Efficient soluble expression and purification of influenza A and B nucleoproteins in E. coli

Viral nucleoprotein (NP) is an abundant essential protein of an influenza virus that has important functional and structural roles. It participates in genomic organization, nuclear trafficking, RNA transcription, and genome replication. From the research point of view, NP is an important protein that is used in the development of new diagnostic methods and vaccination protocols. NP is a promising target for antiviral chemotherapeutic drugs as well. Successful expression of codon-optimized NP genes in E. coli has been reported. In this study, we demonstrated the efficient expression and purification of soluble NPs of influenza A and B viruses in E. coli without the codon-optimization of DNA sequences. This procedure preserves the co-translational protein folding, protein configuration and function. Obtained NPs of influenza A and B viruses were monomers and reacted well with mouse specific antibodies according to Western blot analysis. Our results show that both influenza A and influenza B virus NPs can be efficiently expressed in E. coli without codon-optimization.Viral nucleoprotein (NP) is an abundant essential protein of an influenza virus that has important functional and structural roles. It participates in genomic organization, nuclear trafficking, RNA transcription, and genome replication. From the research point of view, NP is an important protein that is used in the development of new diagnostic methods and vaccination protocols. NP is a promising target for antiviral chemotherapeutic drugs as well. Successful expression of codon-optimized NP genes in E. coli has been reported. In this study, we demonstrated the efficient expression and purification of soluble NPs of influenza A and B viruses in E. coli without the codon-optimization of DNA sequences. This procedure preserves the co-translational protein folding, protein configuration and function. Obtained NPs of influenza A and B viruses were monomers and reacted well with mouse specific antibodies according to Western blot analysis. Our results show that both influenza A and influenza B virus NPs can be efficiently expressed in E. coli without codon-optimization

Эпидемиологические и клинические характеристики острых респираторных инфекций в Санкт-Петербурге в эпидемические сезоны 2017–2018 гг. и 2018–2019 гг.

Objective: to analyze the epidemiological and clinical features of acute respiratory infections occurring during the St. Petersburg 2017–2018 and 2018–2019 epidemic seasons.Materials and methods: the study included 457 patients, treated in St. Petersburg clinics from 2017–2019, displaying symptoms of acute respiratory infection (ARI), including evaluation of their clinical histories. Pathogen types were determined by polymerase chain reaction (PCR). Data analysis was carried out using mathematical statistics methods using the Statistica 10 software package (StatSoft Inc.).Results: in this study, we examined the epidemiological and clinical features of acute respiratory infections in St. Petersburg occurring during two epidemic seasons, 2017–2018 and 2018–2019. The 2017–2018 season was characterized by a prevalence of infections caused by influenza B viruses and influenza A subtype H3N2 viruses. In the 2018–2019 season, there was a greater number of acute respiratory viral infections (ARVIs) and infections caused by influenza A subtype H1N1pdm; influenza B virus was detected only in isolated cases. In the 2017–2018 sore throats and muscle aches were a characteristic symptom of influenza A H1N1pdm infections, of bacterial infections – only sore throats. It was shown that throat pain and vasodilation of the scleral and soft palate vessels were significantly more frequent in the 2017–2018 season, compared to the 2018–2019 season. Cough and redness of the posterior pharyngeal wall were hallmark signs of ARVIs in the 2018–2019 season.Conclusion: according to the data, each epidemic season is characterized not only by its own type-specific acute respiratory infection frequencies, but also by different clinical manifestation frequencies. For global monitoring, treatment effectiveness evaluation, and refined study of acute respiratory infection clinical features, it is advisable to use approaches which incorporate accurate, specific, and rapid molecular biological methods capable of identifying a broad range of pathogens. Цель: сравнительный анализ эпидемиологических данных и клинических характеристик течения острых респираторных инфекций у больных в Санкт-Петербурге в эпидемические сезоны 2017–2018 гг. и 2018–2019 гг.Материалы и методы: в исследование включено 457 пациентов, находившихся на лечении в клиниках Санкт-Петербурга в 2017–2019 гг., с симптомами острых респираторных инфекций, проведен анализ эпидемиологических данных и клинических характеристик течения заболевания. Методом полимеразной цепной реакции определен вид патогена. Статистический анализ проведен с использованием методов математической статистики при помощи пакета Statistica 10, StatSoft Inc.Результаты. В сезон 2017–2018 гг. было характерно превалирование инфекций, вызванных вирусами гриппа В и А (H3N2). В сезоне 2018–2019 гг. было отмечено большее число случаев острых респираторных вирусных инфекций, в том числе гриппа, вызванного вирусом гриппа А (H1N1pdm); вирус гриппа В регистрировали в единичных случаях. Характерным симптомом в сезоне 2017–2018 гг. для гриппа А (H1N1pdm) были мышечные боли и боли в горле, для бактериальных инфекций – также боли в горле. Боли в горле и инъекции сосудов склер и мягкого неба достоверно чаще были отмечены в сезоне 2017–2018 гг. по сравнению с сезоном 2018–2019 гг. Кашель и гиперемия задней стенки глотки были характерными симптомами для ОРВИ в сезоне 2018–2019 гг.Заключение: рассмотренные два эпидемических сезона характеризовались не только различной частотой острых респираторных инфекций, но и различной частотой их клинических проявлений. Для глобального мониторинга, оценки эффективности лечения и более детального изучения клинических особенностей острых респираторных инфекций целесообразно использовать точные, специфичные и быстрые молекулярно-биологические методы с большим количеством определяемых патогенов.

APPLICATION OF SOLID-PHASE, ENZYME-LINKED IMMUNOASSAYS IN MICROARRAY FORMATE FOR QUANTITATIVE DETERMINATION OF CYTOKINES

Abstract. Present work describes a multiplex “sandvich” approach to enzyme-linked, solid-phase immunosorbent assay designed in microarray formate, aiming for quantitative evaluation of individual profile of cytokine expression. Its high specificity, throughput and sensitivity (30 to 20,000 pg/mL) are demonstrated with human IL-2, IL-4, IL-10, and TNF assays. This diagnostic technique may be further used both for research applications and in clinical trials

Epidemiological and clinical features of acute respiratory infections occurring in St. Petersburg during the 2017–2018 and 2018–2019 epidemic seasons

Objective: to analyze the epidemiological and clinical features of acute respiratory infections occurring during the St. Petersburg 2017–2018 and 2018–2019 epidemic seasons.Materials and methods: the study included 457 patients, treated in St. Petersburg clinics from 2017–2019, displaying symptoms of acute respiratory infection (ARI), including evaluation of their clinical histories. Pathogen types were determined by polymerase chain reaction (PCR). Data analysis was carried out using mathematical statistics methods using the Statistica 10 software package (StatSoft Inc.).Results: in this study, we examined the epidemiological and clinical features of acute respiratory infections in St. Petersburg occurring during two epidemic seasons, 2017–2018 and 2018–2019. The 2017–2018 season was characterized by a prevalence of infections caused by influenza B viruses and influenza A subtype H3N2 viruses. In the 2018–2019 season, there was a greater number of acute respiratory viral infections (ARVIs) and infections caused by influenza A subtype H1N1pdm; influenza B virus was detected only in isolated cases. In the 2017–2018 sore throats and muscle aches were a characteristic symptom of influenza A H1N1pdm infections, of bacterial infections – only sore throats. It was shown that throat pain and vasodilation of the scleral and soft palate vessels were significantly more frequent in the 2017–2018 season, compared to the 2018–2019 season. Cough and redness of the posterior pharyngeal wall were hallmark signs of ARVIs in the 2018–2019 season.Conclusion: according to the data, each epidemic season is characterized not only by its own type-specific acute respiratory infection frequencies, but also by different clinical manifestation frequencies. For global monitoring, treatment effectiveness evaluation, and refined study of acute respiratory infection clinical features, it is advisable to use approaches which incorporate accurate, specific, and rapid molecular biological methods capable of identifying a broad range of pathogens

Ceruloplasmin gene expression profile changes in the rat mammary gland during pregnancy, lactation and involution

Copper metabolism disturbances in mammary gland (MG) cells have severe consequences in newborns. The mechanism that controls the balance of copper in the MG has not been thoroughly characterized. Four primary copper homeostasis genes in mammals: (1) ceruloplasmin (Cp) encoding multifunction multicopper blue (ferr)oxidase; (2) CTR1 encoding high affinity copper importer 1; and (3 and 4) two similar genes encoding Cu(I)/Cu(II)-ATPases P1 type (ATP7A and ATP7B) responsible for copper efflux from the cells and metallation of cuproenzymes formed in the Golgi complex are expressed in MG. This study aimed to characterize expression of these genes during pregnancy, lactation and forced involution in the rat MG. We found that Cp anchored to the plasma membrane and ATP7A were expressed during pregnancy and lactation. Soluble Cp and ATP7B were highly expressed in lactating MG decreasing to its ending. CTR1 activity increased during MG growth and reached its maximum at postpartum and then it decreased until the end of lactation. During early forced MG involution, Cp gene expression persisted; while a form of Cp that lacked exon 18 appeared. We suggest that Cp gene expressional changes at the transcriptional and posttranscriptional level reflect various physiological functions of Cp proteins during MG remodeling

MOESM2 of The influenza A virus NS genome segment displays lineage-specific patterns in predicted RNA secondary structure

Additional file 2. List of sequences and RNA second structure for ​Nt 497–564