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We have investigated the effectiveness of a new soybean fertilization technique, named crack fertilization, which involves the application of nodule bacteria on biochar to soybean roots through cracks formed between planting rows during midterm tillage. In the present study, the factors and timing of crack fertilization were investigated in an upland field converted from a paddy. The variables investigated were: 1) crack formation without the application of any agricultural materials, and 2) the application of biochar or 3) nodule bacteria on biochar into cracks. The treatment periods were: 1) before sowing, 2) during midterm tillage, and 3) during both periods. The combination of crack fertilization and reduced tillage was also tested. The method of crack fertilization that increased yield was the combination of crack formation and the application of biochar, and the most effective period for the treatment was before sowing in the reduced tillage field. Seed yields in conventional and reduced tillage fields were comparable in the upland field converted from the paddy. These results suggest that the application of biochar into cracks after scratching the soil surface to remove weeds before sowing is a practical method of increasing soybean yield in upland fields converted from paddies.

During a recent floristic-taxonomic study on the algal flora, including diatoms, from the estuary of the Damietta Branch of the Nile in Egypt, an interesting epilithic diatom species belonging to the genus Seminavis (Naviculaceae) was collected and investigated using both light and scanning electron microscopy. This new diatom species shares morphologically some taxonomic diagnostic features with other related taxa such as S. insignis, S. robusta, and S. ventricosa. However, it still differs by having ventral central striae that are shorter and more or less straight in the middle of the smaller frustules to be clearly radiate in the larger ones and then become geniculate and only radiate near the poles, the central raphe endings are externally more distantly spaced than in the similar species, the elongate central nodule is internally less prominent, and the areola density is much denser. Therefore, we here describe it as Seminavis aegyptiaca sp. nov. Hydrochemical analyses revealed that S. aegyptiaca commonly inhabits typical marine, with a weak tendency towards brackish water, habitats. It was found to be tolerant to meso-eutrophic, nutrient-enriched conditions, based on the data available on seasonal concentrations of N and P compounds. These findings not only contribute to the inventory of Egyptian diatoms, but also increase our understanding of the autecology and distribution of this relatively poorly-known diatom genus.

Manganese (Mn) nodule is one of the ubiquitous polymetallic concretions and mainly consists of Mn − Fe oxi-hydroxide precipitations. A primary oxidation of Mn(II) to MnO2, in which microorganisms may play important roles, is followed by agglomeration of MnO2 into nodules. Celeribater manganoxidans DY25T, belonging to family Rhodobacteraceae, has ability to catalyze the formation of MnO2 [ 1 ]. The concentration of MnO2 formed by harvested cells reached 7.08 μM after suspended in 10 mM HEPES (pH 7.5). Genomic and physiological characteristics of strain DY25T provided a better understanding of its Mn-oxidizing mechanism. Fifteen genes (including four multicopper oxidases) may be involved in Mn(II)-oxidation, whereas only three of them can promote this process. Sulfur-oxidizing activity was detected, which may be associated with manganese oxidation. Genes involved in import and export of primary elemental ingredients (C, N, P and S) and metallic elements (e.g. Mn) were discovered, demonstrating its potential roles in the biogeochemical cycle.

Nonmetallic inclusions play a vital role for graphite nodule nucleation in cast iron with spheroidal graphite (SGI). Therefore, knowledge about composition of heterogeneous nuclei is practically important and can be used to control solidification in SGI castings. Thermodynamic simulations were performed to predict types of nonmetallic precipitates formed in the entire SGI processing, including melt nodularization and inoculation as well as followed melt cooling and casting solidification. To verify simulation results, the experimental SGI heats were performed with variations in nodularization and inoculation practices. Two types of specimens were collected. The first one was direct-quenched from the inoculated melt, and the second one was taken from the castings. An automated SEM/EDX inclusion analysis was used for evaluation of families of nonmetallic inclusions located in the matrix and inside graphite nodules. Nucleation activity of different precipitates was determined from the partitioning of active elements (Mg, Al, Si, Ca, Ce, La, Ti, Zr) between the graphite nuclei and the matrix. Simulation and experimental data were used to discuss a nucleation mechanism of graphite nodules in inoculated SGI. Engineering of nonmetallic inclusions can be used to control SGI solidification.

Polymetallic nodules found on the abyssal plains of the oceans represent one of the slowest known geological processes, and are a source of critical and rare metals for frontier technologies. A quantitative assessment of their occurrence worldwide has been hampered by a research focus on the northeastern Pacific Ocean and the lack of a global open-access data set of nodules. We have compiled a global data set of >10,000 seabed nodule and control samples, and combine it with digital grids of key environmental parameters to generate a predictive machine-learning model of nodule occurrence. In order of decreasing parameter ranking, we find that nodules are associated with very low sedimentation rates (< 0.5 cm/k.y.), moderately high oxygen values (150 and 210 mmol/m3), lithologies of clay followed by calcareous ooze, low summer surface productivity (4500 m, and low total organic carbon content (0.3–0.5 wt%). Competing hypotheses for nodule sustention and thus continued growth on the seafloor are the removal of sediment by bottom-water currents and biological activity. Using a high-resolution eddy-resolving ocean circulation model, we find that the bottom-current speeds over nodule fields are too low (

The synthesis of poly(3-hydroxybutyric acid) (PHB) in nodule bacteria Rhizobium phaseoli grown in submerged culture was studied. The Law method for the determination of PHB was modified, which made it possible to isolate the polymer and to carry out its quantitative assessment directly in the biomass. Only S forms of R. phaseoli cells could synthesize PHB. Its accumulation was more active on carbon-rich medium (by ~50%), and the polymer granules were more clearly visible. The PHB synthesis increased with a decrease in aeration, and the maximum content was reached after 35−40 h of growth. The effect of the oxidative stress caused by t-butyl hydroperoxide and benzyl viologen on the growth of PHB-synthesizing bacteria was studied. Both compounds negatively affected bacterial growth. The growth was reactivated only in PHB-synthesizing S forms. We can conclude that PHB is a protector for nodule bacteria under the conditions of oxidative stress. The maximal PHB content in R. phaseoli cells reached 54% of the dry cell weight, which exceeded the values previously described for rhizobia. This value is close to those reported for the bacterial strains producing the polymer and used in biotechnology. We can conclude that rhizobia grown in the submerged culture can be potential producers of PHB and other polyhydroxyalkanoates.

Xenophyophores, giant deep-sea agglutinated foraminifera, dominate the benthic megafauna in the eastern equatorial Pacific Clarion-Clipperton Zone. This abyssal (>4000 m depth) region hosts major deposits of polymetallic nodules targeted for future seabed mining, an activity that would destroy these highly diverse and delicate protists, particularly those living on the nodules themselves. Since the cell occupies only a small proportion of their test volume, xenophyophores may make a fairly modest contribution to benthic biomass and carbon cycling. Nevertheless, xenophyophore tests can passively enhance particle deposition, concentrate food, and provide habitat structure utilized by diverse organisms. Their destruction could therefore influence the recovery of benthic communities. Species requiring nodule substrates will likely not recover, since nodules take millions of years to form. However, xenophyophores can grow quickly and colonize extensive volcanic ash deposits within years, suggesting that sediment-dwelling species could be among the first large immobile organisms to reappear in mining-impacted areas.

In the above article [1], Figs. 5 and 6 were published incorrectly. The correct figures are given below.

Deep-seabed polymetallic nodule mining can have multiple adverse effects on benthic communities, such as permanent loss of habitat by removal of nodules and habitat modification of sediments. One tool to manage biodiversity risks is the mitigation hierarchy, including avoidance, minimization of impacts, rehabilitation and/or restoration, and offset. We initiated long-term restoration experiments at sites in polymetallic nodule exploration contract areas in the Clarion-Clipperton Zone that were (i) cleared of nodules by a preprototype mining vehicle, (ii) disturbed by dredge or sledge, (iii) undisturbed, and (iv) naturally devoid of nodules. To accommodate for habitat loss, we deployed >2000 artificial ceramic nodules to study the possible effect of substrate provision on the recovery of biota and its impact on sediment biogeochemistry. Seventy-five nodules were recovered after eight weeks and had not been colonized by any sessile epifauna. All other nodules will remain on the seafloor for several years before recovery. Furthermore, to account for habitat modification of the top sediment layer, sediment in an epibenthic sledge track was loosened by a metal rake to test the feasibility of sediment decompaction to facilitate soft-sediment recovery. Analyses of granulometry and nutrients one month after sediment decompaction revealed that sand fractions are proportionally lower within the decompacted samples, whereas total organic carbon values are higher. Considering the slow natural recovery rates of deep-sea communities, these experiments represent the beginning of a ~30-year study during which we expect to gain insights into the nature and timing of the development of hard-substrate communities and the influence of nodules on the recovery of disturbed sediment communities. Results will help us understand adverse long-term effects of nodule removal, providing an evidence base for setting criteria for the definition of "serious harm" to the environment. Furthermore, accompanying research is needed to define a robust ecosystem baseline in order to effectively identify restoration success. Integr Environ Assess Manag 2021;00:1-15. © 2021 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals LLC on behalf of Society of Environmental Toxicology & Chemistry (SETAC).

Background: Cerro de la Estrella (CE) is a natural reserve in Mexico City that suffers from afforestation, and its restoration with Acacia farnesiana is being considered . Question: Will the nodule-forming rhizobia and arbuscular mycorrhizal fungi (AMF) associated with the CE soil support A. farnesiana growth? Study species: Acacia farnesiana (L.) Willd . (Fabaceae). Methods: Mycorrhizal fungi, nodule-forming rhizobia and physicochemical characteristics of the CE soil were studied to determine if they are suitable for improving the growth of Acacia farnesiana . Results: Four different families of AMF were found which generated 13 % mycorrhization with A. farnesiana . However, A. farnesiana from CE did not nodulate, suggesting the lack of native rhizobia. The CE soil has low fertility. Nodules of A. farnesiana were obtained from the soil in Ticuman, Morelos, and 66 rhizobia were isolated from them. Rhizobium isolates were individually added to A. farnesiana grown in the CE soil. Five of the 66 isolates yielded significant differences in shoot dry weight, shoot height, number of nodules, nodulation time and nitrogenase activity compared with the Sinorhizobium americanum CFNEI 156 control strain ( p < 0.05). Three isolates were named as S. americanum ENCBTM1, ENCBTM31 and ENCBTM43, and last two as Sinorhizobium sp. ENCBTM34 and ENCBTM45. Conclusions: CE soil had low fertility and lacked specific rhizobia for A. farnesiana . The individual addition of S. americanum (ENCBTM1, ENCBTM31 or ENCBTM43) or Sinorhizobium sp. (ENCBTM34 or ENCBTM45) improved the growth of A. farnesiana.

N2-fixing bacteria symbiotically dwell inside the root nodules of legume and converts atmospheric N2 to NH3. Unlike high temperature–pressure Haber’s process, this conversion in the nodule is orchestrated by nitrogenase: an enzyme with Fe-S, Fe-Mo, Fe-V cofactor, at its heart. Strategies to increase the nodule population could reduce nitrogen fertilizer use. Here we discovered that pre-treating the chickpea seeds with nano FeS2 (iron pyrite) resulted in a denser root network with larger root nodules. It improved the shoot system and resulted in a higher yield. We observed a higher concentration of Fe, Mo, Mg, P, Ca, Mn, K in the roots of chickpea: possibly emulating nitrogenase.

We employ complementary field and laboratory‐based incubation techniques to explore the geochemical environment where siderite concretions are actively forming and growing, including solid‐phase analysis of the sediment, concretion, and associated pore fluid chemistry. These recently formed siderite concretions allow us to explore the geochemical processes that lead to the formation of this less common carbonate mineral. We conclude that there are two phases of siderite concretion growth within the sediment, as there are distinct changes in the carbon isotopic composition and mineralogy across the concretions. Incubated sediment samples allow us to explore the stability of siderite over a range of geochemical conditions. Our incubation results suggest that the formation of siderite can be very rapid (about two weeks or within 400 hr) when there is a substantial source of iron, either from microbial iron reduction or from steel material; however, a source of dissolved iron is not enough to induce siderite precipitation. We suggest that sufficient alkalinity is the limiting factor for siderite precipitation during microbial iron reduction while the lack of dissolved iron is the limiting factor for siderite formation if microbial sulfate reduction is the dominant microbial metabolism. We show that siderite can form via heated transformation (at temperature 100°C for 48 hr) of calcite and monohydrocalcite seeds in the presence of dissolved iron. Our transformation experiments suggest that the formation of siderite is promoted when carbonate seeds are present.

The release of potentially toxic metals, such as copper (Cu), into the water column is of concern during polymetallic nodule mining. The bioavailability and thus toxicity of Cu is strongly influenced by its speciation which is dominated by organic ligand (L) complexation in seawater, with L-complexes being considered less bioavailable than free Cu2+. The presence of CuL-complexes in deep-sea sediments has, however, not been systematically studied in the context of deep-sea mining. We thus analyzed the Cu-binding L concentration ([L]) in deep-sea pore waters of two polymetallic nodule provinces in the Pacific Ocean, the Peru Basin and the Clarion-Clipperton-Zone, using competitive ligand equilibration–adsorptive stripping voltammetry. The pore-water dissolved Cu concentration ([dCu]) ranged from 3 to 96 nM, generally exceeding bottom water concentrations (4–44 nM). Based on fitting results from ProMCC and Excel, Cu was predominantly complexed by L (3–313 nM) in bottom waters and undisturbed pore waters. We conclude that processes like deep-sea mining are unlikely to cause a release of toxic Cu2+ concentrations ([Cu2+]) to the seawater as > 99% Cu was organically complexed in pore waters and the [Cu2+] was 2+ likely remains beneath toxic thresholds.

The occurrence and genesis of silicified coals in Tanjung Redeb, Embalut, and Loa Kulu have been described and reconstructed by means of petrography and geochemistry. The silicified coals are black, hard, and contain dispersed fine-grained silica minerals. Silica (SiO 2 ) contents vary from 58 to 76%, and LOI (mostly represents organic matter) ranges from 23 to 40% of the bulk silicified coal samples based on the ICP-INAA analysis. Microscopic examination shows the presence of quartz, chalcedony, and amorphous opal. Silicification occurs both as layers and nodules within the coal seams. The layer type is usually thin layers (

The physical and mechanical properties of calcareous concretion soil depend largely on the shape, size and distribution of calcareous concretion soil besides the impact of soil layer. Due to the long time required for clay saturation, consolidation, and shearing, as well as the tedious sample preparation, larger triaxial tests of calcareous nodules have not been carried out. In this chapter, based on the medium triaxial test of calcareous nodules, the strength characteristics of remolded soil samples under different confining pressures are studied. By controlling parameters such as calcareous nodules particle size and water content, the impact of different test conditions on the strength of calcareous nodules is analysed, and the variation rule is analysed. It is concluded that the impact of nodular particle size on soil strength is not obvious at 20% nodular content. The impact of different water content on the strength of soil is more regular, showing hardening characteristics, in line with the natural law.

Manganese nodules are a potential source of critical metals such as Cu, Ni, and Co and are widely distributed on the abyssal plains of the global oceans. A polymetallic nodule metallogenic belt with a heterogeneous and spatially clustered nodule distribution was recently discovered in NW Pacific inter-seamount basin (NPIB) areas. However, the geological processes that regulate the nodule occurrence in that region are unresolved. Here, we report on the characteristics of a high-density field of manganese nodules in the abyssal plain north of Suda Seamount. Ship-borne multibeam bathymetric data reveal a typical seamount sector-collapse topography characterized by radial lineaments of debris channels and ridges formed by rapid debris-avalanche flow. Backscatter data linked with underwater observation indicate that manganese nodules are more concentrated (50%–80% areal coverage) along the main body of the debris apron compared to adjacent neighboring areas (

Myopericytoma is a rare, benign growth characterized by painless lesions with a predilection for the extremities, although they may be found in or on any part of the body. These tumors typically present as a rounded or dome-like non-exophytic lesion and exhibit a benign disease course. Treatment is generally reserved for cosmetic or functional purposes. We present a case of an atypical presentation of an exophytic digital myopericytoma in a 45-year-old female treated with local punch excision.

Deep-sea polymetallic nodules are a mineral resource with potential for commercial development. Due to the unique deep-sea environment in which they are found, specialized technology and equipment are required for their extraction. In this paper, firstly, the development of deep-sea polymetallic nodule mining technology is classified into three stages, and its characteristics are summarized. Moreover, the results from research into deep-sea polymetallic nodule mining technology are analyzed, including proposals for mining systems, research into key technologies, basic scientific problems, and proof of technical feasibility from sea tests. Secondly, the testing of the collector prototype and the environmental impact assessment study of Global Sea Mineral Resources NV, as well as the progress of the deep-sea polymetallic nodule mining test project in China, are introduced. On this basis, the opportunities and challenges brought by the fast-growing demand for electric vehicles to the development of deep-sea polymetallic mining technology is analyzed, and a possible technical scheme for a mining system and the trends in its development towards high reliability and high standards of environmental protection according to the requirements of commercial exploitation are explored. This provides a reference for the research and development of high-efficiency technology and equipment for the mining of deep-sea polymetallic nodules.

Rationale: Ground-glass opacity (GGO)-associated lung cancers are common and radiologically distinct clinical entities known to have an indolent clinical course and superior survival, implying a unique underlying biology. However, the molecular and immune characteristics of GGO-associated lung nodules have not been systemically studied. Objectives: To provide mechanistic insights for the treatment of these radiologically distinct clinical entities. Methods: We initiated a prospective cohort study to collect and characterize pulmonary nodules with GGO components (nonsolid and part-solid nodules) or without GGO components, as precisely quantified by using three-dimensional image reconstruction to delineate the molecular and immune features associated with GGO. Multiomics assessment conducted by using targeted gene panel sequencing, RNA sequencing, TCR (T-cell receptor) sequencing, and circulating tumor DNA detection was performed. Measurements and Main Results: GGO-associated lung cancers exhibited a lower tumor mutation burden than solid nodules. Transcriptomic analysis revealed a less active immune environment in GGO components and immune pathways, decreased expression of immune activation markers, and less infiltration of most immune-cell subsets, which was confirmed by using multiplex immunofluorescence. Furthermore, T-cell repertoire sequencing revealed lower T-cell expansion in GGO-associated lung cancers. HLA loss of heterozygosity was significantly less common in lung adenocarcinomas with GGO components than in those without. Circulating tumor DNA analysis suggested that the release of tumor DNA to the peripheral blood was correlated with the tumor size of non-GGO components. Conclusions: Compared with lung cancers presenting with solid lung nodules, GGO-associated lung cancers are characterized by a less active metabolism and a less active immune microenvironment, which may be the mechanisms underlying their indolent clinical course. Clinical trial registered with www.clinicaltrials.gov (NCT 03320044).

International audience; The increasing demand for metals is pushing forward the progress of deep-sea mining industry. The abyss between the Clarion and Clipperton Fracture Zones (CCFZ), a region holding a higher concentration of minerals than land deposits, is the most targeted area for the exploration of polymetallic nodules worldwide, which may likely disturb the seafloor across large areas and over many years. Effects from nodule extraction cause acute biodiversity loss of organisms inhabiting sediments and polymetallic nodules. Attention to deep-sea ecosystems and their services has to be considered before mining starts but the lack of basic scientific knowledge on the methodologies for the ecological surveys of fauna in the context of deep-sea mining impacts is still scarce. We review the methodology to sample, process and investigate metazoan infauna both inhabiting sediments and nodules dwelling on these polymetallic-nodule areas. We suggest effective procedures for sampling designs, devices and methods involving gear types, sediment processing, morphological and genetic identification including metabarcoding and proteomic fingerprinting, the assessment of biomass, functional traits, fatty acids, and stable isotope studies within the CCFZ based on both first-hand experiences and literature. We recommend multi- and boxcorers for the quantitative assessments of meio- and macrofauna, respectively. The assessment of biodiversity at species level should be focused and/or the combination of morphological with metabarcoding or proteomic fingerprinting techniques. We highlight that biomass, functional traits, and trophic markers may provide critical insights for biodiversity assessments and how statistical modeling facilitates predicting patterns spatially across point-source data and is essential for conservation management.

The interaction between rhizobia and other nodule associated bacteria assist to mitigate nutrient stress in leguminous plants by fixing atmospheric nitrogen and synthesizing plant growth regulators. Beneficial effects of microbial inoculants emphasize the need for further research and their use in modern agriculture. The present work describes the isolation, identification, plant growth promoting properties and phylogenetic analysis of nodule associated bacteria from Mimosa pudica L. Isolation and characterization of nodule associated bacteria were done according to standard procedures. Molecular characterization of the isolates was performed using 16S ribosomal RNA. Plant growth promoting ability was analyzed by quantifying the levels of Indole acetic acid. Evolutionary distance and relatedness was analyzed using neighbor joining method. Rhizobium sp. CU8 and three other co-resident non-rhizobial nodule associated bacteria (Bacillus cereus MY5, Ralstonia pickettii MY1 and Lactococcus lactis MY3) exhibiting nitrogen fixation, plant growth promotion and other potential microbial activities were characterized. Phylogenetic analysis revealed the genetic relatedness and evolutionary significance of Rhizobium sp. CU8 and other co-inhabitant non-rhizobial nodule associated bacteria from M. pudica. Present study identified the four isolates as potential biofertilizers due to their nitrogen fixing and growth promoting characteristics. L. lactis MY3 is the first co-resident nitrogen fixer and plant growth promoter reported from the root nodules of M. pudica.

Siliceous concretions (nodules), from two different geological settings—the Apulian platform margins in Kefalonia island, and the Ionian Basin in Ithaca, Atokos, and Kastos islands—have been studied both in the field and in the laboratory. Nodule cuttings are mainly characterized by the development of a core, around which a ring (rim) has been formed. Mineralogical study, using X-ray powder diffraction (XRPD) analysis, showed that the rim is usually richer in moganite than the core. Homogeneous concretions, without discernible inner core and outer ring, were observed generally in both settings. Mineralogical analysis of the selected siliceous concretions from Kefalonia island showed the presence mostly of quartz and moganite, while calcite either was absent or participated in a few samples in minor/trace abundances. Moganite was generally abundant in all the samples from Kefalonia island. Concretions from the Ionian Basin showed a variation in the quartz, moganite, and calcite contents. Mineralogical differences were recognized both between the different studied geodynamic settings and internally in the same setting, but with different stages of development. The above-mentioned differential diagenesis on nodules evolution could be related to the presence and/or abundance of stylolites, later fluid flows, restrictions from one area to another due to synchronous fault activity, and the composition of substances dissolved in fluids. Moreover, the development of concretions produced secondary fractures in the surrounding area of the nodule-bearing rocks.

Canalicular Adenoma (CA) is a benign accessory salivary gland neoplasm. It represents less than 1% of salivary gland tumors. It occurs generally on the upper lips in female patients over 50 years of age. We report a case of canalicular adenoma that arose from buccal minor salivary glands of the upper lip without any symptomatology. Clinically, CA appears as an often single firm mass of 1cm, sparing the main salivary glands which is not pathognomonic of canalicular adenoma. Treatment of choice is enucleation to rule out many differential diagnoses that may present clinically as an asymptomatic nodule. Recurrences are rare. Histological features are characteristic and usually allow a definitive diagnosis to be made with confidence. However, many differential diagnoses remain to be ruled out.

The detailed mineralogical and microgeochemical characteristics of polymetallic nodules collected from the Interoceanmetal Joint Organization (IOM, Szczecin, Poland) claim area, Eastern Clarion-Clipperton Fracture Zone (CCFZ, Eastern Pacific) were described in this study. The obtained data were applied for the delimitation of nodule growth generations and estimation of the growth ratios (back-stripping using the Co-chronometer method). The applied methods included bulk X-ray powder diffraction (XRD) and electron probe microanalysis (EPMA), providing information about Mn-Fe minerals and clays composing nodules, as well as the geochemical zonation of the growth generations. The analyzed nodules were mostly diagenetic (Mn/Fe &gt, 5), with less influence on the hydrogenous processes, dominated by the presence of 10-Å phyllomanganates represented by todorokite/buserite, additionally mixed with birnessite and vernadite. The specific lithotype (intranodulith), being an integral part of polymetallic nodules, developed as a result of the secondary diagenetic processes of lithification and the cementation of Fe-rich clays (potentially nontronite and Fe-rich smectite), barite, zeolites (Na-phillipsite), bioapatite, biogenic remnants, and detrital material, occurs in holes, microcaverns, and open fractures in between ore colloforms. The contents of ∑(Ni, Cu, and Co) varied from 1.54 to 3.06 wt %. Several remnants of siliceous microorganisms (radiolarians and diatoms) were found to form pseudomorphs. The applied Co-chronometer method indicated that the nodules’ age is mainly Middle Pliocene to Middle Pleistocene, and the growth rates are typical of diagenetic and mixed hydrogenetic–diagenetic (HD) processes. Additionally, few nodules showed suboxic conditions of nucleation. Growth processes in the eastern part of the CCFZ deposit might have been induced with the Plio-Pleistocene changes in the paleooceanographic conditions related to the deglaciation of the Northern Hemisphere.

In the exploration of deep-sea nodule mineral resources, computer vision-based methods have been widely used to assess nodule mineral resources, such as abundance, particle size, coverage and other information. Among them, the separation of nodule minerals from the background is an important part of deep-sea mineral resource assessment. Threshold segmentation, clustering segmentation, multispectral segmentation and other traditional segmentation methods need to rebuild the model from scratch once the distribution of feature space changes. However, the data-driven deep learning method provides a general framework to solve this problem, which has strong generalization and robustness. In order to improve the segmentation performance, a novel deep sea nodule mineral images segmentation algorithm based on Mask R-CNN was proposed. The comparative analysis is also processed on some other different deep learning methods, such as U-Net, and the Generative Adversarial Network. The experimental results show that, the method based on Mask R-CNN is better than the method based on U-Net, improced U-Net and Conditional Generative Adversarial Networks on the dataset.

Deep-sea polymetallic nodule mining research activity has substantially increased in recent years, but the expected level of environmental impact is still being established. One environmental concern is the discharge of a sediment plume into the midwater column. We performed a dedicated field study using sediment from the Clarion Clipperton Fracture Zone. The plume was monitored and tracked using both established and novel instrumentation, including acoustic and turbulence measurements. Our field studies reveal that modeling can reliably predict the properties of a midwater plume in the vicinity of the discharge and that sediment aggregation effects are not significant. The plume model is used to drive a numerical simulation of a commercial-scale operation in the Clarion Clipperton Fracture Zone. Key takeaways are that the scale of impact of the plume is notably influenced by the values of environmentally acceptable threshold levels, the quantity of discharged sediment, and the turbulent diffusivity in the Clarion Clipperton Fracture Zone. The extent of impact by midwater discharge plumes from deep-sea nodule mining will be influenced by the quantity of plume sediment, ocean turbulence and acceptable environmental thresholds, according to numerical modelling and direct experiments

Nitrogen is a paramount important essential element for all living organisms. It has been found to bea crucial structural component of proteins, nucleic acids, enzymes and other cellular constituents which are inevitable for all forms of life. In the atmosphere, the percentage of nitrogen is very high (N2, 78%) compared to other inorganic gases. However, most organisms have practically no direct access to this nitrogen. While plants can not directly uptake nitrogen from atmosphere, they are capable of assimilating other forms of nitrogen, for example ammonium (NH4+) and nitrate (NO3-). For agricultural crop production, artificial fixation of nitrogen is heavily utilized and it is an expensive process that requires high temperatures (at least 400 °C) and pressures (around 200 atm). It has been conspicuously demonstrated that indiscriminate use of fertilizer hampers soil physical, chemical and micro biological properties and also a potential risk to environment e.g. water quality. Besides, chemically manufactured fertilizers are depleted from soils in various ways, for instance; denitrifying bacteria, volatilization, and leaching. Consequently, it results relatively poor availability of nitrogen to get into plants. On the flipside, only 1-2% of the nitrogen fixation in the world occurs through the natural process of lightening. Notably, microbial fixation is well characterized in diazotrophs for example; Rhizobia and Frankia, and blue-green algae. Against the backdrop, we are accentuated on an environmentally friendlyand themost sustainable approach to increase productivity for legume and non-legume crops. Till today, the term biological nitrogen fixation (BNF) has received much attention as a sustainable alternative; this process facilitates atmospheric nitrogen to convert into ammonia by rhizobia in specialized plan organs termed “root nodules”. This review article seeks to better understand plant mechanisms involved in the development of root nodules in soybean. Soybean (Glycine max) is one of the most important oil crops and a source of animal feed protein in the world. It has a salient feature to fix atmospheric nitrogen through symbioses with compatible rhizobia that yields to determinate type nodule (Oldroyd, Murray et al. 2011). Biological nitrogen fixation in soybean nodules reduces the use of chemical nitrogen fertilizers resulting in cost-savings to producers and minimizes environmental damage due to nitrogen run-off. A better understanding of how nodules form and function is important for selection or generation of soybean genotypes with better nitrogen fixation capacity. Soybean nodules originate from root cortex via de novo cell differentiation (Oldroyd 2013). Consequently, two major nodule development zones are formed for instance; the nodule primordium (Npr) in the middle and it is encircled by nodule parenchyma (Npa). At later time point, the Npr gives rise to N-fixation zone and the Npa holds vascular bundles. It is not clear what early signaling pathways driving the conspicuous development of the nodule zones. My research is aimed at filling this knowledge gap by illustrating the molecular signatures that paves the way to cellular differentiation in root nodule development in soybean. Based on initial evidence obtained by the Subramanian lab, we hypothesize that microRNAs (miRNAs) play important regulatory roles in spatio-temporal expression of their target genes during nodule developmental in soybean. For instance, the regulation of auxin sensitivity by miR160 has been found to be crucial for formation of nodule primordia and vasculature in the parenchyma (Marie Turner 2013). Against this backdrop, this review article focused on nuclear and cytoplasmic transcriptome as well as miRNA profiles of parenchyma and primordial tissues and determine the relative abundance and differentially expressed mRNAs and regulatory role of miRNAs in cell differentiation and nodule development. Root nodule a sustainable alternative to fix atmospheric nitrogen Atmospheric nitrogen percentage is very high (N2, 78%) compared to other inorganic gases (Mary Elvira 1932). However, most of the organisms have practically no direct access to this nitrogen. Nevertheless, plants can not directly uptake nitrogen from atmosphere but they are capable of assimilating only very specific forms of nitrogen, for example ammonium (NH4+) and nitrate (NO3-) (Bytnerowicz and Fenn 1996, Peter M. Vitousek 1997) (Sponseller, Gundale et al. 2016). Virtually, nitrogen has been found to be a crucial structural component of proteins, nucleic acids, enzymes, and other cellular constituents which are inevitable for all forms of life (O'Brien, Vega et al. 2016). For agricultural crop production, artificial fixation of nitrogen is heavily utilized. It is an expensive process that requires high temperatures (approx. 400 °C) and pressures (approx. 200 atm) (Witschi 2000). It has been conspicuously demonstrated that indiscriminate use of N fertilizer hampers the diversity of the bacterial community and decreases soil C and N concentrations (Verzeaux, Alahmad et al. 2016). Notably, it has been demonstrated as a potential risk to environment e.g. water quality (Zhao, Sha et al. 2016) (Sponseller, Gundale et al. 2016). Besides, chemically manufactured fertilizers are depleted from soils in various ways, for instance; denitrifying bacteria, volatilization, and leaching (Johnson 1996, Peter M. Vitousek 1997). Consequently, it results relatively poor availability of nitrogen to get into plants. On the flipside, over 90 % of the nitrogen fixation in the world occurs through the natural process of lightening and microorganisms. Furthermore, microbial fixation is well characterized in diazotrophs for example; Rhizobia and Frankia, and blue-green algae (Cheng 2008). It has been demonstrated that Bradyrhizobium strains substantially escalated soybean grain yield, and protein content up to 57% and 26%, respectively (Zimmer, Messmer et al. 2016). Against the backdrop, we are accentuated on an environmentally friendly and a sustainable approach to increase the productivity for legume and non-legume crops. Literature mining depicted that biological nitrogen fixation in soybean nodules reduces the use of chemical nitrogen fertilizers resulting in cost-savings to producers and minimizes environmental damage due to nitrogen run-off. Rhizobia infection leads to the root nodule development In the natural environment, plants are continuously confronted with pathogenic and symbiotic microbes. Symbioses involves mutual exchange of diffusible signal molecules, first endophytic bacteria (rhizobia) are attracted by the plant root exudates flavonoids which are perceived and triggered the bacterial nodulation (nod) genes. Consequently, the bacteria synthesize specific lipochito-oligosaccharides, called nodulation (Nod) factors. This signal is perceived by the LysM receptor like kinase of host plant, it induces the root hair curling, and bacteria get access into the host epidermis through infection threads (ITs) and initiate cell division within the root cortex, leading to the progression of the root nodule meristem. In later stages of the interaction, bacteria are released from the infection threads into the plant cells, surrounded by membrane of plant origin. These bacteria multiply within the host cells and differentiate into the nitrogen fixing bacteroids (Udvardi and Day 1997) (Oldroyd 2013). Till now, integration of genetic and genomic approaches has revealed twenty-six genes to be involved in nodule development of Medicago truncatualaand Lotus japonicum (Kouchi, Imaizumi-Anraku et al. 2010). In addition, deep sequencing of the Medicago truncatularoot transcriptome has uncovered thousands of genes to be induced during Nod factor signaling and its resulting ethylene (ET) biosynthesis throughout the multiple development stages of indeterminate nodule (Larrainzar, Riely et al. 2015). Albeit the molecular mechanism of such regulation is not well understood. There has been a large-scale transcriptome analysis of B. japonicum-inoculated and mock-inoculated soybean root hairs. It has showed that a total of 1,973 soybean genes differentially expressed during root hair infection, particularly NFR5 and NIN genes (Libault, Farmer et al. 2010). Nevertheless, the signaling mechanisms directing the cellular differentiation of nodule are not known. Soybean root nodule organogenesis Soybean (Glycine max) has a genome size of 1.1 to1.5 Gb, it is partially diploidized tetraploid. It is one of the most important oil crops and a source of animal feed protein in the world (soybase.org/sb_about.php). It has a salient feature to fix atmospheric nitrogen through symbioses with compatible rhizobia that yields to determinate type nodule (Udvardi and Day 1997) (Oldroyd, Murray et al. 2011). Notwithstanding of the economic and environmental importance, there has been very few studies about quantitative trait loci (QTL) that controlling BNF traits, for instance nodule number, ration of nodule dry weight with nodule number, and shoot dry weight (SDW). It has been reported via composite interval mapping that approximately six QTLs bears very small effect on BNF traits (Santos, Geraldi et al. 2013). Besides, it has been demonstrated in earlier studies that nodules originate from root cortex via de novo cell differentiation into two different cell types, parenchymal and primordium (Celine Charon 1997) (Oldroyd&Downie 2008; Oldroyd 2013). In addition, early nodulin genes in legume for instance; Enod 40 gene reported to be expressed in root pericycle during the rhizobia infection and later it occupied in the dividing cortical cells (H. Kouchi and S. Hata 1993). Among the two major nodule development zones, the nodule primordium (Npr) in the middle which is encircled by nodule parenchyma (Npa). At later time point, the Npr gives rise to N-fixation zone and the Npa holds vascular bundles. Lately, a β- expansin gene, GmEXPB2 fused with GUS reporter gene which was observed to be preferentially expressed in nodule vascular trace and nodule vascular bundles. It indicated that GmEXPB2 might be crucial for nodule organogenesis. Over expression of GmEXPB2 contrast to suppressed GmEXPB2 transgenic lines found to be escalated nodule number, nodule mass and nitrogenase activity. It further suggested that GmEXPB2 might have influenced over root architecture, nodule formation and development, and profoundly yielding to biological N2 fixation (Li, Zhao et al. 2015). Even though, it is not clear what early signaling pathways driving the conspicuous development of the nodule zones. Against the back drop, to understand the regulation of auxin sensitivity by miR160 which is believed to be crucial for the formation of nodule primordia (Marie Turner 2013). Figure 1 a. Illustrating the progression of root nodule development through Rhizobial bacterial infection in the plant root leading to the determinate nodule (Oldroyd 2013). b. Nodule development zones A. Nodule primordial zone (Enod 40 gene) in the middle B. surrounding parenchyma (Enod 2 gene), differentiated from cortex (collected from Sen Subramanian lab). Regulatory small RNAs biogenesis and its molecular functions Regulatory small RNAs are ranged between 20 to 24 nucleotides which are ubiquitous elements of endogenous plant transcriptomics, a common response to exogenous viral infections and introduced double-stranded RNA (Axtell 2013). Three core enzymes families, for instance; RNAdependent RNA polymerase (RDR), Dicer like (DCL), and Argonaute (AGO) proteins paves the way of small RNA biogenesis and function in plants. Firstly, ribonuclease type III or DICERLIKE1 involves in the yield of a fold-back precursor RNA or primary miRNA (primiRNA) transcripts using an RNA templates in the nuclei. Later, the resulting miRNA-miRNA duplex which is originated in nucleus then translocated into cytoplasm. The guided miRNAmolecule is incorporated into ARGONAUTE (AGO) to form an active RISC complex to specific target RNAs that are complementary to the miRNA, and this process eventually follows up mRNA cleavage, represses the translation of the mRNAs or Chromatin modification. This phenomenon accentuated as an inhibition or silencing of the gene expression, which play a crucial role in the developmental process in plant and animal (Chapman and Carrington 2007) (Axtell 2013). Fig. 2 Regulation of gene expression events via RISC complex (modified from https://www.google.com/?gws_rd=ssl#q=mirna+picture+in+plants accessed on 7th February, 2016) Fig. 3 Gene expression events occurring in typical plant cell (modified https://www.google.com/search?q=transcription+and+translation accessed on 7th February, 2016) It has been found in several studies that most plant miRNAs are non-coding RNA, and small 21-24 nucleotide long (Cuperus, Fahlgren et al. 2011). It requires DCL1-clade DCL for their biogenesis and AGO1-clade AGO for their function (Wu, Zhou et al. 2010, Manavella, Koenig et al. 2012). In rice (Oryza sativa), DCL3 has been reported in the biogenesis of 24nt long miRNA that incorporated in AGO4 to regulate the target gene expression primarily through mRNA cleavage (Wu, Zhou et al. 2010). Argonaute proteins (AGO) form RNA inducing silencing complexes (RISC) with small RNAswhich is known as post-transcriptional gene silencing. It has typically four domains, for instance:N-terminal, PAZ, MID and PIWI domains. The MID-PIWI lobes are belongs to the C-terminus. It has been studied that MID-domains contains the specificity loop to recognize and bind to the 5’-phosphate of smRNAs. The PIWI domains contained the catalytic active site D-E-D-H/D. PAZ domain anchored the 2-nt overhang at the 3’ end of miRNAs. The N-terminal domain involved in the separation of miRNA-miRNA duplex and the slicer activity of the mRNA (Song, Smith et al.2004). There has been an expansion and duplications of AGO family members during plantevolution (Singh, Gase et al. 2015). The functional diversification of AGOs is indicating sRNAdirected regulatory pathways. The binding preference of AGO and sRNA is mainly assigned by the sequence of sRNA. In Arabidopsis, 10 AGO have been extensively studied (Liu et al. 2014). It has been demonstrated that AtAGO10 like AtAGO1, it recognized distinct structural features in miR165/miR166 duplex than involved by AtGO1. AtAGO10 found to regulate shoot apical meristem by decoying miR165/miR166 and subsequent repression of homeodomain-leucinezipper (HD-ZIP) gene expression (Zhu, Hu et al. 2011). Notably, 22 AGO proteins have been reported in Soybean (Glycine max). It has been found that genome duplication in Soybean resulted such a proliferation of AGOs. For example: its genome encodes two copies of AGO1, AGO2, AGO5, AGO4/9, AGO6 and AGO7 (Xiang Liu 2014). However, the molecular function of the plant AGO genes yet not very clear. There are several miRNA families that are conserved across the vast evolutionary distances from flowering plants to mosses (Cuperus, Fahlgren et al. 2011). It has been observed in another study that miRNA, and its target pairing found to be stable for a prolonged periods of plant evolution. On the flip side, another group demonstrated that conserved plant miRNAs and their targets are to somehow flexible. For instance; miR159 is a highly conserved miRNA that targets not only a subset of MYB mRNAs but also observed to target a non MYB mRNA, SGN-U567133 (Buxdorf, Hendelman et al. 2010). A mutant tomato transgenic line (miR159-resistant line) showed higher level of the SGN-U567133 transcript and exhibited defects in leaf and flower development. This result suggests that miR159 involves in a post-transcriptional regulation. Additionally, it is found to be crucial for the normal tomato development. Recently, the identification of miRNAs in the regulation of photoperiodic pathways in soybean have been reported through high throughput sequencing and qRT-PCR. Six libraries were constructed using Illumina Solexa, for instance; 0, 8, and 16 h under short day treatment, similar time points considered for the long the long day treatment. A total of 163 miRNAs families were reported which covered 318 plant miRNAs, and unclassified 81 novel predicted miRNAs. As expected, significant differences in abundance between short day and long day treatment was observed (Wenbin Li 2015). These findings provided evidence of miRNA in the regulation of flowering time that ultimately affects the seed yield and quality of soybean. The complex regulatory network of miRNA-mRNA interactions during viral infection has been revealed via small RNA seq (sRNA), degradome seq, and genome-wide transcriptome analysis. There has been a total of 253 soybean miRNAs found to be two-folds abundance compared with mock-inoculated control demonstrated through sRNA seq analysis. Among them 105 miRNAs were identified as potential targets of 125 transcripts that has been validated by degradome seq analyses. In addition, 2679 genes were detected via genome wide transcriptomic analysis. These genes have been differentially expressed during infection of soybean mosaic virus and among them 71 genes projected to induce in defense response (Hui Chen 2016). These findings suggested the regulatory role miRNA that governed the target gene expression during viral infection. Furthermore, the regulatory role of microRNAs (miRNAs) during Soybean- Bradyrhizobium japonicum mutualistic association was studied first by Subramanian et al. 2008. They sequenced approximately 350000 small RNAs of soybean root sample which were inoculated with B. japonicum. It helps to detect 20 conserved miRNAs loci based on the similarity to miRNAs in another plant species. In addition, 35 novel miRNAs were identified based on potential hairpin forming precursors in Soybean EST as well as shotgun genomic sequences (Subramanian, Fu et al. 2008). These findings advocated the potential role of miRNAs in the regulation of legumerhizobiumsymbiosis. In another study, 120 hairpin-forming precursor genes have been identified in soybean by Turner et al. In addition, they reported three novel miRNAs for instance; miR160, miR164 and miR393 found to be involved in auxin signaling (Turner, Yu et al. 2012). Moreover, the plant hormone auxin is thought to have a pivotal role in nodule organogenesis in determinate and indeterminate type of nodule. It indicates a redundancy and diversity of miRNAs family members that governs the formation of root nodule. It has been illustrated that auxin receptor gene family hushed by over expressed microRNA393. These plant roots found to be hypersensitive to auxin and yielded normal nodule. This observation advocated that only minimal/reduced auxin signaling is required for determinate nodule development. Likewise, overexpressed microRNA160 hushed a set of repressor auxin response transcription factor. These plant roots were hypersensitive to auxin and observed not to be reluctant in epidermal responses to rhizobia. Notably, it yielded to lower sized nodule primordium (Marie Turner 2013). This observation indicated that auxin hypersensitivity inhibits nodule organogenesis Organ specific expression of profile of miRNA and the potential targets were also studied. Two genes (Glyma10g10240 and Glyma17g05920) which were the target of miR169 but detected to be highly expressed in soybean nodule. Likewise, three potential targets of gma-new-miR13587 demonstrated to be highly expressed in the nodules than in the roots. As expected, gma-newmiR13587 found to be poorly expressed in the nodules than in the roots (Turner, Yu et al. 2012). There was an inverse expression pattern observed in between roots and nodules. Li et al., studied the transgene expression of three novel miRNAs namely, miR482, miR1512, and miR1515 in Soybean. They noticed a significant increase of nodule numbers while root length and later root density were normal in all tested miRNA lines. As expected, there were differential expression of these miRNAs in supernodulating and nonnodulating soybean mutants. They reported that 6 novel miRNAs decoyed 22 predicted target genes. And it was estimated via real time polymerase chain reaction and qRT-PCR (Li, Deng et al. 2010). It advocates that miRNAs have the signatory roles in soybean nodule development. Sequencing of small RNAs and Parallel analysis of RNA ends (PARE) libraries revealed to identify 284 nodule miRNAs, more than 500 target genes, and including 178 novel soybean miRNAs. It has been reported that ENOD93 only found to be expressed in nodule tissue not in other plant parts of Soybean. Ectopic expression of miR393j-3p and RNAi silencing approach to ENOD93 expression showed a significant reduction in nodule formation (Zhe Yan 2015). Therefore, this study showed a list of miRNAs and their potential target of nodulation genes. In the model legume (Medicago truncatula), 25 conserved miRNA families and 100 novel miRNA reads were detected by high-throughput sequencing. The expression of MtHAP2-1 (encodes a CCAAT binding transcription factor) to meristematic zones was restricted by miR169a which is found to be critical for the development of indeterminate type of nodule (Combier, Frugier et al. 2006). In another study, HDZIPIII transcripts were inhibited by overexpression of miR166, it dropped the number of symbiotic nodule and lateral root (Boualem, Laporte et al. 2008). To get insights into key genes of nodule zones, transcript profiles of specific cells/tissues were investigated at different time points from indeterminate nodules of M. truncatulausing laser capture micro dissection. It has been demonstrated from the comprehensive gene expression map that selected genes enriched in different cell/tissue types (Limpens, Moling et al. 2013). These findings indicated that organ specific gene expression could be controlled by the presence or absence of miRNAs. Recently, Agrobacterium rhizogenesmediated hairy root transformation has been applied as tool for exploring cell type specific gene expression in tomato. Cell type or tissue specific promoter introduced into INTACT and TRAP constructs via gateway cloning technology to develop binary vectors. INTACT method used to capture biotin tagged nuceli from specific cell types and TRAP method used for profiling of mRNAs or foot printing of individual ribosomes (Ron 2014). TRAP methodology is not required tissue fixation or single cell suspension. It has been successfully used to date in organisms ranging from D. melanogaster to mice and human cultured cells. Multiple ribosomes or Polyribosomes (polysomes) are engaged in translation on a single mRNA. To evaluate the translation state of an mRNA, ribosomal subunits, ribosomes, and polysomes can be isolated from detergent-treated cell extracts (Heiman, Kulicke et al. 2014). In this study, we would perform polysome isolation deploying gene cassettes ENOD40p:HF-GFP-RPL18 for primordial tissues, and ENOD2p:HF-GFP-RPL18 for parenchymal tissues in Glycine max root nodules that express an epitope tagged version of ribosomal protein L18. Over the last one decade, there has been several microarrays-based studies which characterized transcriptional variations deployed in nodule formation. It has been embedded with couple of shortcomings, for instance; relative late time points study, incomplete representation of plant genes,discrimination of close paralogs, and reduced sensitivity. Lately, next generation sequencingtechnology have widened the horizon of transcription analyses in different legume species to detectsymbiosis induced changes in late nodule developmental stages. Against this backdrop, we areaccentuated to reveal early transcriptional changes induced in determinate type of soybean noduleby Bradyrhizobium japonicum. In determinate type of nodule, two major nodule development zones are formed for instance, the nodule primordium (Npr) in the middle and it is encircled by nodule parenchyma (Npa). At later time point, the Npr converted to N-fixation zone and the Npa contained vascular bundles. Of these facts, it is not clear what early signaling pathways driving the conspicuous development of thenodule zones. In this context, mechanisms regulate the distinct gene expression profiles in Npr andNpa cell types has not understood clearly. The proposed research study is aimed at filling this knowledge gap byillustrating the molecular signatures that paves the way to cellular differentiation in root noduledevelopment in soybean considering four different time points (5 dai, 7 dai, 10 dai& 14 dai). The hypothesisis microRNAs(miRNAs) play important regulatory roles in spatio-temporal expression of their target genesduring nodule developmental in soybean. For example, a gradient of microRNA localizationbetween nodule primordium and parenchyma cells could result in distinct differentiation of thesecell types. To test this hypothesis, one has to obtain both cell type-specific miRNA andtranscriptome (miRNA target) profiles. Since, the majority of miRNA regulation occurs in thecytoplasm, we reasoned that comparison of nuclear and ribosomal transcriptome profiles wouldreveal genes whose expression is potentially regulated by post transcriptional mechanisms such asmiRNA cleavage. Combining this information with cell type-specific miRNA profiles, andto test the above hypothesis and identify key miRNA-target pairs important for nodule celldifferentiation. The use of translating ribosome affinity purification (TRAP) of nodule zonecells, namely from parenchyma and primordial tissues, to obtain cytoplasmic transcriptomes data. Techniques to determine cell type specific expression profiles: TRAP methods TRAP is termed translating ribosome affinity purification, combines cell-type-specific transgene expression with affinity purification of translating ribosomes. It supersedes the need for tissue fixation, and facilitates to study the cell type-specific mRNA profiles of any genetically defined cell type. It has been successfully used to date in organisms ranging from D. melanogaster to mice, and human cultured cells. Multiple ribosomes or Polyribosomes (polysomes) are engaged in translation on a single mRNA. To evaluate the translation state of an mRNA, ribosomal subunits, ribosomes, and polysomes can be isolated from detergent-treated cell extracts. In this study, the polysome isolation using gene cassettes ENOD40p:HF-GFP-RPL18 for primordial tissues, and ENOD2p:HF-GFP-RPL18 for parenchymal tissues in Glycine max root nodules that express an epitope tagged version of ribosomal protein L18 RPL18(Heiman, Kulicke et al. 2014, Ron 2014). Relative abundance and differentially expressed mRNAs profile in two different tissue specific zones would help to understand the effect of regulatory role of miRNAs in cell differentiation and nodule development. References: Axtell, M. J. (2013). "Classification and comparison of small RNAs from plants." Annu Rev PlantBiol 64: 137-159. 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This is the first report concerning the sequence of morpho-histological and cytological events occurring during organogenesis from cotyledon-derived meristematic nodules (MNs) in Paeonia ostii ‘Feng Dan’. Sections were made and studies were carried out with dissecting microscope, light microscope, scanning electron microscopy (SEM) and transmission electron microscopy (TEM) observation. Histological studies revealed a complex developmental process of morphogenesis that including five stages: (1) callus originated from cell division in both cambial and cortical regions, and type I - yellow compact callus with densely arranged clumps were identified as embryogenic callus. (2) pre-nodular structure consisted of organization center (a central area of vascularization surrounded by meristematic cell layers) and an epidermis-like layer; (3) independent MNs comprised of organization center, a cortical-like area of parenchymatous cells and an epidermal-like area; (4) nodular clusters displayed vigorously internal meristematic cell division and generated a relative movement towards the nodules periphery, establishing vascular connection with primordia; (5) successive new elongated shoots with complete vascular system and axillary bud primordia were developed. SEM observations showed three types of extracellular matrix (ECM), a smooth membranous layer, fibrillar structures and granular mucilage-like secretions on embryogenic callus, and recorded its dynamic morphological changes. Ultrastructural analysis revealed striking changes of chloroplast morphology and starch content during MNs morphogenesis. This study allows a better understanding of in vitro regeneration via MN culture and provides references for protocol optimization and genetic transformation.

Rhodoliths are free-living and morphologically diverse marine calcareous algae commonly distributed over the continental shelf seafloor. They increase the seabed structural complexity and are of potential value as feeding and reproductive grounds for a myriad of marine fauna. The higher structural seabed complexity within rhodolith beds may also increase benthic diversity by creating microhabitats, but this relationship has been rarely explored within rhodolith beds worldwide. Here we compared benthic macrofaunal (>500 µm) structure on rhodolith beds (nodule epifauna) and within unconsolidated sediments (sediment infauna) under high and low-density beds to test whether rhodolith bed density and nodule morphology influenced macrofaunal assemblages. We observed that macrofaunal density on nodules (2538 ± 288.7 ind·m−2) was 15-fold higher when compared to sediments under those beds (166 ± 38.8 ind·m−2). Rhodolith bed density was positively related to macrofaunal density, composition, and functional diversity on the rhodoliths. Low-density beds (61 ± 27.1 nodules·m−2) with discoid-shape nodules were dominated by peracarid crustaceans whereas high-density beds (204 ± 18.7 nodules·m−2) with spheroidal nodules were dominated by Annelid polychaetes. The sediment macrofauna was also positively influenced by the density of rhodolith nodules, which increased sediment carbonate and organic quality (protein and lipids) under high-density beds. Macrofaunal functional diversity was generally higher on rhodoliths, with low similarity (low nestedness) and high taxa turnover between macrofaunal assemblages of rhodoliths and sediments. These findings indicate that rhodolith beds provide an unique habitat for benthic macrofaunal communities, with exclusive functional and taxonomic richness that are likely not typical in the unconsolidated sediment below these beds in SE Brazil. This study highlights the importance of protecting rhodolith beds from multiple sources of anthropogenic disturbance and exploration on continental shelves.

NODULE-INCEPTION-like proteins (NLPs) are plant specific transcription factors that play significant role in orchestrating nitrogen response. NLPs have been widely studied in vascular plants but very less is known about NLPs in non-vascular bryophytes till date. In current study, first, the in silico tools were employed for identification and characterization of NLPs in model bryophyte Physcomitrella patens genome. Furthermore, the expression profiles of PpNLPs were assessed under variable supply of nitrogen. A total of 6 Physcomitrella patens NLP genes (PpNLPs) were identified that shared resemblance in their physical and chemical attributes with Arabidopsis thaliana NLPs (AtNLPs). PpNLP genes possesses similarities in there iso-electric point and hydropathicity values with those of AtNLPs while gene lengths, protein lengths, and molecular weights were found higher in PpNLPs than AtNLPs. It was further observed that all PpNLPs, except PpNLP6, yield acidic hydrophilic proteins localized in nucleus and share a significant degree of homology in their gene structures and protein motifs with AtNLPs. Phylogenetic analysis indicated that PpNLPs possess significant evolutionary linkage with Arabidopsis thaliana, Oryza sativa, and Zea mays. Protein-protein interaction analysis suggested that PpNLPs possess substantial coordination with nitrogen responsive genes like nitrate reductase. Expressions of all PpNLPs were up-regulated in the availability of nitrate (5 and 10 mM) as sole nitrogen source while no significant increment was observed in the absence (0 mM) of nitrogen. The expression levels increased with increasing retention-time treatment of 0, 6, 12, 24, 48, and 72 hours. Results proposed that NLPs are responsive to as well as significantly regulated by nitrogen supply.

In 2006, the BGR signed a contract with the International Seabed Authority (ISA) for the exploration of polymetallic nodules in the Clarion-Clipperton Zone of the northeast Pacific. During nine expeditions, in particular, “Area E1”, the eastern part of the contract area, with a size of ~60,000 km2, was explored in detail. Here, we outline BGR’s exploration methods and provide resource estimates for Area E1 and three sub-areas. The resource assessment is predominantly based on statistical analyses of data obtained by 12-kHz multibeam bathymetry and backscatter mapping, box core sampling and geochemical analysis of nodules. The main parameter for the assessment is the nodule abundance (kg/m2), as its coefficient of variation (CoV) over the entire eastern contract area is relatively high at 36%. In contrast, the metal contents of nodules show only minor variation, with a CoV of 8% for manganese and 8% for the sum of copper, nickel and cobalt. To estimate mineral resources for the entire Area E1, we used an artificial neural network approach with a multivariate statistical correlation between nodule abundance derived from box cores and hydro-acoustic data. The total estimated resources are 540 ± 189 million tonnes (Mt) of dry nodules, and the total estimated metal contents are 168 Mt of manganese, 7.5 Mt of nickel, 6.3 Mt of copper, 0.9 Mt of cobalt, 0.4 Mt of rare-earth elements and 0.3 Mt of molybdenum. A geostatistical resource estimate of three economically prospective areas with a total size of 4498 km2, intensively sampled by box cores, was carried out using ordinary kriging of nodule abundance and metal grades. Within these three nodule fields, 7.14 Mt of dry nodules are classified as measured mineral resources covering an area of 489 km2. Indicated mineral resources amount to 11.2 Mt, covering an area of 825 km2, and inferred mineral resources of 35.5 Mt of dry nodules were estimated for an area of 3184 km2. In total, the metal contents of the three prospective areas amount to 16.8 Mt of manganese, 0.74 Mt of nickel, 0.63 Mt of copper and 0.09 Mt of cobalt.