Expression of the SARS-CoV-2 receptor-binding domain by live attenuated influenza vaccine virus as a strategy for designing a bivalent vaccine against COVID-19 and influenza.

Influenza and SARS-CoV-2 are two major respiratory pathogens that cocirculate in humans and cause serious illness with the potential to exacerbate disease in the event of co-infection. To develop a bivalent vaccine, capable of protecting against both infections, we inserted the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein into hemagglutinin (HA) molecule or into the open reading frame of the truncated nonstructural protein 1 (NS1) of live attenuated influenza vaccine (LAIV) virus and assessed phenotypic characteristics of the rescued LAIV-RBD viruses, as well as their immunogenicity in mouse and Syrian hamster animal models. A panel of 9 recombinant LAIV-RBD viruses was rescued using the A/Leningrad/17 backbone. Notably, only two variants with RBD insertions into the HA molecule could express sufficient quantities of RBD protein in infected MDCK cells. Intranasal immunization of mice induced high levels of anti-influenza antibody responses in all chimeric LAIV-RBD viruses, which was comparable to the LAIV virus vector. The RBD-specific antibody responses were most pronounced in the variant expressing RBD194 fragment as a chimeric HA protein. This candidate was further tested in Syrian hamsters and was shown to be immunogenic and capable of protecting animals against both infections.

Developing Empirical Latent Profiles of Impulsive Aggression and Mood in Youths across Three Outpatient Samples.

OBJECTIVE: Aggression with impulsivity and reactivity (AIR) may distinguish a subset of youth from those with attention problems, rule-breaking behavior, or mood disorders, potentially with differential treatment response. Yet, DSM-5 and ICD-10 do not include an AIR diagnosis. Thus, we empirically grouped youths into profiles based on AIR, manic, depressive, rule-breaking, and self-harm behaviors; examined which profiles replicated across three samples; and characterized profile sets on demographic and clinical features. METHOD: After harmonizing data from three samples (n = 679, n = 392, n = 634), Latent Profile Analysis (LPA) assigned youth to profiles based on caregiver-reported measures of AIR, manic, depressive, rule-breaking, and self-harm behaviors. Profiles from each sample were grouped into sets based on profile similarity. Analyses tested differences in diagnoses, sex, and race, age, functioning, and mood severity. RESULTS: Eight-profile solutions fit best. Seven profiles replicated across samples: high AIR and self-harm, lower depressive and manic scores; high AIR, manic symptoms, and self-harm; high depression symptoms; three smaller sets with high manic and depressive symptoms and moderate AIR; and two high rates of bipolar diagnoses and family bipolar history. Two sets were high on both AIR and mood symptoms, were the most impaired, and had the highest comorbidity. CONCLUSIONS: Analyses support an empirical definition of AIR, separate from mood disorders. Profile sets distinguished by level of AIR and mood symptoms differed in demographic and diagnostic characteristics as well as functioning. Importantly, a set emerged with high AIR but low mood indicators and with high rates of ADHD and ODD, but not mood disorder.

The Effect of Mice Adaptation Process on the Pathogenicity of Influenza A/South Africa/3626/2013 (H1N1)pdm09 Model Strain.

Influenza virus strain A/South Africa/3626/2013 (H1N1)pdm09 (SA-WT) is a non-mouse-adapted model strain that has naturally high pathogenic properties in mice. It has been suggested that the high pathogenicity of this strain for mice could be due to the three strain-specific substitutions in the polymerase complex (Q687R in PB1, N102T in PB2, and E358E/K heterogeneity in PB2). To evaluate the role of these replacements, SA-WT was passaged five times in mouse lungs, and the genome of the mouse-adapted version of the SA-WT strain (SA-M5) was sequenced. SA-M5 lost E358E/K heterogeneity and retained E358, which is the prevalent amino acid at this position among H1N1pdm09 strains. In addition, in the hemagglutinin of SA-M5, two heterogeneous substitutions (G155G/E and S190S/R) were identified. Both viruses, SA-M5 and SA-WT, were compared for their toxicity, ability to replicate, pathogenicity, and immunogenicity in mice. In mice infected with SA-M5 or SA-WT strains, toxicity, virus titer in pulmonary homogenates, and mouse survival did not differ significantly. In contrast, an increase in the immunogenicity of SA-M5 compared to SA-WT was observed. This increase could be due to the substitutions G155G/E and S190S/R in the HA of SA-M5. The prospects for using SA-M5 in studying the immunogenicity mechanisms were also discussed.

Changes in the concentration of EGFR-mutated plasma DNA in the first hours of targeted therapy allow the prediction of tumor response in patients with EGFR-driven lung cancer.

PURPOSE: This study aimed to analyze changes in the plasma concentration of EGFR-mutated circulating tumor DNA (ctDNA) occurring immediately after the start of therapy with EGFR tyrosine kinase inhibitors (TKIs). METHODS: Serial plasma samples were collected from 30 patients with EGFR-driven non-small cell lung cancer before intake of the first tablet and at 0.5, 1, 2, 3, 6, 12, 24, 36 and 48 h after the start of the therapy. The content of EGFR alleles (exon 19 deletions or L858R) in ctDNA was measured by ddPCR. RESULTS: ctDNA was detected at base-line in 25/30 (83%) subjects. Twelve (50%) out of 24 informative patients showed > 25% reduction of the ctDNA content at 48 h time point; all these patients demonstrated disease control after 4 and 8-12 weeks of therapy. The remaining 12 individuals showed either stable content of EGFR-mutated ctDNA (n = 5) or the elevation of ctDNA concentration (n = 7). 10 of 12 patients with elevated or stable ctDNA level achieved an objective response at 4 weeks, but only 5 of 10 evaluable patients still demonstrated disease control at 8-12 weeks (p = 0.032, when compared to the group with ctDNA decrease). The decline of the amount of circulating EGFR mutant copies at 48 h also correlated with longer progression-free survival (14.7 months vs. 8.5 months, p = 0.013). CONCLUSION: Comparison of concentration of EGFR-mutated ctDNA at base-line and at 48 h after the start of therapy is predictive for the duration of TKI efficacy.

Axonal spheroids in neurodegeneration.

Axonal spheroids are bubble-like biological features that form on most degenerating axons, yet little is known about their influence on degenerative processes. Their formation and growth has been observed in response to various degenerative triggers such as injury, oxidative stress, inflammatory factors, and neurotoxic molecules. They often contain cytoskeletal elements and organelles, and, depending on the pathological insult, can colocalize with disease-related proteins such as amyloid precursor protein (APP), ubiquitin, and motor proteins. Initial formation of axonal spheroids depends on the disruption of axonal and membrane tension governed by cytoskeleton structure and calcium levels. Shortly after spheroid formation, the engulfment signal phosphatidylserine (PS) is exposed on the outer leaflet of spheroid plasma membrane, suggesting an important role for axonal spheroids in phagocytosis and debris clearance during degeneration. Spheroids can grow until they rupture, allowing pro-degenerative factors to exit the axon into extracellular space and accelerating neurodegeneration. Though much remains to be discovered in this area, axonal spheroid research promises to lend insight into the etiologies of neurodegenerative disease, and may be an important target for therapeutic intervention. This review summarizes over 100 years of work, describing what is known about axonal spheroid structure, regulation and function.

Synthesis, in vitro antibacterial activity against Mycobacterium tuberculosis, and reverse docking-based target fishing of 1,4-benzoxazin-2-one derivatives.

Seventeen 1,4-benzoxazin-2-ones bearing the enaminone moiety and three of their analogs were tested for the antibacterial activity against Mycobacterium tuberculosis (H37Rv). Minimal bactericidal concentrations (MBCs) were determined after 41 days of incubation by BACTEC. 1,4-Benzoxazin-2-ones bearing the unsubstituted benzo moiety showed the most promising activities (MBC = 5.00 µg/ml). For most active compounds, antibacterial activities were determined daily during the 41 days. The most promising compound showed a bacteriostatic effect at a concentration of 0.31 µg/ml on Day 4 of incubation, 0.62 µg/ml on Day 6, 2.50 µg/ml on Day 9, and 5.00 µg/ml on Day 41. All studied compounds, along with some of their reported analogs, were docked to 35 proteins of M. tuberculosis to find their potent targets in these organisms. As a result of reverse docking, aspartate 1-decarboxylase, panD, was selected as the most appropriate target. Docking of the most active compounds to mutant panD from pyrazinamide-resistant strains of M. tuberculosis implies that they would not be active against these strains. Considering that most of pyrazinamide clinical resistance cases are due to loss-of-function mutations in pyrazinamidase, pncA, compounds from this study could be useful drugs for the treatment of some cases of pyrazinamide-resistant tuberculosis.

Developing and Validating a Definition of Impulsive/Reactive Aggression in Youth.

The goal of this study is to develop a rational data-driven definition of impulsive/reactive aggression and establish distinctions between impulsive/reactive aggression and other common childhood problems. This is a secondary analysis of data from Assessing Bipolar: A Community Academic Blend (ABACAB; N = 636, ages 5-18), Stanley Medical Research Institute N = 392, ages 5-17), and the Longitudinal Assessment of Manic Symptoms (LAMS; N = 679, ages 6-12) studies, which recruited youths seeking outpatient mental health services in academic medical centers and community clinics. Following Jensen et al.'s (2007) procedure, 3 judges independently rated items from several widely used scales in terms of assessing impulsive/reactive aggression. Principal components analyses (PCA) modeled structure of the selected items supplemented by items related to mood symptoms, rule-breaking behavior, and hyperactivity/impulsivity to better define the boundaries between impulsive/reactive aggression and other common childhood symptoms. In the rational item selection process, there was good agreement among the 3 experts who rated items as characterizing impulsive/reactive aggression or not. PCA favored 5 dimension solutions in all 3 samples. Across all samples, PCA resulted in rule-breaking behavior, aggression-impulsive/reactive (AIR), mania, and depression dimensions; there was an additional hyperactive/impulsive dimension in the LAMS sample and a self-harm dimension in ABACAB and Stanley samples. The dimensions demonstrated good internal consistency; criterion validity coefficients also showed consistency across samples. This study is a step toward developing an empirically derived nosology of impulsive aggression/AIR. Findings support the validity of the AIR construct, which can be distinguished from manic and depressive symptoms as well as rule-breaking behavior.

Synthesis of 1,4-benzothiazinones from acylpyruvic acids or furan-2,3-diones and o-aminothiophenol.

Two synthetic approaches to enaminones fused to 1,4-benzothiazin-2-one moiety, which can be interesting in studies on biological activity, chemosensors, and fluorescence, were developed via the reaction of furan-2,3-diones or acylpyruvic acids in the presence of carbodiimides with o-aminothiophenols. The target enaminones were formed together with pharmaceutically interesting 2-hydroxy-2H-1,4-benzothiazin-3(4H)-ones. A selective synthetic approach to 2-hydroxy-2H-1,4-benzothiazin-3(4H)-ones was developed via the solvent-switchable reaction of furan-2,3-diones with o-aminothiophenol. Preliminary biological assays (antimicrobial, acute toxicity) of the new compounds were carried out.

Role of malaria partners in malaria elimination in Armenia.

Malaria control and preventive activities in the countries of the World Health Organization Region for Europe (WHO/EUR) were strengthened within the framework of the Regional Roll Back Malaria strategy adopted by the member-states at the beginning of the 2000s. A political document "From control to malaria elimination" known as the "Tashkent Declaration" was unanimously endorsed by the member-states of the WHO/EUR with malaria problems in 2005. Since then, considerable progress has been achieved in the countries of the region, signified by the dramatic reduction of malaria incidence in conjunction with the prevention of re-establishment of infection on the territories where malaria was eliminated earlier. Several countries of the region had been certified by the WHO as free of local malaria transmission as a result of the activities of their National Malaria Elimination Programme, Armenia being one of the first in 2011. One of the main lessons learnt during the implementation of the activities by the National Malaria Elimination Programme in Armenia was that the development of an operational plan for malaria elimination required a comprehensive national effort. Full support, both political and financial, from the highest levels of government to smooth coordination between different government ministries, such as Agriculture, Defense, Finance, Health and Policy and Planning and others, was a prerequisite for operational success. The role and place of various partners in the achievement of malaria elimination in the country is discussed in this review.

Annulation of 1H-pyrrole-2,3-diones by thioacetamide: an approach to 5-azaisatins.

A novel approach to 1H-pyrrolo[3,2-c]pyridine-2,3-diones (5-azaisatins) has been developed via an unprecedented annulation of pyrrolo[2,1-c][1,4]benzoxazine-1,2,4-triones by thioacetamide. A new way of C-H functionalization of thioacetamide has been discovered. The reaction proceeds under green catalyst-free conditions.

Facile regiodivergent synthesis of spiro pyrrole-substituted pseudothiohydantoins and thiohydantoins via reaction of [e]-fused 1H-pyrrole-2,3-diones with thiourea.

A highly divergent synthesis of regioisomeric thiohydantoins and pseudothiohydantoins spiro-fused to a pharmacologically valuable pyrrole-2-one fragment involving the reaction of [e]-fused 1H-pyrrole-2,3-diones with thioureas was developed. The obtained spiro pseudothiohydantoin derivatives were found to undergo a pseudothiohydantoin-thiohydantoin rearrangement. The reactions were shown to proceed under catalyst-free conditions in good yields, and the products were isolated without applying preparative chromatography methods.

Filter-Based Protein Digestion (FPD): A Detergent-Free and Scaffold-Based Strategy for TMT Workflows.

High-throughput proteome profiling requires thorough optimization to achieve comprehensive analysis. We developed a filter aided sample preparation (FASP)-like, detergent-free method, termed Filter-Based Protein Digestion (FPD). We compared FPD to protein extraction methods commonly used in isobaric tag-based proteome profiling, namely trichloroacetic acid (TCA) and chloroform-methanol (C-M) precipitation. We divided a mammalian whole cell lysate from the SH-SY5Y neuroblastoma cell line for parallel protein processing with TCA (n = 3), C-M (n = 2), and FPD using either 10 kDa (n = 3) or 30 kDa (n = 3) molecular weight cutoff membranes. We labeled each sample with tandem mass tag (TMT) reagents to construct a TMT11-plex experiment. In total, 8654 proteins were quantified across all samples. Pairwise comparisons showed very little deviation for individual protein abundance measurements between the two FPD methods, whereas TCA and FPD showed the most difference. Specifically, membrane proteins were more readily quantified when samples were processed using TCA precipitation than other methods tested. However, globally, only 4% of proteins differed greater than 4-fold in the most divergent pair of protein extraction methods (i.e., FPD10 and TCA). We conclude that the detergent-free FPD strategy, particularly using the faster-flowing 30 kDa filter, is a seamless alteration to high-throughput TMT workflows.

On the epidemiology of Plasmodium vivax malaria: past and present with special reference to the former USSR.

Presently, many malaria-endemic countries in the world are transitioning towards malaria elimination. Out of the 105 countries with ongoing malaria transmission, 10 countries are classified as being in the pre-elimination phase of malaria control, and 9 countries are in the malaria elimination stage, whereas 7 countries are classified as being in the prevention of introduction phase. Between 2000 and 2015, 17 countries eliminated malaria (i.e., attained zero indigenous cases for 3 years or more). Seven countries were certified by the WHO as having successfully eliminated malaria. The purpose of this review was to analyse the epidemiological characteristics of vivax malaria during the various stages of malaria eradication (elimination) programmes in different countries in the past and present. Experiences of the republics of the former USSR with malaria are interesting, particularly since the data overwhelmingly were published in Russian and might not be known to western readers. Among the most important characteristics of Plasmodium vivax epidemiology at present are changes in the ratio of the short-incubation P. vivax to long-incubation P. vivax, the incidence of severe P. vivax cases, the increased numbers of asymptomatic P. vivax cases, the reduced response to anti-malarials and a few others. Various factors contributing towards the peculiarities of P. vivax epidemiology are discussed.

What Is the Fastest Way to Connect Stations to a Wi-Fi HaLow Network?

Wi-Fi HaLow is an adaptation of the widespread Wi-Fi technology for the Internet of Things scenarios. Such scenarios often involve numerous wireless stations connected to a shared channel, and contention for the channel significantly affects the performance in such networks. Wi-Fi HaLow contains numerous solutions aimed at handling the contention between stations, two of which, namely, the Centralized Authentication Control (CAC) and the Distributed Authentication Control (DAC), address the contention reduction during the link set-up process. The link set-up process is special because the access point knows nothing of the connecting stations and its means of control of these stations are very limited. While DAC is self-adaptive, CAC does require an algorithm to dynamically control its parameters. Being just a framework, the Wi-Fi HaLow standard neither specifies such an algorithm nor recommends which protocol, CAC or DAC, is more suitable in a given situation. In this paper, we solve both issues by developing a novel robust close-to-optimal algorithm for CAC and compare CAC and DAC in a vast set of experiments.

Synthesis of pyrimido[1,6-a]quinoxalines via intermolecular trapping of thermally generated acyl(quinoxalin-2-yl)ketenes by Schiff bases.

Acyl(quinoxalin-2-yl)ketenes generated by thermal decarbonylation of 3-acylpyrrolo[1,2-a]quinoxaline-1,2,4(5H)-triones react regioselectively with Schiff bases under solvent-free conditions to form pyrimido[1,6-a]quinoxaline derivatives in good yields.

Elimination of Plasmodium falciparum malaria in Tajikistan.

Malaria was eliminated in Tajikistan by the beginning of the 1960s. However, sporadic introduced cases of malaria occurred subsequently probably as a result of transmission from infected mosquito Anopheles flying over river the Punj from the border areas of Afghanistan. During the 1970s and 1980s local outbreaks of malaria were reported in the southern districts bordering Afghanistan. The malaria situation dramatically changed during the 1990s following armed conflict and civil unrest in the newly independent Tajikistan, which paralyzed health services including the malaria control activities and a large-scale malaria epidemic occurred with more than 400,000 malaria cases. The malaria epidemic was contained by 1999 as a result of considerable financial input from the Government and the international community. Although Plasmodium falciparum constituted only about 5% of total malaria cases, reduction of its incidence was slower than that of Plasmodium vivax. To prevent increase in P. falciparum malaria both in terms of incidence and territory, a P. falciparum elimination programme in the Republic was launched in 200, jointly supported by the Government and the Global Fund for control of AIDS, tuberculosis and malaria. The main activities included the use of pyrethroids for the IRS with determined periodicity, deployment of mosquito nets, impregnated with insecticides, use of larvivorous fishes as a biological larvicide, implementation of small-scale environmental management, and use of personal protection methods by population under malaria risk. The malaria surveillance system was strengthened by the use of ACD, PCD, RCD and selective use of mass blood surveys. All detected cases were timely epidemiologically investigated and treated based on the results of laboratory diagnosis. As a result, by 2009, P. falciparum malaria was eliminated from all of Tajikistan, one year ahead of the originally targeted date. Elimination of P. falciparum also contributed towards speedy reduction of P. vivax incidence in Tajikistan.

Comprehensive Temporal Protein Dynamics during the Diauxic Shift in Saccharomyces cerevisiae.

Yeast (Saccharomyces cerevisiae) has served as a key model system in biology and as a benchmark for "omics" technology. Although near-complete proteomes of log phase yeast have been measured, protein abundance in yeast is dynamic, particularly during the transition from log to stationary phase. Defining the dynamics of proteomic changes during this transition, termed the diauxic shift, is important to understand the basic biology of proliferative versus quiescent cells. Here, we perform temporal quantitative proteomics to fully capture protein induction and repression during the diauxic shift. Accurate and sensitive quantitation at a high temporal resolution and depth of proteome coverage was achieved using TMT10 reagents and LC-MS3 analysis on an Orbitrap Fusion tribrid mass spectrometer deploying synchronous precursor selection. Triplicate experiments were analyzed using the time-course R package and a simple template matching strategy was used to reveal groups of proteins with similar temporal patterns of protein induction and repression. Within these groups are functionally distinct types of proteins such as those of glyoxylate metabolism and many proteins of unknown function not previously associated with the diauxic shift (e.g. YNR034W-A and FMP16). We also perform a dual time-course experiment to determine Hap2-dependent proteins during the diauxic shift. These data serve as an important basic model for fermentative versus respiratory growth of yeast and other eukaryotes and are a benchmark for temporal quantitative proteomics.

Development of Cross-Reactive Live Attenuated Influenza Vaccine Candidates against Both Lineages of Influenza B Virus.

BACKGROUND: Influenza viruses continue to cause a significant social and economic burden globally. Vaccination is recognized as the most effective measure to control influenza. Live attenuated influenza vaccines (LAIVs) are an effective means of preventing influenza, especially among children. A reverse genetics (RG) system is required to rapidly update the antigenic composition of vaccines, as well as to design LAIVs with a broader spectrum of protection. Such a system has been developed for the Russian LAIVs only for type A strains, but not for influenza B viruses (IBV). METHODS: All genes of the B/USSR/60/69 master donor virus (B60) were cloned into RG plasmids, and the engineered B60, as well as a panel of IBV LAIV reassortants were rescued from plasmid DNAs encoding all viral genes. The engineered viruses were evaluated in vitro and in a mouse model. RESULTS: The B60 RG system was successfully developed, which made it possible to rescue LAIV reassortants with the desired antigenic composition, including hybrid strains with hemagglutinin and neuraminidase genes belonging to the viruses from different IBV lineages. The LAIV candidate carrying the HA of the B/Victoria-lineage virus and NA from the B/Yamagata-lineage virus demonstrated optimal characteristics in terms of safety, immunogenicity and cross-protection, prompting its further assessment as a broadly protective component of trivalent LAIV. CONCLUSIONS: The new RG system for B60 MDV allowed the rapid generation of type B LAIV reassortants with desired genome compositions. The generation of hybrid LAIV reassortants with HA and NA genes belonging to the opposite IBV lineages is a promising approach for the development of IBV vaccines with broad cross-protection.

Sarm1-Dependent Metabolic Reprogramming of Schwann Cells Following Nerve Injury.

Schwann cells (SCs) transition into a repair phenotype after peripheral nerve injury, which is crucial for supporting axon regeneration. However, the early SC injury response preceding the repair state remains poorly understood. Here, we demonstrate that Sarm1, a key regulator of axon degeneration, is expressed in SCs and has a critical role in the early SC injury response. Leveraging the fact that Sarm1 deletion impairs the SC transition to the repair state, we used single-nucleus RNA sequencing to compare the transcriptional responses of wild-type and Sarm1 knockout SCs 24 hours after nerve injury. Remarkably, Sarm1-deficient SCs, unlike wild-type SCs, showed increased expression of genes involved in oxidative phosphorylation and the TCA cycle. These findings were functionally validated, revealing that Sarm1 knockout SCs displayed increased mitochondrial respiration in response to injury. Intriguingly, Sarm1 knockout SCs also exhibited enhanced axon protection compared to wild-type SCs in an in vitro model of axon degeneration. We propose that Sarm1 gates the transition of SCs from a protective, oxidative phosphorylation-dependent state (which we term Protection Associated Schwann Cells or PASCs) to a glycolytic, pro-regenerative repair phenotype after injury. Our findings challenge the prevailing view of Sarm1 as an exclusively axon-autonomous regulator of degeneration and reveal a paradigm shift in understanding the role of Sarm1 in the SC injury response, with broad implications for the treatment of peripheral neuropathies and neurodegenerative diseases.