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This study is done to estimаte in‐hоsрitаl mоrtаlity in раtients with severe асute resрirаtоry syndrоme соrоnаvirus 2 (SАRS‐СоV‐2) strаtified by Vitamin‐D (Vit‐D) levels. Раtients were strаtified ассоrding tо by serum 25‐hydroxy‐vitamin D (25(OH)Vit‐D) levels intо twо grоuрs, that is, 25(OH)Vit‐D less thаn 40 nmol/L аnd 25(OH)Vit‐D greаter thаn 40 nmol/L. А tоtаl оf 231 раtients were inсluded. Оf these, 120 (50.2%) оf the раtients hаd 25(OH)Vit‐D levels greаter thаn 40 nmol/L. The meаn аge wаs 49 ± 17 yeаrs, аnd 67% оf the раtients were mаles. The mediаn length оf оverаll hоsрitаl stаy wаs 18 [6; 53] dаys. The remаining 119 (49.8%) раtients hаd а 25(OH)Vit‐D less thаn 40 nmol/L. Vitamin D levels were seen as deficient in 63% of patients, insufficient in 25% and normal in 12%. Оverаll mоrtаlity wаs 17 раtients (7.1%) but statistically not signifiсаnt among the grоuрs (p = 0.986). The Kарlаn–Meier survivаl аnаlysis shоwed no significance based on an alpha of 0.05, LL = 0.36, df = 1, p = 0.548, indicating Vitamin_D_Levels was not able to adequately predict the hazard of Mortality. In this study, serum 25(OH)Vit‐D levels were found have no significance in terms of predicting the in‐hоsрitаl mortality in раtients with SАRS‐СоV‐2., Highlights This is one among a very few studies which show serum Vitamin‐D levels have no role in predicting the in‐hospital mortality in раtients with SARS‐CoV‐2.

Аbstrасt Introduction This study aims to investigate in‐hоsрitаl mоrtаlity in severe асute resрirаtоry syndrоme соrоnаvirus 2 раtients strаtified by serum ferritin levels. Methods Patients were stratified based on ferritin levels (ferritin levels ≤ 1000 or >1000). Results Approximately 89% (118) of the patients with ferritin levels > 1000 had pneumonia, and 51% (67) had hypertension. Fever (97, 73.5%) and shortness of breath (80, 61%) were two major symptoms among the patients in this group. Logistic regression analysis indicated that ferritin level (odds ratio [OR] = 0.36, 95% confidence interval [CI] = 0.21–0.62; p 1000. Conclusion In this study, higher levels of serum ferritin were found to be an independent predictor of in‐hоsрitаl mоrtаlity.

In this work, we report a new approach for detecting SARS-CoV-2 RBD protein (RBD) using the surface-enhanced Raman spectroscopy (SERS) technique. The optical enhancement was obtained thanks to the preparation of nanostructured Ag/Au substrates. Fabricated Au/Ag nanostructures were used in the SERS experiment for RBD protein detection. SERS substrates show higher capabilities and sensitivity to detect RBD protein in a short time (3 s) and with very low power. We were able to push the detection limit of proteins to a single protein detection level of 1 pM. The latter is equivalent to 1 fM as a detection limit of viruses. Additionally, we have shown that the SERS technique was useful to figure out the presence of RBD protein on antibody functionalized substrates. In this case, the SERS detection was based on protein-antibody recognition, which led to shifts in the Raman peaks and allowed signal discrimination between RBD and other targets such as Bovine serum albumin (BSA) protein. A perfect agreement between a 3D simulated model based on finite element method and experiment was reported confirming the SERS frequency shift potential for trace proteins detection. Our results could open the way to develop a new prototype based on SERS sensitivity and selectivity for rapid detection at a very low concentration of virus and even at a single protein level.

This study is to estimate in‐hospital mortality in severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) patients stratified by hemoglobin (Hb) level. Patients were stratified according to hemoglobin level into two groups, that is, Hb 100 g/L. A total of 6931 patients were included. Of these, 6377 (92%) patients had hemoglobin levels >100 g/L. The mean age was 44 ± 17 years, and 66% of the patients were males. The median length of overall hospital stay was 13 days [2; 31]. The remaining 554 (8%) patients had a hemoglobin level 100 g/L (52, 0.82%). Risk factors associated with increased mortality were determined by multivariate analysis. The Kaplan‐Meier survival analysis showed hemoglobin as a predictor of mortality. Cox proportional hazards regression coefficients for hemoglobin for the HB ≤ 100 category of hemoglobin were significant, B = 2.79, SE = 0.17, and HR = 16.34, p

This research demonstrates an economical and efficient reduction of carbon foot print. Tapioca powder as a source of organic carbon was utilized in the synthesis of carbon dots through optimization of the synthesis parameters such as temperature, dosage and time. Photoluminescent quantum yield (PLQY) was obtained under the visible region of 340 nm at 34.9%, which was achieved without dopants such as sulfur and nitrogen that are popularly used to increase the value of photoluminescence. The characterization of carbon dots such as FTIR and HrTem, were carried out for the analysis of functional groups, particle sizes (1‐5 nm) and shapes (quasi‐spherical). The high carbon‐carbon bonds and oxygen groups detected in FTIR analysis validates the basis of fluorescence of carbon dots and also presence of hydroxyl (OH), carboxylic acids (COO), and other vital functional groups (C=O, C−O‐C, C−H). These characteristics makes tapioca based carbon dots suitable for application in the fields of environmental studies including sensitive detection and absorbance of pollutants in water and bio imaging in health sciences.

Epstein-Barr virus (EBV), is an oncogenic gamma-herpesvirus, which is associated with malignant diseases of B cells, T cells, and epithelial cells. EB viruses have large DNA genomes of more than 170 kb that are difficult to clone and manipulate. Here we describe 2 different approaches for cloning whole EBV genomes of diverse strains for reverse genetics studies. The first approach used CRISPR/Cas9-mediated cloning of the entire EBV genome into a bacterial artificial chromosome (BAC) vector using homologous recombination in B cells. This method allowed the cloning of the type 2 EBV strain Jijoye for the first time, but the BAC-clones are unstable. This strategy is being modified by recoding the homology regions to make the clones more stable. The second approach involves transformation-associated recombination (TAR) cloning of EBV fragments and their assembly in yeast, which will allow for mixing and matching DNA regions from different EBV strains for functional studies. This approach is based on TAR cloning of the EBV genome as 10 overlapping fragments, which average 17 kilobases long, using the natural homologous recombination processes of the yeast. Subsequent assembly of all the overlapping fragments is undertaken in yeast or by Gibson assembly to reconstitute the infectious EBV clone. Two fragments from EBV strains B95-8 and AG876 were captured and isolated successfully, but at low efficiency. We are currently improving the TAR cloning efficiency by increasing the size of the capture homology regions to approximately 500 bp coupled with CRISPR/Cas-9-mediated fragmentation of the EBV genome.

Plant response to salt stress and the mechanism of salt tolerance have received major focus by plant biology researchers. Biotic stresses cause extensive losses in agricultural production globally, but abiotic stress causes significant increase in the methylglyoxal (MG) level of GlyoxalaseI (Gly I). Identification of salt-tolerant genes when characterizing their phenotypes will help to identify novel genes using polymerase chain reaction (PCR) to amplify the DNA coding region for glyoxalase I. This method is specific, requiring only genomic DNA and two pairs of PCR primers, and involving two successive PCR reactions. This method was used rapidly and easily identified glyoxalase I sequences as salt-tolerant genes from Jojoba (Simmondsia chinensis (Link) Schneider). In the present study, the glyoxalase I gene was isolated, amplified by PCR using gene-specific primers and sequenced from the jojoba plant, then compared with other glyoxalase I sequences in other plants and glyoxalase I genes like in Brassica napus, ID: KT720495.1, Brassica juncea ID: Y13239.1, Arachis hypogaea, ID: DQ989209.2, and Arabidopsis thaliana L, ID: AAL84986. The structural gene of glyoxalase I, when sequenced and analyzed, revealed that the uninterrupted open reading frame (ORF) of jojoba Gly I (Jojo-Gly I) spans 775 bp, corresponding to 185 amino acid residues, and shares 45.2% amino acid sequence identity to jojoba (Jojo-Gly I). The cloned ORF, in a multicopy constitutive expression plasmid, complemented the Jojo-Gly I, confirming that the encoded Jojo-Gly I in jojoba showed some homology with other known glyoxalase I sequences of plants.

Purpose Transplantation of organs from animals to humans (xenotransplantation) has been proposed to address the critical shortage of organs for transplantation. The use of genetically engineered organ xenografts, including knock-out (KO) of alpha 1-3 galactosyltransferase (GTKO) along with insertion of a human complement regulator genes (hCD46, hDAF) and other cell surface carbohydrate KOs, has been helpful in circumventing acute and subacute rejection. However, thrombotic complications still remain. Xenografts with additional human transgenes, including hTBM, have been generated to address this barrier. Here, we studied the importance of hTBM in preventing coagulation by comparing two transplanted groups utilizing GE donors with or without hTBM. Methods Specific pathogen-free baboons of either sex weighing 15-30 kg (2-3 years of age) were used as recipients. 6 to 8-week-old genetically modified pigs with hTBM (GTKO.hCD46.hTBM) or without hTBM (triple knock out-GTKO.B4KO.CMAHKO, with or without hCD46 and hDAF) were used as donors, provided by Revivicor. Expression of transgenes were consistent and high level across all pigs. All animals were used in compliance with guidelines provided by the Institutional Animal Care and Use Committee (IACUC). Results GTKO.hCD46.hTBM donors (n=3) were found to have no intracardiac thrombus with survival up to 30 days post-transplant. However, all non-hTBM donors (n=3) were found to have a large intracardiac thrombus burden with propagation into major vascular structures such as the aorta, pulmonary artery and coronary sinus. Gross examination of thrombi indicated acute and subacute components, suggesting early formation of the thrombus postoperatively. Additionally, early clinical signs of thrombus formation included profound hypocalcemia requiring continuous calcium infusions in the first 48-72 hours postoperatively. These phenomena were not prevented by continuous heparin infusion. Histologic examination revealed no signs of rejection. Conclusion hTBM prevents intracardiac thrombus in this model. While immunologic rejection has been circumvented with novel immunosuppression regimens and GTKO, it is likely that hTBM or other transgenes involved in regulation of coagulation may be needed for prevention of thrombotic complications.

The Epstein-Barr virus (EBV) nuclear antigen leader protein (EBNA-LP) is the first viral latency-associated protein produced after EBV infection of resting B cells. Its role in B cell transformation is poorly defined, but it has been reported to enhance gene activation by the EBV protein EBNA2 in vitro. We generated EBNA-LP knockout (LPKO) EBVs containing a STOP codon within each repeat unit of internal repeat 1 (IR1). EBNA-LP-mutant EBVs established lymphoblastoid cell lines (LCLs) from adult B cells at reduced efficiency, but not from umbilical cord B cells, which died approximately two weeks after infection. Adult B cells only established EBNA-LP-null LCLs with a memory (CD27+) phenotype. Quantitative PCR analysis of virus gene expression after infection identified both an altered ratio of the EBNA genes, and a dramatic reduction in transcript levels of both EBNA2-regulated virus genes (LMP1 and LMP2) and the EBNA2-independent EBER genes in the first 2 weeks. By 30 days post infection, LPKO transcription was the same as wild-type EBV. In contrast, EBNA2-regulated cellular genes were induced efficiently by LPKO viruses. Chromatin immunoprecipitation revealed that EBNA2 and the host transcription factors EBF1 and RBPJ were delayed in their recruitment to all viral latency promoters tested, whereas these same factors were recruited efficiently to several host genes, which exhibited increased EBNA2 recruitment. We conclude that EBNA-LP does not simply co-operate with EBNA2 in activating gene transcription, but rather facilitates the recruitment of several transcription factors to the viral genome, to enable transcription of virus latency genes. Additionally, our findings suggest that EBNA-LP is essential for the survival of EBV-infected naïve B cells.

The Epstein-Barr virus (EBV) nuclear antigen leader protein (EBNA-LP) is the first viral latency-associated protein produced after EBV infection of resting B cells. Its role in B cell transformation is poorly defined, but it is reported to enhance gene activation by the EBV protein EBNA2 in vitro.We generated two sets of EBNA-LP knockout (LPKO) EBVs containing a STOP codon within each repeat unit of IR1. Intronic mutations in the first of these knockouts suggested a role for the EBV sisRNAs in transformation. LPKOs with intact introns established lymphoblastoid cell lines (LCLs) from adult B cells at reduced efficiency, but umbilical cord B cells, and naive (IgD+, CD27-) adult B cells consistently died approximately two weeks after infection with LPKO, failing to establish LCLs.Quantitative PCR analysis of virus gene expression after infection identified both an altered ratio of the EBNA genes, and a dramatic reduction in transcript levels of both EBNA2-regulated virus genes (LMP1 and LMP2) and the EBNA2-independent EBER genes, particularly in the first 1-2 weeks. By 30 days post infection, these levels had equalised. In contrast, EBNA2-regulated host genes were induced efficiently by LPKO viruses. Chromatin immunoprecipitation revealed that recruitment of EBNA2 and the host factors EBF1 and RBPJ to all latency promoters tested was severely delayed, whereas these same factors were recruited efficiently to several host genes, some of which exhibited increased EBNA2 recruitment.We conclude that EBNA-LP does not simply co-operate with EBNA2 in activating gene transcription, but rather facilitates the recruitment of several transcription factors to the viral genome, to enable transcription of virus latency genes. Additionally, our findings suggest that different properties of EBV may have differing importance in transforming different B cell subsets.Author summaryEpstein-Barr virus (EBV) infects almost everyone. Once infected, people harbor the virus for life, shedding it in saliva. Infection of children is asymptomatic, but a first infection during adolescence or adulthood can cause glandular fever (mono). EBV is also implicated in several different cancers. EBV infection of B cells (the immune cell that produces antibodies) can drive them to replicate almost indefinitely (‘transformation’), generating cell lines. We have investigated the role of a virus protein – EBNA-LP – which is thought to support gene activation by the essential virus protein EBNA2.We have made an EBV in which the EBNA-LP gene has been disrupted. This virus (LPKO) shows several properties. 1. It is reduced in its ability to transform adult cells, while immature B cells (more frequent in the young) die two weeks after LPKO infection. 2. Some virus genes fail to turn on immediately after LPKO infection. 3. Binding of EBNA2 to these genes is delayed, as is binding of some cellular factors. 4. EBNA-LP does not affect EBNA2-targeted cellular genes in the same way.This shows that EBNA-LP is more important in immature cells, and that it regulates virus genes – but not host genes – more widely than simply through EBNA2.

Epstein-Barr virus (EBV) is a ubiquitous pathogen of humans that can cause several types of lymphoma and carcinoma. Like other herpesviruses, EBV has diversified through both coevolution with its host and genetic exchange between virus strains. Sequence analysis of the EBV genome is unusually challenging because of the large number and lengths of repeat regions within the virus. Here we describe the sequence assembly and analysis of the large internal repeat 1 of EBV (IR1; also known as the BamW repeats) for more than 70 strains. The diversity of the latency protein EBV nuclear antigen leader protein (EBNA-LP) resides predominantly within the exons downstream of IR1. The integrity of the putative BWRF1 open reading frame (ORF) is retained in over 80% of strains, and deletions truncating IR1 always spare BWRF1. Conserved regions include the IR1 latency promoter (Wp) and one zone upstream of and two within BWRF1. IR1 is heterogeneous in 70% of strains, and this heterogeneity arises from sequence exchange between strains as well as from spontaneous mutation, with interstrain recombination being more common in tumor-derived viruses. This genetic exchange often incorporates regions of

The date palm (Phoenix dactylifera L.), famed for its sugar-rich fruits (dates) and cultivated by humans since 4,000 B.C., is an economically important crop in the Middle East, Northern Africa, and increasingly other places where climates are suitable. Despite a long history of human cultivation, the understanding of P. dactylifera genetics and molecular biology are rather limited, hindered by lack of basic data in high quality from genomics and transcriptomics. Here we report a large-scale effort in generating gene models (assembled expressed sequence tags or ESTs and mapped to a genome assembly) for P. dactylifera, using the long-read pyrosequencing platform (Roche/454 GS FLX Titanium) in high coverage. We built fourteen cDNA libraries from different P. dactylifera tissues (cultivar Khalas) and acquired 15,778,993 raw sequencing reads—about one million sequencing reads per library—and the pooled sequences were assembled into 67,651 non-redundant contigs and 301,978 singletons. We annotated 52,725 contigs based on the plant databases and 45 contigs based on functional domains referencing to the Pfam database. From the annotated contigs, we assigned GO (Gene Ontology) terms to 36,086 contigs and KEGG pathways to 7,032 contigs. Our comparative analysis showed that 70.6 % (47,930), 69.4 % (47,089), 68.4 % (46,441), and 69.3 % (47,048) of the P. dactylifera gene models are shared with rice, sorghum, Arabidopsis, and grapevine, respectively. We also assigned our gene models into house-keeping and tissue-specific genes based on their tissue specificity. Electronic supplementary material The online version of this article (doi:10.1007/s11103-012-9924-z) contains supplementary material, which is available to authorized users.

Date palm provides both staple food and gardening for the Middle East and North African countries for thousands of years. Its fruits have diversified significantly, such as nutritional content, size, length, weight color, and ripping process. Dates palm represent an excellent model system for the study of fruit development and diversity of fruit-bearing palm species that produce the most versatile fruit types as compared to other plant families. Using Roche/454 GS FLX instrument, we acquired 7.6 million sequence tags from seven fruiting stages (F1–F7). Over 99% of the raw reads are assembled, and the numbers of isotigs (equivalent to transcription units or unigenes) range from 30,684 to 40,378 during different fruiting stages. We annotated isotigs using BLASTX and BLASTN, and mapped 74% of the isotigs to known functional sequences or genes. Based on gene ontology categorization and pathway analysis, we have identified 10 core cell division genes, 18 ripening related genes, and 7 starch metabolic enzymes, which are involved as nutrition storage and sugar/starch metabolisms. We noticed that many metabolic pathways vary significantly during fruit development, and carbohydrate metabolism (especially sugar synthesis) is particularly prominent during fruit ripening. Transcriptomics study on various fruiting stages of date palm shows complicated metabolic activities during fruit development, ripening, synthesis and accumulation of starch enzymes and other related sugars. Most Genes are highly expressed in early stages of development, while late developmental stages are critical for fruit ripening including most of the metabolism associated ones. Electronic supplementary material The online version of this article (doi:10.1007/s11103-012-9890-5) contains supplementary material, which is available to authorized users.

Epstein-Barr virus (EBV) infects most of the world's population and is causally associated with several human cancers, but little is known about how EBV genetic variation might influence infection or EBV-associated disease. There are currently no published wild-type EBV genome sequences from a healthy individual and very few genomes from EBV-associated diseases. We have sequenced 71 geographically distinct EBV strains from cell lines, multiple types of primary tumor, and blood samples and the first EBV genome from the saliva of a healthy carrier. We show that the established genome map of EBV accurately represents all strains sequenced, but novel deletions are present in a few isolates. We have increased the number of type 2 EBV genomes sequenced from one to 12 and establish that the type 1/type 2 classification is a major feature of EBV genome variation, defined almost exclusively by variation of EBNA2 and EBNA3 genes, but geographic variation is also present. Single nucleotide polymorphism (SNP) density varies substantially across all known open reading frames and is highest in latency-associated genes. Some T-cell epitope sequences in EBNA3 genes show extensive variation across strains, and we identify codons under positive selection, both important considerations for the development of vaccines and T-cell therapy. We also provide new evidence for recombination between strains, which provides a further mechanism for the generation of diversity. Our results provide the first global view of EBV sequence variation and demonstrate an effective method for sequencing large numbers of genomes to further understand the genetics of EBV infection. IMPORTANCE Most people in the world are infected by Epstein-Barr virus (EBV), and it causes several human diseases, which occur at very different rates in different parts of the world and are linked to host immune system variation. Natural variation in EBV DNA sequence may be important for normal infection and for causing disease. Here we used rapid, cost-effective sequencing to determine 71 new EBV sequences from different sample types and locations worldwide. We showed geographic variation in EBV genomes and identified the most variable parts of the genome. We identified protein sequences that seem to have been selected by the host immune system and detected variability in known immune epitopes. This gives the first overview of EBV genome variation, important for designing vaccines and immune therapy for EBV, and provides techniques to investigate relationships between viral sequence variation and EBV-associated diseases.

Date palm (Phoenix dactylifera L.) is a cultivated woody plant species with agricultural and economic importance. Here we report a genome assembly for an elite variety (Khalas), which is 605.4 Mb in size and covers >90% of the genome (~671 Mb) and >96% of its genes (~41,660 genes). Genomic sequence analysis demonstrates that P. dactylifera experienced a clear genome-wide duplication after either ancient whole genome duplications or massive segmental duplications. Genetic diversity analysis indicates that its stress resistance and sugar metabolism-related genes tend to be enriched in the chromosomal regions where the density of single-nucleotide polymorphisms is relatively low. Using transcriptomic data, we also illustrate the date palm’s unique sugar metabolism that underlies fruit development and ripening. Our large-scale genomic and transcriptomic data pave the way for further genomic studies not only on P. dactylifera but also other Arecaceae plants., The date palm is one of the most economically important plants of the palm family. Here, the authors present a high-quality genome assembly of the date palm Phoenix dactylifera, and reveal insights into the unique sugar metabolism underlying fruit ripening.