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The limitation of natural resources coupledwith growing demand for fertilizers to sustain increased crop productivity allyto meet world demand for food intensifies the search for greater efficiency infertilizer use. The objective of this study was to evaluate the response ofphosphorus fertilization on wheat plants, noting its influence on the agronomicand nutritional characteristics of wheat and chemical attributes soil. Theexperiment was conducted in a greenhouse with two sources of singlesuperphosphate (P - 18% P2O5 and top-phos - 22/28% P2O5)and P2O5 five doses (0, 50, 100, 150 and 200 kg ha- 1),2 dm3 vessel. For dryweight of shoot gave maximum point buildup doseof 151.25 kg ha-1 P2O5 and dryweight of roots point was the maximum dose of165 kg ha-1 P2O5 independent of fertilizerused. Theapplication of P2O5 levels with superphosphate fertilizerand top-phos increased levels of pH and available soil P and P content, K and Sin the leaf tissue. It is recommended both simplesuperphosphate with 18% P2O5 as the top-phos, considered with 28% P2O5,for phosphate fertilizer in wheat crop by selecting the fertilizer that providebest relation cost-benefit, in this case the superphosphate.

Alzheimer's disease (AD) is a complex neurodegenerative disorder significantly impairing cognitive faculties, memory, and physical abilities. To characterize the modulation of the gut microbiota in an in vivo AD model, we performed shotgun metagenomics sequencing on 3xTgAD mice at key time points (i.e., 2, 6, and 12 months) of AD progression. Fecal samples from both 3xTgAD and wild-type mice were collected, DNA extracted, and sequenced. Quantitative taxon abundance assessment using MetaPhlAn 4 ensured precise microbial community representation. The analysis focused on species-level genome bins (SGBs) including both known and unknown SGBs (kSGBs and uSGBs, respectively) and also comprised higher taxonomic categories such as family-level genome bins (FGBs), class-level genome bins (CGBs), and order-level genome bins (OGBs). Our bioinformatic results pinpointed the presence of extensive gut microbial diversity in AD mice and showed that the largest proportion of AD- and aging-associated microbiome changes in 3xTgAD mice concern SGBs that belong to the Bacteroidota and Firmicutes phyla, along with a large set of uncharacterized SGBs. Our findings emphasize the need for further advanced bioinformatic studies for accurate classification and functional analysis of these elusive microbial species in relation to their potential bridging role in the gut-brain axis and AD pathogenesis., (© 2024. The Author(s).)

Various human diseases are triggered by molecular alterations influencing the fine-tuned expression and activity of transcription factors, usually due to imbalances in targets including protein-coding genes and non-coding RNAs, such as microRNAs (miRNAs). The transcription factor EB (TFEB) modulates human cellular networks, overseeing lysosomal biogenesis and function, plasma-membrane trafficking, autophagic flux, and cell cycle progression. In endothelial cells (ECs), TFEB is essential for the maintenance of endothelial integrity and function, ensuring vascular health. However, the comprehensive regulatory network orchestrated by TFEB remains poorly understood. Here, we provide novel mechanistic insights into how TFEB regulates the transcriptional landscape in primary human umbilical vein ECs (HUVECs), using an integrated approach combining high-throughput experimental data with dedicated bioinformatics analysis. By analyzing HUVECs ectopically expressing TFEB using ChIP-seq and examining both polyadenylated mRNA and small RNA sequencing data from TFEB-silenced HUVECs, we have developed a bioinformatics pipeline mapping the different gene regulatory interactions driven by TFEB. We show that TFEB directly regulates multiple miRNAs, which in turn post-transcriptionally modulate a broad network of target genes, significantly expanding the repertoire of gene programs influenced by this transcription factor. These insights may have significant implications for vascular biology and the development of novel therapeutics for vascular disease.

Introduction: Human mesenchymal stem cells (hMSCs) sense and respond to biomechanical and biophysical stimuli, yet the involved signaling pathways are not fully identified. The clinical application of biophysical stimulation including pulsed electromagnetic field (PEMF) has gained momentum in musculoskeletal disorders and bone tissue engineering., Methodology: We herein aim to explore the role of PEMF stimulation in bone regeneration by developing trabecular bone-like tissues, and then, culturing them under bone-like mechanical stimulation in an automated perfusion bioreactor combined with a custom-made PEMF stimulator. After selecting the optimal cell seeding and culture conditions for inspecting the effects of PEMF on hMSCs, transcriptomic studies were performed on cells cultured under direct perfusion with and without PEMF stimulation., Results: We were able to identify a set of signaling pathways and upstream regulators associated with PEMF stimulation and to distinguish those linked to bone regeneration. Our findings suggest that PEMF induces the immune potential of hMSCs by activating and inhibiting various immune-related pathways, such as macrophage classical activation and MSP-RON signaling in macrophages, respectively, while promoting angiogenesis and osteogenesis, which mimics the dynamic interplay of biological processes during bone healing., Conclusions: Overall, the adopted bioreactor-based investigation platform can be used to investigate the impact of PEMF stimulation on bone regeneration., Competing Interests: Declaration of competing interest Lia Rimondini reports was provided by University of Eastern Piedmont’Amedeo Avogadro’ Department of Health Sciences. Lia Rimondini reports a relationship with University of Eastern Piedmont’Amedeo Avogadro’ Department of Health Sciences that includes:. Not applicable. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Human cytomegalovirus (HCMV) is an opportunistic pathogen causing severe diseases in immunosuppressed individuals. To replicate its double-stranded DNA genome, HCMV induces profound changes in cellular homeostasis that may resemble senescence. However, it remains to be determined whether HCMV-induced senescence contributes to organ-specific pathogenesis. Here, we show a direct cytopathic effect of HCMV on primary renal proximal tubular epithelial cells (RPTECs), a natural setting of HCMV disease. We find that RPTECs are fully permissive for HCMV replication, which endows them with an inflammatory gene signature resembling the senescence-associated secretory phenotype (SASP), as confirmed by the presence of the recently established SenMayo gene set, which is not observed in retina-derived epithelial (ARPE-19) cells. Although HCMV-induced senescence is not cell-type specific, as it can be observed in both RPTECs and human fibroblasts (HFFs), only infected RPTECs show downregulation of LAMINB1 and KI67 mRNAs, and enhanced secretion of IL-6 and IL-8, which are well-established hallmarks of senescence. Finally, HCMV-infected RPTECs have the ability to trigger a senescence/inflammatory loop in an IL-6-dependent manner, leading to the development of a similar senescence/inflammatory phenotype in neighboring uninfected cells. Overall, our findings raise the intriguing possibility that this unique inflammatory loop contributes to HCMV-related pathogenesis in the kidney., (© 2024. The Author(s).)

Among all cancers, colorectal cancer (CRC) is the 3rd most common and the 2nd leading cause of death worldwide. New therapeutic strategies are required to target cancer stem cells (CSCs), a subset of tumor cells highly resistant to present-day therapy and responsible for tumor relapse. CSCs display dynamic genetic and epigenetic alterations that allow quick adaptations to perturbations. Lysine-specific histone demethylase 1A (KDM1A also known as LSD1), a FAD-dependent H3K4me1/2 and H3K9me1/2 demethylase, was found to be upregulated in several tumors and associated with a poor prognosis due to its ability to maintain CSCs staminal features. Here, we explored the potential role of KDM1A targeting in CRC by characterizing the effect of KDM1A silencing in differentiated and CRC stem cells (CRC-SCs). In CRC samples, KDM1A overexpression was associated with a worse prognosis, confirming its role as an independent negative prognostic factor of CRC. Consistently, biological assays such as methylcellulose colony formation, invasion, and migration assays demonstrated a significantly decreased self-renewal potential, as well as migration and invasion potential upon KDM1A silencing. Our untargeted multi-omics approach (transcriptomic and proteomic) revealed the association of KDM1A silencing with CRC-SCs cytoskeletal and metabolism remodeling towards a differentiated phenotype, supporting the role of KDM1A in CRC cells stemness maintenance. Also, KDM1A silencing resulted in up-regulation of miR-506-3p, previously reported to play a tumor-suppressive role in CRC. Lastly, loss of KDM1A markedly reduced 53BP1 DNA repair foci, implying the involvement of KDM1A in the DNA damage response. Overall, our results indicate that KDM1A impacts CRC progression in several non-overlapping ways, and therefore it represents a promising epigenetic target to prevent tumor relapse., (© 2023. The Author(s).)

Disruption of bacterial quorum sensing (QS) is presented as a promising strategy to overcome clinically relevant and phytopathogenic bacteria. This work presents α-alkylidene δ-lactones as new chemical scaffolds that inhibit the biosynthesis of violacein in the biosensor strain Chromobacterium CV026. Three molecules displayed higher than 50% violacein reduction when tested at concentrations lower than 625 µM. The most active α-alkylidene δ-lactone inhibited the hydrolysis of chitin concomitantly with the inhibition of violacein production in CV026, suggesting the disruption of its QS machinery. Further, RT-qPCR and competition experiments showed this molecule to be a transcriptional inhibitor of the QS-regulated operon vioABCDE . Docking calculations suggested a good correlation between binding affinity energies and inhibition effects, with all molecules positioned within the CviR autoinducer-binding domain (AIBD). The most active lactone yielded the best binding affinity energy, most probably due to its unprecedented binding with the AIBD. Our results show α-alkylidene δ-lactones as promising chemical scaffolds for the development of new QS inhibitors affecting LuxR/LuxI-systems., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Study Question: Can chromosomal abnormalities beyond copy-number aneuploidies (i.e. ploidy level and microdeletions (MDs)) be detected using a preimplantation genetic testing (PGT) platform?, Summary Answer: The proposed integrated approach accurately assesses ploidy level and the most common pathogenic microdeletions causative of genomic disorders, expanding the clinical utility of PGT., What Is Known Already: Standard methodologies employed in preimplantation genetic testing for aneuploidy (PGT-A) identify chromosomal aneuploidies but cannot determine ploidy level nor the presence of recurrent pathogenic MDs responsible for genomic disorders. Transferring embryos carrying these abnormalities can result in miscarriage, molar pregnancy, and intellectual disabilities and developmental delay in offspring. The development of a testing strategy that integrates their assessment can resolve current limitations and add valuable information regarding the genetic constitution of embryos, which is not evaluated in PGT providing new level of clinical utility and valuable knowledge for further understanding of the genomic causes of implantation failure and early pregnancy loss. To the best of our knowledge, MDs have never been studied in preimplantation human embryos up to date., Study Design, Size, Duration: This is a retrospective cohort analysis including blastocyst biopsies collected between February 2018 and November 2021 at multiple collaborating IVF clinics from prospective parents of European ancestry below the age of 45, using autologous gametes and undergoing ICSI for all oocytes. Ploidy level determination was validated using 164 embryonic samples of known ploidy status (147 diploids, 9 triploids, and 8 haploids). Detection of nine common MD syndromes (-4p=Wolf-Hirschhorn, -8q=Langer-Giedion, -1p=1p36 deletion, -22q=DiGeorge, -5p=Cri-du-Chat, -15q=Prader-Willi/Angelman, -11q=Jacobsen, -17p=Smith-Magenis) was developed and tested using 28 positive controls and 97 negative controls. Later, the methodology was blindly applied in the analysis of: (i) 100 two pronuclei (2PN)-derived blastocysts that were previously defined as uniformly euploid by standard PGT-A; (ii) 99 euploid embryos whose transfer resulted in pregnancy loss., Participants/materials, Setting, Methods: The methodology is based on targeted next-generation sequencing of selected polymorphisms across the genome and enriched within critical regions of included MD syndromes. Sequencing data (i.e. allelic frequencies) were analyzed by a probabilistic model which estimated the likelihood of ploidy level and MD presence, accounting for both sequencing noise and population genetics patterns (i.e. linkage disequilibrium, LD, correlations) observed in 2504 whole-genome sequencing data from the 1000 Genome Project database. Analysis of phased parental haplotypes obtained by single-nucleotide polymorphism (SNP)-array genotyping was performed to confirm the presence of MD., Main Results and the Role of Chance: In the analytical validation phase, this strategy showed extremely high accuracy both in ploidy classification (100%, CI: 98.1-100%) and in the identification of six out of eight MDs (99.2%, CI: 98.5-99.8%). To improve MD detection based on loss of heterozygosity (LOH), common haploblocks were analyzed based on haplotype frequency and LOH occurrence in a reference population, thus developing two further mathematical models. As a result, chr1p36 and chr4p16.3 regions were excluded from MD identification due to their poor reliability, whilst a clinical workflow which incorporated parental DNA information was developed to enhance the identification of MDs. During the clinical application phase, one case of triploidy was detected among 2PN-derived blastocysts (i) and one pathogenic MD (-22q11.21) was retrospectively identified among the biopsy specimens of transferred embryos that resulted in miscarriage (ii). For the latter case, family-based analysis revealed the same MD in different sibling embryos (n = 2/5) from non-carrier parents, suggesting the presence of germline mosaicism in the female partner. When embryos are selected for transfer based on their genetic constitution, this strategy can identify embryos with ploidy abnormalities and/or MDs beyond aneuploidies, with an estimated incidence of 1.5% (n = 3/202, 95% CI: 0.5-4.5%) among euploid embryos., Limitations, Reasons for Caution: Epidemiological studies will be required to accurately assess the incidence of ploidy alterations and MDs in preimplantation embryos and particularly in euploid miscarriages. Despite the high accuracy of the assay developed, the use of parental DNA to support diagnostic calling can further increase the precision of the assay., Wider Implications of the Findings: This novel assay significantly expands the clinical utility of PGT-A by integrating the most common pathogenic MDs (both de novo and inherited ones) responsible for genomic disorders, which are usually evaluated at a later stage through invasive prenatal testing. From a basic research standpoint, this approach will help to elucidate fundamental biological and clinical questions related to the genetics of implantation failure and pregnancy loss of otherwise euploid embryos., Study Funding/competing Interest(s): No external funding was used for this study. S.C., M.F., F.C., P.Z., I.P., L.G., C.P., M.P., D.B., J.J.-A., D.B.-J., J.M.-V., and C.R. are employees of Igenomix and C.S. is the head of the scientific board of Igenomix. A.C. and L.P. are employees of JUNO GENETICS. Igenomix and JUNO GENETICS are companies providing reproductive genetic services., Trial Registration Number: N/A., (© The Author(s) 2023. Published by Oxford University Press on behalf of European Society of Human Reproduction and Embryology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Costimulatory molecules of the CD28 family play a crucial role in the activation of immune responses in T lymphocytes, complementing and modulating signals originating from the T-cell receptor (TCR) complex. Although distinct functional roles have been demonstrated for each family member, the specific signaling pathways differentiating ICOS- from CD28-mediated costimulation during early T-cell activation are poorly characterized. In the present study, we have performed RNA-Seq-based global transcriptome profiling of anti-CD3-treated naïve CD4 + T cells upon costimulation through either inducible costimulator (ICOS) or CD28, revealing a set of signaling pathways specifically associated with each signal. In particular, we show that CD3/ICOS costimulation plays a major role in pathways related to STAT3 function and osteoarthritis (OA), whereas the CD3/CD28 axis mainly regulates p38 MAPK signaling. Furthermore, we report the activation of distinct immunometabolic pathways, with CD3/ICOS costimulation preferentially targeting glycosaminoglycans (GAGs) and CD3/CD28 regulating mitochondrial respiratory chain and cholesterol biosynthesis. These data suggest that ICOS and CD28 costimulatory signals play distinct roles during the activation of naïve T cells by modulating distinct sets of immunological and immunometabolic genes., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Gigliotti, Boggio, Favero, Incarnato, Santoro, Oliviero, Rojo, Zucchelli, Persichetti, Baldanzi, Dianzani and Corà.)

The key role played by host-microbiota interactions on human health, disease onset and progression, and on host response to treatments has increasingly emerged in the latest decades. Indeed, dysbiosis has been associated to several human diseases such as obesity, diabetes, cancer and also neurodegenerative disease, such as Parkinson, Huntington and Alzheimer's disease (AD), although whether causative, consequence or merely an epiphenomenon is still under investigation. In the present study, we performed a metabologenomic analysis of stool samples from a mouse model of AD, the 3xTgAD. We found a significant change in the microbiota of AD mice compared to WT, with a longitudinal divergence of the F/B ratio, a parameter suggesting a gut dysbiosis. Moreover, AD mice showed a significant decrease of some amino acids, while data integration revealed a dysregulated production of desaminotyrosine (DAT) and dihydro-3-coumaric acid. Collectively, our data show a dysregulated gut microbiota associated to the onset and progression of AD, also indicating that a dysbiosis can occur prior to significant clinical signs, evidenced by early SCFA alterations, compatible with gut inflammation., Competing Interests: The authors have declared that no competing interests exist.

Bacterial endophytes were isolated from nodules of pea and fava bean. The strains were identified and characterized for plant beneficial activities (phosphate solubilization, synthesis of indole acetic acid and siderophores) and salt tolerance. Based on these data, four strains of Rahnella aquatilis and three strains of Serratia plymuthica were selected. To shed light on the mechanisms underlying salt tolerance, the proteome of the two most performant strains (Ra4 and Sp2) grown in the presence or not of salt was characterized. The number of proteins expressed by the endophytes was higher in the presence of salt. The modulated proteome consisted of 302 (100 up-regulated, 202 down-regulated) and 323 (206 up-regulated, 117 down-regulated) proteins in Ra4 and Sp2, respectively. Overall, proteins involved in abiotic stress responses were up-regulated, while those involved in metabolism and flagellum structure were down-regulated. The main up-regulated proteins in Sp2 were thiol: disulfide interchange protein DsbA, required for the sulfur binding formation in periplasmic proteins, while in Ra4 corresponded to the soluble fraction of ABC transporters, having a role in compatible solute uptake. Our results demonstrated a conserved response to salt stress in the two taxonomically related species.

Approximately 50% of colorectal cancer (CRC) patients still die from recurrence and metastatic disease, highlighting the need for novel therapeutic strategies. Drug repurposing is attracting increasing attention because, compared to traditional de novo drug discovery processes, it may reduce drug development periods and costs. Epidemiological and preclinical evidence support the antitumor activity of antipsychotic drugs. Herein, we dissect the mechanism of action of the typical antipsychotic spiperone in CRC. Spiperone can reduce the clonogenic potential of stem-like CRC cells (CRC-SCs) and induce cell cycle arrest and apoptosis, in both differentiated and CRC-SCs, at clinically relevant concentrations whose toxicity is negligible for non-neoplastic cells. Analysis of intracellular Ca 2+ kinetics upon spiperone treatment revealed a massive phospholipase C (PLC)-dependent endoplasmic reticulum (ER) Ca 2+ release, resulting in ER Ca 2+ homeostasis disruption. RNA sequencing revealed unfolded protein response (UPR) activation, ER stress, and induction of apoptosis, along with IRE1-dependent decay of mRNA (RIDD) activation. Lipidomic analysis showed a significant alteration of lipid profile and, in particular, of sphingolipids. Damage to the Golgi apparatus was also observed. Our data suggest that spiperone can represent an effective drug in the treatment of CRC, and that ER stress induction, along with lipid metabolism alteration, represents effective druggable pathways in CRC.

Known multiple sclerosis (MS) susceptibility variants can only explain half of the disease's estimated heritability, whereas low-frequency and rare variants may partly account for the missing heritability. Thus, here we sought to determine the occurrence of rare functional variants in a large Italian MS multiplex family with five affected members. For this purpose, we combined linkage analysis and next-generation sequencing (NGS)-based whole exome and whole genome sequencing (WES and WGS, respectively). The genetic burden attributable to known common MS variants was also assessed by weighted genetic risk score (wGRS). We found a significantly higher burden of common variants in the affected family members compared to that observed among sporadic MS patients and healthy controls (HCs). We also identified 34 genes containing at least one low-frequency functional variant shared among all affected family members, showing a significant enrichment in genes involved in specific biological processes-particularly mRNA transport-or neurodegenerative diseases. Altogether, our findings point to a possible pathogenic role of different low-frequency functional MS variants belonging to shared pathways. We propose that these rare variants, together with other known common MS variants, may account for the high number of affected family members within this MS multiplex family.

Study Question: What is the clinical validity and utility of preconception Expanded Carrier Screening (ECS) application on the management of prospective parents?, Summary Answer: The high detection rate of at-risk couples (ARCs) and the high proportion opting for IVF/preimplantation genetic testing (PGT) treatment demonstrate the clinical utility of ECS in the preconception space in IVF and general population., What Is Known Already: About 2-4% of couples are at risk of conceiving a child with an autosomal recessive or X-linked genetic disorder. In recent years, the increasing cost-effectiveness of genetic diagnostic techniques has allowed the creation of ECS panels for the simultaneous detection of multiple recessive disorders. Comprehensive preconception genetic screening holds the potential to significantly improve couple's genetic risk assessment and reproductive planning to avoid detectable inheritable genetic offspring., Study Design, Size, Duration: A total of 3877 individuals without a family history of genetic conditions were analyzed between January 2017 and January 2020. Of the enrolled individuals, 1212 were gamete donors and 2665 were patients planning on conceiving from both the IVF and the natural conception group. From the non-donor cohort, 1133 were analyzed as individual patients, while the remaining ones were analyzed as couples, for a total of 766 couples., Participants/materials, Setting, Methods: A focused ECS panel was developed following American College of Obstetrics and Gynecology ACOG-recommended criteria (prevalence, carrier rate, severity), including highly penetrant severe childhood conditions. Couples were defined at-risk when both partners carried an autosomal recessive pathogenic/likely pathogenic variant (PLP) on the same gene or when the woman was a carrier of an X-linked PLP variant. ARC detection rate defined the clinical validity of the ECS approach. Clinical utility was evaluated by monitoring ARCs reproductive decision making., Main Results and the Role of Chance: A total of 402 individuals (10.4%) showed PLP for at least one of the genes tested. Among the 766 couples tested, 173 showed one carrier partner (22.6%), whereas 20 couples (2.6%) were found to be at increased risk. Interestingly, one ARC was identified as a result of cascade testing in the extended family of an individual carrying a pathogenic variant on the Survival Of Motor Neuron 1SMN1 gene. Of the identified ARCs, 5 (0.7%) were at risk for cystic fibrosis, 5 (0.7%) for fragile X syndrome, 4 (0.5%) for spinal muscular atrophy, 4 (0.5%) for Beta-Thalassemia/Sickle Cell Anemia, 1 (0.1%) for Smith-Lemli-Opitz Syndrome and 1 (0.1%) for Duchenne/Becker Dystrophy. Fifteen ARCs were successfully followed up from both the IVF and the natural conception groups. All of these (15/15) modified their reproductive planning by undergoing ART with Preimplantation Genetic Testing for Monogenic disease and Aneuploidies (PGT-M and PGT-A). To date, 6/15 (40%) couples completed their PGT cycle with euploid/unaffected embryos achieving a pregnancy after embryo transfer and three of them have already had an unaffected baby., Limitations, Reasons for Caution: The use of a limited panel of core gene-disease pairs represents a limitation on the research perspective as it can underestimate the rate of detectable carriers and ARCs in this cohort of prospective parents. Expanding the scope of ECS to a larger panel of conditions is becoming increasingly feasible, thanks to a persistent technological evolution and progressive cataloging of gene-disease associations., Wider Implications of the Findings: These results highlight the potential clinical validity and utility of ECS in reducing the risk of a pregnancy affected by a detectable inheritable genetic condition. The steady reduction in the costs of genetic analyses enables the expansion of monogenic testing/screening applications at the preimplantation stage, thus, providing valid decisional support and reproductive autonomy to patients, particularly in the context of IVF., Study Funding/competing Interest(s): No external funding was used for this study. A.C., M.F., S.C., M.P., L.G., and C.P. are employees of Igenomix Italy. C.S. is the head of the scientific board of Igenomix., Trial Registration Number: N/A., (© The Author(s) 2021. Published by Oxford University Press on behalf of European Society of Human Reproduction and Embryology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Clonal hematopoiesis of indeterminate potential (CHIP) is suspected of being a risk factor for patients with cancer. This study aimed to assess the clinical consequences of CHIP in patients with lymphoma intended for high-dose chemotherapy and autologous stem-cell transplantation (ASCT) in a population-based setting. We identified 892 lymphoma patients who had undergone stem cell harvest at all transplant centers in Denmark. A total of 565 patients had an available harvest sample, which was analysed for CHIP by next-generation sequencing, and the median follow-up was 9.1 years. Of the patients who were intended for immediate ASCT, 25.5% (112/440) carried at least one CHIP mutation. In contrast to previous single-center studies CHIP was not associated with inferior overall survival (OS) in multivariate analyses. However, patients with mutations in genes of the DNA repair pathway (PPM1D, TP53, RAD21, BRCC3) had a significant inferior OS (HR after 1 year of follow-up 2.79, 95% confidence interval 1.71-4.56; p < 0.0001), which also was evident in multivariate analysis (p = 0.00067). These patients had also increased rates of therapy-related leukemia and admission to intensive care. Furthermore, in patients who did not undergo immediate ASCT, a significant inferior OS of individuals with DNA repair mutations was also identified (p = 0.003).

The dynamic and hierarchical structure of rivers, together with disruption of the natural river continuum by human activities, makes it difficult to identify and locate sources of nutrient pollution affecting receiving waters and observe its dispersion, thus impairing monitoring efforts. The identification of reliable indicators of anthropogenic nitrogen inputs in catchments is therefore key to achieving effective management of polluted rivers. We tested the capacity of N isotopic signatures (δ 15 N) of epilithon and snails to provide useful indications of organic and inorganic anthropogenic N inputs in three Mediterranean rivers differing in terms of surrounding land use and physicochemical conditions. We used a combined approach based on (i) analysis of nutrient concentrations in water, (ii) CORINE land cover classification and drainage patterns in catchments and (iii) isotopic analysis of river biota to verify whether isotopic variations were indicative of anthropic activities in the watershed, the associated alteration of water quality, and the consequent impact on snail abundance and diversity. Variation in the δ 15 N of epilithon within and between rivers reflected localised and diffuse N inputs from inorganic and organic sources. Negative epilithon δ 15 N values (<0‰) indicated inorganic pollution from agriculture. Values between 4‰ and 8‰ and those above 8‰ respectively indicated moderate organic pollution from urban areas, and high organic pollution, mostly from waste waters. The diversity and abundance of snails decreased with increasing water pollution. While their isotopic variations reflected between-river differences, they failed to indicate within-river variations in anthropogenic N inputs, since the proportion of epilithon in their diet varied along the rivers. Concluding, epilithon was a reliable indicator of anthropogenic N sources across a wide range of nutrient concentrations and anthropogenic inputs, and the proposed approach allowed us to determine the nature of nitrogen pollutants, their sources, location and dispersion along rivers embedded in complex human landscapes., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Clonal hematopoiesis (CH) of indeterminate potential (CHIP) is defined by mutations in myeloid cancer-associated genes with a variant allele frequency of at least 2%. Recent studies have suggested a possible genetic predisposition to CH. To further explore this phenomenon, we conducted a population-based study of 594 twins from 299 pairs aged 73 to 94 years, all with >20 years' follow-up. We sequenced DNA from peripheral blood with a customized 21-gene panel at a median coverage of 6179X. The casewise concordance rates for mutations were calculated to assess genetic predisposition. Mutations were identified in 214 (36%) of the twins. Whereas 20 twin pairs had mutations within the same genes, the exact same mutation was only observed in 2 twin pairs. No significant difference in casewise concordance between monozygotic and dizygotic twins was found for any specific gene, subgroup, or CHIP mutations overall, and no significant heritability could be detected. In pairs discordant for CHIP mutations, we tested if the affected twin died before the unaffected twin, as a direct measurement of the association of having CH when controlling for familial factors. A total of 127 twin pairs were discordant for carrying a mutation, and in 61 (48%) cases, the affected twin died first (P = .72). Overall, we did not find a genetic predisposition to CHIP mutations in this twin study. The previously described negative association of CHIP mutations on survival could not be confirmed in a direct comparison among twin pairs that were discordant for CHIP mutations., (© 2020 by The American Society of Hematology.)

2-methyl-5,6,7,8-tetrahydro-2H-chromen-4(3H)-one (called 6-oxo) is presented as a new AI-1 quorum sensing inhibitor for Vibrio harveyi. The development of a chemical process to afford traceable materials for new biological assays demands the development of analytical methods to ensure their purity and quality. This work describes the use of quantitative 1 H nuclear magnetic resonance (NMR) spectroscopy (qNMR) to assess the purity of a sample of 6-oxo (99.88%) and a sample of its major process impurity (E)-1-(2-hydroxycyclohex-2-en-1-yl)but-2-en-1-one (called HCB; 98.28%). To explore the scope of the use of qNMR to quantify the amount of low-content components in samples related to the chemical process for 6-oxo synthesis, this work also determined the amount of 6-oxo in two HCB samples: (a) the high-purity HCB sample described above and (b) a crude HCB sample collected during the chemical process. Despite the complexity of the crude sample, the amount of 6-oxo was readily assessed and could help to estimate the extent to which 6-oxo was already formed during the HCB synthesis. This information can help the understanding of how the process parameters can be modified to improve the performance of the whole process, by controlling the reaction mechanisms working at each step of this chemical process. In this context, our results reinforce qNMR as a complementary analytical tool for the quantification of the main component found in a sample, contributing to the standardization of reference materials and thus allowing the development of analytical methods for process control and traceability of the samples used for biological assays., (© 2019 John Wiley & Sons, Ltd.)

Identifying the earliest somatic changes in prostate cancer can give important insights into tumor evolution and aids in stratifying high- from low-risk disease. We integrated whole genome, transcriptome and methylome analysis of early-onset prostate cancers (diagnosis ≤55 years). Characterization across 292 prostate cancer genomes revealed age-related genomic alterations and a clock-like enzymatic-driven mutational process contributing to the earliest mutations in prostate cancer patients. Our integrative analysis identified four molecular subgroups, including a particularly aggressive subgroup with recurrent duplications associated with increased expression of ESRP1, which we validate in 12,000 tissue microarray tumors. Finally, we combined the patterns of molecular co-occurrence and risk-based subgroup information to deconvolve the molecular and clinical trajectories of prostate cancer from single patient samples., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Pigmented Bowen disease (pBD) is an uncommon variant of squamous cell carcinoma in situ. Sometimes it can show clinical and dermoscopic features that are seen in other pigmented lesions of the skin and mucosa, making the diagnosis difficult. We report six cases of pBD occurring on the anogenital area, and discuss the importance of dermoscopy for improving the diagnostic accuracy in pBD., (© 2018 British Association of Dermatologists.)

Background: De novo assembly of RNA-seq data allows the study of transcriptome in absence of a reference genome either if data is obtained from a single organism or from a mixed sample as in metatranscriptomics studies. Given the high number of sequences obtained from NGS approaches, a critical step in any analysis workflow is the assembly of reads to reconstruct transcripts thus reducing the complexity of the analysis. Despite many available tools show a good sensitivity, there is a high percentage of false positives due to the high number of assemblies considered and it is likely that the high frequency of false positive is underestimated by currently used benchmarks. The reconstruction of not existing transcripts may false the biological interpretation of results as - for example - may overestimate the identification of "novel" transcripts. Moreover, benchmarks performed are usually based on RNA-seq data from annotated genomes and assembled transcripts are compared to annotations and genomes to identify putative good and wrong reconstructions, but these tests alone may lead to accept a particular type of false positive as true, as better described below., Results: Here we present a novel methodology of de novo assembly, implemented in a software named STAble (Short-reads Transcriptome Assembler). The novel concept of this assembler is that the whole reads are used to determine possible alignments instead of using smaller k-mers, with the aim of reducing the number of chimeras produced. Furthermore, we applied a new set of benchmarks based on simulated data to better define the performance of assembly method and carefully identifying true reconstructions. STAble was also used to build a prototype workflow to analyse metatranscriptomics data in connection to a steady state metabolic modelling algorithm. This algorithm was used to produce high quality metabolic interpretations of small gene expression sets obtained from already published RNA-seq data that we assembled with STAble., Conclusions: The presented results, albeit preliminary, clearly suggest that with this approach is possible to identify informative reactions not directly revealed by raw transcriptomic data.

The first genomic scar-based homologous recombination deficiency (HRD) measures were produced using SNP arrays. As array-based technology has been largely replaced by next generation sequencing approaches, it has become important to develop algorithms that derive the same type of genomic scar scores from next generation sequencing (whole exome "WXS", whole genome "WGS") data. In order to perform this analysis, we introduce here the scarHRD R package and show that using this method the SNP array-based and next generation sequencing-based derivation of HRD scores show good correlation (Pearson correlation between 0.73 and 0.87 depending on the actual HRD measure) and that the NGS-based HRD scores distinguish similarly well between BRCA mutant and BRCA wild-type cases in a cohort of triple-negative breast cancer patients of the TCGA data set., Competing Interests: N.J.B., A.C.E., and Zo.S are listed as co-inventors on a patent on telomeric allelic imbalance, which is owned by Children’s Hospital Boston and licensed to Myriad Genetics. The remaining authors declare no competing interests.

This paper reports the development, modelling and application of a semi-random multicore fibre (MCF) design for adaptive multiphoton endoscopy. The MCF was constructed from 55 sub-units, each comprising 7 single mode cores, in a hexagonally close-packed lattice where each sub-unit had a random angular orientation. The resulting fibre had 385 single mode cores and was double-clad for proximal detection of multiphoton excited fluorescence. The random orientation of each sub-unit in the fibre reduces the symmetry of the positions of the cores in the MCF, reducing the intensity of higher diffracted orders away from the central focal spot formed at the distal tip of the fibre and increasing the maximum size of object that can be imaged. The performance of the MCF was demonstrated by imaging fluorescently labelled beads with both distal and proximal fluorescence detection and pollen grains with distal fluorescence detection. We estimate that the number of independent resolution elements in the final image - measured as the half-maximum area of the two-photon point spread function divided by the area imaged - to be ~3200.

Nuclear mutations are well known to drive tumor incidence, aggression and response to therapy. By contrast, the frequency and roles of mutations in the maternally inherited mitochondrial genome are poorly understood. Here we sequence the mitochondrial genomes of 384 localized prostate cancer patients, and identify a median of one mitochondrial single-nucleotide variant (mtSNV) per patient. Some of these mtSNVs occur in recurrent mutational hotspots and associate with aggressive disease. Younger patients have fewer mtSNVs than those who diagnosed at an older age. We demonstrate strong links between mitochondrial and nuclear mutational profiles, with co-occurrence between specific mutations. For example, certain control region mtSNVs co-occur with gain of the MYC oncogene, and these mutations are jointly associated with patient survival. These data demonstrate frequent mitochondrial mutation in prostate cancer, and suggest interplay between nuclear and mitochondrial mutational profiles in prostate cancer.In prostate cancer, the role of mutations in the maternally-inherited mitochondrial genome are not well known. Here, the authors demonstrate frequent, age-dependent mitochondrial mutation in prostate cancer. Strong links between mitochondrial and nuclear mutational profiles are associated with clinical aggressivity.

Neurofibromatosis type 1 (NF1) is a genetic disorder commonly associated with an increased risk for development of malignancy, including skin cancers. Herein we describe a case of invasive melanoma occurring in a patient with NF1 and discuss the association between these two diseases, highlighting the importance of comparative clinical and dermoscopic approaches in this category of patients in which the detection of melanoma can be difficult because of the presence of multiple skin tumors., Competing Interests: Competing interests: The authors have no conflicts of interest to disclose.

Embryonic poly(A)-binding protein (EPAB)-deficient mice are infertile due to defects in both the oocyte and the somatic cells of the ovary. Since EPAB is oocyte specific, the abnormalities in the somatic compartment of Epab−/− mice are likely due to factors inherent to the oocyte. Herein, we investigated whether oocyte–somatic communication is disrupted as a result of EPAB deficiency. We found that gap junctions are disrupted at the late preantral stage of folliculogenesis in Epab−/– mice and remain disrupted in cumulus-enclosed oocytes (COCs) from antral follicles. Consistent with the timing of gap junction dysfunction, F-actin staining of transzonal processes (TZPs) is lower in Epab−/− follicles at the late preantral stage and completely absent in Epab−/− COCs. Epab−/− oocytes express significantly lower levels of the junction protein E-cadherin, which is likely to be a contributing factor leading to premature TZP retraction. Overall, these results demonstrate that EPAB is important for oocyte–somatic communication by maintaining TZPs and gap junctions at the preantral stage of folliculogenesis.

Chromosomal instability (CIN) contributes to cancer evolution, intratumor heterogeneity, and drug resistance. CIN is driven by chromosome segregation errors and a tolerance phenotype that permits the propagation of aneuploid genomes. Through genomic analysis of colorectal cancers and cell lines, we find frequent loss of heterozygosity and mutations in BCL9L in aneuploid tumors. BCL9L deficiency promoted tolerance of chromosome missegregation events, propagation of aneuploidy, and genetic heterogeneity in xenograft models likely through modulation of Wnt signaling. We find that BCL9L dysfunction contributes to aneuploidy tolerance in both TP53-WT and mutant cells by reducing basal caspase-2 levels and preventing cleavage of MDM2 and BID. Efforts to exploit aneuploidy tolerance mechanisms and the BCL9L/caspase-2/BID axis may limit cancer diversity and evolution., (Copyright © 2017 The Francis Crick Institute. Published by Elsevier Inc. All rights reserved.)