The use of quantitative enzyme-linked immunosorbent assay for the determination of S-antigen concentration in whole-virion inactivated adsorbed coronavirus vaccines

The severe consequences and high mortality of COVID-19 prompted the development of a wide range of preventive vaccines. The first vaccines to be tested were developed in China and formulated as inactivated SARS-CoV-2 adsorbed on aluminium hydroxide. One of the quality indicators for inactivated adsorbed vaccines is the degree of adsorption, which can be used to control the content not only of non-adsorbed antigen, but also of specific antigen in one dose of a vaccine.The aim of the study was to investigate the possibility of desorbing SARS-CoV-2 antigen from formulated adsorbed vaccines and the possibility of measuring its concentration using the BioScan-SARS-CoV-2 (S) ELISA kit for SARS-CoV-2 S-protein content determination.Materials and methods: the study used four batches of BBIBP-CorV by CNBG, Sinopharm (China) and three batches of CoronaVac by Sinovac Biotech (China). The authors desorbed SARS-CoV-2 S antigen in accordance with monograph FS.3.3.1.0029.15 of the State Pharmacopoeia of the Russian Federation (Ph. Rus.), edition XIV, and quantified it using the BioScan-SARS-CoV-2 (S) ELISA kit by Bioservice Biotechnology Co. Ltd. (Russia).Results: mean S-antigen concentrations in the desorbed samples ranged from 61 to 129 ng/mL for BBIBP-CorV and from 461 to 533 ng/mL for CoronaVac.Conclusions: the study demonstrated the possibility of specific SARS-CoV-2 antigen desorption from the surface of aluminium hydroxide using the Ph. Rus. method, as well as the possibility of S-antigen quantification in desorbed medicinal products and supernatants using the BioScan-SARS-CoV-2 (S) ELISA kit. The authors observed 3.6- to 8.7-fold difference between the S-antigen concentrations of the desorbed preparations by the two manufacturers

Alternaria leaf blight of clusterbean

The annual  legume  crop clusterbean (Cyamopsis tetragonoloba  (L.) Taub.) is a promising  crop for cultivation  in the south of Russia. In 2018, phytosanitary examinations of clusterbean fields (13 collection accessions) were conducted in five VIR branches (Krasnodar Territory, Dagestan,  Astrakhan and Volgograd Regions). At all points,  several types of leaf spots were observed on clusterbean leaves and symptoms of plant damage by fungi of the genus  Alternaria Nees prevailed.  Using microbiological  methods and  Next-Generation Sequencing (NGS) of the  nuclear  ribosomal internal  transcribed spacer  two (ITS2), the  species  composition of micromycetes was identified.  It was found  that the  micromycetes Alternaria spp. in different  ecological  and  geographical conditions of the  south  of Russia cause two main types of leaf spots: the typical (beige and brown round spots, usually accompanied by concentric zonality) and brown  spot (small brown  bulging  merging spots). Overwhelmingly  the damage to leaf tissues is caused  by the fungus  A. tenuissima (Nees & T. Nees : Fr.) Wiltshire. A quite  numerous accompanying mycoflora (first of all, fungi of the genus  Fusarium Link) was also detected. A. cyamopsidis Rangaswami & A.V. Rao, one of the most  harmful guar pathogens in the countries  where the main acreage is located  (India, Pakistan, USA), was not found on clusterbean fields in Russia. The accessions  of clusterbean differ in degree of damage by A. tenuissima that causes Alternaria leaf blight. Differential interaction of parasite  and plant  host was revealed.  Therefore, to prevent epiphytotics, varieties protected by non-identical resistance genes  should  be grown.  At all VIR branches, accessions  k-52568 (Argentina) and k-52569 (Pakistan) were weakly damaged by Alternaria leaf blight, and some  accessions  were resistant only in the environmental conditions of one or two experimental stations. The accessions  studied were heterogeneous  in pathogen resistance,  which  allows selecting  disease-resistant lines from most  of the  collection  accessions.  Thus, in various VIR branches, plants  without symptoms of disease  were selected from accessions  k-52571, k-52573 and k-52580, and seeds were collected to create new donors of disease resistance

The problem of SARS-CoV-2 virus resistance to direct-acting antivirals

INTRODUCTION. The high prevalence of mutations in the SARS-CoV-2 genome raises particular concerns about the resistance of the virus to current antiviral therapy, including inhibitors of the main protease, or 3C-like protease (3CLpro), and RNA-dependent RNA polymerase (RdRp).AIM. This study aimed to analyse the prevalence, spectrum, and causes of SARS-CoV-2 mutations conferring resistance to approved and pipeline RdRp and 3CLpro inhibitors on the basis of clinical, virological, and genotypic data.DISCUSSION. The authors have analysed the prevalence of SARS-CoV-2 mutations conferring resistance to antivirals (RdRp inhibitors, including remdesivir and molnupiravir, and 3CLpro inhibitors, including paxlovid) in 2021–2024. The results suggest that certain mutations existed prior to the use of these antivirals. The prevalence of resistance-conferring mutations does not exceed 0.5% of the global population. However, the results of clinical and experimental studies demonstrate the possibility of a more than 200-fold reduction in susceptibility to medicinal products and, in particular, the emergence of multidrug-resistant variants. This is especially important for immunocompromised patients. SARS-CoV-2 can persist in such patients for many months, during which spontaneous or selection-driven mutations can render antiviral therapy ineffective. This would create a risk of spreading drug-resistant variants and/or a risk of adverse outcomes for patients.CONCLUSIONS. As COVID-19 treatment coverage increases, there may be a rise in drug-resistant variants of the virus. The presented data indicate the need for genomic epidemiological surveillance, including an analysis of potential targets for medicinal products based on clinical observations. In the future, surveillance data may determine the treatment strategy and the need to develop new antivirals (RdRp and protease inhibitors) adjusted to resistant SARS-CoV-2 variants

Применение количественного иммуноферментного анализа для определения концентрации S-антигена в цельновирионных инактивированных адсорбированных коронавирусных вакцинах

The severe consequences and high mortality of COVID-19 prompted the development of a wide range of preventive vaccines. The first vaccines to be tested were developed in China and formulated as inactivated SARS-CoV-2 adsorbed on aluminium hydroxide. One of the quality indicators for inactivated adsorbed vaccines is the degree of adsorption, which can be used to control the content not only of non-adsorbed antigen, but also of specific antigen in one dose of a vaccine.The aim of the study was to investigate the possibility of desorbing SARS-CoV-2 antigen from formulated adsorbed vaccines and the possibility of measuring its concentration using the BioScan-SARS-CoV-2 (S) ELISA kit for SARS-CoV-2 S-protein content determination.Materials and methods: the study used four batches of BBIBP-CorV by CNBG, Sinopharm (China) and three batches of CoronaVac by Sinovac Biotech (China). The authors desorbed SARS-CoV-2 S antigen in accordance with monograph FS.3.3.1.0029.15 of the State Pharmacopoeia of the Russian Federation (Ph. Rus.), edition XIV, and quantified it using the BioScan-SARS-CoV-2 (S) ELISA kit by Bioservice Biotechnology Co. Ltd. (Russia).Results: mean S-antigen concentrations in the desorbed samples ranged from 61 to 129 ng/mL for BBIBP-CorV and from 461 to 533 ng/mL for CoronaVac.Conclusions: the study demonstrated the possibility of specific SARS-CoV-2 antigen desorption from the surface of aluminium hydroxide using the Ph. Rus. method, as well as the possibility of S-antigen quantification in desorbed medicinal products and supernatants using the BioScan-SARS-CoV-2 (S) ELISA kit. The authors observed 3.6- to 8.7-fold difference between the S-antigen concentrations of the desorbed preparations by the two manufacturers.Тяжелые последствия заболевания, вызываемые вирусом SARS-CoV-2, а также большое количество случаев заболевания с летальным исходом, обусловили разработку целого ряда профилактических вакцин. Первые вакцины, об испытаниях которых было заявлено, были разработаны в Китае и представляли собой инактивированный вирус SARS-CoV-2, адсорбированный на гидроокиси алюминия. Для инактивированных адсорбированных вакцин одним из показателей качества является полнота сорбции. Определение этого параметра позволяет не только контролировать количество несорбированного антигена, но и количество специфического антигена в дозе.Цель работы: изучение возможности проведения десорбции антигена вируса SARS-CoV-2 в готовых лекарственных формах адсорбированных вакцин и определение концентрации антигена вируса с использованием набора реагентов «БиоСкан-SARS-CoV-2 (S)» для количественного определения S-белка вируса SARS-CoV-2 методом иммуноферментного анализа (ИФА).Материалы и методы: в работе были использованы образцы четырех серий вакцины BBIBP-CorV (CNBG, Sinopharm, Китай) и трех серий вакцины CoronaVac (Sinovac Biotech, Китай). Десорбцию антигена проводили в соответствии с ФС.3.3.1.0029.15 Государственной фармакопеи Российской Федерации XIV издания (ГФ РФ XIV), а количественное определение S-антигена вируса SARS-CoV-2 – с использованием набора реагентов для ИФА «БиоСкан-SARS-CoV-2 (S)» (АО БТК «Биосервис», Россия).Результаты: в исследованных образцах четырех серий вакцины BBIBP-CorV концентрация S-антигена в десорбированных образцах варьировала в среднем от 61 до 129 нг/мл, а в образцах трех серий вакцины CoronaVac – от 461 до 533 нг/мл.Выводы: была показана возможность десорбции специфического антигена вируса SARS-CoV-2 с гидроокиси алюминия по методике ФС.3.3.1.0029.15 ГФ РФ XIV. Показана возможность количественной оценки содержания S-антигена в десорбированном препарате и в супернатанте с использованием набора реагентов для ИФА «БиоСкан-SARS-CoV-2 (S)». Разница в концентрациях S-антигена в десорбированных препаратах между двумя разными производителями составляла от 3,6 до 8,7 раза

Molecular epidemiological study of clinical cases of acute hepatitis E in Belarus

Relevance. The frequency of occurrence of anamnestic antibodies to the hepatitis E virus (HEV) in the general population of the Republic of Belarus is 7.3%, which is clearly not consistent with the low incidence of hepatitis E (HE). Most of primary HEV infections remain undiagnosed. The intensive epidemic process of HEV in the Belarusian population is hidden. Conducting epidemiological studies, including genotyping of HEV sequences isolated on the territory of the republic, makes it possible to more accurately characterize the sources of HEV infection and the mechanisms of its transmission. Aim molecular epidemiological study of two cases of acute hepatitis E detected in patients from Belarus. Materials and methods. During 20212022, samples of biological material were obtained from two patients undergoing treatment with an established diagnosis of acute hepatitis E. Serum samples were tested to detect antibodies to HEV using enzyme immunoassay, HEV RNA was detected in fecal samples using nested RT-PCR. The nucleotide sequence was determined by an automatic sequencer using the Sanger method. Analysis of nucleotide sequences, their genotyping, and calculation of evolutionary distances were performed using MEGA X software. Results. The HEV sequence isolated from a pregnant woman who had an epidemiological episode of alimentary contact with raw pork meat is clustered into a common phylogenetic clade with HEV sequence obtained from the patient from Belarus with a history of kidney transplantation and HEV sequences isolated from a domestic pigs. The HEV sequence isolated from a patient with a history of travel to Pakistan belongs to the HEV genotype 1 and joins a clade of HEV sequences isolated in Pakistan, India, Nepal and Mongolia

РЕЗУЛЬТАТЫ ИСПОЛЬЗОВАНИЯ СОФОСБУВИРА В КОМБИНАЦИИ С ЛЕДИПАСВИРОМ ИЛИ ДАКЛАТАСВИРОМ ДЛЯ ЛЕЧЕНИЯ ХРОНИЧЕСКОГО ГЕПАТИТА С В РЕСПУБЛИКЕ БЕЛАРУСЬ

To evaluate the efficacy of therapy with sofosbuvir in combination with ledipasvir or daclatasvir, the results of treatment of 299 patients with chronic hepatitis C, including 128 non-responders to combined interferon plus ribavirin therapy, who have prognostically unfavorable single nucleotide polymorphisms 39743165T> G (rs8099917) and 39738787C> T (rs12979860) of interleukin-28B gene, were analyzed. 57 people had liver cirrhosis. 80,9% (242) had genotype 1 of hepatitis C virus, 5% (15) – genotype 2, 13,7% (41) – genotype 3, and 0,4% (1) – genotype 4.All 299 patients, who adhered to the recommendations of the European Association for the Study of the Liver 2016 and the American Association for the Study of the Liver Disease 2017, achieved a sustained virologic response 12 weeks after the end of therapy. The clinical case of treatment failure, associated with the lack of confirmation of the elimination of hepatitis C virus by means of highly sensitive polymerase chain reaction methods and with the later identified amino acid substitution in position Y93H of NS5A (resistance to NS5A inhibitors), is shown.It is necessary to carry out monitoring of effectiveness of therapy only by means of highly sensitive polymerase chain reaction (from 10 ME/ml). If the virus elimination delays in patients with advanced stages of liver fibrosis it is needed to use the prolonged schemes of treatment. Repeated treatment of patients with existence of a mutation of Y93H requires the use of new NS5A inhibitors or combined drugs.С целью оценки эффективности терапии препаратами софосбувир в комбинации с ледипасвиром или даклатасвиром были проанализированы исходы лечения 299 пациентов с хроническим гепатитом С, включая 128 ранее неудачно пролеченных интерферонами и рибавирином, а также имеющих прогностически неблагоприятные однонуклеотидные полиморфизмы 39743165T>G (rs8099917) и 39738787C>T (rs12979860) гена интерлейкин-28В. У 57 пациентов был диагностирован цирроз печени, 128 пациентов были ранее неудачно пролечены интерферонами и рибавирином. У 80,9% (242) был первый генотип вируса, у 5% (15) – второй, у 13,7% (41) – третий, у 0,4% (1) – четвертый.В результате исследования было отмечено, что все 299 пациентов, которые придерживались рекомендаций European Association for the Study of the Liver 2015, 2016 и American Association for the Study of the Liver Disease 2017, достигли устойчивого вирусологического ответа через 12 недель после окончания терапии. Приведен клинический случай неудачного лечения, связанный с отсутствием подтверждения элиминации вируса гепатита С при помощи высокочувствительных методов полимеразной цепной реакции и с позднее выявленной аминокислотной заменой в позиции Y93H NS5A (резистентность к ингибиторам NS5A).Мониторинг эффективности терапии необходимо проводить только при помощи высокочувствительных методик полимеразной цепной реакции (от 10 МЕ/мл) и при задержке элиминации вируса у пациентов с продвинутыми стадиями фиброза печени использовать пролонгированные схемы лечения. Для повторного лечения пациентов с наличием мутации Y93Н необходимы новые препараты ингибиторов NS5A или комбинированных препаратов.