CHARACTERIZATION OF AVIAN INFLUENZA H5N8 VIRUS STRAINS THAT CAUSED THE OUTBREAKS IN THE RUSSIAN FEDERATION IN 2016–2017

Objective of the study is to investigate biological properties of avian influenza virus strains that caused the outbreaks in Russia in 2016–2017.Materials and methods. The study was performed using advanced virological and molecular-biological methods in state-of-the-art equipment.Results and conclusion. In 2016, the outbreaks among wild birds and poultry caused by highly pathogenic avian influenza H5N8 virus have occurred in the territory of the Russian Federation. In May, 2016 an outbreak of H5N8 among wild birds was registered in the territory of the Republic of Tyva. In October-November, 2016 influenza virus H5N8 was isolated in the territory of the Republics of Tatarstan and Kalmykia, Krasnodar and Astrakhan Regions of Russia. In 2017 avian influenza H5N8 has become widespread in European part of Russia and caused multiple outbreaks among wild birds and poultry. Results of the investigations of the isolated strains show that all of them are highly pathogenic and belong to the clade 2.3.4.4. Molecular-genetic and virological analysis has revealed the differences between the viruses isolated in 2016–2017 and the virus of the same clade 2.3.4.4 that was isolated in 2014

Thyroid cancer risk in Belarus among children and adolescents exposed to radioiodine after the Chornobyl accident

BACKGROUND: Previous studies showed an increased risk of thyroid cancer among children and adolescents exposed to radioactive iodines released after the Chornobyl (Chernobyl) accident, but the effects of screening, iodine deficiency, age at exposure and other factors on the dose-response are poorly understood. METHODS: We screened 11 970 individuals in Belarus aged 18 years or younger at the time of the accident who had estimated (131)I thyroid doses based on individual thyroid activity measurements and dosimetric data from questionnaires. The excess odds ratio per gray (EOR/Gy) was modelled using linear and linear-exponential functions. RESULTS: For thyroid doses \u3c5 \u3eGy, the dose-response was linear (n=85; EOR/Gy=2.15, 95% confidence interval: 0.81-5.47), but at higher doses the excess risk fell. The EOR/Gy was significantly increased among those with prior or screening-detected diffuse goiter, and larger for men than women, and for persons exposed before age 5 than those exposed between 5 and 18 years, although not statistically significant. A somewhat higher EOR/Gy was estimated for validated pre-screening cases. CONCLUSION: 10-15 years after the Chornobyl accident, thyroid cancer risk was significantly increased among individuals exposed to fallout as children or adolescents, but the risk appeared to be lower than in other Chornobyl studies and studies of childhood external irradiation

A Universal System for Highly Efficient Cardiac Differentiation of Human Induced Pluripotent Stem Cells That Eliminates Interline Variability

The production of cardiomyocytes from human induced pluripotent stem cells (hiPSC) holds great promise for patient-specific cardiotoxicity drug testing, disease modeling, and cardiac regeneration. However, existing protocols for the differentiation of hiPSC to the cardiac lineage are inefficient and highly variable. We describe a highly efficient system for differentiation of human embryonic stem cells (hESC) and hiPSC to the cardiac lineage. This system eliminated the variability in cardiac differentiation capacity of a variety of human pluripotent stem cells (hPSC), including hiPSC generated from CD34(+) cord blood using non-viral, non-integrating methods.We systematically and rigorously optimized >45 experimental variables to develop a universal cardiac differentiation system that produced contracting human embryoid bodies (hEB) with an improved efficiency of 94.7±2.4% in an accelerated nine days from four hESC and seven hiPSC lines tested, including hiPSC derived from neonatal CD34(+) cord blood and adult fibroblasts using non-integrating episomal plasmids. This cost-effective differentiation method employed forced aggregation hEB formation in a chemically defined medium, along with staged exposure to physiological (5%) oxygen, and optimized concentrations of mesodermal morphogens BMP4 and FGF2, polyvinyl alcohol, serum, and insulin. The contracting hEB derived using these methods were composed of high percentages (64-89%) of cardiac troponin I(+) cells that displayed ultrastructural properties of functional cardiomyocytes and uniform electrophysiological profiles responsive to cardioactive drugs.This efficient and cost-effective universal system for cardiac differentiation of hiPSC allows a potentially unlimited production of functional cardiomyocytes suitable for application to hPSC-based drug development, cardiac disease modeling, and the future generation of clinically-safe nonviral human cardiac cells for regenerative medicine

Оценка гуморального иммунного ответа экспериментальных животных на введение рекомбинантного эктодомена поверхностного S-гликопротеина вируса SARS-CoV-2 с ИСКОМ-адъювантом

Scientific relevance. The use of recombinant antigens in vaccine production is limited because vaccines based on such antigens tend to have low immunogenicity. However, a COVID-19 vaccine that combines recombinant SARS-CoV-2 spike glycoprotein as its antigen and virus-like immune-stimulating complexes (ISCOMs) as its adjuvant (Nuvaxovid) induces a protective virus-neutralising response. The State Research Center of Virology and Biotechnology “Vector” (hereinafter, Vector) has developed the ISCOM adjuvant Matrix-V, which plays a key role in inducing virus-neutralising antibodies. Studying Matrix-V will provide for the wide use of recombinant antigens combined with this adjuvant in the development and production of novel Russian vaccines.Aim. This study aimed to evaluate the humoral immune responses of experimental animals to intramuscular injections of a complex combining the recombinant Wuhan-type SARS-CoV-2 spike RBD antigen and the virus-like ISCOM adjuvant containing Quillaja saponaria saponins.Materials and methods. The Matrix-V ISCOM adjuvant was produced using Vector’s proprietary technology, which involves cross-flow filtration through Sartorius VivaFlow cassettes. To determine the saponin and residual detergent concentrations in Matrix-V, the authors conducted high-performance liquid chromatography. Having produced the recombinant SARS-CoV-2 RBD antigen, the authors used electron microscopy to analyse the ultrastructure of the ISCOM–antigen complex. In the study of the ISCOM–antigen complex, 25 female Balb/c mice (5 groups) and 15 male and female outbred guinea pigs (3 groups) received two intramuscular injections with a 14-day interval. Serum tests relied on virus neutralisation (VN) and enzyme-linked immunosorbent assay (ELISA) methods and used antigens of 8 SARS-CoV-2 variants (State Collection of Viruses and Rickettsia, Vector). The authors used Statistica 10 to analyse the results.Results. Two injections of the SARS-CoV-2 RBD antigen (mice: 7 μg, guinea pigs: 1 μg) alone did not induce statistically significant virus-neutralising antibody responses, as shown by the VN results. Two injections of the SARS-CoV-2 RBD antigen (mice: 7 μg, guinea pigs: 1 μg) adjuvanted with Matrix-V (25 μg) resulted in geometric mean antibody titres of 1:83–1:178 (mice) and 1:174–1:587 (guinea pigs) in the VN tests with the Wuhan variant. One injection of the antigen (1 μg or 7 μg) with Matrix-V (25 μg) induced antibodies only in individual cases, as demonstrated by the VN and/or ELISA results. The most intensive immune response was observed in ELISA tests with the Delta variant after two injections of the Ecto-S-Wuhan (1 μg) and Matrix-V (25 μg) complex. Immune responses did not differ between the group that received two injections of the Ecto-S-Wuhan antigen (1 μg) without the ISCOM adjuvant and the negative control group (titres below 1:100; p=0.95). Two injections of the SARS-CoV-2 RBD antigen (7 μg) without the ISCOM adjuvant induced antibodies in mice (titres between 1:248 and 1:1477).Conclusions. Two intramuscular injections of the complex containing the recombinant SARS-CoV-2 RBD antigen and the Matrix-V ISCOM adjuvant induce virus-neutralising antibodies. The approach proposed by the authors has the potential for use in the development of immunobiological medicinal products to prevent and treat a wide range of infectious diseases.Актуальность. Использование рекомбинантных антигенов в вакцинах ограничено низкой иммуногенностью таких препаратов. Однако вакцина от COVID-19 (Nuvaxovid), содержащая не только рекомбинантный антиген гликопротеина Spike вируса SARS-CoV-2, но и вирусоподобный иммуностимулирующий комплекс ИСКОМ-адъювант, индуцирует выработку протективного вируснейтрализующего ответа. Исследование разработанного в ФБУН ГНЦ ВБ «Вектор» Роспотребнадзора ИСКОМ-адъюванта «Матрикс-В», играющего ключевую роль в формировании вируснейтрализующего иммунного ответа, позволит широко использовать рекомбинантные антигены в комплексе с адъювантом для разработки и производства новых отечественных вакцин.Цель. Оценка гуморального иммунного ответа на внутримышечное введение животным комплекса рекомбинантного антигена RBD поверхностного Spike-гликопротеина SARS-CoV-2 (штамм «Ухань») и полученных вирусоподобных ИСКОМ с сапонинами Quillaja saponaria.Материалы и методы. ИСКОМ-адъювант «Матрикс-В» получали по разработанной в ФБУН ГНЦ ВБ «Вектор» Роспотребнадзора технологии с применением метода тангенциальной фильтрации в системе Sartorious VivaFlow. Оценивали концентрацию сапонинов и детергента в препарате методом высокоэффективной жидкостной хроматографии. Получен рекомбинантный антиген RBD SARS-CoV-2, проведено электронно-микроскопическое исследование ультраструктуры комплекса «ИСКОМ — антиген». Комплексами «ИСКОМ — антиген» внутримышечно двукратно (через 14 сут) иммунизировали 25 (5 групп) мышей линии Balb/c и 15 (3 группы) разнополых аутбредных морских свинок. Образцы сыворотки крови исследовали в реакции нейтрализации и методом ИФА с антигенами 8 штаммов вируса SARS-CoV-2 (Государственная коллекция возбудителей вирусных инфекций и риккетсиозов ФБУН ГНЦ ВБ «Вектор» Роспотребнадзора). Результаты обрабатывались статистически в программе Statistica 10.Результаты. Двукратное введение антигена RBD SARS-CoV-2 морским свинкам (по 1 мкг) и мышам (по 7 мкг) не приводило в реакции нейтрализации к достоверному формированию вируснейтрализующего ответа. В реакции нейтрализации со штаммом «Ухань» средние геометрические значения титров антител сыворотки крови животных, двукратно иммунизированных комплексом RBD SARS-CoV-2 (мыши — по 7 мкг, морские свинки — по 1 мкг) + Матрикс-В (по 25 мкг), были в пределах 1:83–1:178 и 1:174–1:587 соответственно. В группах животных, однократно иммунизированных комплексом (1 мкг и 7 мкг антигена + Матрикс-В) антитела определялись в реакции нейтрализации (и/или ИФА) только в единичных случаях. В опытах ИФА наиболее интенсивный ответ (на антиген штамма Delta) получен после двукратной иммунизации комплексом Ecto-S-Wuhan (1 мкг) + Матрикс-В (25 мкг). Ответ на двукратное введение антигена Ecto-S-Wuhan (1 мкг) без ИСКОМ-адъюванта (p=0,95) не отличался от отрицательного контроля (значение титра менее 1:100). Двукратное введение мышам антигена RBD SARS-CoV-2 (по 7 мкг) приводило к формированию антител (значения титров 1:248–1:1477).Выводы. Комплекс рекомбинантного антигена RBD SARS-CoV-2 и адъюванта «Матрикс-В» при двукратном внутримышечном введении индуцирует выработку вируснейтрализующих антител. Предложенный подход перспективен для разработки иммунобиологических препаратов профилактики и терапии широкого спектра инфекционных заболеваний

Distribution and diet of juvenile Patagonian toothfish on the South Georgia and Shag Rocks shelves (Southern Ocean)

The distribution and diet of juvenile (<750 mm) Patagonian toothfish are described from 4 annual trawl surveys (2003-06) around the island of South Georgia in the Atlantic sector of the Southern Ocean. Recruitment of toothfish varies inter-annually, and a single large cohort dominated during the four years surveyed. Most juveniles were caught on the Shag Rocks shelf to the NW of South Georgia, with fish subsequently dispersing to deeper water around both the South Georgia and Shag Rocks shelves. Mean size of juvenile toothfish increased with depth of capture. Stomach contents analysis was conducted on 795 fish that contained food remains and revealed that juvenile toothfish are essentially piscivorous, with the diet dominated by notothenid fish. The yellow-finned notothen, Patagonotothen guntheri, was the dominant prey at Shag Rocks whilst at South Georgia, where P. guntheri is absent, the dominant prey were Antarctic krill and notothenid fish. The diet changed with size, with an increase in myctophid fish and krill as toothfish grow and disperse. The size of prey also increased with fish size, with a greater range of prey sizes consumed by larger fish