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Background: The key issue in the development of novel antimicrobials is a rapid expansion of new bacterial strains resistant to current antibiotics. Indeed, World Health Organization has reported that bacteria commonly causing infections in hospitals and in the community, e.g. E. Coli, K. pneumoniae and S. aureus, have high resistance vs the last generations of cephalosporins, carbapenems and fluoroquinolones. During the past decades, only few successful efforts to develop and launch new antibacterial medications have been performed. This study aims to identify new class of antibacterial agents using novel high-throughput screening technique. Methods: We have designed library containing 125K compounds not similar in structure (Tanimoto coeff.< 0.7) to that published previously as antibiotics. The HTS platform based on double reporter system pDualrep2 was used to distinguish between molecules able to block translational machinery or induce SOS-response in a model E. coli system. MICs for most active chemicals in LB and M9 medium were determined using broth microdilution assay. Results: In an attempt to discover novel classes of antibacterials, we performed HTS of a large-scale small molecule library using our unique screening platform. This approach permitted us to quickly and robustly evaluate a lot of compounds as well as to determine the mechanism of action in the case of compounds being either translational machinery inhibitors or DNA-damaging agents/replication blockers. HTS has resulted in several new structural classes of molecules exhibiting an attractive antibacterial activity. Herein, we report as promising antibacterials. Two most active compounds from this series showed MIC value of 1.2 (5) and 1.8 μg/mL (6) and good selectivity index. Compound 6 caused RFP induction and low SOS response. In vitro luciferase assay has revealed that it is able to slightly inhibit protein biosynthesis. Compound 5 was tested on several archival strains and exhibited slight activity against gram-negative bacteria and outstanding activity against S. aureus. The key structural requirements for antibacterial potency were also explored. We found, that the unsubstituted carboxylic group is crucial for antibacterial activity as well as the presence of bulky hydrophobic substituents at phenyl fragment. Conclusion: The obtained results provide a solid background for further characterization of the 5'- (carbonylamino)-2,3'-bithiophene-4'-carboxylate derivatives discussed herein as new class of antibacterials and their optimization campaign.

Aim and Objective:Antibiotic resistance is a serious constraint to the development of new effective antibacterials. Therefore, the discovery of the new antibacterials remains one of the main challenges in modern medicinal chemistry. This study was undertaken to identify novel molecules with antibacterial activity.Materials and Methods:Using our unique double-reporter system, in-house large-scale HTS campaign was conducted for the identification of antibacterial potency of small-molecule compounds. The construction allows us to visually assess the underlying mechanism of action. After the initial HTS and rescreen procedure, luciferase assay, C14-test, determination of MIC value and PrestoBlue test were carried out.Results:HTS rounds and rescreen campaign have revealed the antibacterial activity of a series of Nsubstituted triazolo-azetidines and their isosteric derivatives that has not been reported previously. Primary hit-molecule demonstrated a MIC value of 12.5 µg/mL against E. coli Δ tolC with signs of translation blockage and no SOS-response. Translation inhibition (26%, luciferase assay) was achieved at high concentrations up to 160 µg/mL, while no activity was found using C14-test. The compound did not demonstrate cytotoxicity in the PrestoBlue assay against a panel of eukaryotic cells. Within a series of direct structural analogues bearing the same or bioisosteric scaffold, compound 2 was found to have an improved antibacterial potency (MIC=6.25 µg/mL) close to Erythromycin (MIC=2.5-5 µg/mL) against the same strain. In contrast to the parent hit, this compound was more active and selective, and provided a robust IP position.Conclusion:N-substituted triazolo-azetidine scaffold may be used as a versatile starting point for the development of novel active and selective antibacterial compounds.

The present report describes our efforts to identify new structural classes of compounds having promising antibacterial activity using previously published double-reporter system pDualrep2. This semi-automated high-throughput screening (HTS) platform has been applied to perform a large-scale screen of a diverse small-molecule compound library. We have selected a set of more than 125,000 molecules and evaluated them for their antibacterial activity. On the basis of HTS results, eight compounds containing 2-pyrazol-1-yl-thiazole scaffold exhibited moderate-to-high activity against ΔTolC Escherichia coli. Minimum inhibitory concentration (MIC) values for these molecules were in the range of 0.037–8 μg ml−1. The most active compound 8 demonstrated high antibacterial potency (MIC = 0.037 μg ml−1), that significantly exceed that measured for erythromycin (MIC = 2.5 μg ml−1) and was comparable with the activity of levofloxacin (MIC = 0.016 μg ml−1). Unfortunately, this compound showed only moderate selectivity toward HEK293 eukaryotic cell line. On the contrary, compound 7 was less potent (MIC = 0.8 μg ml−1) but displayed only slight cytotoxicity. Thus, 2-pyrazol-1-yl-thiazoles can be considered as a valuable starting point for subsequent optimization and morphing.

Paramushir virus belongs to Sakhalin virus genogroup within Orthonairovirus genus and is one of the poorly studied viruses with unknown pathogenicity. At the moment, only one nearly complete sequence of Paramushir virus genome, isolated in 1972, is available. Two new strains of PARV were isolated in 2015 from a sample collected at the Tyuleniy Island in the Okhotsk Sea and sequenced using a combination of high throughput sequencing and specific multiplex PCR. Both strains are closely related to the early sequenced PARV strain LEIV–1149 K. The signs of intersegment reassortment and probable recombination were revealed, which point to a high variability potential of Paramushir virus and may lead to the formation of strains with novel properties, different from those of the predecessors. The new data regarding Paramushir virus can promote a better understanding of the diversity and relations within Orthonairovirus genus and help define intragenic demarcation criteria, which have not yet been established.

SARS-CoV-2 has spread rapidly across the globe, with most nations failing to prevent or substantially delay its introduction. While many countries have imposed some limitations on trans-border passenger traffic, the effect of these measures on the spread of COVID-19 strains remains unclear. Here, we report an analysis of whole-genome sequencing of 3206 SARS-CoV-2 samples from 78 regions of Russia covering the period between March and November 2020. We describe recurring imports of multiple COVID-19 strains throughout this period, giving rise to 457 uniquely Russian transmission lineages, as well as repeated cross-border transmissions of local circulating variants out of Russia.

The rapid development of high-throughput sequencing (HTS) methods offers new opportunities for food quality control and identification of food components using the DNA barcoding approach (metabarcoding in cases of complex mixes). However, the protocols of DNA barcoding applied to food analysis are not yet fully established; testing and optimization are required to achieve the highest accuracy and cost efficiency. We report here a comparative study of the two most widely used sequencing platforms - Illumina and Ion Torrent - for composition analysis of herbal teas, and show that both technologies yield congruent results, both qualitatively and quantitatively. They have revealed the substitution of fireweed (Epilobium angustifolium L.) by Lythrum sp. in one of the samples. It was confirmed by classic methods of botanical analysis (anatomy and palynology). In most samples, undeclared components have been detected, such as bindweed (Convolvulus) and ragweed (Ambrosia), which are known toxic and allergy-causing plants.

Advances in the next generation sequencing (NGS) technologies have significantly increased our ability to detect new viral pathogens and systematically determine the spectrum of viruses prevalent in various biological samples. In addition, this approach has also helped in establishing the associations of viromes with many diseases. However, unlike the metagenomic studies using16SrRNA for the detection of bacteria, it is impossible to create universal oligonucleotides to target all known and novel viruses, owing to their genomic diversity and variability. On the other hand, sequencing the entire genome is still expensive and has relatively low sensitivity for such applications. The existing approaches for the design of oligonucleotides for targeted enrichment are usually involved in the development of primers for the PCR-based detection of particular viral species or genera, but not for families or higher taxonomic orders. In this study, we have developed a computational pipeline for designing the oligonucleotides capable of covering a significant number of known viruses within various taxonomic orders, as well as their novel variants. We have subsequently designed a genus-specific oligonucleotide panel for targeted enrichment of viral nucleic acids in biological material and demonstrated the possibility of its application for virus detection in bird samples. We have tested our panel using a number of collected samples and have observed superior efficiency in the detection and identification of viral pathogens. Since a reliable, bioinformatics-based analytical method for the rapid identification of the sequences was crucial, an NGS-based data analysis module was developed in this study, and its functionality in the detection of novel viruses and analysis of virome diversity was demonstrated.

Plants are widely used for food and beverage preparation, most often in the form of complex mixtures of dried and ground parts, such as teas, spices or herbal medicines. Quality control of such products is important due to the potential health risks from the presence of unlabelled components or absence of claimed ones. A promising approach to analyse such products is DNA metabarcoding due to its high resolution and sensitivity. However, this method’s application in food analysis requires several methodology optimizations in DNA extraction, amplification and library preparation. In this study, we present such optimizations. The most important methodological outcomes are the following: 1) the DNA extraction method greatly influences amplification success, 2) the main problem for the application of metabarcoding is DNA purity, not integrity or quantity, and 3) the “non-amplifiable” samples can be amplified with polymerases resistant to inhibitors. Using this optimized workflow, we analysed a broad set of plant products (teas, spices and herbal remedies) using two NGS platforms. The analysis revealed the problem of both the presence of extraneous components and the absence of labelled ones. Notably, for teas, no correlation was found between the price and either the absence of labelled components or presence of unlabelled ones, for spices, a negative correlation was found between the price and presence of unlabelled components.

A series of 5-oxo-4H-pyrrolo[3,2-b]pyridine derivatives was identified as novel class of highly potent antibacterial agents during an extensive large-scale high-throughput screening (HTS) program utilizing a unique double-reporter system—pDualrep2. The construction of the reporter system allows us to perform visual inspection of the underlying mechanism of action due to two genes—Katushka2S and RFP—which encode the proteins with different imaging signatures. Antibacterial activity of the compounds was evaluated during the initial HTS round and subsequent rescreen procedure. The most active molecule demonstrated a MIC value of 3.35 µg/mL against E. coli with some signs of translation blockage (low Katushka2S signal) and no SOS response. The compound did not demonstrate cytotoxicity in standard cell viability assay. Subsequent structural morphing and follow-up synthesis may result in novel compounds with a meaningful antibacterial potency which can be reasonably regarded as an attractive starting point for further in vivo investigation and optimization.

Introduction: A variety of organic compounds has been reported to have antibacterial activity. However, antimicrobial resistance is one of the main problems of current anti-infective therapy, and the development of novel antibacterials is one of the main challenges of current drug discovery. Methods: Using our previously developed dual-reporter High-Throughput Screening (HTS) platform, we identified a series of furanocoumarins as having high antibacterial activity. The construction of the reporter system allows us to differentiate three mechanisms of action for the active compounds: inhibition of protein synthesis (induction of Katushka2S), DNA damaging (induction of RFP) or other (inhibition of bacterial growth without reporter induction). Results: Two primary hit-molecules of furanocoumarin series demonstrated relatively low MIC values comparable to that observed for Erythromycin (Ery) against E. coli and weakly induced both reporters. Dose-dependent translation inhibition was shown using in vitro luciferase assay, however it was not confirmed using C14-test. A series of close structure analogs of the identified hits was obtained and investigated using the same screening platform. Compound 19 was found to have slightly lower MIC value (15.18 µM) and higher induction of Katushka2S reporter in contrast to the parent structures. Moreover, translation blockage was clearly identified using both in vitro luciferase assay and C14 test. The standard cytotoxicity test revealed a relatively low cytotoxicity of the most active molecules. Conclusion: High antibacterial activity in combination with low cytotoxicity was demonstrated for a series of furanocoumarins. Further optimization of the described structures may result in novel and attractive lead compounds with promising antibacterial efficiency.

Highlights • LASV RT-qPCR diagnostic sensitivity was 96.4% & specificity 100%. • Limit of detection ranged from 103- 105copies/ml with lineages I-IV. • Assay can be used with whole blood & serum samples from humans and rodent tissues., Lassa fever is a severe viral hemorrhagic illness caused by Lassa virus. Based on estimates, the number of LASV infections ranges from 300,000 to 500,000 cases in endemic areas with a fatality rate of 1%. Development of fast and sensitive tools for the control and prevention of Lassa virus infection as well as for clinical diagnostics of Lassa fever are crucial. Here we reported development and evaluation of a one-step quantitative RT-qPCR assay for the Lassa virus detection – LASV-Fl. This assay is suitable for the detection of lineages I-IV of Lassa virus. The limit of detection of the assay ranged from 103 copies/ml to 105 copies/ml and has 96.4% diagnostic sensitivity, whereas analytical and diagnostic specificities both were 100%. Serum, whole blood and tissue are suitable for use with the assay. The assay contains all the necessary components to perform the analysis, including an armored positive control (ARC+) and an armored internal control (IC). The study was done during the mission of specialized anti-epidemic team of the Russian Federation (SAET) in the Republic of Guinea in 2015-2018. Based on sequencing data, LASV-specific assay was developed using synthetic MS2-phage-based armored RNA particles, RNA from Lassa virus strain Josiah, and further, evaluated in field conditions using samples from patients and Mastomys natalensis rodents.

Introduction: Non-structural 5A (NS5A) protein has achieved a considerable attention as an attractive target for the treatment of hepatitis C (HCV). A number of novel NS5A inhibitors have been reported to date. Several drugs having favorable ADME properties and mild side effects were launched into the pharmaceutical market. For instance, daclatasvir was launched in 2014, elbasvir is currently undergoing registration, ledipasvir was launched in 2014 as a fixed-dose combination with sofosbuvir (NS5B inhibitor). Areas covered: Thomson integrity database and SciFinder database were used as a valuable source to collect the patents on small-molecule NS5A inhibitors. All the structures were ranked by the date of priority. Patent holder and antiviral activity for each scaffold claimed were summarized and presented in a convenient manner. A particular focus was placed on the best-in-class bis-pyrrolidine-containing NS5A inhibitors. Expert opinion: Several first generation NS5A inhibitors have recently progressed into advanced clinical trials and showed superior efficacy in reducing viral load in infected subjects. Therapy schemes of using these agents in combination with other established antiviral drugs with complementary mechanisms of action can address the emergence of resistance and poor therapeutic outcome frequently attributed to antiviral drugs.