CRISPR editing of CCR5 and HIV-1 facilitates viral elimination in antiretroviral drug-suppressed virus-infected humanized mice.

Treatment of HIV-1ADA-infected CD34+ NSG-humanized mice with long-acting ester prodrugs of cabotegravir, lamivudine, and abacavir in combination with native rilpivirine was followed by dual CRISPR-Cas9 C-C chemokine receptor type five (CCR5) and HIV-1 proviral DNA gene editing. This led to sequential viral suppression, restoration of absolute human CD4+ T cell numbers, then elimination of replication-competent virus in 58% of infected mice. Dual CRISPR therapies enabled the excision of integrated proviral DNA in infected human cells contained within live infected animals. Highly sensitive nucleic acid nested and droplet digital PCR, RNAscope, and viral outgrowth assays affirmed viral elimination. HIV-1 was not detected in the blood, spleen, lung, kidney, liver, gut, bone marrow, and brain of virus-free animals. Progeny virus from adoptively transferred and CRISPR-treated virus-free mice was neither detected nor recovered. Residual HIV-1 DNA fragments were easily seen in untreated and viral-rebounded animals. No evidence of off-target toxicities was recorded in any of the treated animals. Importantly, the dual CRISPR therapy demonstrated statistically significant improvements in HIV-1 cure percentages compared to single treatments. Taken together, these observations underscore a pivotal role of combinatorial CRISPR gene editing in achieving the elimination of HIV-1 infection.

Inheritance and molecular mapping of solitary/cluster fruit-bearing habit in Luffa.

Fruiting behaviour and sex form are important goals for Luffa breeders and this study aimed to shed light upon inheritance patterns for both these traits. The hermaphrodite form of Luffa acutangula (known as Satputia) is an underutilized vegetable with a unique clustered fruiting habit. Its desirable traits, such as plant architecture, earliness, as well as contrasting traits like unique clustered fruiting, bisexual flower, and crossability with Luffa acutangula (monoecious ridge gourd with solitary fruits), make it a potential source for trait improvement and mapping of desirable traits in Luffa. In the present study, we have elucidated the inheritance pattern of fruiting behaviour in Luffa using F2 mapping population generated from a cross between Pusa Nutan (Luffa acutangula, monoecious, solitary fruiting) × DSat-116 (Luffa acutangula, hermaphrodite, cluster fruiting). In F2 generation, the observed distribution of plant phenotypes fitted in the expected ratio of 3:1 (solitary vs cluster) for fruit-bearing habit. This is the first report of monogenic recessive control for cluster fruit-bearing habit in Luffa. Herein, we designate for the first time the gene symbol cl for cluster fruit bearing in Luffa. Linkage analysis revealed that SRAP marker ME10 EM4-280 was linked to the fruiting trait at the distance of 4.6 cM from the Cl locus. In addition, the inheritance pattern of hermaphrodite sex form in Luffa was also studied in the F2 population of Pusa Nutan × DSat-116 that segregated into 9:3:3:1 ratio (monoecious:andromonoecious:gynoecious:hermaphrodite), suggesting a digenic recessive control of hermaphrodite sex form in Luffa, which was further confirmed by the test cross. The inheritance and identification of molecular marker for cluster fruiting trait provides a basis for breeding in Luffa species.

HIV-1 Nef hijacks both exocytic and endocytic pathways of host intracellular trafficking through differential regulation of Rab GTPases.

BACKGROUND: HIV-1 Nef regulates several cellular functions in an infected cell which results in viral persistence and AIDS pathogenesis. The currently understood molecular mechanism(s) underlying Nef-dependent cellular function(s) are unable to explain how events are coordinately regulated in the host cell. Intracellular membranous trafficking maintains cellular homeostasis and is regulated by Rab GTPases - a member of the Ras superfamily. RESULTS: In the current study, we tried to decipher the role of Nef on the Rab GTPases-dependent complex and vesicular trafficking. Expression profiling of Rabs in Nef-expressing cells showed that Nef differentially regulates the expression of individual Rabs in a cell-specific manner. Further analysis of Rabs in HIV-1NL4-3 or ΔNef infected cells demonstrated that the Nef protein is responsible for variation in Rabs expression. Using a panel of competitive peptide inhibitors against Nef, we identified the critical domain of HIV-1 Nef involved in modulation of Rabs expression. The molecular function of Nef-mediated upregulation of Rab5 and Rab7 and downregulation of Rab11 increased the transport of SERINC5 from the cell surface to the lysosomal compartment. Moreover, the Nef-dependent increase in Rab27 expression assists exosome release. Reversal of Rabs expression using competitive inhibitors against Nef and manipulation of Rabs expression reduced viral release and infectivity of progeny virions. CONCLUSION: This study demonstrates that Nef differentially regulates the expression of Rab proteins in HIV-1 infected cells to hijack the host intracellular trafficking, which augments viral replication and HIV-1 pathogenesis. SIGNIFICANCE: Our study emphasized the indispensable role of HIV-1 protein Nef on various aspects of the intracellular trafficking regulated by Rabs GTPases, which explained how HIV-1 Nef may hijack membrane trafficking pathways in infected cells.

Cajanus platycarpus Flavonoid 3'5' Hydroxylase_2 (CpF3'5'H_2) Confers Resistance to Helicoverpa armigera by Modulating Total Polyphenols and Flavonoids in Transgenic Tobacco.

Pod borer Helicoverpa armigera, a polyphagus herbivorous pest, tremendously incurs crop damage in economically important crops. This necessitates the identification and utility of novel genes for the control of the herbivore. The present study deals with the characterization of a flavonoid 3'5' hydroxylase_2 (F3'5'H_2) from a pigeonpea wild relative Cajanus platycarpus, possessing a robust chemical resistance response to H. armigera. Though F3'5'H_2 displayed a dynamic expression pattern in both C. platycarpus (Cp) and the cultivated pigeonpea, Cajanus cajan (Cc) during continued herbivory, CpF3'5'H_2 showed a 4.6-fold increase vis a vis 3-fold in CcF3'5'H_2. Despite similar gene copy numbers in the two Cajanus spp., interesting genic and promoter sequence changes highlighted the stress responsiveness of CpF3'5'H_2. The relevance of CpF3'5'H_2 in H. armigera resistance was further validated in CpF3'5'H_2-overexpressed transgenic tobacco based on reduced leaf damage and increased larval mortality through an in vitro bioassay. As exciting maiden clues, CpF3'5'H_2 deterred herbivory in transgenic tobacco by increasing total flavonoids, polyphenols and reactive oxygen species (ROS) scavenging capacity. To the best of our knowledge, this is a maiden attempt ascertaining the role of F3'5'H_2 gene in the management of H. armigera. These interesting leads suggest the potential of this pivotal branch-point gene in biotic stress management programs.

Bedside Chest Ultrasound in Postoperative Pediatric Cardiac Surgery Patients: Comparison With Bedside Chest Radiography.

OBJECTIVE: The primary objective was to study the degree of agreement between the chest ultrasound (CUS) studies and chest x-ray (CXR) studies in postoperative pediatric cardiac surgical patients regarding the diagnosis of thoracic abnormalities, and also to compare the diagnostic performance of CUS in reference to CXR for the detection of thoracic abnormalities. The secondary objective was to compare the necessity for interventions done on the basis of CUS and CXR findings in the postoperative setting. DESIGN: A prospective observational study. SETTING: At a postoperative pediatric cardiac surgical intensive care unit in a tertiary-care center. PARTICIPANTS: One hundred sixty patients between the age of 2 months to 18 years undergoing elective cardiac surgery for various congenital heart diseases. INTERVENTIONS: After obtaining permission from the institutional ethics committee, 160 pediatric cardiac surgical patients were studied prospectively in the postoperative period. On the day of surgery (postoperative day [POD] 0), bedside CXR was done in the immediate postoperative period. After bedside CXR, CUS examination was performed and then interpreted by the principal investigator. The CXR was interpreted by the surgical team. Provisional diagnosis was made by the principal investigator and surgical team. Any intervention required was decided based on CXR or CUS findings or both. The procedure was repeated in the morning of POD 1. MEASUREMENTS AND MAIN RESULTS: The degree of agreement between CUS studies and CXR studies in detecting abnormalities was evaluated by Cohen's kappa (k) statistics. The diagnostic performance of CUS was compared with that of CXR using sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and diagnostic accuracy. Overall, kappa analysis (k) showed substantial agreement between the findings of the CUS and CXR studies (k = 0.749). The diagnostic performance of CUS, as compared with CXR, was found to have a sensitivity of 96.9%, specificity of 84.75%, PPV of 73.4%, NPV of 98.43%, and diagnostic accuracy of 88.44%. In 94 abnormal findings, the interventions were done based on CUS or CXR findings or both. Overall, there was a substantial agreement (k = 0.787) between CUS and CXR regarding the necessity for interventions. CONCLUSIONS: The degree of agreement between CUS and CXR studies was substantial for atelectasis, interstitial edema, and diaphragmatic weakness. The degree of agreement between CUS and CXR studies was almost perfect for pneumothorax and fair for pleural effusion. More CUS studies detected intrathoracic pathologies than CXR studies. The CUS also detected abnormalities earlier than CXR and was found to be useful for the early institution of intervention therapy in patients with interstitial edema and atelectasis. It would be reasonable to conclude that CUS may be considered in some instances as an alternative to CXR.

CRISPR-Cas9 directed genome engineering for enhancing salt stress tolerance in rice.

Crop productivity in rice is harshly limited due to high concentration of salt in the soil. To understand the intricacies of the mechanism it is important to unravel the key pathways operating inside the plant cell. Emerging state-of-the art technologies have provided the tools to discover the key components inside the plant cell for salt tolerance. Among the molecular entities, transcription factors and/or other important components of sensing and signaling cascades have been the attractive targets and the role of NHX and SOS1 transporters amply described. Not only marker assisted programs but also transgenic approaches by using reverse genetic strategies (knockout or knockdown) or overexpression have been extensively used to engineer rice crop. CRISPR/Cas is an attractive paradigm and provides the feasibility for manipulating several genes simultaneously. Here, in this review we highlight some of the molecular entities that could be potentially targeted for generating rice amenable to sustain growth under high salinity conditions by employing CRISPR/Cas. We also try to address key questions for rice salt stress tolerance other than what is already known.

Humanized Mice for Infectious and Neurodegenerative disorders.

Humanized mice model human disease and as such are used commonly for research studies of infectious, degenerative and cancer disorders. Recent models also reflect hematopoiesis, natural immunity, neurobiology, and molecular pathways that influence disease pathobiology. A spectrum of immunodeficient mouse strains permit long-lived human progenitor cell engraftments. The presence of both innate and adaptive immunity enables high levels of human hematolymphoid reconstitution with cell susceptibility to a broad range of microbial infections. These mice also facilitate investigations of human pathobiology, natural disease processes and therapeutic efficacy in a broad spectrum of human disorders. However, a bridge between humans and mice requires a complete understanding of pathogen dose, co-morbidities, disease progression, environment, and genetics which can be mirrored in these mice. These must be considered for understanding of microbial susceptibility, prevention, and disease progression. With known common limitations for access to human tissues, evaluation of metabolic and physiological changes and limitations in large animal numbers, studies in mice prove important in planning human clinical trials. To these ends, this review serves to outline how humanized mice can be used in viral and pharmacologic research emphasizing both current and future studies of viral and neurodegenerative diseases. In all, humanized mouse provides cost-effective, high throughput studies of infection or degeneration in natural pathogen host cells, and the ability to test transmission and eradication of disease.

COVID-19 ARDS: A Multispecialty Assessment of Challenges in Care, Review of Research, and Recommendations.

Physicians and care providers are familiar with the management of ARDS, however, when it occurs as a sequalae of COVID-19, it has different features and there remains uncertainty on the consensus of management. To answer this question on how it compares and contrasts with ARDS from other causes, the authors reviewed the published literature and management guidelines as well as their own clinical experience while managing patients with COVID-19 ARDS. For research, a PubMed search was conducted on 01.04.2021 using the systematic review filter to identify articles that were published using MeSH terms COVID-19 and ARDS. Systematic reviews or meta-analyses were selected from a systematic search for literature containing diagnostic, prognostic and management strategies in MEDLINE/PubMed. Those were compared and reviewed to the existing practices by the various treating specialists and recommendations were made. Specifically, the COVID-19 ARDS, its risk factors and pathophysiology, lab diagnosis, radiological findings, rational of recommendation of drugs proposed so far, oxygenation and ventilation strategies and the psychological ramifications of the disease were. discussed. Because of the high mortality in mechanically ventilated patients, the above recommendations and findings direct the potential for improvement in the management of patients with COVID-19 ARDS.

The mixed lineage kinase-3 inhibitor URMC-099 improves therapeutic outcomes for long-acting antiretroviral therapy.

During studies to extend the half-life of crystalline nanoformulated antiretroviral therapy (nanoART) the mixed lineage kinase-3 inhibitor URMC-099, developed as an adjunctive neuroprotective agent was shown to facilitate antiviral responses. Long-acting ritonavir-boosted atazanavir (nanoATV/r) nanoformulations co-administered with URMC-099 reduced viral load and the numbers of HIV-1 infected CD4+ T-cells in lymphoid tissues more than either drug alone in infected humanized NOD/SCID/IL2Rγc-/- mice. The drug effects were associated with sustained ART depots. Proteomics analyses demonstrated that the antiretroviral responses were linked to affected phagolysosomal storage pathways leading to sequestration of nanoATV/r in Rab-associated recycling and late endosomes; sites associated with viral maturation. URMC-099 administered with nanoATV induced a dose-dependent reduction in HIV-1p24 and reverse transcriptase activity. This drug combination offers a unique chemical marriage for cell-based viral clearance. From the Clinical Editor: Although successful in combating HIV-1 infection, the next improvement in antiretroviral therapy (nanoART) would be to devise long acting therapy, such as intra-cellular depots. In this report, the authors described the use of nanoformulated antiretroviral therapy given together with the mixed lineage kinase-3 inhibitor URMC-099, and showed that this combination not only prolonged drug half-life, but also had better efficacy. The findings are hoped to be translated into the clinical setting in the future.

Biochemical and biologic characterization of exosomes and microvesicles as facilitators of HIV-1 infection in macrophages.

Exosomes and microvesicles (MV) are cell membranous sacs originating from multivesicular bodies and plasma membranes that facilitate long-distance intercellular communications. Their functional biology, however, remains incompletely understood. Macrophage exosomes and MV isolated by immunoaffinity and sucrose cushion centrifugation were characterized by morphologic, biochemical, and molecular assays. Lipidomic, proteomic, and cell biologic approaches uncovered novel processes by which exosomes and MV facilitate HIV-1 infection and dissemination. HIV-1 was "entrapped" in exosome aggregates. Robust HIV-1 replication followed infection with exosome-enhanced fractions isolated from infected cell supernatants. MV- and exosome-facilitated viral infections are affected by a range of cell surface receptors and adhesion proteins. HIV-1 containing exosomes readily completed its life cycle in human monocyte-derived macrophages but not in CD4(-) cells. The data support a significant role for exosomes as facilitators of viral infection.

Accelerated Neuroimmune Dysfunction in Aged HIV-1-Infected Humanized Mice.

Disordered immunity, aging, human immunodeficiency virus type one (HIV-1) infection, and responses to antiretroviral therapy are linked. However, how each factor is linked with the other(s) remains incompletely understood. It has been reported that accelerated aging, advanced HIV-1 infection, inflammation, and host genetic factors are associated with host cellular, mitochondrial, and metabolic alterations. However, the underlying mechanism remains elusive. With these questions in mind, we used chronically HIV-1-infected CD34-NSG humanized mice (hu-mice) to model older people living with HIV and uncover associations between HIV-1 infection and aging. Adult humanized mice were infected with HIV-1 at the age of 20 weeks and maintained for another 40 weeks before sacrifice. Animal brains were collected and subjected to transcriptomics, qPCR, and immunofluorescence assays to uncover immune disease-based biomarkers. CD4+ T cell decline was associated with viral level and age. Upregulated C1QA, CD163, and CXCL16 and downregulated LMNA and CLU were identified as age-associated genes tied to HIV-1 infection. Ingenuity pathway analysis affirmed links to innate immune activation, pyroptosis signaling, neuroinflammation, mitochondrial dysfunction, cellular senescence, and neuronal dysfunction. In summary, CD34-NSG humanized mice are identified as a valuable model for studying HIV-1-associated aging. Biomarkers of immune senescence and neuronal signaling are both age- and virus-associated. By exploring the underlying biological mechanisms that are linked to these biomarkers, interventions for next generation HIV-1-infected patients can be realized.

The genome of flax (Linum usitatissimum) assembled de novo from short shotgun sequence reads.

Flax (Linum usitatissimum) is an ancient crop that is widely cultivated as a source of fiber, oil and medicinally relevant compounds. To accelerate crop improvement, we performed whole-genome shotgun sequencing of the nuclear genome of flax. Seven paired-end libraries ranging in size from 300 bp to 10 kb were sequenced using an Illumina genome analyzer. A de novo assembly, comprised exclusively of deep-coverage (approximately 94× raw, approximately 69× filtered) short-sequence reads (44-100 bp), produced a set of scaffolds with N(50) =694 kb, including contigs with N(50)=20.1 kb. The contig assembly contained 302 Mb of non-redundant sequence representing an estimated 81% genome coverage. Up to 96% of published flax ESTs aligned to the whole-genome shotgun scaffolds. However, comparisons with independently sequenced BACs and fosmids showed some mis-assembly of regions at the genome scale. A total of 43384 protein-coding genes were predicted in the whole-genome shotgun assembly, and up to 93% of published flax ESTs, and 86% of A. thaliana genes aligned to these predicted genes, indicating excellent coverage and accuracy at the gene level. Analysis of the synonymous substitution rates (K(s) ) observed within duplicate gene pairs was consistent with a recent (5-9 MYA) whole-genome duplication in flax. Within the predicted proteome, we observed enrichment of many conserved domains (Pfam-A) that may contribute to the unique properties of this crop, including agglutinin proteins. Together these results show that de novo assembly, based solely on whole-genome shotgun short-sequence reads, is an efficient means of obtaining nearly complete genome sequence information for some plant species.

HIV-1 Tat-mediated induction of platelet-derived growth factor in astrocytes: role of early growth response gene 1.

HIV-associated neurologic disorders (HAND) are estimated to affect almost 60% of HIV-infected individuals. HIV encephalitis, the pathologic correlate of the most severe form of HAND, is often characterized by glial activation, cytokine-chemokine dysregulation, and neuronal damage and loss. However, the severity of HIV encephalitis correlates better with glial activation rather than viral load. Using the macaque model, it has been demonstrated that SIV encephalitis correlates with increased expression of the mitogen platelet-derived growth factor (PDGF) B chain in the brain. The goal of this study was to explore the role of PDGF-B chain in HIV-associated activation and proliferation of astrocytes. Specifically, the data demonstrate that exposure of rat and human astrocytes to the HIV-1 protein Tat resulted in the induction of PDGF at both the mRNA and protein levels. Furthermore, PDGF-BB induction was regulated by activation of ERK1/2 and JNK signaling pathways and the downstream transcription factor early growth response 1. Chromatin immunoprecipitation assays demonstrated binding of Egr-1 to the PDGF-B promoter. Exposure of astrocytes to PDGF-BB in turn led to increased proliferation and the release of proinflammatory cytokines MCP-1 and IL-1ß. Because astrogliosis is linked to disease severity, understanding its regulation by PDGF-BB could aid in the development of therapeutic intervention strategies for HAND.

Cloning and Molecular Characterization of the phlD Gene Involved in the Biosynthesis of "Phloroglucinol", a Compound with Antibiotic Properties from Plant Growth Promoting Bacteria Pseudomonas spp.

phlD is a novel kind of polyketide synthase involved in the biosynthesis of non-volatile metabolite phloroglucinol by iteratively condensing and cyclizing three molecules of malonyl-CoA as substrate. Phloroglucinol or 2,4-diacetylphloroglucinol (DAPG) is an ecologically important rhizospheric antibiotic produced by pseudomonads; it exhibits broad spectrum anti-bacterial and anti-fungal properties, leading to disease suppression in the rhizosphere. Additionally, DAPG triggers systemic resistance in plants, stimulates root exudation, as well as induces phyto-enhancing activities in other rhizobacteria. Here, we report the cloning and analysis of the phlD gene from soil-borne gram-negative bacteria-Pseudomonas. The full-length phlD gene (from 1078 nucleotides) was successfully cloned and the structural details of the PHLD protein were analyzed in-depth via a three-dimensional topology and a refined three-dimensional model for the PHLD protein was predicted. Additionally, the stereochemical properties of the PHLD protein were analyzed by the Ramachandran plot, based on which, 94.3% of residues fell in the favored region and 5.7% in the allowed region. The generated model was validated by secondary structure prediction using PDBsum. The present study aimed to clone and characterize the DAPG-producing phlD gene to be deployed in the development of broad-spectrum biopesticides for the biocontrol of rhizospheric pathogens.

Humanized Mice for Studies of HIV-1 Persistence and Elimination.

A major roadblock to achieving a cure for human immunodeficiency virus type one (HIV-1) is the persistence of latent viral infections in the cells and tissue compartments of an infected human host. Latent HIV-1 proviral DNA persists in resting memory CD4+ T cells and mononuclear phagocytes (MPs; macrophages, microglia, and dendritic cells). Tissue viral reservoirs of both cell types reside in the gut, lymph nodes, bone marrow, spleen, liver, kidney, skin, adipose tissue, reproductive organs, and brain. However, despite the identification of virus-susceptible cells, several limitations persist in identifying broad latent reservoirs in infected persons. The major limitations include their relatively low abundance, the precise identification of latently infected cells, and the lack of biomarkers for identifying latent cells. While primary MP and CD4+ T cells and transformed cell lines are used to interrogate mechanisms of HIV-1 persistence, they often fail to accurately reflect the host cells and tissue environments that carry latent infections. Given the host specificity of HIV-1, there are few animal models that replicate the natural course of viral infection with any precision. These needs underlie the importance of humanized mouse models as both valuable and cost-effective tools for studying viral latency and subsequently identifying means of eliminating it. In this review, we discuss the advantages and limitations of humanized mice for studies of viral persistence and latency with an eye toward using these models to test antiretroviral and excision therapeutics. The goals of this research are to use the models to address how and under which circumstances HIV-1 latency can be detected and eliminated. Targeting latent reservoirs for an ultimate HIV-1 cure is the task at hand.

Phylogenomic Analysis of micro-RNA Involved in Juvenile to Flowering-Stage Transition in Photophilic Rice and Its Sister Species.

Vegetative to reproductive phase transition in phototropic plants is an important developmental process and is sequentially mediated by the expression of micro-RNA MIR172. To obtain insight into the evolution, adaptation, and function of MIR172 in photophilic rice and its wild relatives, we analyzed the genescape of a 100 kb segment harboring MIR172 homologs from 11 genomes. The expression analysis of MIR172 revealed its incremental accumulation from the 2-leaf to 10-leaf stage, with maximum expression coinciding with the flag-leaf stage in rice. Nonetheless, the microsynteny analysis of MIR172s revealed collinearity within the genus Oryza, but a loss of synteny was observed in (i) MIR172A in O. barthii (AA) and O. glaberima (AA); (ii) MIR172B in O. brachyantha (FF); and (iii) MIR172C in O. punctata (BB). Phylogenetic analysis of precursor sequences/region of MIR172 revealed a distinct tri-modal clade of evolution. The genomic information generated in this investigation through comparative analysis of MIRNA, suggests mature MIR172s to have evolved in a disruptive and conservative mode amongst all Oryza species with a common origin of descent. Further, the phylogenomic delineation provided an insight into the adaptation and molecular evolution of MIR172 to changing environmental conditions (biotic and abiotic) of phototropic rice through natural selection and the opportunity to harness untapped genomic regions from rice wild relatives (RWR).

The plant specialized metabolite epicatechin- 3-gallate (EC3G) perturbs lipid metabolism and attenuates fat accumulation in pigeonpea pod borer, Helicoverpa armigera.

Control of pod borer Helicoverpa armigera, a notorious polyphagous pest requires paramount attention with focus on environment-friendly management approaches. Overproduction of catechins (epigallocatechin-EGC and epicatechin-3-gallate-EC3G) in the pod borer-resistant pigeonpea wild relative, Cajanus platycarpus during continued herbivory prodded us to assess their underlying molecular effect on H. armigera. Significant reduction in larval and pupal growth parameters was observed when reared on artificial diet incorporated with 100 ppm EC3G vis a vis 100 ppm EGC and EGC + EC3G. Comparative RNAseq analyses of larvae that fed on normal and EC3G-incorporated diet revealed 62 differentially expressed genes dominated by detoxification and lipid metabolism. While lipase and fatty acid-binding protein 2-like were up-regulated, delta9-FADS-like involved in fatty acid synthesis was downregulated, indicating effect of EC3G on fat metabolism. Validation of RNAseq data by qPCR; midgut glutathione-S-transferase and esterase assays depicted increased lipolysis and reduced lipogenesis in EC3G-fed larvae. Additionally, differential accumulation of stearic acid and oleic acid in EC3G-fed and control larvae/adults ascertained perturbation in lipogenesis. Supported by modelling, molecular docking and simulations, we demonstrate the possible involvement of the insect adipokinetic hormone receptor (AKHR) in the EC3G-mediated response. The study demonstrates plant specialized metabolite EC3G as a potential candidate for H. armigera control.

Drought and Oxidative Stress in Flax (Linum usitatissimum L.) Entails Harnessing Non-Canonical Reference Gene for Precise Quantification of qRT-PCR Gene Expression.

Flax (Linum usitatissimum L.) is a self-pollinating, annual, diploid crop grown for multi-utility purposes for its quality oil, shining bast fiber, and industrial solvent. Being a cool (Rabi) season crop, it is affected by unprecedented climatic changes such as high temperature, drought, and associated oxidative stress that, globally, impede its growth, production, and productivity. To precisely assess the imperative changes that are inflicted by drought and associated oxidative stress, gene expression profiling of predominant drought-responsive genes (AREB, DREB/CBF, and ARR) was carried out by qRT-PCR. Nevertheless, for normalization/quantification of data obtained from qRT-PCR results, a stable reference gene is mandatory. Here, we evaluated a panel of four reference genes (Actin, EF1a, ETIF5A, and UBQ) and assessed their suitability as stable reference genes for the normalization of gene expression data obtained during drought-induced oxidative stress in flax. Taking together, from the canonical expression of the proposed reference genes in three different genotypes, we report that EF1a as a stand-alone and EF1a and ETIF5A in tandem are suitable reference genes to be used for the real-time visualization of cellular impact of drought and oxidative stress on flax.

Multipolymer microsphere delivery of SARS-CoV-2 antigens.

Effective antigen delivery facilitates antiviral vaccine success defined by effective immune protective responses against viral exposures. To improve severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) antigen delivery, a controlled biodegradable, stable, biocompatible, and nontoxic polymeric microsphere system was developed for chemically inactivated viral proteins. SARS-CoV-2 proteins encapsulated in polymeric microspheres induced robust antiviral immunity. The viral antigen-loaded microsphere system can preclude the need for repeat administrations, highlighting its potential as an effective vaccine. STATEMENT OF SIGNIFICANCE: Successful SARS-CoV-2 vaccines were developed and quickly approved by the US Food and Drug Administration (FDA). However, each of the vaccines requires boosting as new variants arise. We posit that injectable biodegradable polymers represent a means for the sustained release of emerging viral antigens. The approach offers a means to reduce immunization frequency by predicting viral genomic variability. This strategy could lead to longer-lasting antiviral protective immunity. The current proof-of-concept multipolymer study for SARS-CoV-2 achieve these metrics.

Generation of High-Value Genomic Resource in Rice: A "Subgenomic Library" of Low-Light Tolerant Rice Cultivar Swarnaprabha.

Rice is the major staple food crop for more than 50% of the world's total population, and its production is of immense importance for global food security. As a photophilic plant, its yield is governed by the quality and duration of light. Like all photosynthesizing plants, rice perceives the changes in the intensity of environmental light using phytochromes as photoreceptors, and it initiates a morphological response that is termed as the shade-avoidance response (SAR). Phytochromes (PHYs) are the most important photoreceptor family, and they are primarily responsible for the absorption of the red (R) and far-red (FR) spectra of light. In our endeavor, we identified the morphological differences between two contrasting cultivars of rice: IR-64 (low-light susceptible) and Swarnaprabha (low-light tolerant), and we observed the phenological differences in their growth in response to the reduced light conditions. In order to create genomic resources for low-light tolerant rice, we constructed a subgenomic library of Swarnaprabha that expedited our efforts to isolate light-responsive photoreceptors. The titer of the library was found to be 3.22 × 105 cfu/mL, and the constructed library comprised clones of 4-9 kb in length. The library was found to be highly efficient as per the number of recombinant clones. The subgenomic library will serve as a genomic resource for the Gramineae community to isolate photoreceptors and other genes from rice.