A nidovirus perspective on SARS-CoV-2

Two pandemics of respiratory distress diseases associated with zoonotic introductions of the species Severe acute respiratory syndrome-related coronavirus in the human population during 21st century raised unprecedented interest in coronavirus research and assigned it unseen urgency. The two viruses responsible for the outbreaks, SARS-CoV and SARS-CoV-2, respectively, are in the spotlight, and SARSCoV-2 is the focus of the current fast-paced research. Its foundation was laid down by studies of many coronaand related viruses that collectively form the vast order Nidovirales. Comparative genomics of nidoviruses played a key role in this advancement over more than 30 years. It facilitated the transfer of knowledge from characterized to newly identified viruses, including SARS-CoV and SARS-CoV-2, as well as contributed to the dissection of the nidovirus proteome and identification of patterns of variations between different taxonomic groups, from species to families. This review revisits selected cases of protein conservation and variation that define nidoviruses, illustrates the remarkable plasticity of the proteome during nidovirus adaptation, and asks questions at the interface of the proteome and processes that are vital for nidovirus reproduction and could inform the ongoing research of SARS-CoV-2.(c) 2020 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).Molecular basis of virus replication, viral pathogenesis and antiviral strategie

Определение концентрации холестерина и триглицеридов в экстраклеточных везикулах сыворотки крови с помощью коммерческих наборов

Exosomes and microvesicles, collectively referred to as small extracellular vesicles (sEV) are vesicles with an average size of about 100-150 nm. Currently, the role of sEV in various aspects of signaling in the body is being actively investigated; in addition, sEV can often serve as markers of various pathologies. The active study of the sEV composition is continuing. In this study we have demonstrated that in sEV it is possible to determine cholesterol and triglycerides concentration by using commercial kits designed for serum. The technique was tested on sEV from the blood of patients diagnosed with depression and on healthy volunteers. No differences were found in the concentration of cholesterol and triglycerides in mEV from the blood serum of depressed patients and the control group. The concentration of cholesterol and triglycerides in the samples is several times higher than the sensitivity threshold of the methods set by the manufacturer of the kits.Экзосомы и микровезикулы, совместно называемые малые экстраклеточные везикулы (мЭВ), представляют собой везикулы со средним размером около 100-150 нм. В настоящее время активно исследуется роль мЭВ в самых разных аспектах сигналинга в организме, кроме того, часто мЭВ могут служить маркерами разных патологий. Продолжается активное изучение состава мЭВ. В данной работе мы показали, что в мЭВ можно определить концентрацию холестерина и триглицеридов с помощью коммерческих наборов, предназначенных для сыворотки крови. Методика была опробована на мЭВ из крови пациентов с диагнозом депрессия и на здоровых добровольцах. Различий в концентрации холестерина и триглицеридов в мЭВ из сыворотки крови пациентов с депрессией и контрольной группы найдено не было. Концентрация холестерина и триглицеридов в образцах в несколько раз превосходит порог чувствительности методов, установленный производителем наборов

A planarian nidovirus expands the limits of RNA genome size

Molecular basis of virus replication, viral pathogenesis and antiviral strategie

PCR assays for detection of human astroviruses: In silico evaluation and design, and in vitro application to samples collected from patients in the Netherlands

Molecular basis of virus replication, viral pathogenesis and antiviral strategie

A nidovirus perspective on SARS-CoV-2

Two pandemics of respiratory distress diseases associated with zoonotic introductions of the species Severe acute respiratory syndrome-related coronavirus in the human population during 21st century raised unprecedented interest in coronavirus research and assigned it unseen urgency. The two viruses responsible for the outbreaks, SARS-CoV and SARS-CoV-2, respectively, are in the spotlight, and SARS-CoV-2 is the focus of the current fast-paced research. Its foundation was laid down by studies of many corona- and related viruses that collectively form the vast order Nidovirales. Comparative genomics of nidoviruses played a key role in this advancement over more than 30 years. It facilitated the transfer of knowledge from characterized to newly identified viruses, including SARS-CoV and SARS-CoV-2, as well as contributed to the dissection of the nidovirus proteome and identification of patterns of variations between different taxonomic groups, from species to families. This review revisits selected cases of protein conservation and variation that define nidoviruses, illustrates the remarkable plasticity of the proteome during nidovirus adaptation, and asks questions at the interface of the proteome and processes that are vital for nidovirus reproduction and could inform the ongoing research of SARS-CoV-2. © 2020 The Author

Winter resistance of sour cherry and sweet cherry flower buds in extreme winter 2009-2010 and 2020-2021

This article is devoted to the problem of field winter hardiness of sour cherry and sweet cherry flower buds as the main element of winter hardiness of fruit plants in the Oryol region. The issue is considered from the point of view of winter hardiness in extremely cold winters of 2009-2010. and 2020-2021, which allow you to more accurately determine the level of winter hardiness of each individual variety, which is rarely possible in mild comfortable winters. More than 30 varieties of sour cherries and sweet cherries from the gene pool of the All-Russian Research Institute of Fruit Crop Breeding were taken as objects of study. In both cultures, a number of varieties with high winter hardiness of flower buds were identified – sour cherry varieties Griot Ostgeimsky, Volochaevka, Otrada, Shokoladnitsa, Otrada, Uifekhertoy Furtosh and sweet cherry varieties – Zarya Vostoka and Orlovskaya Yantarnaya

Чувствительность экстраклеточных везикул из сыворотки крови человека к различным детергентам

Blood exosomes and microvesicles, collectively known as small extracellular vesicles (sEV), are vesicles about 100-150 nm in size. Small EV are involved in various aspects of signaling in the body; in addition, they can serve as markers of various pathologies. For biochemical studies, vesicle solubilization is often required. We tested the ability of various detergents to dissolve membranes of the sEV. Small EV were isolated from the blood serum of healthy volunteers by gel filtration on Sepharose CL-2B and tried to solubilize them using the anionic detergent DOC (sodium deoxycholate), non-ionic detergent Brij 35 (polyoxyethyleneglycol dodecyl ether), zwitterionic detergent CHAPS (3 - [(3-chloramidopropyl) dimethylammonio] -1-propanesulfonate), and cationic detergent CTAB (cetyl trimethylammonium bromide). The concentration of sEV in the solution was determined by dynamic light scattering. We find DOC is the most effective for sEV solubilization.Экзосомы и микровезикулы, совместно называемые малые экстраклеточные везикулы (мЭВ), представляют собой везикулы размером около 100-150 нм. Малые ЭВ принимают участие в самых разных аспектах сигналинга в организме; кроме того, они могут служить маркерами разных патологий. Для биохимических исследований зачастую требуется солюбилизация везикул. Мы проверили способность различных детергентов растворять мембраны мЭВ. Малые ЭВ выделяли из сыворотки крови здоровых добровольцев с помощью гель-фильтрации на Sepharose CL-2B и пытались их солюбилизировать с помощью анионного детергента DOC (дезоксихолат натрия), неионного детергента Brij 35 (додециловый эфир полиэтиленгликоля), цвиттерионного детергента CHAPS (3-[(3-хлорамидопропил) диметиламмонио]-1-пропансульфонат), катионного детергента CTAB (цетил триметиламмония бромид). Концентрацию мЭВ в растворе определяли с помощью динамического светорассеяния. Самым эффективным детергентом для солюбилизации мЭВ из сыворотки крови оказался DOC

Оптимизация иммуноферментного метода для определения цилиарного нейротрофического фактора в слезной жидкости человека

The concentration of ciliary neurotrophic factor (CNTF) was measured in lacrimal fluid (LF) using Human CNTF Quantikine ELISA kit (“R&D Systems”, USA) on a ChemWell 2910 automatic analyzer (“Awareness Technology Inc.”, USA). We initially attempted to use commercial kits, designed for serum and plasma CNTF detection, to quantify lacrimal CNTF. The results, however, were rarely above the minimum detection level of the kits, most likely due to matrix complexity and low concentrations of CNTF in diluted LF (LF had to be diluted because of the small volume of collected samples). The optimal sensitivity and the lowest background for the best minimum quantifiable value were determined empirically. Phosphate buffer solution containing 1% bovine serum albumin was selected as an optimal diluent for CNTF measurements in small fluid samples. A standard curve was produced using the calibrating solutions 0-250 pg/ml. Acid treatment of LF samples before the analysis allowed to increase the detectable concentration of the CNTF two-fold. The 1:3 dilution was selected based on the available volume of collected LF and a reasonable variation coefficient. The described protocol allowed to develop a sandwich ELISA optimized for lacrimal CNTF.Определение цилиарного нейротрофического фактора – ciliary neurotrophic factor (CNTF) в слёзной жидкости (СЖ) человека рутинными лабораторными методами затруднено из-за его низкой концентрации в структурах глаза и недостаточного количества материала для анализа. Были подобраны условия для определения CNTF в СЖ твердофазным методом иммуноферментного анализа (ИФА) на основе набора Human CNTF Quantikine ELISA Kit (“R&D Systems”, США) с использованием автоматического иммуноферментного анализатора ChemWell 2910 Combi (“Awareness Technology Inc.”, США). Предварительная кислотная обработка проб СЖ позволила вдвое повысить концентрацию CNTF. Полностью автоматизированная система проведения анализа и выбор разбавителя с низкой оптической плотностью бланка (0.011) обеспечили линейную зависимость калибровочного графика от 0 до 250 пг/мл с возможностью его использования для расчета концентрации CNTF в области низких значений. Рабочие характеристики оптимизированной тест-системы ИФА (линейность метода, предел обнаружения, воспроизводимость, интерференция матрицы) в указанном диапазоне концентраций соответствовали критериям приемлемости для используемого метода определения. Разведение образцов СЖ в четыре раза, совмещённое с кислотной обработкой проб, оказалось оптимальным для получения стабильных результатов при проведении анализа. В этом случае концентрация CNTF в пробе составляла в среднем 10.4±0.4 пг/мл с коэффициентом вариации 4.2% и близостью определяемых значений к ожидаемым величинам на уровне 104%

LAMPA, LArge Multidomain Protein Annotator, and its application to RNA virus polyproteins

Motivation: To facilitate accurate estimation of statistical significance of sequence similarity in profile-profile searches, queries should ideally correspond to protein domains. For multidomain proteins, using domains as queries depends on delineation of domain borders, which may be unknown. Thus, proteins are commonly used as queries that complicate establishing homology for similarities close to cutoff levels of statistical significance.Results: In this article, we describe an iterative approach, called LAMPA, LArge Multidomain Protein Annotator, that resolves the above conundrum by gradual expansion of hit coverage of multidomain proteins through re-evaluating statistical significance of hit similarity using ever smaller queries defined at each iteration. LAMPA employs TMHMM and HHsearch for recognition of transmembrane regions and homology, respectively. We used Pfam database for annotating 2985 multidomain proteins (polyproteins) composed of >1000 amino acid residues, which dominate proteomes of RNA viruses. Under strict cutoffs, LAMPA outperformed HHsearch-mediated runs using intact polyproteins as queries by three measures: number of and coverage by identified homologous regions, and number of hit Pfam profiles. Compared to HHsearch, LAMPA identified 507 extra homologous regions in 14.4% of polyproteins. This Pfam-based annotation of RNA virus polyproteins by LAMPA was also superior to RefSeq expert annotation by two measures, region number and annotated length, for 69.3% of RNA virus polyprotein entries. We rationalized the obtained results based on dependencies of HHsearch hit statistical significance for local alignment similarity score from lengths and diversities of query-target pairs in computational experiments

The Missing Link: How Exosomes and miRNAs can Help in Bridging Psychiatry and Molecular Biology in the Context of Depression, Bipolar Disorder and Schizophrenia

MicroRNAs (miRNAs) only recently have been recognized as promising molecules for both fundamental and clinical neuroscience. We provide a literature review of miRNA biomarker studies in three most prominent psychiatric disorders (depression, bipolar disorder and schizophrenia) with the particular focus on depression due to its social and healthcare importance. Our search resulted in 191 unique miRNAs across 35 human studies measuring miRNA levels in blood, serum or plasma. 30 miRNAs replicated in more than one study. Most miRNAs targeted neuroplasticity and neurodevelopment pathways. Various limitations do not allow us to make firm conclusions on clinical potential of studied miRNAs. Based on our results we discuss the rationale for future research investigations of exosomal mechanisms to overcome methodological caveats both in studying etiology and pathogenesis, and providing an objective back-up for clinical decisions. © 2019, Springer Science+Business Media, LLC, part of Springer Nature