inertia analysis identified significant co-structures between Frankia and AMF communities in the rhizosphere of sea buckthorn. We conclude that at the large scale, there are certain linkages between nitrogen-fixing bacteria and the AMF expressed in the distributional pattern. [ABSTRACT FROM AUTHOR]

The health and functioning of reef-building corals is dependent on a balanced association with prokaryotic and eukaryotic microbes. The coral skeleton harbours numerous endolithic microbes, but their diversity, ecological roles and responses to environmental stress, including ocean acidification ( OA), are not well characterized. This study tests whether pH affects the diversity and structure of prokaryotic and eukaryotic algal communities associated with skeletons of Porites spp. using targeted amplicon (16S r RNA gene, UPA and tufA) sequencing. We found that the composition of endolithic communities in the massive coral Porites spp. inhabiting a naturally high pCO2 reef (avg. pCO2 811 μatm) is not significantly different from corals inhabiting reference sites (avg. pCO2 357 μatm), suggesting that these microbiomes are less disturbed by OA than previously thought. Possible explanations may be that the endolithic microhabitat is highly homeostatic or that the endolithic micro-organisms are well adapted to a wide pH range. Some of the microbial taxa identified include nitrogen-fixing bacteria ( Rhizobiales and cyanobacteria), algicidal bacteria in the phylum Bacteroidetes, symbiotic bacteria in the family Endozoicomoniaceae, and endolithic green algae, considered the major microbial agent of reef bioerosion. Additionally, we test whether host species has an effect on the endolithic community structure. We show that the endolithic community of massive Porites spp. is substantially different and more diverse than that found in skeletons of the branching species Seriatopora hystrix and Pocillopora damicornis. This study reveals highly diverse and structured microbial communities in Porites spp. skeletons that are possibly resilient to OA. [ABSTRACT FROM AUTHOR]

BACKGROUND Chickpea ( Cicer arietinum L.) is a widely cropped pulse and an important source of proteins for humans. In Mediterranean regions it is predicted that drought will reduce soil moisture and become a major issue in agricultural practice. Nitrogen (N)-fixing bacteria and arbuscular mycorrhizal (AM) fungi have the potential to improve plant growth and drought tolerance. The aim of the study was to assess the effects of N-fixing bacteria and AM fungi on the growth, grain yield and protein content of chickpea under water deficit. RESULTS Plants inoculated with Mesorhizobium mediterraneum or Rhizophagus irregularis without water deficit and inoculated with M. mediterraneum under moderate water deficit had significant increases in biomass. Inoculation with microbial symbionts brought no benefits to chickpea under severe water deficit. However, under moderate water deficit grain crude protein was increased by 13%, 17% and 22% in plants inoculated with M. mediterraneum, R. irregularis and M. mediterraneum + R. irregularis, respectively. CONCLUSION Inoculation with N-fixing bacteria and AM fungi has the potential to benefit agricultural production of chickpea under water deficit conditions and to contribute to increased grain protein content. © 2017 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

The abundance of nitrogen (N)-fixing plants in ecosystems where phosphorus (P) limits plant productivity poses a paradox because N fixation entails a high P cost. One explanation for this paradox is that the N-fixing strategy allows greater root phosphatase activity to enhance P acquisition from organic sources, but evidence to support this contention is limited., We measured root phosphomonoesterase ( PME) activity of 10 N-fixing species, including rhizobial legumes and actinorhizal Allocasuarina species, and eight non-N-fixing species across a retrogressive soil chronosequence showing a clear shift from N to P limitation of plant growth and representing a strong natural gradient in P availability., Legumes showed greater root PME activity than non-legumes, with the difference between these two groups increasing markedly as soil P availability declined. By contrast, root PME activity of actinorhizal species was always lower than that of co-occurring legumes and not different from non-N-fixing plants., The difference in root PME activity between legumes and actinorhizal plants was not reflected in a greater or similar reliance on N fixation for N acquisition by actinorhizal species compared to co-occurring legumes., Synthesis. Our results support the idea that N-fixing legumes show high root phosphatase activity, especially at low soil P availability, but suggest that this is a phylogenetically conserved trait rather than one directly linked to their N-fixation capacity. [ABSTRACT FROM AUTHOR]

Small non-coding RNAs (sRNAs) are expected to have pivotal roles in the adaptive responses underlying symbiosis of nitrogen-fixing rhizobia with legumes. Here, we provide primary insights into the function and activity mechanism of the Sinorhizobium meliloti trans-sRNA NfeR1 (Nodule Formation Efficiency RNA). Northern blot probing and transcription tracking with fluorescent promoter-reporter fusions unveiled high nfeR1 expression in response to salt stress and throughout the symbiotic interaction. The strength and differential regulation of nfeR1 transcription are conferred by a motif, which is conserved in nfeR1 promoter regions in α-proteobacteria. NfeR1 loss-of-function compromised osmoadaptation of free-living bacteria, whilst causing misregulation of salt-responsive genes related to stress adaptation, osmolytes catabolism and membrane trafficking. Nodulation tests revealed that lack of NfeR1 affected competitiveness, infectivity, nodule development and symbiotic efficiency of S. meliloti on alfalfa roots. Comparative computer predictions and a genetic reporter assay evidenced a redundant role of three identical NfeR1 unpaired anti Shine-Dalgarno motifs for targeting and downregulation of translation of multiple mRNAs from transporter genes. Our data provide genetic evidence of the hyperosmotic conditions of the endosymbiotic compartments. NfeR1-mediated gene regulation in response to this cue could contribute to coordinate nutrient uptake with the metabolic reprogramming concomitant to symbiotic transitions. [ABSTRACT FROM AUTHOR]

The genome of Azoarcus olearius DQS-4T, a N2-fixing Betaproteobacterium isolated from oil-contaminated soil in Taiwan, was sequenced and compared with other Azoarcus strains. The genome sequence showed high synteny with Azoarcus sp. BH72, a model endophytic diazotroph, but low synteny with five non-plant-associated strains ( Azoarcus CIB, Azoarcus EBN1, Azoarcus KH32C, A. toluclasticus MF63T and Azoarcus PA01). Average Nucleotide Identity (ANI) revealed that DQS-4T shares 98.98% identity with Azoarcus BH72, which should now be included in the species A. olearius. The genome of DQS-4T contained several genes related to plant colonization and plant growth promotion, such as nitrogen fixation, plant adhesion and root surface colonization. In accordance with the presence of these genes, DQS-4T colonized rice ( Oryza sativa) and Setaria viridis, where it was observed within the intercellular spaces and aerenchyma mainly of the roots. Although they promote the growth of grasses, the mechanism(s) of plant growth promotion by A. olearius strains is unknown, as the genomes of DQS-4T and BH72 do not contain genes for indole acetic acid (IAA) synthesis nor phosphate solubilization. In spite of its original source, both the genome and behaviour of DQS-4T suggest that it has the capacity to be an endophytic, nitrogen-fixing plant growth-promoting bacterium. [ABSTRACT FROM AUTHOR]

The conversion of atmospheric nitrogen into valuable compounds, that is, so-called nitrogen fixation, is gaining increased interest, owing to the essential role in the nitrogen cycle of the biosphere. Plasma technology, and more specifically gliding arc plasma, has great potential in this area, but little is known about the underlying mechanisms. Therefore, we developed a detailed chemical kinetics model for a pulsed-power gliding-arc reactor operating at atmospheric pressure for nitrogen oxide synthesis. Experiments are performed to validate the model and reasonable agreement is reached between the calculated and measured NO and NO2 yields and the corresponding energy efficiency for NO x formation for different N2/O2 ratios, indicating that the model can provide a realistic picture of the plasma chemistry. Therefore, we can use the model to investigate the reaction pathways for the formation and loss of NO x. The results indicate that vibrational excitation of N2 in the gliding arc contributes significantly to activating the N2 molecules, and leads to an energy efficient way of NO x production, compared to the thermal process. Based on the underlying chemistry, the model allows us to propose solutions on how to further improve the NO x formation by gliding arc technology. Although the energy efficiency of the gliding-arc-based nitrogen fixation process at the present stage is not comparable to the world-scale Haber-Bosch process, we believe our study helps us to come up with more realistic scenarios of entering a cutting-edge innovation in new business cases for the decentralised production of fertilisers for agriculture, in which low-temperature plasma technology might play an important role. [ABSTRACT FROM AUTHOR]