Envelope Protein-Targeting Zika Virus Entry Inhibitors.

Zika virus (ZIKV; family, Flaviviridae), which causes congenital Zika syndrome, Guillain-Barré Syndrome, and other severe diseases, is transmitted mainly by mosquitoes; however, the virus can be transmitted through other routes. Among the three structural and seven nonstructural proteins, the surface envelope (E) protein of ZIKV plays a critical role in viral entry and pathogenesis, making it a key target for the development of effective entry inhibitors. This review article describes the life cycle, genome, and encoded proteins of ZIKV, illustrates the structure and function of the ZIKV E protein, summarizes E protein-targeting entry inhibitors (with a focus on those based on natural products and small molecules), and highlights challenges that may potentially hinder the development of effective inhibitors of ZIKV infection. Overall, the article will provide useful guidance for further development of safe and potent ZIKV entry inhibitors targeting the viral E protein.

A Unique mRNA Vaccine Elicits Protective Efficacy against the SARS-CoV-2 Omicron Variant and SARS-CoV.

The highly pathogenic coronaviruses SARS-CoV-2 and SARS-CoV have led to the COVID-19 pandemic and SARS outbreak, respectively. The receptor-binding domain (RBD) of the spike (S) protein of SARS-CoV-2, particularly the Omicron variant, has frequent mutations, resulting in the reduced efficiency of current COVID-19 vaccines against new variants. Here, we designed two lipid nanoparticle-encapsulated mRNA vaccines by deleting the mutant RBD of the SARS-CoV-2 Omicron variant (SARS2-S (RBD-del)) or by replacing this mutant RBD with the conserved and potent RBD of SARS-CoV (SARS2-S (SARS-RBD)). Both mRNA vaccines were stable at various temperatures for different time periods. Unlike SARS2-S (RBD-del) mRNA, SARS2-S (SARS-RBD) mRNA elicited effective T-cell responses and potent antibodies specific to both SARS-CoV-2 S and SARS-CoV RBD proteins. It induced strong neutralizing antibodies against pseudotyped SARS-CoV-2 and SARS-CoV infections and protected immunized mice from the challenge of the SARS-CoV-2 Omicron variant and SARS-CoV by significantly reducing the viral titers in the lungs after Omicron challenge and by completely preventing SARS-CoV-induced weight loss and death. SARS2-S (SARS-RBD)-immunized serum antibodies protected naïve mice from the SARS-CoV challenge, with its protective efficacy positively correlating with the neutralizing antibody titers. These findings indicate that this mRNA vaccine has the potential for development as an effective vaccine against current and future SARS-CoV-2 variants and SARS-CoV.

Successful interventional treatment of huge hepatic haemangioma in a neonate following failed medical approach.

An outborn male term neonate presented with a complaint of respiratory distress since birth on day 9 of life. On examination, baby was having tachypnoea, tachycardia and hepatomegaly. The baby was delivered at term gestation and cried immediately after birth. The chest X-ray showed cardiomegaly. The abdomen ultrasound showed a complex cystic vascular lesion suggestive of hepatic haemangioma. The echocardiography showed an atrial septal defect. The baby was initially treated conservatively along with specific treatment (steroids and propranolol) for haemangioma for 6 weeks. However, the symptoms persisted and there was non-resolution, therefore, particle embolisation of the right hepatic artery was performed. Subsequently, it resulted in complete resolution of the lesion.

Understanding the mechanism of pathogenicity through interactome studies between Arachis hypogaea L. and Aspergillus flavus.

Aspergillus flavus (A. flavus) infects the peanut seeds during pre-and post-harvest stages, causing seed quality destruction for humans and livestock consumption. Even though many resistant varieties were developed, the molecular mechanism of defense interactions of peanut against A. flavus still needs further investigation. Hence, an interologous host-pathogen protein interaction (HPPI) network was constructed to understand the subcellular level interaction mechanism between peanut and A. flavus. Out of the top 10 hub proteins of both organisms, protein phosphatase 2C and cyclic nucleotide-binding/kinase domain-containing protein and different ribosomal proteins were identified as candidate proteins involved in defense. Functional annotation and subcellular localization based characterization of HPPI identified protein SGT1 homolog, calmodulin and Rac-like GTP-binding proteins to be involved in defense response against fungus. The relevance of HPPI in infectious conditions was assessed using two transcriptome data which identified the interplay of host kinase class R proteins, bHLH TFs and cell wall related proteins to impart resistance against pathogen infection. Further, the pathogenicity analysis identified glycogen phosphorylase and molecular chaperone and allergen Mod-E/Hsp90/Hsp1 as potential pathogen targets to enhance the host defense mechanism. Hence, the computationally predicted host-pathogen PPI network could provide valuable support for molecular biology experiments to understand the host-pathogen interaction. SIGNIFICANCE: Protein-protein interactions execute significant cellular interactions in an organism and are influenced majorly by stress conditions. Here we reported the host-pathogen protein-protein interaction between peanut and A. flavus, and a detailed network analysis based on function, subcellular localization, gene co-expression, and pathogenicity was performed. The network analysis identified key proteins such as host kinase class R proteins, calmodulin, SGT1 homolog, Rac-like GTP-binding proteins bHLH TFs and cell wall related to impart resistance against pathogen infection. We observed the interplay of defense related proteins and cell wall related proteins predominantly, which could be subjected to further studies. The network analysis described in this study could be applied to understand other host-pathogen systems generally.

Computational identification and characterization of vascular wilt pathogen (Fusarium oxysporum f. sp. lycopersici) CAZymes in tomato xylem sap.

Fusarium oxysporum f. sp. lycopersici is a devastating plant pathogenic fungi known for wilt disease in the tomato plant and secrete cell wall degrading enzymes. These enzymes are collectively known as carbohydrate-active enzymes (CAZymes), crucial for growth, colonization and pathogenesis. Therefore, the present study was aimed to identify and annotate pathogen CAZymes in the xylem sap of a susceptible tomato variety using downstream proteomics and meta servers. Further, structural elucidation and conformational stability analysis of the selected CAZyme families were done through homology modeling and molecular dynamics simulation. Among all the fungal proteins identified, the carbohydrate metabolic process was found to be enriched. Most of the annotated CAZymes belonged to the hydrolase and oxidoreductase families, and 90% were soluble and extracellular. Moreover, using a publically available interactome database, interactions were observed between the families acting on chitin, hemicellulose and pectin. Subsequently, important catalytic residues were identified in the candidate CAZymes belonging to carbohydrate esterase (CE8) and glycosyl hydrolase (GH18 and GH28). Further, essential dynamics after molecular simulation of 100 ns revealed the overall behavior of these CAZymes with distinct global minima and transition states in CE8. Thus, our study identified some of the CAZyme families that assist in pathogenesis and growth through host cell wall deconstruction with further structural insight into the selected CAZyme families.Communicated by Ramaswamy H. Sarma.

Identification of lncRNA and weighted gene coexpression network analysis of germinating Rhizopus delemar causing mucormycosis.

Rhizopus delemar, an opportunistic fungal pathogen, causes a highly fatal disease, mucormycosis. Spore germination is a crucial mechanism for disease pathogenesis. Thus, exploring the molecular mechanisms of fungal germination would underpin our knowledge of such transformation and, in turn, help control mucormycosis. To gain insight into the developmental process particularly associated with cell wall modification and synthesis, weighted gene co-expression network analysis (WGCNA) was performed including both coding and non-coding transcripts identified in the current study, to find out the module of interest in the germination stages. The module-trait relationship identified a particular module to have a high correlation only at the resting phase and further analysis revealed the module to be enriched for protein phosphorylation, carbohydrate metabolic process, and cellular response to stimulus. Moreover, co-expression network analysis of highly connected nodes revealed cell wall modifying enzymes, especially those involved in mannosylation, chitin-glucan crosslinking, and polygalacturonase activities co-expressing and interacting with the novel lncRNAs among which some of them predicted to be endogenous target mimic (eTM) lncRNAs. Hence, the present study provides an insight into the onset of spore germination and the information on the novel non-coding transcripts with key cell wall-related enzymes as potential targets against mucormycosis.

Maternal and neonatal peripartum factors associated with late initiation of breast feeding in Bangladesh: a secondary analysis.

OBJECTIVES: Late initiation of breast feeding (LIBF) is associated with increased neonatal mortality and morbidity. This study aimed to assess the association between intrapartum, early postpartum and neonatal factors, and LIBF in Bangladesh. DESIGN, SETTING AND PARTICIPANTS: In this analysis, we used data from the mothers participating in a cluster-randomised controlled trial (Rang-Din Nutrition Study) conducted in rural northwest Bangladesh. Mothers (n=3594) were interviewed about the time of initiation of breast feeding, and peripartum maternal and neonatal complications within the first 72 hours of delivery. LIBF was defined as initiation of breast feeding 1 hour after birth. Factors associated with LIBF were identified by multivariable logistic regression analysis. MAIN OUTCOME MEASURES: Prevalence and associated factors of LIBF. RESULTS: The prevalence of LIBF was 18.5%. Factors significantly associated with LIBF in multivariable logistic regression were assisted vaginal delivery (adjusted OR (AOR) 2.17, 95% CI 1.44 to 3.27); delivery by caesarean section (AOR 9.67, 95% CI 7.21 to 12.96); maternal health problems during childbirth (AOR 1.61, 95% CI 1.30 to 2.00); preterm newborns (AOR 1.39, 95% CI 1.09 to 1.78); newborns moved slowly immediately after birth (AOR 1.43, 95% CI 1.05 to 1.94); and sick newborns (AOR 1.60, 95% CI 1.12 to 2.29). CONCLUSIONS: Findings from this study suggest that to reduce LIBF, peripartum maternal and neonatal complications should be prevented and treated. TRIAL REGISTRATION NUMBER: NCT01715038.

Prevalence of Infant and Young Child Feeding Practices and Differences in Estimates of Minimum Dietary Diversity Using 2008 and 2021 Definitions: Evidence from Bangladesh.

Background: Infant and young child feeding (IYCF) practices directly impact the health of <2-y-old children. Minimum dietary diversity (MDD) is an IYCF indicator to assess feeding practices of children aged 6-23 mo. The definition of MDD has recently been updated by the WHO and UNICEF, substituting "≥4 out of 7 food groups" (MDD-7FG) with "≥5 out of 8 food groups" (MDD-8FG). Objectives: The goals of this study were to estimate the prevalence of IYCF indicators and identify the implications of the change in the prevalence of MDD at the national and regional levels of Bangladesh. Methods: This study used data from the National Food Security and Nutrition Surveillance 2018-2019 round. A total of 1992 children aged 0-23 mo were included in this analysis. IYCF indicators and MDD were calculated according to the WHO-UNICEF guidelines. The difference between the prevalence of MDD-7FG and MDD-8FG is presented as percentage points. Results: The prevalence of early initiation of breastfeeding was 43.8%, and exclusive breastfeeding was 56.2%. Approximately 55% of children maintained MDD (MDD-7FG), 48% received minimum meal frequency, and 28% received a minimum acceptable diet. Compared with MDD-7FG, the prevalence of MDD-8FG was lower among 6-23-mo-old children. The difference between MDD prevalence (MDD-8FG vs. MDD-7FG) was high for boys (44.0% vs. 53.2%), children aged 12-23 mo (53.4% vs. 63.4%), in urban areas (30.2% vs. 42.4%), in the Dhaka administrative division (42.0% vs. 56.3%), among uneducated mothers (37.1% vs. 47.1%), in households with ≤4 members (44.3% vs. 55%), and for middle-class households (40.3% vs. 57.6%). Conclusions: The new method led to a decrease in the prevalence of MDD in Bangladesh. As the country prepares to implement the new indicator, it is critical to disseminate the new knowledge and its positive implication for improved child feeding and nutrition.

Immune response gene coexpression network analysis of Arachis hypogaea infected with Aspergillus flavus.

Aspergillus flavus (A. flavus) infection and aflatoxin contamination is a major bottleneck for peanut cultivation and value chain industry. In this study, a transcriptomic network study was conducted by retrieving publically available RNA-seq datasets of resistant and susceptible peanut varieties infected by A. flavus separately to understand the peanut defense mechanism against A. flavus. The gene expression analysis revealed differentially expressed genes (DEGs) in response to the different levels of infection and coexpression network of DEGs deciphered hub genes involved in the immune process in resistant and susceptible varieties. The interplay of resistance conferring genes and cell wall related genes was observed through functional enrichment analysis in response to pathogen infection and identified few key genes such as Protein P21, R genes, Pattern Recognition Receptor genes, Pectinesterases, Laccase and Thaumatin-like protein 1b as candidate genes in imparting immune response against A. flavus.

Genome-wide annotation, comparison and functional genomics of carbohydrate-active enzymes in legumes infecting Fusarium oxysporum formae speciales.

Fusarium wilt caused by soil borne ascomycetes fungi Fusarium oxysporum which has host-specific forms known as formae speciales (ff. spp.), apparently requires plant cell wall degrading enzymes (PCWDE) for successful invasion. In this study, 12 F. oxysporum ff. spp. were taken for genome-wide annotation and comparative analysis of CAZymes, with an assessment of secretory PCWDE and orthologues identification in the three legumes infecting ff. spp.  Further, transcriptomic analysis in two legumes infecting ff. spp. using publically available data was also done. The comparative studies showed Glycoside hydrolase (GH) families to be abundant and Principle Component Analysis (PCA) formed two distinct clusters of ff. spp. based on the CAZymes modules and families. Nearly half of the CAZymes in the legumes infecting ff. spp. coded for signal peptides. The orthologue clusters of secretory CAZymes common in all the three legume infecting ff. spp. mostly belonged to families of AA9, GH28, CE5 and PL1 and the expression analysis revealed the abundant PCWDE were differentially expressed in these legumes infecting ff. spp.  Therefore, this study gave an insight into the distribution of CAZymes especially extracellular PCWDE in legumes infecting ff. spp. with further shedding light onto some of the key PCWDE families through differential expression analysis.

Glycosyltransferases from oat (Avena) implicated in the acylation of avenacins.

Plants produce a huge array of specialized metabolites that have important functions in defense against biotic and abiotic stresses. Many of these compounds are glycosylated by family 1 glycosyltransferases (GTs). Oats (Avena spp.) make root-derived antimicrobial triterpenes (avenacins) that provide protection against soil-borne diseases. The ability to synthesize avenacins has evolved since the divergence of oats from other cereals and grasses. The major avenacin, A-1, is acylated with N-methylanthranilic acid. Previously, we have cloned and characterized three genes for avenacin synthesis (for the triterpene synthase SAD1, a triterpene-modifying cytochrome P450 SAD2, and the serine carboxypeptidase-like acyl transferase SAD7), which form part of a biosynthetic gene cluster. Here, we identify a fourth member of this gene cluster encoding a GT belonging to clade L of family 1 (UGT74H5), and show that this enzyme is an N-methylanthranilic acid O-glucosyltransferase implicated in the synthesis of avenacin A-1. Two other closely related family 1 GTs (UGT74H6 and UGT74H7) are also expressed in oat roots. One of these (UGT74H6) is able to glucosylate both N-methylanthranilic acid and benzoic acid, whereas the function of the other (UGT74H7) remains unknown. Our investigations indicate that UGT74H5 is likely to be key for the generation of the activated acyl donor used by SAD7 in the synthesis of the major avenacin, A-1, whereas UGT74H6 may contribute to the synthesis of other forms of avenacin that are acylated with benzoic acid.

Gene expression enabling synthetic diversification of natural products: chemogenetic generation of pacidamycin analogs.

Introduction of prnA, the halogenase gene from pyrrolnitrin biosynthesis, into Streptomyces coeruleorubidus resulted in efficient in situ chlorination of the uridyl peptide antibotic pacidamycin. The installed chlorine provided a selectably functionalizable handle enabling synthetic modification of the natural product using mild cross-coupling conditions in crude aqueous extracts of the culture broth.

An expeditious route to fluorinated rapamycin analogues by utilising mutasynthesis.

Rapamycin is a drug with several important clinical uses. Its complex structure means that total synthesis of this natural product and its analogues is demanding and lengthy. A more expeditious approach is to utilise biosynthesis to enable the generation of otherwise synthetically intractable analogues. In order to achieve this, rules governing biosynthetic precursor substrate preference must be established. Through determining these rules and synthesising and administering suitable substrate precursors, we demonstrate the first generation of fluorinated rapamycin analogues. Here we report the generation of six new fluororapamycins.

A serine carboxypeptidase-like acyltransferase is required for synthesis of antimicrobial compounds and disease resistance in oats.

Serine carboxypeptidase-like (SCPL) proteins have recently emerged as a new group of plant acyltransferases. These enzymes share homology with peptidases but lack protease activity and instead are able to acylate natural products. Several SCPL acyltransferases have been characterized to date from dicots, including an enzyme required for the synthesis of glucose polyesters that may contribute to insect resistance in wild tomato (Solanum pennellii) and enzymes required for the synthesis of sinapate esters associated with UV protection in Arabidopsis thaliana. In our earlier genetic analysis, we identified the Saponin-deficient 7 (Sad7) locus as being required for the synthesis of antimicrobial triterpene glycosides (avenacins) and for broad-spectrum disease resistance in diploid oat (Avena strigosa). Here, we report on the cloning of Sad7 and show that this gene encodes a functional SCPL acyltransferase, SCPL1, that is able to catalyze the synthesis of both N-methyl anthraniloyl- and benzoyl-derivatized forms of avenacin. Sad7 forms part of an operon-like gene cluster for avenacin synthesis. Oat SCPL1 (SAD7) is the founder member of a subfamily of monocot-specific SCPL proteins that includes predicted proteins from rice (Oryza sativa) and other grasses with potential roles in secondary metabolism and plant defense.

Real time HR-MAS NMR: application in reaction optimization, mechanism elucidation and kinetic analysis for heterogeneous reagent catalyzed small molecule chemistry.

A novel application of in situ (1)H high-resolution magic angle spinning (HR-MAS) NMR technique for real-time monitoring of H(2)SO(4)-silica promoted formation of 2, 2-disubstituted quinozolin-4(3H)-ones is reported. The detailed NMR spectroscopic data led to elucidation of the mechanism, reaction optimization, kinetics and quantitative analysis of the product accurately and efficiently. The translation of the optimized parameters obtained by (1)H HR-MAS NMR in the wet laboratory provided similar results. It is proposed that (1)H HR-MAS has a potential utility for optimization of various organic transformations in solid supported catalyzed reactions.

Development of fluorescent aryltryptophans by Pd mediated cross-coupling of unprotected halotryptophans in water.

A convenient and high yielding procedure for the Suzuki-Miyaura cross-coupling of unprotected bromo- and chlorotryptophans in water provides fluorescent aryltryptophans.

A convenient one-step synthesis of L-aminotryptophans and improved synthesis of 5-fluorotryptophan.

A one-pot biotransformation for the generation of a series of L-aminotryptophans using a readily prepared protein extract containing tryptophan synthase is reported. The extract exhibits remarkable stability upon freeze-drying, and may be stored and used for long periods after its preparation without significant loss of activity.