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Very little information is available for Mediterranean areas about the soil N dynamics and crop N use efficiency during the transition phase from conventional tillage (CT) to no-tillage (NT). Hence, a 2-yr experiment was conducted under semiarid Mediterranean conditions in three sites to study how soil N dynamics, crop N uptake, grain yield, and N use efficiency vary with N-fertilization rate and crop genotype in the switch year from CT to NT. Treatments consisted of two tillage systems (CT and NT), five N-fertilization rates (0, 40, 80, 120, and 160 kg N ha−1), and two durum wheat (Triticum durum Desf.) genotypes (one modern variety and one old landrace). Irrespective of the genotype, NT reduced compared to CT both wheat N uptake and grain yield under low soil N availability. The greater soil N supply under CT was the main reason for this outcome, so much so that the differences between CT and NT for these traits gradually decreased with the increase of N fertilization rate, practically disappearing at 80 kg N ha−1). The analysis of the N use efficiency components corroborated this hypothesis showing clearly that the yield advantage observed in CT depended mainly on the increase in N supply in soil under CT than NT condition. The two wheat genotypes responded similarly to varying soil tillage system. However, the adverse effects of NT practice were more evident in the modern variety than the old landrace. This study ultimately indicates that in the Mediterranean areas the switch year from CT to NT regime is rather delicate. Given that the lack of soil cultivation considerably reduces the soil N availability, hence, using NT technique alone as a substitute for CT is not agronomically feasible. Instead, an optimal application of NT is achievable by acting simultaneously on other factors of the cropping management, particularly the N-fertilization strategy, to maximize the crop N use efficiency and increase crop yield, which are essential requirements for a more sustainable agriculture.

Conservative tillage techniques have several agro-ecological benefits for organic farming. The application of these techniques, however, can create quite a few challenges due to the increased weed competition. Here, we report the results of an organic field experiment in which the responses of wheat and weeds to no tillage (NT) were evaluated compared with conventional tillage (CT). We also tested the hypothesis that, under NT, moving up the sowing date, compared with using the ordinary sowing date for the study area, can result in increased competitiveness of the crop against weeds. Two wheat genotypes, a modern variety and an ancient landrace, were tested.Substantial reductions in grain yield and protein content were observed in wheat under NT than under CT when the ordinary sowing date was used. This was mainly due to the considerable increase in weed biomass under NT. The tillage system also altered the composition of weed flora, with some species favored under NT and others under CT. In general, early sowing mitigated the detrimental effect of NT on yield. The two genotypes responded differently to the treatments. The early sowing in the modern variety reduced but did not eliminate the advantages of CT over NT, whereas no appreciable differences in grain yield were observed between CT and NT in the landrace.Our results show clearly that, under organic management, using NT alone as a substitute for CT is not agronomically feasible. Moving up the sowing date and using a competitive genotype can help mitigate the negative effects of NT, but surely a more effective application of NT could be achieved by acting simultaneously on other factors of the cropping management system (e.g. crop rotation, fertilization strategy, type of seeder). © 2022 The Authors. Journal of The Science of Food and Agriculture published by John WileySons Ltd on behalf of Society of Chemical Industry.

The salinity of soil is a relevant environmental problem around the world, with climate change raising its relevance, particularly in arid and semiarid areas. Arbuscular Mycorrhizal Fungi (AMF) positively affect plant growth and health by mitigating biotic and abiotic stresses, including salt stress. The mechanisms through which these benefits manifest are, however, still unclear. This work aimed to identify key genes involved in the response to salt stress induced by AMF using RNA-Seq analysis on durum wheat (Triticum turgidum L. subsp. durum Desf. Husn.). Five hundred sixty-three differentially expressed genes (DEGs), many of which involved in pathways related to plant stress responses, were identified. The expression of genes involved in trehalose metabolism, RNA processing, vesicle trafficking, cell wall organization, and signal transduction was significantly enhanced by the AMF symbiosis. A downregulation of genes involved in both enzymatic and non-enzymatic oxidative stress responses as well as amino acids, lipids, and carbohydrates metabolisms was also detected, suggesting a lower oxidative stress condition in the AMF inoculated plants. Interestingly, many transcription factor families, including WRKY, NAC, and MYB, already known for their key role in plant abiotic stress response, were found differentially expressed between treatments. This study provides valuable insights on AMF-induced gene expression modulation and the beneficial effects of plant-AMF interaction in durum wheat under salt stress.

Collateral arteries bridge opposing artery branches, forming a natural bypass that can deliver blood flow downstream of an occlusion. Inducing coronary collateral arteries could treat cardiac ischemia, but more knowledge on their developmental mechanisms and functional capabilities is required. Here we used whole-organ imaging and three-dimensional computational fluid dynamics modeling to define spatial architecture and predict blood flow through collaterals in neonate and adult mouse hearts. Neonate collaterals were more numerous, larger in diameter and more effective at restoring blood flow. Decreased blood flow restoration in adults arose because during postnatal growth coronary arteries expanded by adding branches rather than increasing diameters, altering pressure distributions. In humans, adult hearts with total coronary occlusions averaged 2 large collaterals, with predicted moderate function, while normal fetal hearts showed over 40 collaterals, likely too small to be functionally relevant. Thus, we quantify the functional impact of collateral arteries during heart regeneration and repair–a critical step toward realizing their therapeutic potential.

Cell-free RNA from liquid biopsies can be analyzed to determine disease tissue of origin. We extend this concept to identify cell types of origin using the Tabula Sapiens transcriptomic cell atlas as well as individual tissue transcriptomic cell atlases in combination with the Human Protein Atlas RNA consensus dataset. We define cell type signature scores, which allow the inference of cell types that contribute to cell-free RNA for a variety of diseases.

INTRODUCTION: Although the genome is often called the blueprint of an organism, it is perhaps more accurate to describe it as a parts list composed of the various genes that may or may not be used in the different cell types of a multicellular organism. Although nearly every cell in the body has essentially the same genome, each cell type makes different use of that genome and expresses a subset of all possible genes. This has motivated efforts to characterize the molecular composition of various cell types within humans and multiple model organisms, both by transcriptional and proteomic approaches. We created a human reference atlas comprising nearly 500,000 cells from 24 different tissues and organs, many from the same donor. This atlas enabled molecular characterization of more than 400 cell types, their distribution across tissues, and tissue-specific variation in gene expression. RATIONALE: One caveat to current approaches to make cell atlases is that individual organs are often collected at different locations, collected from different donors, and processed using different protocols. Controlled comparisons of cell types between different tissues and organs are especially difficult when donors differ in genetic background, age, environmental exposure, and epigenetic effects. To address this, we developed an approach to analyzing large numbers of organs from the same individual. RESULTS: We collected multiple tissues from individual human donors and performed coordinated single-cell transcriptome analyses on live cells. The donors come from a range of ethnicities, are balanced by gender, have a mean age of 51 years, and have a variety of medical backgrounds. Tissue experts used a defined cell ontology terminology to annotate cell types consistently across the different tissues, leading to a total of 475 distinct cell types with reference transcriptome profiles. The full dataset can be explored online with the cellxgene tool. Data were collected for the bladder, blood, bone marrow, eye, fat, heart, kidney, large intestine, liver, lung, lymph node, mammary, muscle, pancreas, prostate, salivary gland, skin, small intestine, spleen, thymus, tongue, trachea, uterus, and vasculature. Fifty-nine separate specimens in total were collected, processed, and analyzed, and 483,152 cells passed quality control filtering. On a per-compartment basis, the dataset includes 264,824 immune cells, 104,148 epithelial cells, 31,691 endothelial cells, and 82,478 stromal cells. Working with live cells, as opposed to isolated nuclei, ensured that the dataset includes all mRNA transcripts within the cell, including transcripts that have been processed by the cell’s splicing machinery, thereby enabling insight into variation in alternative splicing. The Tabula Sapiens also provided an opportunity to densely and directly sample the human microbiome throughout the gastrointestinal tract. The intestines from two donors were sectioned into five regions: the duodenum, jejunum, ileum, and ascending and sigmoid colon. Each section was transected, and three to nine samples were collected from each location, followed by amplification and sequencing of the 16S ribosomal RNA gene. CONCLUSION: The Tabula Sapiens has revealed discoveries relating to shared behavior and subtle, organ-specific differences across cell types. We found T cell clones shared between organs and characterized organ-dependent hypermutation rates among B cells. Endothelial cells and macrophages are shared across tissues, often showing subtle but clear differences in gene expression. We found an unexpectedly large and diverse amount of cell type–specific RNA splice variant usage and discovered and validated many previously undefined splices. The intestinal microbiome was revealed to have nonuniform species distributions down to the 3-inch (7.62-cm) length scale. These are but a few examples of how the Tabula Sapiens represents a broadly useful reference to deeply understand and explore human biology at cellular resolution.

Microplastics are recognized as a factor of global change contaminating many environmental compartments. Agricultural soils are very likely to receive microplastic contamination and are of particular concern due to their role in food production. Microplastic fibers have already been shown to be able to affect soil properties, but their effect on different soil types is poorly understood. Moreover, limited information is available on how the presence of this pollutant can affect soil water erosion processes, which are extremely important issues in many environments. In the light of this, we performed two experiments (carried out on a microscale) to investigate how the presence of polyester microplastic fibers affects soil physical and hydrological parameters and processes such as aggregate formation and soil erosion in three different agricultural soil types (a Vertisol, an Entisol, and an Alfisol). Our data show that the effects of polyester microplastic fibers on soil physical parameters and erosion are strongly dependent on soil type. We found that microplastic fiber contamination can affect soil bulk density, capacitive indicators of soil physical quality, and decrease the formation of new aggregates (labile in the incubation period applied in our experiments) but did not affect their stability in water. However, we found that polyester microplastic fibers reduced soil loss and sediment concentration, especially in the most erodible soils. In this paper, we provide some hypotheses, but certainly future data are still needed to confirm or disprove our hypotheses. Overall, our results highlight the importance of broadly exploring soil properties, such as texture, mineralogy, organic carbon content, etc., to better understand how the various soil types respond to microplastic fiber contamination.

Microplastic contamination, like other global change factors, can induce effects on ecosystem functions and processes, affecting various soil biophysical properties. However, effects of such contaminants on nutrient cycles in agroecosystems are still poorly understood. We here performed two pot experiments to investigate the effect of polyester microplastic fibers (PMFs) on soil physical properties, nitrogen cycle, and plant performance in a maize-based agroecosystem. Moreover, we followed the N loss via leaching in soil contaminated or not with PMFs by simulating heavy rainfall events that mimic a future scenario of climate change. Our results show that soil contaminated with PMFs (at a concentration of 0.5% w/w) can jeopardize agroecosystem sustainability by affecting soil physical properties and in particular soil macro- and microporosity, the nitrogen cycle, and plant performance. In particular, we found that soil PMF contamination limited crop growth and N uptake by circa 30%, and consequently increased N loss via leaching. Overall, our findings show that soil contamination with PMFs may pose problems to future agricultural challenges like food security and environmental protection.

Rationale: Coronary artery disease is the leading cause of death worldwide, but there are currently no methods to stimulate artery growth or regeneration in diseased hearts. Studying how arteries are built during development could illuminate strategies for re-building these vessels during ischemic heart disease. We previously found that Dach1 deletion in mouse embryos resulted in small coronary arteries. However, it was not known whether Dach1 gain-of-function would be sufficient to increase arterial vessels and whether this could benefit injury responses. Objective: We investigated how Dach1 overexpression in endothelial cells affected transcription and artery differentiation, and how it influenced recovery from myocardial infarction. Methods and Results: Dach1 was genetically overexpressed in coronary endothelial cells in either developing or adult hearts using ApjCreER. This increased the length and number of arterial end branches expanded arteries during development, in both the heart and retina, by inducing capillary endothelial cells to differentiate and contribute to growing arteries. Single-cell RNA sequencing of endothelial cells undergoing Dach1 -induced arterial specification indicated that it potentiated normal artery differentiation, rather than functioning as a master regulator of artery cell fate. Single-cell RNA sequencing also showed that normal arterial differentiation is accompanied by repression of lipid metabolism genes, which were also repressed by Dach1. In adults, Dach1 overexpression did not cause a statistically significant change artery structure before injury, but increased the number of perfused arteries in the injury zone post-myocardial infarction. Conclusions: Our data demonstrate that increasing Dach1 is a novel method for driving artery specification and extending arterial branches, which could be explored as a means of mitigating the effects of coronary artery disease.

Many aspects concerning the role of arbuscular mycorrhizal (AM) fungi in plant nutrient uptake from organic sources remain unclear. Here, we investigated the contribution of AM symbiosis to N and P uptake by durum wheat after the addition of a high C:N biomass to a P-limited soil. Plants were grown in pots in the presence or absence of a multispecies AM inoculum, with (Org) or without (Ctr) the addition of 15N-labelled organic matter (OM). A further treatment, in which 15N was applied in mineral form (Ctr+N) in the same amount as that supplied in the Org treatment, was also included. Inoculation with AM had positive effects on plant growth in both control treatments (Ctr and Ctr+N), mainly linked to an increase in plant P uptake. The addition of OM, increasing the P available in the soil for the plants, resulted in a marked decrease in the contribution of AM symbiosis to plant growth and nutrient uptake, although the percentage of mycorrhization was higher in the Org treatment than in the controls. In addition, mycorrhization drastically reduced the recovery of 15N from the OM added to the soil whereas it slightly increased the N recovery from the mineral fertiliser. This suggests that plants and AM fungi probably exert a differential competition for different sources of N available in the soil. On the whole, our results provide a contribution to a better understanding of the conditions under which AM fungi can play an effective role in mitigating the negative effects of nutritional stresses in plants. Supplementary Information The online version contains supplementary material available at 10.1007/s00572-021-01031-8.

Coronary artery disease (CAD) is the leading cause of death worldwide, but there are currently no available methods to stimulate growth or regeneration of artery networks in diseased hearts. Studying how arteries are built during embryonic development could illuminate strategies for re-building these vessels in the setting of ischemic heart disease. We previously found, using loss-of-function experiments, that the transcription factorDach1is required for coronary artery development in mouse embryos. Here, we report thatDach1overexpression in endothelial cells (ECs) extended coronary arteries and improved survival and heart function in adult mice following myocardial infarction (MI).Dach1overexpression increased the length and number of arterial end branches, in both heart and retinal vasculature, by causing additional capillary ECs to differentiate into arterial ECs and contribute to growing arteries. Single-cell RNA sequencing (scRNAseq) of ECs undergoingDach1-induced arterial specification indicated that it potentiated normal artery differentiation, rather than functioning as a master regulator of artery cell fate. ScRNAseq also showed that normal arterial differentiation is accompanied by repression of lipid metabolism genes, which were also repressed by Dach1 prior to arterialization. Together, these results demonstrate that increasing the expression level ofDach1is a novel pathway for driving specification of artery ECs and extending arterial vessels, which could be explored as a means of increasing artery coverage to mitigate the effects of CAD.

Plant performance is strongly dependent on nitrogen (N), and thus increasing N nutrition is of great relevance for the productivity of agroecosystems. The effects of arbuscular mycorrhizal (AM) fungi on plant N acquisition are debated because contradictory results have been reported. Using 15N-labeled fertilizers as a tracer, we evaluated the effects of AM fungi on N uptake and recovery from mineral or organic sources in durum wheat. Under sufficient N availability, AM fungi had no effects on plant biomass but increased N concentrations in plant tissue, plant N uptake, and total N recovered from the fertilizer. In N-deficient soil, AM fungi led to decreased aboveground biomass, which suggests that plants and AM fungi may have competed for N. When the organic source had a low carbon-to-nitrogen ratio, AM fungi favored both plant N uptake and N recovery. In contrast, when the organic source had a high carbon-to-nitrogen ratio, a clear reduction in N recovery from the fertilizer was observed. Overall, the results indicate an active role of arbuscular mycorrhizae in favoring plant N-related traits when N is not a limiting factor and show that these fungi help in N recovery from the fertilizer. These results hold great potential for increasing the sustainability of durum wheat production.

International audience; Arteries and veins are specified by antagonistic transcriptional programs. However, during development and regeneration, new arteries can arise from pre-existing veins through a poorly understood process of cell fate conversion. Here, using single-cell RNA sequencing and mouse genetics, we show that vein cells of the developing heart undergo an early cell fate switch to create a pre-artery population that subsequently builds coronary arteries. Vein cells underwent a gradual and simultaneous switch from venous to arterial fate before a subset of cells crossed a transcriptional threshold into the pre-artery state. Before the onset of coronary blood flow, pre-artery cells appeared in the immature vessel plexus, expressed mature artery markers, and decreased cell cycling. The vein-specifying transcription factor COUP-TF2 (also known as NR2F2) prevented plexus cells from overcoming the pre-artery threshold by inducing cell cycle genes. Thus, vein-derived coronary arteries are built by pre-artery cells that can differentiate independently of blood flow upon the release of inhibition mediated by COUP-TF2 and cell cycle factors.

During development, the formation of a mature, well-functioning heart requires transformation of the ventricular wall from a loose trabecular network into a dense compact myocardium at mid-gestation. Failure to compact is associated in humans with congenital diseases such as left ventricular non-compaction (LVNC). The mechanisms regulating myocardial compaction are however still poorly understood. Here, we show that deletion of the Ino80 chromatin remodeler in vascular endothelial cells prevents ventricular compaction in the developing mouse heart. This correlates with defective coronary vascularization, and specific deletion of Ino80 in the two major coronary progenitor tissues—sinus venosus and endocardium—causes intermediate phenotypes. In vitro, endothelial cells promote myocardial expansion independently of blood flow in an Ino80-dependent manner. Ino80 deletion increases the expression of E2F-activated genes and endothelial cell S-phase occupancy. Thus, Ino80 is essential for coronary angiogenesis and allows coronary vessels to support proper compaction of the heart wall., Heart development requires compaction of the ventricular wall into a dense myocardium at mid-gestation. Here, Rhee and colleagues show that the chromatin remodeller Ino80 is critical for the formation of the coronary vasculature, and show that coronary vessels are needed for successful cardiac compaction during embryonic development.

The introduction of legumes into crop sequences and the reduction of tillage intensity are both proposed as agronomic practices to mitigate the soil degradation and negative impact of agriculture on the environment. However, the joint effects of these practices on nitrous oxide (NO) and ammonia (NH) emissions from soil remain unclear, particularly concerning semiarid Mediterranean areas. In the frame of a long-term field experiment (23 years), a 2-year study was performed on the faba bean (Vicia faba L.) to evaluate the effects of the long-term use of no tillage (NT) compared to conventional tillage (CT) on yield and NO and NH emissions from a Vertisol in a semiarid Mediterranean environment. Changes induced by the tillage system in soil bulk density, water filled pore space (WFPS), organic carbon (TOC) and total nitrogen (TN), denitrifying enzyme activity (DEA), and bacterial gene (16S, amoA, and nosZ) abundance were measured as parameters potentially affecting N gas emissions. No tillage, compared with CT, significantly increased the faba bean grain yield by 23%. The tillage system had no significant effect on soil NH emissions. Total NO emissions, averaged over two cropping seasons, were higher in NT than those in CT plots (2.58 vs 1.71 kg NO-N ha, respectively; P < 0.01). In addition, DEA was higher in NT compared to that in CT (74.6 vs 18.6 μg NO-N kg h; P < 0.01). The higher NO emissions in NT plots were ascribed to the increase of soil bulk density and WFPS, bacteria (16S abundance was 96% higher in NT than that in CT) and N cycle genes (amoA and nosZ abundances were respectively 154% and 84% higher in NT than that in CT). The total NO emissions in faba bean were similar to those measured in other N-fertilized crops. In conclusion, a full evaluation of NT technique, besides the benefits on soil characteristics (e.g. TOC increase) and crop yield, must take into account some criticisms related to the increase of NO emissions compared to CT., We thank Beatriz Moreno (CSIC-EEZ) and Vincenzo Cannella (Università degli Studi di Palermo) for technical advice and support. This work was funded by the Italian Ministry for Education, University, and Research (MIUR) to Fondazione Angelo e Salvatore Lima Mancuso – PON/01_01145 Project – ISCOCEM.

The endocardium is a specialized endothelium that lines the inner surface of the heart. Functional studies in mice and zebrafish have established that the endocardium is a source of instructive signals for the development of cardiac structures, including the heart valves and chambers. Here, we characterized the NOTCH-dependent endocardial secretome by manipulating NOTCH activity in mouse embryonic endocardial cells (MEEC) followed by mass spectrometry-based proteomics. We profiled different sets of soluble factors whose secretion not only responds to NOTCH activation but also shows differential ligand specificity, suggesting that ligand-specific inputs may regulate the expression of secreted proteins involved in different cardiac development processes. NOTCH signaling activation correlates with a transforming growth factor-β2 (TGFβ2)-rich secretome and the delivery of paracrine signals involved in focal adhesion and extracellular matrix (ECM) deposition and remodeling. In contrast, NOTCH inhibition is accompanied by the up-regulation of specific semaphorins that may modulate cell migration. The secretome protein expression data showed a good correlation with gene profiling of RNA expression in embryonic endocardial cells. Additional characterization by in situ hybridization in mouse embryos revealed expression of various NOTCH candidate effector genes (Tgfβ2, Loxl2, Ptx3, Timp3, Fbln2, and Dcn) in heart valve endocardium and/or mesenchyme. Validating these results, mice with conditional Dll4 or Jag1 loss-of-function mutations showed gene expression alterations similar to those observed at the protein level in vitro These results provide the first description of the NOTCH-dependent endocardial secretome and validate MEEC as a tool for assaying the endocardial secretome response to a variety of stimuli and the potential use of this system for drug screening. We thank C. Martí Gómez-Aldaraví for help with graphic representation and critical reading of the manuscript, and S. Bartlett for English editing. RTC is supported by a Foundation La Caixa PhD fellowship (Ref LCF/BQ/ES15/10360023). LLZ is supported by a Ramón y Cajal postdoctoral contract (Ref: RYC-2016-20917). JLdlP is funded by grants SAF2016-78370-R, CB16/11/00399 (CIBER CV), and RD16/0011/0021 (TERCEL) from the Ministerio de Ciencia, Innovación y Universidades, and grants from the Fundación BBVA (Ref.: BIO14_298) and Fundación La Marató TV3 (Ref.: 20153431). JV is supported by grants BIO2015-67580-P and CB16/11/00277 (CIBER CV) from the Ministerio de Ciencia, Innovación y Universidades, and Carlos III Institute of Health-Fondo de Investigación Sanitaria (Grant ProteoRed-PRB3-IPT17/0019-ISCIII-SGEFI/ERDF), the Fundación La Marató TV3 (Ref. 122/C/2015) and “La Caixa” Banking Foundation (project code HR17-00247). The cost of this publication was supported in part with funds from the ERDF. The CNIC is supported by the Ministerio de Ciencia, Innovación y Universidades and the Pro CNIC Foundation, and is a Severo Ochoa Center of Excellence (SEV-2015-0505). Sí

Arbuscular mycorrhizal fungi (AMF) are soil microrganisms that establish symbiosis with plants positively influencing their resistance to abiotic stresses. The aim of this work was to identify wheat miRNAs differentially regulated by water deficit conditions in presence or absence of AMF treatment. Small RNA libraries were constructed for both leaf and root tissues considering four conditions: control (irrigated) or water deficit in presence/absence of mycorrhizal (AMF) treatment. A total of 12 miRNAs were significantly regulated by water deficit in leaves: five in absence and seven in presence of AMF treatment. In roots, three miRNAs were water deficit-modulated in absence of mycorrhizal treatment while six were regulated in presence of it. The most represented miRNA family was miR167 that was regulated by water deficit in both leaf and root tissues. Interestingly, miR827-5p was differentially regulated in leaves in the absence of mycorrhizal treatment while it was water deficit-modulated in roots irrespective of AMF treatment. In roots, water deficit repressed miR827-5p, miR394, miR6187, miR167e-3p, and miR9666b-3p affecting transcription, RNA synthesis, protein synthesis, and protein modifications. In leaves, mycorrhizae modulated miR5384-3p and miR156e-3p affecting trafficking and cell redox homeostasis. DNA replication and transcription regulation should be targeted by the repression of miR1432-5p and miR166h-3p. This work provided interesting insights into the post-transcriptional mechanisms of wheat responses to water deficit in relation to mycorrhizal symbiosis.

Determining the best conservation agriculture practices for increasing soil organic carbon (C) and hence soil quality is of paramount importance in the semi-arid Mediterranean environment, where soils are experiencing a continuous decline in organic matter. Therefore, the aim of this long-term study was to assess the combined effects of tillage system and crop sequence on soil organic C and biochemical properties of soil generally used as indicators of soil quality. After 23 years of continuous application of contrasting tillage systems (conventional tillage [CT], vs. no tillage [NT]) and crop sequences (wheat monoculture vs. wheat-faba bean rotation), soil samples were collected from topsoil (0–15 cm) and subsoil (15–30 cm) at three different times during a cropping year. Soil samples were analyzed for total and labile organic C pools, microbial biomass C (MBC) and microbial biomass N, basal respiration, and the abundance of main microbial groups by phospholipid fatty acids. Long-term NT increased total organic C (TOC) at a yearly rate of 0.17 g kg−1. This in turn stimulated microbial biomass, in particular Gram-negative bacteria. This suggests a higher soil quality in NT, as was confirmed by the increase in MBC/TOC and the decrease in stress indices. In contrast, no differences were observed with regard to fungal biomass. These findings suggest the need to reconsider the role of specific bacterial groups in organic C accumulation in soils of semiarid environments. It is interesting that the effects of long-term NT varied widely by crop sequence, whereas in CT changes in biochemical characteristics and in the main microbial groups due to crop sequence were modest. Thus, the interaction among various aspects of agronomic management modulates the effects of substrate quality on chemical and biological properties of soil.

In organic farming, the adoption of the conventional tillage (CT) technique is considered by many farmers to be necessary to control weeds. Such tillage system, in fact, permits to bury weed seeds deep in the soil by means of soil inversion with moldboard plowing and to eliminate the weed plants that gradually emerge by means of the secondary tillage operations. However, it is also true that intensive tillage progressively reduces the soil organic matter content and the stability of soil aggregates, thus increasing the risk of soil erosion (Six et al. 2000). This is in contrast with one of the basic principles of organic agriculture, which is the conservation of soil fertility. Alternatively to CT, the no tillage (NT) technique can maintain or even enhance soil fertility by increasing C storage, soil biological activity, and soil aggregate stability, but, as a matter of fact, its application relies on herbicide use as the primary weed control mechanism (Gattinger et al. 2011). In the light of these considerations, efforts must be made to revisit the NT technique to make it applicable in organic farming. Without prejudice to the fact that this challenge should be addressed through a systemic approach (Peigné et al. 2007), one possible option could be to take advantage of the possibility given by the NT technique to sow the crop in an earlier period than what usually the farmer does when adopts the CT technique. Anticipating the sowing time would allow operating when most of the weed plants are still poorly developed, so that the sowing operation itself can kill many of them. Moreover, sowing early, when temperatures are still relatively mild, could accelerate the initial growth, thus reducing the period during which the crop is particularly vulnerable to weed competition. Usually, early sowing in the CT systems is not possible since a proper seedbed preparation needs time so that clods formed as a result of plowing could be broken down by natural weathering processes and by one or more secondary tillage operations. Therefore, an experiment was performed under organic management to study the effects of NT compared to CT on durum wheat (Triticum durum Desf.) grain yield, and to verify whether early sowing under NT conditions, compared to sowing at the ordinary time for the study area, can provide an advantage to the crop by increasing its competitiveness against weeds. Furthermore, the above effects were investigated on two durum wheat genotypes highly different for pheno-morphological and agronomic characteristics, assuming for them different competitiveness against weeds.

Durum wheat is the keystone of the agro-ecosystems in the arable land of the Mediterranean environments and an important part of its area falls within organic farms. For this crop competition exerted by weeds for the use of resources (natural and auxiliary) can determine drastic yield and quality reductions (Ruisi et al., 2015). In organic farming such critical issue is often addressed through a remodelling of several techniques such as soil tillage management, sowing time, plant density and genotype choice. With regard to the latter, there is a growing interest by organic farmers towards the old varieties as they, compared to the modern varieties, have a definitely greater competitive weed abilities thanks to some morpho-physiological plant traits (establishment speed, tillering capacity, plant height) (Röös et al., 2018); moreover, the old varieties/landraces are often characterized by a greater protein and gluten content and for peculiar sensory properties (Newton et al., 2010; Vita et al., 2016). On the other hand, the new varieties have a much higher production potential and technological characteristics of the grain often more responsive to the needs of the processing industry (De Vita et al., 2007). This study, carried out in a organic farming system, aimed to answer the following questions: 1) can the mixture of old and modern durum wheat varieties offer advantages over the monovarietal crop, combining the qualities of the different genotypes? 2) Which mixing ratio should be used in order to maximize the potential advantages of the mixture?

Intercropping is an agricultural practice that can offer several benefits allowing a better native resources use efficiency and, consequently, a restraint of the auxiliary inputs and often a greater production compared to the monocultures (Brooker et al. 2015). Several authors observed that, in a legume/non-legume mixture, one of the benefits could be the N transfer (up to 80 % of the non-legume N demand; Thilakarathna et al. 2016). The transfer may occur via different pathways: legume rhizodeposition, plant tissue decomposition and direct transfer through arbuscular mycorrhizal fungi (AMF) (Bedoussac et al. 2015). The latter, can simultaneously establish symbiotic relationship with different plant species creating a common mycorrhizal network, which serve as a preferential pathway for exchange among plants (He et al. 2003). However, contrasting results have been reported about the contribution of the AMF on N transfer; for instance, Li et al. (2009) showed that N transfer from mung bean to rice increased from 5.4% to 15.7% due to hyphal linkage, whereas Ikram et al. (1994) showed no significant differences with or without AMF inoculum. This experiment aimed to investigate the role of AMF on N transfer from faba bean to durum wheat grown in mixture, using the stem 15N injecting method.

Three obligately heterofermentative lactic acid bacteria (LAB) strains (Lactobacillus sanfranciscensis PON100336, Leuconostoc citreum PON10079 and Weissella cibaria PON10030) were used in this study as a multi-species starter culture for sourdough production. The starter inoculum was prepared and propagated in sterile semolina extract (SSE) broth. Acidification kinetics, microbiological counts detected on specific media for sourdough LAB, polymorphic profile comparison and species-specific PCRs evidenced a stability of the liquid inoculum over time determining its suitability for direct addition to semolina. In order to validate this innovative method for the production of durum wheat (Triticum durum Desf) sourdoughs, 15 semolinas (from ten old and five modern genotypes cultivated in Sicily, southern Italy) were used to prepare the SSEs and to produce sourdoughs and finally breads. Chemical and microbiological analyses of the sourdoughs and the evaluation of the quality parameters (weight loss, height, crumb and crust colour, image analysis and volatile organic compound generation) of the resulting breads indicated that the direct addition of the liquid inocula propagated in SSE is a valuable method to stabilise the production of sourdoughs. The differences registered during the technological characterisation of the breads were underlined by the sensory tests and the multivariate analysis and are mainly imputable to the type of semolina.

Rationale: The Notch signaling pathway is crucial for primitive cardiac valve formation by epithelial–mesenchymal transition, and NOTCH1 mutations cause bicuspid aortic valve; however, the temporal requirement for the various Notch ligands and receptors during valve ontogeny is poorly understood. Objective: The aim of this study is to determine the functional specificity of Notch in valve development. Methods and Results: Using cardiac-specific conditional targeted mutant mice, we find that endothelial/endocardial deletion of Mib1-Dll4-Notch1 signaling, possibly favored by Manic-Fringe, is specifically required for cardiac epithelial–mesenchymal transition. Mice lacking endocardial Jag1 , Notch1 , or RBPJ displayed enlarged valve cusps, bicuspid aortic valve, and septal defects, indicating that endocardial Jag1 to Notch1 signaling is required for post–epithelial–mesenchymal transition valvulogenesis. Valve dysmorphology was associated with increased mesenchyme proliferation, indicating that Jag1-Notch1 signaling restricts mesenchyme cell proliferation non–cell autonomously. Gene profiling revealed upregulated Bmp signaling in Jag1 -mutant valves, providing a molecular basis for the hyperproliferative phenotype. Significantly, the negative regulator of mesenchyme proliferation, Hbegf , was markedly reduced in Jag1 -mutant valves. Hbegf expression in embryonic endocardial cells could be readily activated through a RBPJ-binding site, identifying Hbegf as an endocardial Notch target. Accordingly, addition of soluble heparin-binding EGF-like growth factor to Jag1 -mutant outflow tract explant cultures rescued the hyperproliferative phenotype. Conclusions: During cardiac valve formation, Dll4-Notch1 signaling leads to epithelial–mesenchymal transition and cushion formation. Jag1-Notch1 signaling subsequently restrains Bmp-mediated valve mesenchyme proliferation by sustaining Hbegf-EGF receptor signaling. Our studies identify a mechanism of signaling cross talk during valve morphogenesis involved in the origin of congenital heart defects associated with reduced NOTCH function.

Heart muscle maintains blood circulation, while skeletal muscle powers skeletal movement. Despite having similar myofibrilar sarcomeric structures, these striated muscles differentially express specific sarcomere components to meet their distinct contractile requirements. The mechanism responsible is still unclear. We show here that preservation of the identity of the two striated muscle types depends on epigenetic repression of the alternate lineage gene program by the chromatin remodeling complex Chd4/NuRD. Loss of Chd4 in the heart triggers aberrant expression of the skeletal muscle program, causing severe cardiomyopathy and sudden death. Conversely, genetic depletion of Chd4 in skeletal muscle causes inappropriate expression of cardiac genes and myopathy. In both striated tissues, mitochondrial function was also dependent on the Chd4/NuRD complex. We conclude that an epigenetic mechanism controls cardiac and skeletal muscle structural and metabolic identities and that loss of this regulation leads to hybrid striated muscle tissues incompatible with life. We thank Drs. Richard Harvey (University of Sydney, Australia) and Michael Schneider (Imperial College, London, UK) for providing Nkx2.5cre/+ and α-MHCcre/+ transgenic lines, respectively, Dr. S. Bartlett for English editing, and Gemma Benito and Ana Guío for technical assistance. J.M.R. is supported by the Spanish Ministry of Economy and Competitiveness (Ministerio de Economía y Competitividad; SAF 2012 34296 and SAF 2015 63633-R), the Spanish Ministry of Health (Ministerio de Sanidad y Consumo) Red de Investigación Cardiovascular (RIC; grant RD06/0042/0022), and the Fundación La Marató TV3 (264/C/2012). The RIC also supports J.L.P. (RD12/0042/0005) and L.J.J.-B. (RD12/0042/0056). RIC grants are partially funded by FEDER funds. P.M.-C., J.A.E., and J.L.P. are supported by SAF2012-38547, SAF2015-67369-R, SAF2012-1207, and SAF2013-45543-R, respectively. E.P. and P.M.-C. were also supported by SAF2015-67369-R. NGS experiments were performed at the CNIC Genomics Unit. IF of adult hearts was performed at the Advanced Fluorescence Microscopy Unit of (IBMB-CSIC) Barcelona. The CNIC is supported by the Spanish Ministry of Economy and Competitiveness and the Pro-CNIC Foundation and is a Severo Ochoa Center of Excellence (MINECO award SEV-2015-0505). R.A.-P. was supported by a Ramón y Cajal grant and is a holder of a Marie Curie grant (RA-P:UEO/MCA1108). M.Z. was a holder of a Marie Curie Outgoing International Fellowship (MOIF-CT-2006-039327), and P.G.A. was supported by a Ramón y Cajal grant from the Spanish Ministry of Education, Science and Sport.

Arbuscular mycorrhizal fungi (AMF) can play a key role in natural and agricultural ecosystems affecting plant nutrition, soil biological activity and modifying the availability of nutrients by plants. This research aimed at expanding the knowledge of the role played by AMF in the uptake of macro- and micronutrients and N transfer (using a 15 N stem-labelling method) in a faba bean/wheat intercropping system. It also investigates the role of AMF in biological N fixation (using the natural isotopic abundance method) in faba bean grown in pure stand and in mixture. Finally, it examines the role of AMF in driving competition and facilitation between faba bean and wheat. Durum wheat and faba bean were grown in pots (five pots per treatment) as sole crops or in mixture in the presence or absence of AMF. Root colonisation by AMF was greater in faba bean than in wheat and increased when species were mixed compared to pure stand (particularly for faba bean). Mycorrhizal symbiosis positively influenced root biomass, specific root length, and root density and increased the uptake of P, Fe, and Zn in wheat (both in pure stand and in mixture) but not in faba bean. Furthermore, AMF symbiosis increased the percentage of N derived from the atmosphere in the total N biomass of faba bean grown in mixture (+20%) but not in pure stand. Nitrogen transfer from faba bean to wheat was low (2.5–3.0 mg pot -1 ); inoculation with AMF increased N transfer by 20%. Overall, in terms of above- and belowground growth and uptake of nutrients, mycorrhization favoured the stronger competitor in the mixture (wheat) without negatively affecting the companion species (faba bean). Results of this study confirm the role of AMF in driving biological interactions among neighbouring plants. and analysis, decision to publish, or preparation of the manuscript.

During vertebrate cardiac development NOTCH signaling activity in the endocardium is essential for the crosstalk between endocardium and myocardium that initiates ventricular trabeculation and valve primordium formation. This crosstalk leads later to the maturation and compaction of the ventricular chambers and the morphogenesis of the cardiac valves, and its alteration may lead to disease. Although endocardial NOTCH signaling has been shown to be crucial for heart development, its physiological role in the myocardium has not been clearly established. Here we have used a genetic strategy to evaluate the role of NOTCH in myocardial development. We have inactivated the unique and ubiquitous NOTCH effector RBPJ in the early cardiomyocytes progenitors, and examined its consequences in cardiac development and function. Our results demonstrate that mice withcTnT-Cre-mediated myocardial-specific deletion ofRbpjdevelop to term, with homozygous mutant animals showing normal expression of cardiac development markers, and normal adult heart function. Similar observations have been obtained afterNotch1deletion withcTnT-Cre. We have also deletedRbpjin both myocardial and endocardial progenitor cells, using theNkx2.5-Credriver, resulting in ventricular septal defect (VSD), double outlet right ventricle (DORV), and bicuspid aortic valve (BAV), due to NOTCH signaling abrogation in the endocardium of cardiac valves territories. Our data demonstrate that NOTCH-RBPJ inactivation in the myocardium does not affect heart development or adult cardiac function.

In this chapter we illustrate the signalling interactions of the endocardium with the other cardiac tissues to coordinate cardiac development. First, we describe the developmental origins of the endocardium. Then we focus on the Notch pathway because of its unique signalling activity in the endocardium, and briefly describe the elements of this signalling mechanism and the key cardiogenic processes that require endocardial Notch signalling: patterning of the early embryonic endocardium into prospective territories for valves and ventricular chambers, early valve formation, ventricular trabeculation, and compaction. Finally, we discuss how Notch dysfunction in the endocardium results in cardiac structural malformations that can lead to congenital heart disease.

Summary Knowledge of the effects of agricultural practices on weed seedbank dynamics is essential for predicting future problems in weed management. This article reports data relative to weed seedbank structure after 18 years of continuous application of conventional tillage (CT, based on mouldboard ploughing) or no tillage (NT) within three crop sequences (continuous wheat, WW; wheat–faba bean, WF; and wheat–berseem clover, WB). Tillage system did not affect the size of the total weed seedbank, but altered both its composition and the distribution of seeds within the soil profile. In particular, the adoption of CT favoured some species (mainly Polygonum aviculare), whereas the continuous use of NT favoured other species (Papaver rhoeas, Phalaris spp. and Lactuca serriola). The effects of tillage system on weed seedbank size and composition were less pronounced in the WB cropping system than in either the WW or WF. Compared with WF and WB, WW resulted in an increase in total weed seedbank density (about 16 000 seedlings m−2 in WW, compared with 10 000 and 6000 seedlings m−2 in WF and WB, respectively) and a reduction in weed diversity, with a strong increase in some species (e.g. Polygonum aviculare). Our results for the effect of NT application on weed seedbank size and composition suggest that farmers should only apply such a conservative technique within an appropriate crop sequence.

This 2-year study aimed to verify whether the continuous application of no tillage (NT) for over 20 years, in comparison with conventional tillage (CT), affects nitrous oxide (NO) and ammonia (NH) emissions from a Vertisol and, if so, whether such an effect varies with crop sequence (continuous wheat, WW and wheat after faba bean, FW). To shed light on the mechanisms involved in determining N-gas emissions, soil bulk density, water filled pore space (WFPS), some carbon (C) and nitrogen (N) pools, denitrifying enzyme activity (DEA), and nitrous oxide reductase gene abundance (nosZ gene) were also assessed at 0–15 and 15–30 cm soil depth. Tillage system had no significant effect on total NH emissions. On average, total NO emissions were higher under NT (2.45 kg NO-N ha) than CT (1.72 kg NO-N ha), being the differences between the two tillage systems greater in FW than WW. The higher NO emissions in NT treatments were ascribed to the increased bulk density, WFPS, and extractable organic C under NT compared to CT, all factors that generally promote the production of NO. Moreover, compared to CT, NT enhanced the potential DEA (114 vs 16 μg N kg h) and nosZ gene abundance (116 vs 69 copy number mg dry soil) in the topsoil. Finally, NT compared to CT led to an average annual increase in C stock of 0.70 Mg C ha year. Though NT can increase the amount os soil organic matter so storing CO into soil, some criticisms related to the increase of NO emission arise, thereby suggesting the need for defining management strategies to mitigate such a negative effect., We thank Beatriz Moreno (CSIC-EEZ) and Vincenzo Cannella(Università degli Studi di Palermo) for technical advice and support.This work was funded by the Italian Ministry for Education, University and Research (MIUR) to Fondazione Angelo e Salvatore Lima Mancuso (Università degli Studi di Palermo)–PON/01_01145 Project–ISCOCEM

Sufficient blood flow to tissues relies on arterial blood vessels, but the mechanisms regulating their development are poorly understood. Many arteries, including coronary arteries of the heart, form through remodeling of an immature vascular plexus in a process triggered and shaped by blood flow. However, little is known about how cues from fluid shear stress are translated into responses that pattern artery development. Here, we show that mice lacking endothelial Dach1 had small coronary arteries, decreased endothelial cell polarization, and reduced expression of the chemokine Cxcl12. Under shear stress in culture, Dach1 overexpression stimulated endothelial cell polarization and migration against flow, which was reversed upon CXCL12/CXCR4 inhibition. In vivo, DACH1 was expressed during early arteriogenesis but was down in mature arteries. Mature artery-type shear stress (high, uniform laminar) specifically down-regulated DACH1, while the remodeling artery-type flow (low, variable) maintained DACH1 expression. Together, our data support a model in which DACH1 stimulates coronary artery growth by activating Cxcl12 expression and endothelial cell migration against blood flow into developing arteries. This activity is suppressed once arteries reach a mature morphology and acquire high, laminar flow that down-regulates DACH1. Thus, we identified a mechanism by which blood flow quality balances artery growth and maturation.

Autoimmune diseases are a family of more than 80 chronic, and often disabling, illnesses that develop when underlying defects in the immune system lead the body to attack its own organs, tissues, and cells. Diagnosis of autoimmune pathologies is based on research and identification of antinuclear antibodies (ANA) through indirect immunofluorescence (IIF) method and is performed by analyzing patterns and fluorescence intensity. We propose here a method to automatically classify the centromere pattern based on the grouping of centromeres on the cells through a clustering K-means algorithm. The described method was tested on a public database (MIVIA). The results of the test showed an Accuracy equal to (92.0 ± 1.0)%. Comparing our results with the results obtained on the MIVIA database it is possible to note that our method has a performance comparable with the three best values obtained. Indeed, the method here proposed allows an automatic segmentation and counting of the cells in the images, while the participants to the contest received the training set with the original images of the cells already segmented by specialists.

SUMMARYArbuscular mycorrhizal (AM) fungi increase nitrogen (N) uptake by their host plants, but their role in plant N capture from soil organic material is still unclear. In particular, it is not clear if AM fungi compete with the host plant for the N coming from the decomposing organic matter (OM), especially when the AM extraradical mycelium (ERM) and plant roots share the same soil volume. The goal of the present research was to study the effects of AM fungi on wheat N capture after the addition of 15N-labelled OM to soil. Durum wheat (Triticum durum) was grown under controlled conditions in a sand:soil mix and the following treatments were applied: (1) AM inoculation with Glomus mosseae and uninoculated control; and (2) soil amended with 15N-enriched maize leaves and unamended soil. The addition of OM reduced plant growth and N uptake. The AM fungi increased both plant growth and N uptake compared with uninoculated control plants and the effect was enhanced when wheat was grown in soil amended with OM compared with the unamended control. Although AM fungi increased soil N mineralization rates and total plant N uptake, they strongly reduced wheat N recovery from OM, suggesting that AM fungi have marked effects on competition between plants and bacteria for the different N sources in soil.

Left ventricular noncompaction (LVNC) causes prominent ventricular trabeculations and reduces cardiac systolic function. The clinical presentation of LVNC ranges from asymptomatic to heart failure. We show that germline mutations in human MIB1 (mindbomb homolog 1), which encodes an E3 ubiquitin ligase that promotes endocytosis of the NOTCH ligands DELTA and JAGGED, cause LVNC in autosomal-dominant pedigrees, with affected individuals showing reduced NOTCH1 activity and reduced expression of target genes. Functional studies in cells and zebrafish embryos and in silico modeling indicate that MIB1 functions as a dimer, which is disrupted by the human mutations. Targeted inactivation of Mib1 in mouse myocardium causes LVNC, a phenotype mimicked by inactivation of myocardial Jagged1 or endocardial Notch1. Myocardial Mib1 mutants show reduced ventricular Notch1 activity, expansion of compact myocardium to proliferative, immature trabeculae and abnormal expression of cardiac development and disease genes. These results implicate NOTCH signaling in LVNC and indicate that MIB1 mutations arrest chamber myocardium development, preventing trabecular maturation and compaction. 28.054 JCR (2013) Q1, 2/291 Biochemistry & molecular biology, 3/185 Cell biology, 1/122 Medicine, research & experimental