ОЦЕНКА ИММУНОГЕННОЙ АКТИВНОСТИ КЛОНИРОВАННОГО ШТАММА WA РОТАВИРУСА А ЧЕЛОВЕКА

Introduction. Rotovirus infection (RVI) caused by the dsRNA-containing virus from genus Rotavirus, Reoviridae family, belonging to group A (RVA), is the cause of severe diarrhea in human and other mammalian species. Vaccination is the most effective way to reduce the incidence of RVI. At present, the effectiveness of using gnotobiotic piglets as a universal model for reproducing human rotavirus infection and assessing the quality of RVI vaccine preparations has been experimentally proven. Goals and objectives. Evaluation of immunogenic activity of the cloned RVA Wa strain in the new-born Vietnamese pot-bellied piglets trial. Material and methods. Development of viral preparations of the cloned human Wa strain PBA, development of human RVA rVP6, ELISA, polymerase chain reaction with reverse transcription, immunization and experimental infection of newborn piglets. Results. The article presents the results of the experiment on double immunization of newborn piglets with native virus preparations with the infection activity 5.5 lg TCID50/ml, 3 cm3 per dose, HRV with adjuvant 500 µg per dose and mock preparation (control group) followed with experimental inoculation of all animals with virulent virus strain Wa G1P[8] human RVA with infectious activity of 5.5 lg TCID50/ml in 5 cm3 dose. Development of clinical signs of disease and animal death were observed only in control group. RT-PCR system to detect RVA RNA in rectal swabs, samples of small intestine and peripheral lymph nodes was developed. ELISA based on obtained human RVA rVP6 was developed and results on RVA-specific IgG-antibodies in serum samples of experimental piglets are presented. Conclusion. In the course of the research, a high immunogenic activity of the native and purified virus of the cloned Wa RVA strain Wa was established and the possibility of its use as the main component of the RVI vaccine was confirmed. The possibility of using conventional newborn pigs instead of gnotobiotic piglets as an experimental model was demonstrated.Введение. Ротавирусная инфекция (РВИ), этиологическим агентом которой является дцРНК-содержащий вирус рода Rotavirus семейства Reoviridae, относящийся к группе А (РВА), служит причиной тяжёлых диарей у человека и различных видов млекопитающих. Вакцинопрофилактика - наиболее эффективный способ снижения уровня заболеваемости РВИ. В настоящее время экспериментально доказана эффективность использования поросят-гнотобиотов в качестве универсальной модели для воспроизведения РВИ человека и оценки качества вакцинных препаратов против РВИ. Цели и задачи. Оценка иммуногенной активности клонированного штамма Wa РВА в опыте на новорождённых поросятах вьетнамской вислобрюхой породы. Материал и методы. Получение клонированного штамма Wa РВА человека, rVP6 РВА человека. Иммуноферментный анализ и полимеразная цепная реакция с обратной транскрипцией для выявления РВА человека. Иммунизация и экспериментальное заражение новорожденных поросят. Результаты. В статье представлены результаты опыта по двукратной иммунизации новорождённых поросят препаратами нативного вируса с инфекционной активностью 5,5 lg ТЦИД50/мл в дозе 3 см3, HRV с адъювантом в дозе 500 мкг и плацебо (контроль) с последующим экспериментальным заражением всех животных вирусом вирулентного штамма Wa G1P[8] РВА человека с инфекционной активностью 5,5 lg ТЦИД50/мл в дозе 5 см3. Клинические признаки болезни и гибель наблюдали только у контрольных животных. Разработан метод детекции РНК РВА с помощью ПЦР с обратной транскрипцией (ОТ-ПЦР) в ректальных смывах, пробах тонкого отдела кишечника и периферических лимфатических узлах. На основе полученного rVP6 РВА человека разработан метод выявления специфических IgG-антител к РВА с помощью иммуноферментного анализа (ИФА), и приведены результаты выявления РВА в сыворотке крови подопытных поросят. Заключение. В ходе проведённых исследований установлена высокая иммуногенная активность нативного и очищенного вируса клонированного штамма Wa РВА и подтверждена возможность его применения в качестве основного компонента вакцины против РВИ. Показана возможность использования конвенционных новорождённых поросят вместо поросят-гнотобиотов в качестве опытной модели

The DNA of Bacteria of the World Ocean and the Earth in Cosmic Dust at the International Space Station

Cosmic dust samples from the surface of the illuminator of the International Space Station (ISS) were collected by a crew member during his spacewalk. The sampler with tampon in a vacuum container was delivered to the Earth. Washouts from the tampon's material and the tampon itself were analyzed for the presence of bacterial DNA by the method of nested PCR with primers specific to DNA of the genus Mycobacteria, DNA of the strains of capsular bacteria Bacillus, and DNA encoding 16S ribosomal RNA. The results of amplification followed by sequencing and phylogenetic analysis indicated the presence of the bacteria of the genus Mycobacteria and the extreme bacterium of the genus Delftia in the samples of cosmic dust. It was shown that the DNA sequence of one of the bacteria of the genus Mycobacteria was genetically similar to that previously observed in superficial micro layer at the Barents and Kara seas' coastal zones. The presence of the wild land and marine bacteria DNA on the ISS suggests their possible transfer from the stratosphere into the ionosphere with the ascending branch of the global electric circuit. Alternatively, the wild land and marine bacteria as well as the ISS bacteria may all have an ultimate space origin. © 2018 T. V. Grebennikova et al

The DNA of Bacteria of the World Ocean and the Earth in Cosmic Dust at the International Space Station

Cosmic dust samples from the surface of the illuminator of the International Space Station (ISS) were collected by a crew member during his spacewalk. The sampler with tampon in a vacuum container was delivered to the Earth. Washouts from the tampon's material and the tampon itself were analyzed for the presence of bacterial DNA by the method of nested PCR with primers specific to DNA of the genus Mycobacteria, DNA of the strains of capsular bacteria Bacillus, and DNA encoding 16S ribosomal RNA. The results of amplification followed by sequencing and phylogenetic analysis indicated the presence of the bacteria of the genus Mycobacteria and the extreme bacterium of the genus Delftia in the samples of cosmic dust. It was shown that the DNA sequence of one of the bacteria of the genus Mycobacteria was genetically similar to that previously observed in superficial micro layer at the Barents and Kara seas' coastal zones. The presence of the wild land and marine bacteria DNA on the ISS suggests their possible transfer from the stratosphere into the ionosphere with the ascending branch of the global electric circuit. Alternatively, the wild land and marine bacteria as well as the ISS bacteria may all have an ultimate space origin. © 2018 T. V. Grebennikova et al

Nanodispersions of polyelectrolytes based on humic substances: Isolation, physico-chemical characterization and evaluation of biological activity

Natural polyelectrolytes, including in the form of complexes with colloidal particles, are increasingly used in pharmacy due to the possibility of regulated attachment of medicinal substances and their targeted delivery to the target organ. However, the formation, stability, and molecularmass characteristics of polyelectrolyte nanodispersions (ND) vary depending on the nature and composition of the medium of their origin. This is due to the lack of standardized approaches to quality control and regulatory documentation for most natural ND. In this paper, we first introduced the isolation, followed by investigations into their physico-chemical properties and bioactivity. Using the dried droplet method, we were able to detect the “coffee ring effect”. Fractographic studies of the surface structure of EHA and FA dried samples using SEM showed its heterogeneity and the presence of submicron particles encapsulated in the internal molecular cavities of polyelectrolyte. FTIR spectroscopy revealed the ND chemical structure of benzo-α-pyron and benzo-γ-pyron, consisting of nanoparticles and a branched frame part. The main elements detected by X-ray fluorescence in humic substance extract and fulvic acid include Si, P, S, K, Ca, Mn, Fe, Cu, Zn, whereas Fe is in high concentrations. The UV-spectra and fluorescent radiation demonstrated the possibility of studying the effect of the fulvate chromone structure on its optical properties. It is shown that dilution of the initial solutions of polyelectrolytes 1:10 contributes to the detection of smaller nanoparticles and an increase in the absolute value of the negative ζ-potential as a factor of ND stability. A study of the EHS effect on the SARS-CoV-2 virus infectious titer in the Vero E6 cell showed the effective against virus both in the virucidal scheme (the SI is 11.90–22.43) and treatment/prevention scheme (the SI is 34.85–57.33). We assume that polyelectrolyte ND prevent the binding of the coronavirus spike glycoprotein to the receptor. Taking into account the results obtained, we expect that the developed approach can become unified for the standardization of the ND natural polyelectrolytes complex, which has great prospects for use in pharmacy and medicine as a drug with antiviral activity. © 2021 by the authors. Licensee MDPI, Basel, Switzerland