Molecular genetics and genomics of the ABO blood group system

The A and B oligosaccharide antigens of the ABO blood group system are produced from the common precursor, H substance, by enzymatic reactions catalyzed by A and B glycosyltransferases (AT and BT) encoded by functional A and B alleles at the ABO genetic locus, respectively. In 1990, my research team cloned human A, B, and O allelic cDNAs. We then demonstrated this central dogma of ABO and opened a new era of molecular genetics. We identified four amino acid substitutions between AT and BT and inactivating mutations in the O alleles, clarifying the allelic basis of ABO. We became the first to achieve successful ABO genotyping, discriminating between AA and AO genotypes and between BB and BO, which was impossible using immunohematological/serological methods. We also identified mutations in several subgroup alleles and also in the cis-AB and B(A) alleles that specify the expression of the A and B antigens by single alleles. Later, other scientists interested in the ABO system characterized many additional ABO alleles. However, the situation has changed drastically in the last decade, due to rapid advances in next-generation sequencing (NGS) technology, which has allowed the sequencing of several thousand genes and even the entire genome in individual experiments. Genome sequencing has revealed not only the exome but also transcription/translation regulatory elements. RNA sequencing determines which genes and spliced transcripts are expressed. Because more than 500,000 human genomes have been sequenced and deposited in sequence databases, bioinformaticians can retrieve and analyze this data without generating it. Now, in this era of genomics, we can harness the vast sequence information to unravel the molecular mechanisms responsible for important biological phenomena associated with the ABO polymorphism. Two examples are presented in this review: the delineation of the ABO gene evolution in a variety of species and the association of single nucleotide variant (SNV) sites in the ABO gene with diseases and biological parameters through genome-wide association studies (GWAS).Copyright © Annals of Blood. All rights reserved.

Plasma markers of Covid-19 severity: A pilot study

SARS-CoV-2 infected patients show heterogeneous clinical presentations ranging from mild symptoms to severe respiratory failure and death. Consequently, various markers reflect certain disease presentations. Our cohort included moderate (n = 10) and severe (n = 10) COVID-19 patients, and 10 healthy controls. We determined plasma levels of nine acute phase proteins by nephelometry, full-length (M65), caspase-cleaved (M30) cytokeratin 18, and ADAMTS13 (a disintegrin-like and metalloprotease with thrombospondin type-1 motif 13) by ELISA. In addition, we examined whole plasma N-glycosylation by capillary gel electrophoresis coupled to laser-induced fluorescence detection. When compared to healthy controls, COVID-19 patients had significantly lower concentrations of ADAMTS13 and albumin (ALB) but higher M30, M65, alpha-1-acid glycoprotein, alpha1-antitrypsin (AAT), ceruloplasmin, haptoglobin, and highsensitivity C-reactive protein. The concentrations of alpha1-antichymotrypsin, alpha2-macroglobulin and serum amyloid A proteins did not differ. We found significantly higher levels of AAT and M65 but lower ALB in severe compared to moderate COVID-19 patients. N-glycan analysis of the serum proteome revealed increased levels of oligomannose and sialylated di-antennary glycans, while the non-sialylated di-antennary glycan A2G2 significantly decreased in COVID-19 patients compared to controls. COVID-19-associated changes in levels and N-glycosylation of specific plasma proteins highlight involvement of different pathophysiological mechanisms and grant further investigations.

Totally-green cellulosic fiber with prominent sustained antibacterial and antiviral properties for potential use in spunlaced non-woven fabric production.

The worldwide spread of COVID-19 has put a higher requirement for personal medical protective clothing, developing protective clothing with sustained antibacterial and antiviral performance is the priority for safe and sustaining application. For this purpose, we develop a novel cellulose based material with sustained antibacterial and antiviral properties. In the proposed method, the chitosan oligosaccharide (COS) was subjected to a guanylation reaction with dicyandiamide in the presence of Scandium (III) triflate; because of the relatively lower molecular weight and water solubility of the COS, GCOS (guanylated chitosan oligosaccharide) with high substitution degree (DS) could be successfully synthetized without acid application. In this instance, the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of the GCOS were only 1/8 and 1/4 of that of COS. The introduction of GCOS onto the fiber endowed the fiber with extremely high antibacterial and antiviral performance, showing 100% bacteriostatic rate against Staphylococcus aureus and Escherichia coli and 99.48% virus load reduction of bacteriophage MS2. More importantly, the GCOS modified cellulosic fibers (GCOS-CFs) exhibit excellent sustained antibacterial and antiviral properties; namely, 30 washing cycles had negligible effect on the bacteriostatic rate (100%) and inhibition rate of bacteriophage MS2 (99.0%). Moreover, the paper prepared from the GCOS-CFs still exhibited prominent antibacterial and antiviral activity; inferring that the sheeting forming, press, and drying process have almost no effect on the antibacterial and antiviral performances. The insensitive of antibacterial and antiviral activity to water washing (spunlace) and heat (drying) make the GCOS-CFs a potential material applicable in the spunlaced non-woven fabric production.

Inhibitory activity of a sulfated oligo-porphyran from Pyropia yezoensis against SARS-CoV-2.

COVID-19 caused by SARS-CoV-2 has spread around the world at an unprecedented rate. A more homogeneous oligo-porphyran with mean molecular weight of 2.1 kD, named OP145, was separated from Pyropia yezoensis. NMR analysis showed OP145 was mainly composed of →3)-ß-d-Gal-(1 â†’ 4)-α-l-Gal (6S) repeating units with few replacement of 3,6-anhydride, and the molar ratio was 1:0.85:0.11. MALDI-TOF MS revealed OP145 contained mainly tetrasulfate-oligogalactan with Dp range from 4 to 10 and with no more than two 3,6-anhydro-α-l-Gal replacement. The inhibitory activity of OP145 against SARS-CoV-2 was investigated in vitro and in silico. OP145 could bind to Spike glycoprotein (S-protein) through SPR result, and pseudovirus tests confirmed that OP145 could inhibite the infection with an EC50 of 37.52 µg/mL. Molecular docking simulated the interaction between the main component of OP145 and S-protein. All the results indicated that OP145 had the potency to treat and prevent COVID-19.

Anti-SARS-CoV-2 and anticoagulant properties of Pentacta pygmaea fucosylated chondroitin sulfate depend on high molecular weight structures.

Fucosylated chondroitin sulfate (FucCS) is a unique marine glycosaminoglycan that exhibits diverse biological functions including antiviral and anticoagulant activity. In previous work, the FucCS derived from Pentacta pygmaea (PpFucCS) showed moderate anticoagulant effect but high inhibitory activity against the Wuhan strain of severe acute respiratory syndrome coronavirus (SARS-CoV-2). In this study, we perform free-radical depolymerization of PpFucCS by the copper-based Fenton method to generate low molecular weight (MW) oligosaccharides. PpFucCS oligosaccharides were structurally analyzed by 1H nuclear magnetic resonance spectroscopy and used to conduct structure-activity relationship studies regarding their effects against SARS-CoV-2 and clotting. Anticoagulant properties were measured by activated partial thromboplastin time, protease (factors Xa and IIa) inhibition by serine protease inhibitors [antithrombin (AT) and heparin cofactor II (HCII)], and competitive surface plasmon resonance (SPR) assay using AT, HCII and IIa. Anti-SARS-CoV-2 properties were measured by concentration-response inhibitory curves of HEK-293 T-hACE2 cells infected with a baculovirus pseudotyped SARS-CoV-2 Delta variant spike (S)-protein and competitive SPR assays using multiple S-proteins [Wuhan, N501Y (Alpha), K417T/E484K/N501Y (Gamma), L542R (Delta) and Omicron (BA.2 subvariant)]. Cytotoxicity of native PpFucCS and oligosaccharides was also assessed. The PpFucCS-derived oligosaccharide fraction of the highest MW showed great anti-SARS-CoV-2 Delta activity and reduced anticoagulant properties. Results have indicated no cytotoxicity and MW-dependency on both anti-SARS-CoV-2 and anticoagulant effects of PpFucCS as both actions were reduced accordingly to the MW decrease of PpFucCS. Our results demonstrate that the high MW structures of PpFucCS is a key structural element to achieve the maximal anti-SARS-CoV-2 and anticoagulant effects.

Antibacterial and antiviral chitosan oligosaccharide modified cellulosic fibers with durability against washing and long-acting activity.

The worldwide outbreak of SARS-CoV-2 has attracted extensive attention to antibacterial and antivirus materials. Cellulose is the most potential candidate for the preparation of green, environmentally friendly antibacterial and antiviral materials. Herein, modified cellulosic fibers with sustained antibacterial and antiviral performance was prepared by introducing chitosan oligosaccharide onto the fibers. The two-step method is proved to be more effective than the one-step method for enhanced chitosan oligosaccharide loadings and antibacterial and antiviral activity. In this instance, the modified fibers with 61.77 mg/g chitosan oligosaccharide loadings can inhibit Staphylococcus aureus and Escherichia coli by 100 % after contacting with bacteria for 12 h and reduce the bacteriophage MS2 by 99.19 % after 1 h of contact. More importantly, the modified fibers have washing durable antibacterial and antiviral activity; the modified fibers have 100 % antibacterial and 98.38 % antiviral activity after 20 washing cycles. Benefiting from the excellent performance of the individual fibers, the paper prepared from the modified fibers show great antibacterial (100 %) and antiviral performance (99.01 %) and comparable mechanical strength. The modified fibers have potential applications in the manufacture of protective clothing and protective hygiene products.

Glyfinder and Glycoprotein Builder: Online Tools for Finding and Modeling Glycoproteins in the PDB

Tens of thousands of 3D structures of oligosaccharides have been deposited into the Protein Databank (PDB), representing hundreds of thousands of hours of effort by crystallographers. Yet, despite the critical importance of these structures in furthering the development of glycomimetic drugs, in explaining the activity of glycan-processing enzymes, and in providing a deeper understanding of the properties of glycoproteins and vaccines, they remain unnecessarily difficult to locate within the PDB. Part of this is due to limitations in searching for oligosaccharides on the PDB website, even after a recent carbohydrate remediation project completed by the PDB. While several databases have been reported that contain carbohydrate structural information extracted from the PDB, few offer flexible search capabilities and even fewer provide independent assessment of data quality. Here we present the GlyFinder and GlyProbity webtools (glycam.org/gf) and illustrate their application to locating oligosaccharides, carbohydrate derivatives, and glycoproteins stored in the PDB. We highlight the utility of curating the data on the basis of the theoretical conformational (CHI) energies [1] of the glycosidic linkages and illustrate how the deposited data can be employed to generate 3D models of glycoproteins, including the SARS-CoV-2 Spike protein [2].

Therapeutic potential of plant secondary metabolites in treatment of respiratory viral diseases: A review

Respiratory viral infections are a major public health concern because of their global occurrence, ease of spread and considerable morbidity and mortality. Medical treatments for viral respiratory diseases primarily involve providing relief from symptoms like pain and discomfort rather than treating the infection. Very few antiviral medications have been approved with restrictive usage, high cost, unwanted side effects and limited availability. Plants with their unique metabolite composition and high remedial values offer unique preventive and therapeutic efficacy in treatment of viral infections. The present review is focused on the types and mode of action of plant secondary metabolites that have been used successfully ί in the treatment of infections caused by respiratory viruses like Influenza, SARS, MERS, RSV etc. Plant metabolites such as phenolics, alkaloids, terpenoids and oligosaccharides inhibit attachment and entry of the virus. Others such as flavonoids, viz quercetin and baicalein, alkaloids viz sanguinarine, berberine and emetine, specific lipids and fatty acids prevent viral replication and protein synthesis. These metabolites have the potential to be used as lead molecules that can be optimized to develop potent drugs for effectively combating pandemics caused by respiratory viruses.

Fractionation of sulfated galactan from the red alga Botryocladia occidentalis separates its anticoagulant and anti-SARS-CoV-2 properties.

Sulfation pattern and molecular weight (MW) play a key role in the biological actions of sulfated glycans. Besides anticoagulant effects, certain sulfated glycans can also exhibit anti-SARS-CoV-2 properties. To develop a more selective antiviral carbohydrate, an efficient strategy to separate these two actions is required. In this work, low MW fractions derived from the red alga Botryocladia occidentalis sulfated galactan (BoSG) were generated, structurally characterized, and tested for activity against SARS-CoV-2 and blood coagulation. The lowest MW fraction was found to be primarily composed of octasaccharides of monosulfated monosaccharides. Unlike heparin or native BoSG, we found that hydrolyzed BoSG products had weak anticoagulant activities as seen by aPTT and inhibitory assays using purified cofactors. In contrast, lower MW BoSG-derivatives retained anti-SARS-CoV-2 activity using SARS-CoV-2 spike (S)-protein pseudotyped lentivirus vector in HEK-293T-hACE2 cells monitored by GFP. Surface plasmon resonance confirmed that longer chains are necessary for BoSG to interact with coagulation cofactors but is not required for interactions with certain S-protein variants. We observed distinct affinities of BoSG derivatives for the S-proteins of different SARS-CoV-2 strains, including WT, N501Y (Alpha), K417T/E484K/N501Y (Gamma), and L542R (Delta) mutants, and stronger affinity for the N501Y-containing variants. Docking of the four possible monosulfated BoSG disaccharides in interactions with the N501Y mutant S-protein predicted potential binding poses of the BoSG constructs and favorable binding in close proximity to the 501Y residue. Our results demonstrate that depolymerization and fractionation of BoSG are an effective strategy to segregate its anticoagulant property from its anti-SARS-CoV-2 action.

Nutritional status and the critically Ill patient: Gut microbiota and immuno-nutrition in I.C.U. At the time of SARS-COV 2 pandemic

Background: Gut microbiota is a complex ecosystem of bacteria, viruses, archaea, protozoa and yeasts in our intestine. It has several functions, including maintaining human body equilibrium. Microbial “dysbiosis” can be responsible for outbreak of local and systemic infections, especially in critically ill patients. Methods: to build a narrative review, we performed a Pubmed, Medline and EMBASE search for English language papers, reviews, meta-analyses, case series and randomized controlled trials (RCTs) by keywords and their associations: critically ill patient;nutrition;gut microbiota;probiotics;gut virome;SARS-COV 2. Results: Over the antibiotic-based “selective decontamination”, potentially responsible for drug-resistant microorganisms development, there is growing interest of scientists and the pharmaceutical industry for pre-, probiotics and their associations as safe and reliable remedies restoring gut microbial “eubiosis”. Very first encouraging evidences link different gut microbiota profiles with SARS-COV 2 disease stage and gravity. Thus, there is frame for a probiotic therapeutic approach of COVID-19. Conclusions: gut microbiota remodulation seems to be a promising and safe therapeutic approach to prevent local and systemic multi-resistant bug infections in the intensive care unit (ICU) patients. This approach deserves more and more attention at the time of SARS-COV 2 pandemic.