Impacts of in-vitro zebularine treatment on genome-wide DNA methylation and transcriptomic profiles in Salix purpurea L.

Renewable energy resources such as biomass are crucial for a sustainable global society. Trees are a major source of lignocellulosic biomass, which can vary in response to different environmental factors owing to epigenetic regulation, such as DNA C-methylation. To investigate the effects of DNA methylation on plant development and wood formation, and its impacts on gene expression, with a focus on secondary cell wall (SCW)-associated genes, Salix purpurea plantlets were cloned from buds derived from a single hybrid tree for both treatment and control conditions. For the treatment condition, buds were exposed to 50 µM zebularine in vitro and a combined strategy of whole-genome bisulfite sequencing (WGBS) and RNA-seq was employed to examine the methylome and transcriptome profiles of different tissues collected at various time points under both conditions. Transcriptomic and methylome data revealed that most of the promoter and gene body demethylation had no marked effects on the expression profiles of genes. Nevertheless, gene expression tended to decrease with the increased methylation levels of genes with highly methylated promoters. Results indicated that demethylation is less evident in centromeric regions and sex chromosomes. Promoters of secondary cell wall-associated genes, such as 4-coumarate-CoA ligase-like and Rac-like GTP-binding protein RHO, were differentially methylated in the secondary xylem samples collected from two-month potted treated plants compared to control samples. Our results provide novel insights into DNA methylation and gene expression landscapes and a basis for investigating the epigenetic regulation of wood formation in S. purpurea as a model plant for bioenergy species.

Drought-induced molecular changes in crown of various barley phytohormone mutants.

One of the main signal transduction pathways that modulate plant growth and stress responses, including drought, is the action of phytohormones. Recent advances in omics approaches have facilitated the exploration of plant genomes. However, the molecular mechanisms underlying the response in the crown of barley, which plays an essential role in plant performance under stress conditions and regeneration after stress treatment, remain largely unclear. The objective of the present study was the elucidation of drought-induced molecular reactions in the crowns of different barley phytohormone mutants. We verified the hypothesis that defects of gibberellins, brassinosteroids, and strigolactones action affect the transcriptomic, proteomic, and hormonal response of barley crown to the transitory drought influencing plant development under stress. Moreover, we assumed that due to the strong connection between strigolactones and branching the hvdwarf14.d mutant, with dysfunctional receptor of strigolactones, manifests the most abundant alternations in crowns and phenotype under drought. Finally, we expected to identify components underlying the core response to drought which are independent of the genetic background. Large-scale analyses were conducted using gibberellins-biosynthesis, brassinosteroids-signaling, and strigolactones-signaling mutants, as well as reference genotypes. Detailed phenotypic evaluation was also conducted. The obtained results clearly demonstrated that hormonal disorders caused by mutations in the HvGA20ox2, HvBRI1, and HvD14 genes affected the multifaceted reaction of crowns to drought, although the expression of these genes was not induced by stress. The study further detected not only genes and proteins that were involved in the drought response and reacted specifically in mutants compared to the reaction of reference genotypes and vice versa, but also the candidates that may underlie the genotype-universal stress response. Furthermore, candidate genes involved in phytohormonal interactions during the drought response were identified. We also found that the interplay between hormones, especially gibberellins and auxins, as well as strigolactones and cytokinins may be associated with the regulation of branching in crowns exposed to drought. Overall, the present study provides novel insights into the molecular drought-induced responses that occur in barley crowns.

Incidence and Risk Factors of Cholestasis in Newborns with Hemolytic Disease-A Case-Control Study.

Background/Objectives: One of the rare causes of cholestasis may be hemolytic disease of the fetus and newborn (HDFN). Methods: We retrospectively analyzed 88 medical records of HDFN newborns with cholestasis and 186 records of children with HDFN without cholestasis and conducted an observational, case-control, retrospective study. Results: Factors influencing the risk of cholestasis were lower gestational age at birth (36.83 ± 1.9 vs. 37.57 ± 1.8, p = 0.002), Rh or Kidd HDFN (80.7% vs. 53.2%), and the need for intrauterine transfusion (27.3 vs. 11.8%). The subjects had lower hemoglobin concentrations at birth (14.01 ± 3.8 vs. 16.39 ± 2.8 g/dL) and during whole hospital stay, higher cord blood total bilirubin concentration (4.26 ± 1.8 vs. 2.39 ± 1.4 mg/dL), higher maximum bilirubin concentration (15.27 ± 5.8 vs. 10.24 ± 3.4 mg/dL), and more frequent liver ultrasound abnormalities (19.9 vs. 6.3%). They also required more extended hospitalization due to higher rates of postnatal blood transfusion (33 vs. 3.8%), more frequent need for exchange transfusion (8.8% vs. 2.2%), more extended time and higher risk of phototherapy (94.3 vs. 59.1%), and higher usage of immunoglobulins (55.7 vs. 8.1%), parenteral nutrition (45.5 vs. 12.9%), and antibiotics (14.8 vs. 4.8%). Conclusions: The risk factors for cholestasis in children with HDFN are lower gestational age at delivery, Rh and Kidd serological type of HDFN, and the need for intrauterine transfusions.

Severe Cholestasis in Neonates with Hemolytic Disease of the Fetus and Newborn-A Case Report.

Hemolytic disease of the fetus and newborn (HDFN) may cause severe cholestasis with direct bilirubin concentrations reaching up to 50 times the upper limit of normal. This case report describes twins whose highest direct bilirubin concentrations were 32.2 mg/dL and 50.2 mg/dL, with no significant signs of hepatic impairment. The index pregnancy was complicated by Rhesus factor immunization with anti-D antibodies present in maternal serum, which caused fetal anemia requiring intrauterine blood transfusions. Complementary tests demonstrated Rhesus D alloimmunization as the sole cause of cholestasis. To the best of our knowledge, this is the first study to describe such elevated direct bilirubin concentrations caused by HDFN.

The impact of multiple abiotic stresses on ns-LTP2.8 gene transcript and ns-LTP2.8 protein accumulation in germinating barley (Hordeum vulgare L.) embryos.

Abiotic stresses occur more often in combination than alone under regular field conditions limiting in more severe way crop production. Stress recognition in plants primarily occurs in the plasma membrane, modification of which is necessary to maintain homeostasis in response to it. It is known that lipid transport proteins (ns-LTPs) participate in modification of the lipidome of cell membranes. Representative of this group, ns-LTP2.8, may be involved in the reaction to abiotic stress of germinating barley plants by mediating the intracellular transport of hydrophobic particles, such as lipids, helping to maintain homeostasis. The ns-LTP2.8 protein was selected for analysis due to its ability to transport not only linear hydrophobic molecules but also compounds with a more complex spatial structure. Moreover, ns-LTP2.8 has been qualified as a member of pathogenesis-related proteins, which makes it particularly important in relation to its high allergenic potential. This paper demonstrates for the first time the influence of various abiotic stresses acting separately as well as in their combinations on the change in the ns-LTP2.8 transcript, ns-LTP2.8 protein and total soluble protein content in the embryonal axes of germinating spring barley genotypes with different ns-LTP2.8 allelic forms and stress tolerance. Tissue localization of ns-LTP2.8 transcript as well as ns-LTP2.8 protein were also examined. Although the impact of abiotic stresses on the regulation of gene transcription and translation processes remains not fully recognized, in this work we managed to demonstrate different impact on applied stresses on the fundamental cellular processes in very little studied tissue of the embryonal axis of barley.

Temperature drift during environmental radiation monitoring and its unfolding.

The aim of the presented paper is to examine the temperature drift and its unfolding for an environmental monitor equipped with a LaBr3(Ce) detector. It is known that temperature could influence energy, shape, and efficiency calibration. Consequently, when ambient temperature changes, the full energy absorption peak moves in the resulting spectrum. Research consists of experimental and analytical parts. During research in the climatic chamber, the dependence on temperature of energy, shape, and efficiency calibration was completed. The numerical method allows generating gamma spectra for theoretical so-called binary temperature variation.

Single molecule real-time sequencing data sets of Hypericum perforatum L. plantlets and cell suspension cultures.

Hypericum is a large genus that includes more than 500 species of pharmacological, ecological and conservation value. Although latest advances in sequencing technologies were extremely exploited for generating and assembling genomes of many living organisms, annotated whole genome sequence data is not publicly available for any of the Hypericum species so far. Bioavailability of secondary metabolites varies for different tissues and the data derived from different cultures will be a valuable tool for comparative studies. Here, we report the single molecule real-time sequencing (SMRT) data sets of Hypericum perforatum L. plantlets and cell suspension cultures for the first time. Sequencing data from cell suspension cultures yielded more than 33,000 high-quality transcripts from 20 Gb of raw data, while more than 55,000 high-quality transcripts were obtained from 35 Gb of raw data from plantlets. This dataset is a valuable tool for comparative transcriptomic analysis and will help to understand the unknown biosynthetic pathways of high medicinal value in the Hypericum genus.

Liver Dysfunction with Severe Cholestasis and Coagulation Disorders in the Course of Hemolytic Disease of the Newborn Requiring Chelation Therapy-A Case Report and Review of the Literature.

Severe hemolytic disease of the fetus and newborn (HDFN) requiring intrauterine transfusions (IUTs) may cause iron accumulation, resulting in liver damage, which may lead to cholestasis and coagulation disorders. In this article, we reported a case of a female neonate who underwent chelation therapy with a positive outcome, and we reviewed the English and Polish literature on chelation therapy in HDFN available in PubMed. The patient with maximum ferritin concentration above 33,511.2 ng/mL developed liver dysfunction with coagulation disorders requiring multiple transfusions of fresh frozen plasma (FFP), Octaplex® and cryoprecipitate, and hypoalbuminemia treated with numerous albumin infusions. Furthermore, severe cholestasis was observed with direct bilirubin levels up to 33.14 mg/dL. Additionally, the child developed transient myelosuppression with neutropenia, thrombocytopenia, and low reticulocyte count due to several blood transfusions. The differential diagnosis tests were conducted to rule out any causes of hepatic failure other than hemolytic disease of the newborn. This case proves that adequate treatment of severe HDFN with anemia requiring IUT and hepatic failure can lead to positive outcomes with no long-term consequences.

Evaluation of internal exposure of nuclear medicine staff working with radioiodine in Poland.

The iodine-131 (I-131) content in the thyroid of staff members working with this radionuclides has been measured with about 500 employees in about 25 hospital's departments of nuclear medicine performing therapy and diagnosis of thyroid disease in Poland. The measurements were performed with portable detection unit for in situ measurements of radioiodine. This is consist with scintillation detector sodium iodine activated by thallium (NaI(Tl)) - battery-powered and portable tube base Multichannel Analyzer Canberra UniSPEC. Based on direct measurements of the iodine content, the effective dose equivalent for workers due to inhalation of I-131 was estimated. All individuals actively working with iodine show measurable amounts of this isotopes in their thyroids. The average measured activity in the thyroid of the nuclear medicine staff was found to be equal at average 550 Bq within the range 70 Bq-2.5 kBq. There is no apparent correlation between the measured I-131 levels and risk categories. Nevertheless the technical and nuclear medicine staff show higher I-131 thyroid level comparing to hospital services staff. Calculated maximum committed effective dose for particular exposed person is <10% of 20 mSv/year. Int J Occup Med Environ Health. 2023;36(5):587-95.

Basilar tip fenestration giving rise to Percheron's and mesencephalic arteries.

The basilar bifurcation region is a common site for intracranial aneurysms, as well as it gives rise to a group of perforating arteries that supply the mesencephalon and the thalamus. Complex vascular microanatomy poses a diagnostic and therapeutic challenge for neurosurgeons, neuroradiologists and neurologists. In this paper, we present a previously unreported case of basilar tip fenestration that gave rise to five perforating arteries: the artery of Percheron and four mesencephalic arteries. Due to invaluable clinical significance, the possibility of such a variant must be considered during performing various neurovascular procedures, since e.g., embolization of the fenestration misdiagnosed as an aneurysm would inevitably lead to severe neurological complications (consciousness disturbances, quadriplegia, and sensory loss). Comprehensive knowledge of the neuroanatomy and neuroembryology is crucial to safe execution of intracranial interventions.

Point-of-Care Verification of Blood Culture Volume in Neonates: A Feasibility Trial.

BACKGROUND: Blood cultures remain the gold standard for the diagnosis of sepsis. However, volumes of blood submitted for cultures often do not match the recommended values. We propose a simple intervention aimed to verify the volume of blood sampled using a scale. This study was undertaken in preparation for a future, multicenter, pre- and post-intervention trial. Our primary objective was to test the feasibility (uptake and retention) of this future intervention. MATERIALS AND METHODS: This study was conducted at a neonatal department in Warsaw, Poland, over a period of eight months (May to December 2020). Before starting the study, we undertook an educational intervention focused on obtaining adequate blood volumes for culture. The culture bottles that were weighed in advance were distributed in all blood collection areas. Blood volume was verified by weighing the bottle immediately after blood inoculation. The calculated value was communicated to the collecting clinician and recorded. The primary outcome measure was the percentage of blood culture submissions for which the blood volume inoculated into the bottles was determined by weighing. RESULTS: During the study period, 244 blood samples were collected for culture, out of which 205 samples were weighed (84.0%, CI95 [78.8% to 88.4%]). This high proportion remained stable throughout the study period. We have not observed any adverse events related to the study. CONCLUSIONS: The point-of-care verification of blood culture volume using a scale was feasible to implement. Since we have met our pre-established criterion for success, a future, definitive trial is likely to proceed.

Global Proteome Profiling Revealed the Adaptive Reprogramming of Barley Flag Leaf to Drought and Elevated Temperature.

Plants, as sessile organisms, have developed sophisticated mechanisms to survive in changing environments. Recent advances in omics approaches have facilitated the exploration of plant genomes; however, the molecular mechanisms underlying the responses of barley and other cereals to multiple abiotic stresses remain largely unclear. Exposure to stress stimuli affects many proteins with regulatory and protective functions. In the present study, we employed liquid chromatography coupled with high-resolution mass spectrometry to identify stress-responsive proteins on the genome-wide scale of barley flag leaves exposed to drought, heat, or both. Profound alterations in the proteome of genotypes with different flag leaf sizes were found. The role of stress-inducible proteins was discussed and candidates underlying the universal stress response were proposed, including dehydrins. Moreover, the putative functions of several unknown proteins that can mediate responses to stress stimuli were explored using Pfam annotation, including calmodulin-like proteins. Finally, the confrontation of protein and mRNA abundances was performed. A correlation network between transcripts and proteins performance revealed several components of the stress-adaptive pathways in barley flag leaf. Taking the findings together, promising candidates for improving the tolerance of barley and other cereals to multivariate stresses were uncovered. The presented proteomic landscape and its relationship to transcriptomic remodeling provide novel insights for understanding the molecular responses of plants to environmental cues.

The Effects of Growth Modification on Pollen Development in Spring Barley (Hordeum vulgare L.) Genotypes with Contrasting Drought Tolerance.

Drought stress inducing pollen sterility can reduce crop yield worldwide. The regulatory crosstalk associated with the effects of drought on pollen formation at the cellular level has not been explored in detail so far. In this study, we performed morphological and cytoembryological analysis of anther perturbations and examined pollen development in two spring barley genotypes that differ in earliness and drought tolerance. The Syrian breeding line CamB (drought-tolerant) and the European cultivar Lubuski (drought-sensitive) were used as experimental materials to analyze the drought-induced changes in yield performance, chlorophyll fluorescence kinetics, the pollen grain micromorphology and ultrastructure during critical stages of plant development. In addition, fluctuations in HvGAMYB expression were studied, as this transcription factor is closely associated with the development of the anther. In the experiments, the studied plants were affected by drought, as was confirmed by the analyses of yield performance and chlorophyll fluorescence kinetics. However, contrary to our expectations, the pollen development of plants grown under specific conditions was not severely affected. The results also suggest that growth modification, as well as the perturbation in light distribution, can affect the HvGAMYB expression. This study demonstrated that the duration of the vegetation period can influence plant drought responses and, as a consequence, the processes associated with pollen development as every growth modification changes the dynamics of drought effects as well as the duration of plant exposition to drought.

Genome-wide association mapping in elite winter wheat breeding for yield improvement.

Increased grain yield (GY) is the primary breeding target of wheat breeders. We performed the genome-wide association study (GWAS) on 168 elite winter wheat lines from an ongoing breeding program to identify the main determinants of grain yield. Sequencing of Diversity Array Technology fragments (DArTseq) resulted in 19,350 single-nucleotide polymorphism (SNP) and presence-absence variation (PAV) markers. We identified 15 main genomic regions located in ten wheat chromosomes (1B, 2B, 2D, 3A, 3D, 5A, 5B, 6A, 6B, and 7B) that explained from 7.9 to 20.3% of the variation in grain yield and 13.3% of the yield stability. Loci identified in the reduced genepool are important for wheat improvement using marker-assisted selection. We found marker-trait associations between three genes involved in starch biosynthesis and grain yield. Two starch synthase genes (TraesCS2B03G1238800 and TraesCS2D03G1048800) and a sucrose synthase gene (TraesCS3D03G0024300) were found in regions of QGy.rut-2B.2, QGy.rut-2D.1, and QGy.rut-3D, respectively. These loci and other significantly associated SNP markers found in this study can be used for pyramiding favorable alleles in high-yielding varieties or to improve the accuracy of prediction in genomic selection.

The effects of heading time on yield performance and HvGAMYB expression in spring barley subjected to drought.

In the lifetime of a plant, flowering is not only an essential part of the reproductive process but also a critical developmental stage that can be vulnerable to environmental stresses. To ensure survival during drought, plants accelerate the flowering process, and this response is known as "drought escape." HvGAMYB-transcription factor associated, among others, with flowering process and anther development in barley-has also an important role in developmental modification and yield performance in plants subjected to stressed conditions. Due to the fact that information about the mechanisms associated both with the flowering acceleration and the anther or pollen disruption is limited, the exploration of the potential HvGAMYB role in flower development may shed light on pollen and spike morphology formations in plants grown under unfavorable water conditions. The aim of this study was to characterize differences in responses to drought among early- and late-heading barley genotypes. These two subgroups of plants-differentiated in terms of phenology-were analyzed, and traits linked to plant phenotype, physiology, and yield were investigated. In our study, the drought stress reactions of two barley subgroups showed a wide range of diversity in terms of yield performance, anther morphology, chlorophyll fluorescence kinetics, and pollen viability. The studied plants exhibited different yield performances under control and drought conditions. Moreover, the random distribution of genotypes on the biplot showing variability of OJIP parameters in the second developmental point of our investigation revealed that prolonged drought stress caused that among early- and late-heading plants, the studied genotypes exhibited different responses to applied stress conditions. The results of this study also showed that the HvGAMYB expression level was correlated positively with traits associated with lateral spike morphology in the second developmental point of this investigation, which showed that this association occurred only under prolonged drought and highlighted the drought stress duration effect on the HvGAMYB expression level.

Genome-wide association study of agronomical and root-related traits in spring barley collection grown under field conditions.

The root system is a key component for plant survival and productivity. In particular, under stress conditions, developing plants with a better root architecture can ensure productivity. The objectives of this study were to investigate the phenotypic variation of selected root- and yield-related traits in a diverse panel of spring barley genotypes. By performing a genome-wide association study (GWAS), we identified several associations underlying the variations occurring in root- and yield-related traits in response to natural variations in soil moisture. Here, we report the results of the GWAS based on both individual single-nucleotide polymorphism markers and linkage disequilibrium (LD) blocks of markers for 11 phenotypic traits related to plant morphology, grain quality, and root system in a group of spring barley accessions grown under field conditions. We also evaluated the root structure of these accessions by using a nondestructive method based on electrical capacitance. The results showed the importance of two LD-based blocks on chromosomes 2H and 7H in the expression of root architecture and yield-related traits. Our results revealed the importance of the region on the short arm of chromosome 2H in the expression of root- and yield-related traits. This study emphasized the pleiotropic effect of this region with respect to heading time and other important agronomic traits, including root architecture. Furthermore, this investigation provides new insights into the roles played by root traits in the yield performance of barley plants grown under natural conditions with daily variations in soil moisture content.

The eINTACT system dissects bacterial exploitation of plant osmosignalling to enhance virulence.

Bacteria inject effector proteins into host cells to manipulate cellular processes that promote disease. Since bacteria deliver minuscule amounts of effectors only into targeted host cells, it is technically challenging to capture effector-dependent cellular changes from bulk-infected host tissues. Here, we report a new technique called effector-inducible isolation of nuclei tagged in specific cell types (eINTACT), which facilitates affinity-based purification of nuclei from Arabidopsis plant cells that have received Xanthomonas bacterial effectors. Analysis of purified nuclei reveals that the Xanthomonas effector XopD manipulates the expression of Arabidopsis abscisic acid signalling-related genes and activates OSCA1.1, a gene encoding a calcium-permeable channel required for stomatal closure in response to osmotic stress. The loss of OSCA1.1 causes leaf wilting and reduced bacterial growth in infected leaves, suggesting that OSCA1.1 promotes host susceptibility. eINTACT allows us to uncover that XopD exploits host OSCA1.1/abscisic acid osmosignalling-mediated stomatal closure to create a humid habitat that favours bacterial growth and opens up a new avenue for accurately elucidating functions of effectors from numerous gram-negative plant bacteria in native infection contexts.

Genome-Wide Association Analysis for Hybrid Breeding in Wheat.

Disclosure of markers that are significantly associated with plant traits can help develop new varieties with desirable properties. This study determined the genome-wide associations based on DArTseq markers for six agronomic traits assessed in eight environments for wheat. Moreover, the association study for heterosis and analysis of the effects of markers grouped by linkage disequilibrium were performed based on mean values over all experiments. All results were validated using data from post-registration trials. GWAS revealed 1273 single nucleotide polymorphisms with biologically significant effects. Most polymorphisms were predicted to be modifiers of protein translation, with only two having a more pronounced effect. Markers significantly associated with the considered set of features were clustered within chromosomes based on linkage disequilibrium in 327 LD blocks. A GWAS for heterosis revealed 1261 markers with significant effects.

Effects of Early versus Standard Central Line Removal on the Growth of Preterm Infants with Very Low Birth Weight: A Non-Inferiority, Randomized Clinical Trial.

Very preterm infants are usually supported by parenteral nutrition delivered through central lines (CLs) while progressing with enteral intake, although the optimal time point for their removal is unclear. This study evaluated the impact of the CL discontinuation time on the short-term growth outcomes of preterm infants. A non-inferiority, parallel-group, randomized controlled trial was conducted in four neonatal intensive care units in Poland. Preterm infants with very low birth weight (VLBW) without congenital abnormalities were eligible. Patients were allocated to discontinue central access at an enteral feeding volume of 100 mL/kg/day (intervention group) or 140 mL/kg/day (control group). The study's primary outcome was weight at 36 weeks' postmenstrual age, with a non-inferiority margin of -210 g. Overall, 211 patients were allocated to the intervention or control groups between January 2019 and February 2021, of which 101 and 100 were eligible for intention-to-treat analysis, respectively. The mean weight was 2232 g and 2200 g at 36 weeks' postmenstrual age in the intervention and control groups, respectively. The mean between-group difference was 32 g (95% confidence interval, -68 to 132; p = 0.531), which did not cross the specified margin of non-inferiority. No intervention-related adverse events were observed. Early CL removal was non-inferior to the standard type for short-term growth outcomes in VLBW infants.