Exploring Chemical Space to Identify Partial Binders Against hMPV Nucleocapsid Protein.

Human metapneumovirus (hMPV) has gained prominence in recent times as the predominant etiological agent of acute respiratory tract infections. This virus targets children, the elderly, and individuals with compromised immune systems. Given the protracted duration of hMPV transmission, it is probable that the majority of children will have acquired the virus by the age of 5. In individuals with compromised immune systems, recurrence of hMPV infection is possible. As hMPV matures, it remains latent from the time of acquisition. The genome of hMPV encompasses a pivotal protein referred to as the nucleocapsid protein (N). This protein assumes the form of a left-handed helical nucleocapsid, enveloping the viral RNA genome. The primary function of this structure is to protect nucleases, rendering it a potentially promising target for therapeutic advancements. The present study employs a methodology that involves structure-based virtual screening, followed by molecular dynamics simulation at a 250-ns time scale, to identify potential natural molecules or their derivatives from the ZINC Database. These molecules are investigated for their binding properties against the hMPV nucleoprotein. Based on an evaluation of the docking score, binding site interaction, and molecular dynamics studies, it has been found that two naturally occurring molecules, namely M1 (ZINC85629735) and M3 (ZINC85569125), have shown notable docking scores of -9.6 and -10.7 kcal/mol, acceptable RMSD, RMSF, Rg, and so on calculated from molecular dynamics trajectory associated with MMGBSA binding energy of -81.94 and -99.63 kcal/mol, respectively. These molecules have shown the highest binding affinity toward nucleocapsid protein and demonstrated promising attributes as potential binders against hMPV.

Antibacterial and antioxidant assay of novel heteroaryl-substituted methane derivatives synthesized via ceric ammonium nitrate (CAN) catalyzed one-pot green approach.

Individually, hydroxycoumarin and amino pyrimidine derivatives are of significant biological interest owing to their importance in drugs and pharmaceuticals. To access their combined biological impact into one molecule, we designed a novel, one-pot green approach for synthesizing trisubstituted methanes. A series of new heteroaryl-substituted methanes have been synthesized and subjected to in vitro antibacterial and antioxidant evaluation. Tests against clinical isolates of Escherichia coli (gram-negative) and Staphylococcus aureus showed potent activity of the derivatives 4a, 4b, 4d, 4e, 4f, 4l, and 5 against the former, and 4a, 4e, 4j, and 4l against the later one. Further, antioxidant assay for these TRSMs was also studied where 4a, 4b, 4f, 4j, and 4l exhibited the most promising results. These preliminary bioassay evaluations strongly suggest the promise and scope of these molecules in medical science. A one pot methodology for the synthesis of coumarin and uracil tethered trisubstituted methanes has been reported. The synthesized derivatives were further analyzed for their antibacterial and antioxidant properties to explore their medicinal applications. The salient features of this methodology are operational simplicity, short reaction time, good to moderate yields of the products, easy purification method. Biochemical assay of the synthesized TRSMs a reveals their utility in medicinal & pharma industry.

CRISPR-based point-of-care diagnostics incorporating Cas9, Cas12, and Cas13 enzymes advanced for SARS-CoV-2 detection.

An outbreak of the novel beta coronavirus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) first came to light in December 2019, which has unfolded rapidly and turned out to be a global pandemic. Early prognosis of viral contamination involves speedy intervention, disorder control, and good-sized management of the spread of disease. Reverse transcription-polymerase chain reaction, considered the gold standard test for detecting nucleic acids and pathogen diagnosis, provides high sensitivity and specificity. However, reliance on high-priced equipped kits, associated reagents, and skilled personnel slow down sickness detection. Lately, the improvement of clustered regularly interspaced short palindromic repeat (CRISPR)-Cas (CRISPR-associated protein)-based diagnostic systems has reshaped molecular diagnosis due to their low cost, simplicity, speed, efficiency, high sensitivity, specificity, and versatility, which is vital for accomplishing point-of-care diagnostics. We reviewed and summarized CRISPR-Cas-based point-of-care diagnostic strategies and research in these paintings while highlighting their characteristics and challenges for identifying SARS-CoV-2.

Cellular responses to proteostasis perturbations reveal non-optimal feedback in chaperone networks.

The proteostasis network (PN) comprises a plethora of proteins that are dedicated to aid in protein folding and maintenance; some with overlapping functions. Despite this, there are multiple pathophysiological states associated with depletion of chaperones. This is counter-intuitive, assuming cells have the ability to re-program transcriptional outputs in accordance with its proteostasic limitations. Here, we have used S. cerevisiae to understand how cells respond to different types of proteostasis impairments. We monitored the proteostasis status and transcriptome of single deletions of fourteen different Protein Quality Control (PQC) genes. In most cases, cellular response did not activate proteostasis components or pathways that could either complement the function of the missing PQC gene or restore proteostasis. Over-expression of alternate machineries could restore part of the proteostasis defect in two representative PQC gene deletion strains. We posit that S. cerevisiae inherently lacks the ability to sense and respond optimally to defects in proteostasis caused due to deletion of specific PQC components.

Generation of a novel chimeric PALFn antigen of Bacillus anthracis and its immunological characterization in mouse model.

Bacillus anthracis chimeric molecule PALFn, comprising the immunodominant domains of protective antigen (PA) and lethal factor (LF), has been developed in the past and has been shown to confer enhanced protection against anthrax in mouse model when challenged with anthrax lethal toxin (LeTx). However, the immunological correlates for this chimeric antigen, both in terms of humoral as well as cell-mediated immune responses, have not been described in detail. To address this gap, we have determined the immunological responses both at humoral as well as cellular levels for the protection conferred by the novel chimeric antigen PALFn constructed in our laboratory in comparison to PA antigen. The biological functionality of the chimeric antigen was ascertained by the trypsin digestion assay. The trypsin cleavage activated the functionality of PALFn and rendered it to interact and bind with the LF molecule. Similarly, the LFn component in the chimera could independently interact and bind to the trypsin-activated wild-type PA. Further, it was observed that the PALFn-immunized mice sera could readily react to both PA and LF antigens while PA-immunized mice sera showed reaction to PA and PALFn alone and not to the individual LF antigen. The in vitro toxin neutralizing ability of PALFn antisera on macrophage cell line J774.1 was robust but with 1.3-fold lesser titer than PA-immunized antisera. PALFn-immunized mouse splenocytes showed a significant lymphocyte proliferation when stimulated with PALFn. There was a remarkable increase in the level of interleukin-2 (IL-2), interleukin-4 (IL-4), interleukin 10 (IL-10), interferon-γ (IFN- γ), and tumor necrosis factor α (TNFα) from PALFn- and PA-stimulated splenocytes. In addition, there was a significant increase in antigen-specific CD4+ and CD8+ T-cell counts from both PALFn- and PA-immunized mouse splenocytes. The results clearly demonstrate the ability of chimeric molecule PALFn in eliciting robust humoral and cell-mediated immune responses in mouse model that is parallel to the wild-type PA but has additional anti-LF antibody response. Considering the enhanced protection offered by the chimera PALFn, we can conclude that it can be a better alternative to the wild-type PA-based recombinant vaccine against anthrax.

A Simple, Rapid, and Effective Method for Tumor Xenotransplantation Analysis in Transparent Zebrafish Embryos.

In vivo studies of tumor behavior are a staple of cancer research; however, the use of mice presents significant challenges in cost and time. Here, we present larval zebrafish as a transplant model that has numerous advantages over murine models, including ease of handling, low expense, and short experimental duration. Moreover, the absence of an adaptive immune system during larval stages obviates the need to generate and use immunodeficient strains. While established protocols for xenotransplantation in zebrafish embryos exist, we present here an improved method involving embryo staging for faster transfer, survival analysis, and the use of flow cytometry to assess disease burden. Embryos are staged to facilitate rapid cell injection into the yolk of the larvae and cell marking to monitor the consistency of the injected cell bolus. After injection, embryo survival analysis is assessed up to 7 days post injection (dpi). Finally, disease burden is also assessed by marking transferred cells with a fluorescent protein and analysis by flow cytometry. Flow cytometry is enabled by a standardized method of preparing cell suspensions from zebrafish embryos, which could also be used in establishing the primary culture of zebrafish cells. In summary, the procedure described here allows a more rapid assessment of the behavior of tumor cells in vivo with larger numbers of animals per study arm and in a more cost-effective manner.

Enhanced electrochemical performance of Ce-MOF/h-CeO2 composites for high-capacitance energy storage applications.

The escalating demand for energy storage underscores the significance of supercapacitors as devices with extended lifespans, high energy densities, and rapid charge-discharge capabilities. Ceria (CeO2), known for its exceptional properties and dual oxidation states, emerges as a potent material for supercapacitor electrodes. This study enhances its capacitance by integrating it with Metal-Organic Frameworks (MOFs), carbon-rich compounds noted for their good conductivity. In our research, hollow ceria (h-ceria) is synthesized via hydrothermal methods and amalgamated with Ce-MOF, employing 2,6-dinaphthalene dicarboxylic acid as a ligand, to fabricate Ce-MOF@h-CeO2 composites. The structural and morphological characteristics of the composite are methodically examined using X-ray Diffraction (XRD), Field Emission Scanning Electron Microscopy (FE-SEM), and Fourier-Transform Infrared (FT-IR) spectroscopy. The band gap of the materials is ascertained through UV-Diffuse Reflectance Spectroscopy (UV-DRS). Electrochemical behavior and redox properties of the Ce-MOF composites are explored using Cyclic Voltammetry (CV), Galvanostatic Charge and Discharge (GCD), and Electrochemical Impedance Spectroscopy (EIS), providing insights into the material's stability. Electrochemical characterization of the composite reveals maximum specific capacitance, energy density and power density are 2643.78 F g-1 at a scan rate of 10 mV s-1, 249.22 W h kg-1, and 7.9 kW kg-1, respectively. Additionally, the specific capacitance of Ce-MOF synthesized with a 2,6-dinaphthalene dicarboxylic acid (NDC) ligand reaches 995.59 F g-1, surpassing that of Ce-MOF synthesized using a 1,3,5-tricarboxylic acid (H3BTC) ligand. These findings highlight the promising economic potential of high-performance, environmentally sustainable, and cost-effective energy storage devices. The innovative Ce-MOF@h-CeO2 composite materials at the core of this research pave the way for advancing the field of energy storage solutions.

Recent Advancement in Multicomponent Synthesis of Fused Coumarin Derivatives.

The synthesis of coumarin derivatives has been an essential topic since its discovery in 1820. In bioactive compounds, the coumarin moiety serves as a backbone, as many such bioactive compounds with the coumarin moiety play a significant role in their bioactivities. Given this moiety's relevance, several researchers are developing fused-coumarin derivatives to create new drugs. Mostly the approach done for this purpose was a multicomponent reaction based. Over the years, the multicomponent reaction has gained enormous popularity, and this approach has evolved as a replacement for conventional synthetic methods. Because of all these perspectives, we have reported the various fused-coumarin derivatives synthesized using multicomponent reactions in recent years.

Drug Utilization Research and Predictors of Outcomes in the Intensive Care Unit of a Tertiary Care Hospital: A Prospective Observational Study.

BACKGROUND: Multiple drugs are commonly prescribed to intensive care unit (ICU) patients owing to the disease profile, multiple organ dysfunction, prophylaxis, management of stress ulcers, nosocomial infections, etc. This study aimed to evaluate the drug utilization patterns and factors influencing mortality and duration of stay in ICU patients.  Methodology: A prospective observational study was conducted in the ICU of our tertiary care hospital, Postgraduate Institute of Medical Sciences, Rohtak. Data was collected from treatment charts of patients using a structured pretested proforma. World Health Organization Anatomical Therapeutic Chemical/Defined Daily Dose (WHO ATC/DDD) methodology and core prescribing indicators were used to assess drug utilization data. The effect of different variables on mortality and duration of stay in the ICU was evaluated using regression analysis. RESULTS: An average of 8.78 drugs were prescribed per patient. Among the 922 prescriptions, anti-infectives, anti-inflammatory drugs, and drugs acting on the gastrointestinal tract were the most frequent medication classes prescribed. Polypharmacy and trade name prescribing were common. For most of the drugs, the prescribed daily dose corresponded to the WHO-DDD except ceftriaxone and levofloxacin. Age, presence of cardiac disorders, and Glasgow Coma Scale (GCS) score at admission directly correlated with mortality while the use of diuretics had a negative correlation with the duration of ICU stay.  Conclusions: There is a need to rationalize drug therapy in the ICU with regard to limiting polypharmacy and emphasizing generic drug name prescribing and adherence to the essential drug list. Antibiotic prescription patterns, in particular, deserve a special focus keeping in mind the multitude of factors demanding aggressive antibiotic use in critically ill intensive care patients.

Ribosomal protein control of hematopoietic stem cell transformation through direct, non-canonical regulation of metabolism.

We report here that expression of the ribosomal protein, RPL22, is frequently reduced in human myelodysplastic syndrome (MDS) and acute myelogenous leukemia (AML); reduced RPL22 expression is associated with worse outcomes. Mice null for Rpl22 display characteristics of an MDS-like syndrome and develop leukemia at an accelerated rate. Rpl22-deficient mice also display enhanced hematopoietic stem cell (HSC) self-renewal and obstructed differentiation potential, which arises not from reduced protein synthesis but from increased expression of the Rpl22 target, ALOX12, an upstream regulator of fatty acid oxidation (FAO). The increased FAO mediated by Rpl22-deficiency also persists in leukemia cells and promotes their survival. Altogether, these findings reveal that Rpl22 insufficiency enhances the leukemia potential of HSC via non-canonical de-repression of its target, ALOX12, which enhances FAO, a process that may serve as a therapeutic vulnerability of Rpl22 low MDS and AML leukemia cells. Highlights: RPL22 insufficiency is observed in MDS/AML and is associated with reduced survivalRpl22-deficiency produces an MDS-like syndrome and facilitates leukemogenesisRpl22-deficiency does not impair global protein synthesis by HSCRpl22 controls leukemia cell survival by non-canonical regulation of lipid oxidation eTOC: Rpl22 controls the function and transformation potential of hematopoietic stem cells through effects on ALOX12 expression, a regulator of fatty acid oxidation.

Recent Advancement in the One-pot Synthesis of the Tri-substituted Methanes (TRSMs) and their Biological Applications.

The history of tri-substituted methanes (TRSMs) in chemical industries is much older. Tri-substituted methanes were previously used as dyes in the chemical industries. Still, there is a significant surge in researchers' interest in them due to their wide range of bioactivities. Trisubstituted methane derivatives show a wide range of biological activities like anti-tumor, antimicrobial, antibiofilm, antioxidant, anti-inflammatory, anti-arthritic activities. Due to the wide range of medicinal applications shown by tri-substituted methanes, most of the methodologies reported in the literature for the synthesis of TRSMs are focused on the one-pot method. This review explored the recently reported one-pot processes for synthesizing trisubstituted methanes and their various medicinal applications. Based on the substitution attached to the -CH carbon, this review categorizes them into two major classes: (I) symmetrical and (II) unsymmetrical trisubstituted methanes. In addition, this review gives an insight into the growing opportunities for the construction of trisubstituted scaffolds via one-pot methodologies. To the best of our knowledge, no one has yet reported a review on the one-pot synthesis of TRSMs. Therefore, here we present a brief literature review of the synthesis of both symmetrical and unsymmetrical TRSMs covering various one-pot methodologies along with their medicinal applications.

Recent advances in understanding the role of proteostasis.

Maintenance of a functional proteome is achieved through the mechanism of proteostasis that involves precise coordination between molecular machineries assisting a protein from its conception to demise. Although each organelle within a cell has its own set of proteostasis machinery, inter-organellar communication and cell non-autonomous signaling bring forth the multidimensional nature of the proteostasis network. Exposure to extrinsic and intrinsic stressors can challenge the proteostasis network, leading to the accumulation of aberrant proteins or a decline in the proteostasis components, as seen during aging and in several diseases. Here, we summarize recent advances in understanding the role of proteostasis and its regulation in aging and disease, including monogenetic and infectious diseases. We highlight some of the emerging as well as unresolved questions in proteostasis that need to be addressed to overcome pathologies associated with damaged proteins and to promote healthy aging.

Consumers discard a lot more food than widely believed: Estimates of global food waste using an energy gap approach and affluence elasticity of food waste.

This work provides an internationally comparable consumer food waste dataset based on food availability, energy gap and consumer affluence. Such data can be used for constructing meaningful and internationally comparable metrics on food waste, such as those for Sustainable Development Goal 12. The data suggests that consumer food waste follows a linear-log relationship with consumer affluence and starts to emerge when consumers reach a threshold of approximately $6.70/day/capita level of expenditure. These findings also imply that most empirical models overestimate consumption by not accounting for the possibility of food waste in their analysis. The results also show that the most widely cited global estimate of food waste is underestimated by a factor greater than 2 (214 Kcal/day/capita versus 527 Kcal/day/capita). Comparison with estimates of US consumer food waste based on national survey data shows this approach can reasonably reproduce the results without needing extensive data from national surveys.

Uterine composite tumor composed of leiomyosarcoma and embryonal rhabdomyosarcoma with immature cartilage.

SUMMARY: We report a rare composite uterine mesenchymal neoplasm in a 66-year-old woman composed of approximately equal amounts of leiomyosarcoma and embryonal rhabdomyosarcoma, the latter containing foci of immature cartilage. Immunohistochemically, both components were positive with desmin. The leiomyosarcomatous element was positive with alpha smooth muscle actin and h-caldesmon and the rhabdomyosarcomatous component with myogenin and myo-D1. In a review of the literature, we have identified only a single example of a similar case. This is likely to represent rhabdomyosarcomatous differentiation in a leiomyomatous neoplasm or alternatively bidirectional differentiation from an uncommitted mesenchymal stem cell.

In vitro cytotoxic potential of Polyalthia longifolia on human cancer cell lines and induction of apoptosis through mitochondrial-dependent pathway in HL-60 cells.

Polyalthia longifolia is a lofty evergreen tree found in India and Sri Lanka. We are reporting first time the anticancer potential of P. longifolia leaves extract (A001) and its chloroform fraction (F002). Both inhibited cell proliferation of various human cancer cell lines in which colon cancer cells SW-620 showed maximum inhibition with IC(50) value 6.1 microg/ml. Furthermore, F002 induce apoptosis in human leukemia HL-60 cells as measured by several biological end points. F002 induce apoptotic bodies formation, DNA ladder, enhanced annexin-V-FITC binding of the cells, increased sub-G(0) DNA fraction, loss of mitochondrial membrane potential (DeltaPsi(mt)), release of cytochrome c, activation of caspase-9, caspase-3, and cleavage of poly ADP ribose polymerase (PARP) in HL-60 cells. All the above parameters revealed that F002-induced apoptosis through the mitochondrial-dependent pathway in HL-60 cells.

Anthrax lethal toxin (LeTx) neutralization by PA domain specific antisera.

Anthrax associated causalities in humans and animals are implicated mainly due to the action of two exotoxins that are secreted by the bacterium Bacillus antharcis during the infection. These exotoxins comprise of three protein components namely protective antigen (PA), lethal factor (LF) and edema factor (EF). The protective antigen is the common toxin component required to form both lethal toxin (LeTx) and edema toxin (EdTx). The LeTx is formed, when PA combines with LF and EdTx is formed when PA combines with EF. Therapeutic interventions aiming to neutralize these key effectors of anthrax pathology would therefore, provide an effective means to counter the toxicity imposed by the anthrax toxins on the host. The present work describes the lethal toxin neutralization potential of polyclonal antisera developed against the individual domains of the protective antigen component of the anthrax toxin. The individual domains were produced as recombinant proteins in E. coli and validated with peptide mass fingerprinting by MALDI-TOF analysis and corresponding mice polyclonal antisera by western blotting. Each domain specific antibody titre and isotype was ascertained by ELISA. The isotyping revealed the predominance of IgG1 isotype. The toxin neutralizing potential of these domain specific antisera were evaluated by in-vitro cell viability MTT assay, employing J774.1 mouse macrophage cell line against LeTx (0.25 µg ml-1 PA and 0.125 µg ml-1 LF concentrations). Among the four domain specific antisera, the antiserum against PA domain IV could neutralize LeTx with high efficiency. No significant neutralization of LeTx was observed with other domain specific antibodies. Results indicate that antibodies to r-PA domain IV could be explored further as therapeutic anti toxin molecule along with appropriate antibiotic regimens against anthrax.

Soluble Expression and Characterization of Biologically Active Bacillus anthracis Protective Antigen in Escherichia coli.

Bacillus anthracis secretory protein protective antigen (PA) is primary candidate for subunit vaccine against anthrax. Attempts to obtain large quantity of PA from Escherichia coli expression system often result in the formation of insoluble inclusion bodies. Therefore, it is always better to produce recombinant proteins in a soluble form. In the present study, we have obtained biologically active recombinant PA in small scale E. coli shake culture system using three different expression constructs. The PA gene was cloned in expression vectors bearing trc, T5, and T7 promoters and transformed into their respective E. coli hosts. The growth conditions were optimized to obtain maximum expression of PA in soluble form. The expression construct PA-pET32c in DE3-pLysS E. coli host resulted in a maximum production of soluble PA (15 mg L(-1)) compared to other combinations. Purified PA was subjected to trypsin digestion and binding assay with lethal factor to confirm the protein's functionality. Biological activity was confirmed by cytotoxicity assay on J774.1 cells. Balb/c mice were immunized with PA and the immunogenicity was tested by ELISA and toxin neutralization assay. This study highlights the expression of soluble and biologically active recombinant PA in larger quantity using simpler E. coli production platform.

Prevalence of HIV and VDRL seropositivity in blood donors of Delhi.

Blood transfusion has been the transmission mechanism in 15 per cent of total patients infected with human immunodeficiency virus (HIV). A few reports are available regarding the trend of HIV seropositivity in northern India. Prevalence of VDRL (venereal disease research laboratory) reactivity varies from 0.8-15 per cent in blood donors. We present data on the prevalence and trends of infection with HIV 1 and 2 and VDRL reactivity in blood donors in Delhi. Between 2000-2002, a total of 76089 (voluntary and replacement) donors were screened. Majority (82.4%) were replacement donors. Seropositivity for HIV and VDRL was seen in 0.54 and 2.6 per cent of donors respectively. The percentage of seropositivity for HIV and VDRL was significantly higher in replacement donors (P<0.001). It is suggested that extensive donor selection and a voluntary donor service would reduce the number of infectious donors significantly. Non-renumerated repeat voluntary donor services are urgently required to lower the prevalence of transmissible infections. While the need to change to a voluntary donor service and devising effective donor screening cannot be over emphasized, there is also a need to mandate HIV antigen detection in India.

Markers for transfusion-associated hepatitis in north Indian blood donors: prevalence and trends.

Transfusion-associated hepatitis is a great problem in developing countries including India due to endemic hepatitis infections and a lack of voluntary donors, trained personnel, and funds. The prevalence of post-transfusion hepatitis B and C in India is about 1-5% and 1%, respectively. A total of 128,589 blood donors were screened for hepatitis B surface antigen (HBsAg) and 76,089 donors were screened for anti-hepatitis C virus (HCV) from 1997 - 2002. Data were tabulated annually. Out of the total 83.6% were replacement donors. Our study concluded that the prevalence of HBsAg and antibodies for HCV ranged between 1.7 - 2.2% and 0.25 - 0.9%, respectively among all of the donors. Seropositivity was definitely higher in replacement donors than in voluntary donors. Based on these results, we recognize an urgent need to establish a non-remunerated voluntary donor base in India. A stringent deferral system should be developed. The use of sensitive laboratory tests and the addition of core antigen (anti-HBc) to the mandatory screening test list would further reduce the incidence of post-transfusion hepatitis.