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High-throughput sequencing-based methods for bulked segregant analysis (BSA) allow for the rapid identification of genetic markers associated with traits of interest. BSA studies have successfully identified qualitative (binary) and quantitative trait loci (QTLs) using QTL mapping. However, most require population structures that fit the models available and a reference genome. Instead, high-throughput short-read sequencing can be combined with BSA of k-mers (BSA-k-mer) to map traits that appear refractory to standard approaches. This method can be applied to any organism and is particularly useful for species with genomes diverged from the closest sequenced genome. It is also instrumental when dealing with highly heterozygous and potentially polyploid genomes without phased haplotype assemblies and for which a single haplotype can control a trait. Finally, it is flexible in terms of population structure. Here, we apply the BSA-k-mer method for the rapid identification of candidate regions related to seed spot and seed size in diploid potato. Using a mixture of F1 and F2 individuals from a cross between 2 highly heterozygous parents, candidate sequences were identified for each trait using the BSA-k-mer approach. Using parental reads, we were able to determine the parental origin of the loci. Finally, we mapped the identified k-mers to a closely related potato genome to validate the method and determine the genomic loci underlying these sequences. The location identified for the seed spot matches with previously identified loci associated with pigmentation in potato. The loci associated with seed size are novel. Both loci are relevant in future breeding toward true seeds in potato.

The purpose of this study is to understand the reasons why Japanese higher education institutions show a lack of digital marketing activities. Specifically, we focused on the challenges faced by institutions with management and business faculties when implementing digital marketing. To achieve this purpose, the research invited 167 Japanese higher education institutions to respond to a self-administered online questionnaire followed by semi-structured interviews. 49 valid responses were obtained and then 11 interviews were performed. After analyzing the data collected using text coding, the results indicate the emergence of nine digital marketing enablers and barriers, namely training & knowledge, tools, financial resources & measurement results, human resources, leadership, process, internal public relations, organization strategy and external stakeholders. A comparison between the various factors shows a significant dominance of barriers with respect to enablers, reflecting the concerns of Japanese higher education institutions with respect to digital marketing. In addition, the interviews reveal the existence of four environmental factors which have motivated Japanese higher education institutions to adopt digital marketing.

Minichromosomes are small, sometimes circular, rearranged chromosomes consisting of one centromere and short chromosomal arms formed by treatments that break DNA, including plant transformation. Minichromosomes have the potential to serve as vectors to quickly move valuable genes across a wide range of germplasm, including into adapted crop varieties. To realize this potential, minichromosomes must be reliably generated, easily manipulated, and stably inherited. Here we show a reliable method for minichromosome formation in haploids resulting from CENH3-mediated genome elimination, a process that generates genome instability and karyotypic novelty specifically on one parental genome. First, we identified 2 out of 260 haploids, each containing a single-copy minichromosome originating from centromeric regions of chromosomes 1 and 3, respectively. The chromosome 1 minichromosome we characterized did not pair at meiosis but displayed consistent transmission over nine selfing generations. Next, we demonstrated that CENH3-based haploid induction can produce minichromosomes in a targeted manner. Haploid inducers carrying a selectable pericentromeric marker were used to isolate additional chromosome-specific minichromosomes, which occurred in 3 out of 163 haploids. Our findings document the formation of heritable, rearranged chromosomes, and we provide a method for convenient minichromosome production.

Urine-derived renal epithelial cells (URECs) are highly voided after kidney transplant and express typical kidney markers, including markers of kidney epithelial progenitor cells. Recently URECs have shown promising immunomodulatory properties when cultured with Peripheral Blood Mononuclear Cells (PBMCs), promoting an increase in the T regulatory cells. In vivo, kidney cells are highly exposed to damage associated molecules during both acute and chronic kidney injury. Neutrophil gelatinase-associated lipocalin (NGAL) is one of the most -known early marker of acute and chronic kidney damage. However, its role on the evolution of renal damage has not yet been fully described, nor has its impact on the characteristics of renal-derived cells during in vitro culture. The aim of this study is to investigate the effect of NGAL on the characteristics of URECs isolated after kidney transplant, by exposing these cells to the treatment with NGAL during in vitro culture and evaluating its effect on UREC viability, proliferation, and immunomodulatory potential. The exposure of URECs to NGAL reduced their viability and proliferative capacity, promoting the onset of apoptosis. The immunomodulatory properties of URECs were partially inhibited by NGAL, without affecting the increase of Treg cells observed during UREC-PBMCs coculture. These results suggest that the exposure to NGAL may compromise some features of kidney stem and specialized cell types, reducing their viability, increasing apoptosis, and partially altering their immunomodulatory properties. Thus, NGAL could represent a target for approaches acting on its inhibition or reduction to improve functional recovery.

IntroductionOwing to advances in high-throughput genome sequencing, QTL-Seq mapping of salt tolerance traits is a major platform for identifying soil-salinity tolerance QTLs to accelerate marker-assisted selection for salt-tolerant rice varieties. We performed QTL-BSA-Seq in the seedling stage of rice from a genetic cross of the extreme salt-sensitive variety, IR29, and “Jao Khao” (JK), a Thai salt-tolerant variety.MethodsA total of 462 F2 progeny grown in soil and treated with 160 mM NaCl were used as the QTL mapping population. Two high- and low-bulk sets, based on cell membrane stability (CMS) and tiller number at the recovery stage (TN), were equally sampled. The genomes of each pool were sequenced, and statistical significance of QTL was calculated using QTLseq and G prime (G′) analysis, which is based on calculating the allele frequency differences or Δ(SNP index).ResultsBoth methods detected the overlapping interval region, wherein CMS-bulk was mapped at two loci in the 38.41–38.85 Mb region with 336 SNPs on chromosome 1 (qCMS1) and the 26.13–26.80 Mb region with 1,011 SNPs on chromosome 3 (qCMS3); the Δ(SNP index) peaks were −0.2709 and 0.3127, respectively. TN-bulk was mapped at only one locus in the overlapping 38.26–38.95 Mb region on chromosome 1 with 575 SNPs (qTN1) and a Δ(SNP index) peak of −0.3544. These identified QTLs in two different genetic backgrounds of segregating populations derived from JK were validated. The results confirmed the colocalization of the qCMS1 and qTN1 traits on chromosome 1. Based on the CMS trait, qCMS1/qTN1 stably expressed 6%–18% of the phenotypic variance in the two validation populations, while qCMS1/qTN1 accounted for 16%–20% of the phenotypic variance in one validation population based on the TN trait.ConclusionThe findings confirm that the CMS and TN traits are tightly linked to the long arm of chromosome 1 rather than to chromosome 3. The validated qCMS-TN1 QTL can be used for gene/QTL pyramiding in marker-assisted selection to expedite breeding for salt resistance in rice at the seedling stage.

Human babesiosis is an emerging zoonotic disease; diffused especially in some regions of the United States, it has been less frequently observed in other continents, including Europe. Serological surveys suggest that babesiosis could be more frequent than expected in European countries, representing an emerging health-issue and a possible harm, especially in immunocompromised populations. Only one case of human babesiosis has been reported in Italy and data about the diffusion of the pathogen in this country are scant. We conducted a multicentric serological survey in 5 centers of North-Eastern Italy, aimed to detect the seroprevalence of Babesia spp. antibodies in 3 groups of immunocompromised patients: people living with HIV (PLHIV), rheumatologic patients undergoing immunosuppressive therapies and patients undergoing renal transplant. Among the 433 enrolled patients, 3 (0.7%) tested positive for Babesia spp. serology. All positive patients belonged to the PLHIV group, with a seroprevalence of 1.7% (3/180) in this population; the three serologically positive patients were all asymptomatic. They were all enrolled in the provinces of Bolzano and Trento, where seroprevalences of 3.1% and 3.6% were recorded, respectively. Our results suggest that further research is needed on this field, awareness should be raised toward the human disease in Europe, especially in immunocompromised patients, and this emerging health issue should be analyzed in a One-Health perspective to be fully understood.

The chromosome axis plays a crucial role in meiotic recombination. Here, we study the function of ASY1, the Arabidopsis homolog of the yeast chromosome axis-associated component Hop1. Specifically, we characterized cross-over (CO) distribution in female and male meiosis by deep sequencing of the progeny of an allelic series of asy1 mutants. Combining data from nearly 1,000 individual plants, we find that reduced ASY1 functionality leads to genomic instability and sometimes drastic genomic rearrangements. We further observed that COs are less frequent and appear in more distal chromosomal regions in plants with no or reduced ASY1 functionality, consistent with previous analyses. However, our sequencing approach revealed that the reduction in CO number is not as dramatic as suggested by cytological analyses. Analysis of double mutants of asy1 with mutants with three other CO factors, MUS81, MSH4, and MSH5, as well as the determination of foci number of the CO regulator MLH1 demonstrates that the majority of the COs in asy1, similar to the situation in the wildtype (WT), largely belong to the class I, which are subject to interference. However, these COs are redistributed in asy1 mutants and typically appear much closer than in the WT. Hence, ASY1 plays a key role in CO interference that spaces COs along a chromosome. Conversely, since a large proportion of chromosomes do not receive any CO, we conclude that CO assurance, the process that ensures the obligatory assignment of one CO per chromosome, is also affected in asy1 mutants.

Chromoanagenesis is a catastrophic event that involves localized chromosomal shattering and reorganization. In this study, we report a case of chromoanagenesis resulting from defective meiosis in the MEIOTIC ASYNAPTIC MUTANT 1 (asy1) background in Arabidopsis thaliana. We provide a detailed characterization of the genomic structure of this individual with a severely shattered segment of chromosome 1. We identified 260 novel DNA junctions in the affected region, most of which affect gene sequence on 1 or both sides of the junction. Our results confirm that asy1-related defective meiosis is a potential trigger for chromoanagenesis. This is the first example of chromoanagenesis associated with female meiosis and indicates the potential for genome evolution during oogenesis. Chromoanagenesis is a complex and catastrophic event that results in severely restructured chromosomes. It has been identified in cancer cells and in some plant samples, after specific triggering events. Here, we identified this kind of genome restructuring in a mutant that exhibits defective meiosis in the model plant system Arabidopsis thaliana.

Diverse morphological, cellular and physiological changes occur during seed maturation in Bixa orellana when the seed tissues form specialized cell glands that produce reddish latex with high bixin amounts. Transcriptomic profiling during seed development in three B. orellana accessions (P12, N4 and N5) with contrasting morphologic characteristics showed enrichment in pathways of triterpenes, sesquiterpenes, and cuticular wax biosynthesis. WGCNA allows groups of all identified genes in six modules the module turquoise, the largest and highly correlated with the bixin content. The high number of genes in this module suggests a diversification of regulatory mechanisms for bixin accumulation with the genes belonging to isoprene, triterpenes and carotene pathways, being more highly correlated with the bixin content. Analysis of key genes of the mevalonate (MVA) and the 2C-methyl-D-erythritol-4-phosphate (MEP) pathways revealed specific activities of orthologs of BoHMGR, BoFFP, BoDXS, and BoHDR. This suggests that isoprenoid production is necessary for compounds included in the reddish latex of developing seeds. The carotenoid-related genes BoPSY2, BoPDS1 and BoZDS displayed a high correlation with bixin production, consistent with the requirement for carotene precursors for apocarotenoid biosynthesis. The BoCCD gene member (BoCCD4-4) and some BoALDH (ALDH2B7.2 and ALDH3I1) and BoMET (BoSABATH1 and BoSABATH8) gene members were highly correlated to bixin in the final seed development stage. This suggested a contributing role for several genes in apocarotenoid production. The results revealed high genetic complexity in the biosynthesis of reddish latex and bixin in specialized seed cell glands in different accessions of B. orellana suggesting gene expression coordination between both metabolite biosynthesis processes.

Abstract Background The endocannabinoid (eCB) system and the serotonin (5-HT) are both implicated in the severity of the depression. 5-HT is synthesized from the amino acid tryptophan (Trp), which is also a precursor for kynurenine (Kyn) whose production is increased at the expense of 5-HT in depressed patients. No clinical studies have investigated the crosstalk between the eCB system and the Trp/5-HT/Kyn pathways. Here, we hypothesized that the eCB system is associated with an enhanced Kyn production in relation to the severity of depressive symptoms. Methods Eighty-two subjects (51 patients with a diagnosis of depressive disorder (DSM-5) and 31 healthy volunteers), were assessed with the Montgomery-Åsberg Depression Rating Scale (MADRS), Beck Depression Scale, and Global Clinical Impression. Serum concentrations of eCBs (N-arachidonoylethanolamine (AEA) and 2-arachidonoylglycerol (2-AG)); structurally related fatty acyl compounds 2-oleoylglycerol (2-OG), oleoylethanolamide (OEA), and palmitoylethanolamide (PEA); Trp, Kyn, Kyn/Trp ratio (an index of Trp degradation into Kyn) and 5-HT were also determined. Results Following a principal component analysis including the severity of depression, Kyn and the Kyn/Trp ratio appear to be directly associated with 2-AG, AEA, and PEA. Interestingly, these biomarkers also permitted to distinguish the population into two main clusters: one of individuals having mild/severe depressive symptoms and the other with an absence of depressive symptoms. Using parametric analysis, higher serum levels of 2-AG, Kyn, and the ratio Kyn/Trp and lower levels of Trp and 5-HT were found in individuals with mild/severe depressive symptoms than in those without depressive symptoms. While in asymptomatic people, PEA was directly associated to Trp, and OEA indirectly linked to 5-HT, in individuals with depressive symptoms, these correlations were lost, and instead, positive correlations between AEA and 2-AG, PEA and AEA, and PEA vs 2-AG and OEA concentrations were found. Conclusions Parametric and non-parametric analyses suggest a possible association between eCBs, tryptophan/kynurenine biomarkers, and severity of depression, confirming a likely interplay among inflammation, stress, and depression. The enhanced relationships among the biomarkers of the 2-AG and AEA pathways and related lipids seen in individuals with depressive symptoms, but not in asymptomatics, suggest an altered metabolism of the eCB system in depression.

Gene regulatory networks that act upstream of skeletal muscle fate determinants are distinct in different anatomical locations. Despite recent efforts, a clear understanding of the cascade of events underlying the emergence and maintenance of the stem cell pool in specific muscle groups remains unresolved and debated. Here, we invalidated Pitx2 with multiple Cre-driver mice prenatally, postnatally, and during lineage progression. We showed that this gene becomes progressively dispensable for specification and maintenance of the muscle stem (MuSC) cell pool in extraocular muscles (EOMs) despite being, together with Myf5, a major upstream regulator during early development. Moreover, constitutive inactivation of Pax7 postnatally led to a greater loss of MuSCs in the EOMs compared to the limb. Thus, we propose a relay between Pitx2, Myf5 and Pax7 for EOM stem cell maintenance. We demonstrate also that MuSCs in the EOMs adopt a quiescent state earlier that those in limb muscles and do not spontaneously proliferate in the adult, yet EOMs have a significantly higher content of Pax7+ MuSCs per area pre- and post-natally. Finally, while limb MuSCs proliferate in the mdx mouse model for Duchenne muscular dystrophy, significantly less MuSCs were present in the EOMs of the mdx mouse model compared to controls, and they were not proliferative. Overall, our study provides a comprehensive in vivo characterisation of MuSC heterogeneity along the body axis and brings further insights into the unusual sparing of EOMs during muscular dystrophy.

Objectives: In immunocompromised patients, asymptomatic Leishmania infection can reactivate, and evolve to severe disease. To date, no test is considered the gold standard for the identification of asymptomatic Leishmania infection. A combination of methods was employed to screen for Leishmania infection in patients undergoing kidney transplant (KT). Methods: We employed polymerase chain reaction for the detection of parasitic DNA in peripheral blood, Western blot to identify serum immunoglobulin G and whole blood assay to detect cytokines/chemokines after stimulation of whole blood with parasitic antigen. Results: One-hundred twenty patients residing in Italy were included in the study at the time of KT. Each patient that tested positive to at least one test was considered as Leishmania positive. Fifty out of 120 patients (42%) tested positive for one or more tests. The detection of specific cell-mediated response (32/111, 29%) was the most common marker of Leishmania infection, followed by a positive serology (24/120, 20%). Four patients (3%) harbored parasitic DNA in the blood. Conclusion: Our findings underline the high prevalence of asymptomatic Leishmania infection in patients undergoing KT in Italy, who are potentially at-risk for parasite reactivation and can benefit from an increased vigilance. Understanding the clinical relevance of these findings deserves further studies.

Hyperspectral imaging holds significant promise in remote sensing applications, particularly for land cover and land-use classification, thanks to its ability to capture rich spectral information. However, leveraging hyperspectral data for accurate segmentation poses critical challenges, including the curse of dimensionality and the scarcity of ground truth data, that hinder the accuracy and efficiency of machine learning approaches. This paper presents a holistic approach for adaptive optimized hyperspectral-based land cover and land-use segmentation using automated machine learning (AutoML). We address the challenges of high-dimensional hyperspectral data through a revamped machine learning pipeline, thus emphasizing feature engineering tailored to hyperspectral classification tasks. We propose a framework that dissects feature engineering into distinct steps, thus allowing for comprehensive model generation and optimization. This framework incorporates AutoML techniques to streamline model selection, hyperparameter tuning, and data versioning, thus ensuring robust and reliable segmentation results. Our empirical investigation demonstrates the efficacy of our approach in automating feature engineering and optimizing model performance, even without extensive ground truth data. By integrating automatic optimization strategies into the segmentation workflow, our approach offers a systematic, efficient, and scalable solution for hyperspectral-based land cover and land-use classification.