Bacillibactin class siderophores produced by the endophyte Bacillus subtilis NPROOT3 as antimycobacterial agents.

The bacterial endophytes isolated from the halophyte Salicornia brachiata were explored for the antimicrobial potential to discover novel microbial inhibitors that combat multidrug resistance. Upon investigation, ethyl acetate extract of the endophyte Bacillus subtilis NPROOT3 displayed significant potency against Mycobacterium smegmatis MTCC6 as well as Mycobacterium tuberculosis H37Rv strain. Further investigation of ethyl acetate crude extract by repeated chromatographic separations followed by characterization using UV, HR-ESI-MS, MALDI-MS, MALDI-MS/MS, CD, and NMR spectroscopy yielded a series of five known siderophores, namely, SVK21 (1), bacillibactin C (2), bacillibactin B (3), tribenglthin A (4), and bacillibactin (5). A total of two out of five compounds, 4 (MIC 38.66 µM) and 5 (MIC 22.15 µM) exhibited significant inhibition against the strain M. smegmatis MTCC6 comparable with positive control rifampicin (MIC 12.15 µM). None of these five bacillibactin molecules are previously reported to exhibit bioactivity against Mycobacterium sp. Herein for the first time, all the compounds were screened for their antibacterial activities against a panel of bacterial pathogens of humans. Furthermore, the probable mechanism of action of bacillibactin compounds for their antimycobacterial activity is also discussed. The findings of this study open up a new chemotype for inhibition of the Mycobacterium sp. and other multidrug-resistant pathogens.

Bacillinaphthin A: A New Naphthohydroquinone from the Endophyte Bacillus subtilis NPROOT3.

The present study explores the endophyte associated with the halophyte Salicornia brachiata for uncovering new biologically important compounds. Thus, HPLC-PDA guided chemical investigation of the ethyl acetate extract of the Bacillus subtilis NPROOT3 led to the isolation of a new compound named bacillinaphthin A (1) along with previously known rubinaphthin A (2). The structure of 2 was determined by a comparison of HR-ESI-MS, 1 H and 13 C nuclear magnetic resonances (NMR) with those of reported data, whereas the structure of new compound 1 was elucidated by interpretation of 1D- and 2D-NMR and MS data. Bacillinaphthin (1) and rubinaphthin (2) feature 1,4-dihydroxy-2-naphthoic acid derivatives which have been isolated herein for the first time from the genus Bacillus. Bacillinaphthin (1) is a new congener of 2 with an additional succinic acid side chain attached to the sugar moiety. Production of succinoglycan compounds was reported to regulate symbiosis, hence the isolation of 1 exhibits an example of chemical ecology between the halophyte and its endophyte. In silico tools were used to assess the bioactive potential of both isolated molecules.

Indazole-based microtubule-targeting agents as potential candidates for anticancer drugs discovery.

Tremendous research is focused on developing novel drug candidates targeting microtubules to inhibit their function in several cellular processes, including cell division. In this regard, several indazole derivatives were sought to target the colchicine binding site on the ß-tubulin, a crucial protein required to form microtubules, to develop microtubule targeting agents. Even though there are several reviews on the indazole-based compounds, none of them focused on using indazole scaffold to develop microtubule targeting agents. Therefore, this review aims to present the advances in research on compounds containing indazole scaffolds as microtubule targeting agents based on the articles published in the last two decades. Among the articles reviewed, we found that compounds 6 and 7 showed the lowest IC50 values of 0.6 âˆ¼ 0.9 nM in the cell line studies, making them the strongest indazole derivatives that target microtubules. The compounds 30, 31, 37 (IC50 = ∼ 1 nM) and compounds 8, 38 (IC50 = ∼ 2 nM) have proved to be potent microtubule inhibitors. The compounds 18, 31, 44, 45 also showed strong anticancer activity (IC50 = ∼ 8 nM). It is important to notice that except for compounds 9, 12, 13, 15, and SRF, the top activity compounds including 6, 7, 8, 10, 11, 30, 31, 37, 44, and 45 contain 3,4,5­trimethoxyphenyl substitution similar to that of colchicine. Therefore, it appears that the 3,4,5­trimethoxyphenyl substituent on the indazole scaffold is crucial for targeting CBS.

Synthesis and evaluation of 2-aryl-1H-benzo[d]imidazole derivatives as potential microtubule targeting agents.

Microtubule targeting agents (MTAs) are the potential drug candidates for anticancer drug discovery. Disrupting the microtubule formation or inhibiting the de-polymerization process by a synthetic molecule can lead to an excellent anticancer drug candidate. Here, we present the 2,5-substituted-1H-benzo[d]imidazole derivatives as potential colchicine, nocodazole binding site targeting agents. About 20 benzimidazole derivatives were synthesized with 82.0%-94.0% yield using mild reaction conditions. The synthesized compounds showed moderate to excellent anticancer activity established in three cell lines, including Hela cells, A549 cells, MRC-5 cells. The compounds B15, B16, B19, and B20 are the potential candidates with the IC50 values <15 µM in the three different cell lines. In MTT assay, compounds B15, B16, B19, and B20 showed excellent antiproliferation activity indicated by IC50 values in the range of 5.3 ± 0.21 to 18.1 ± 0.32 µM using HeLa and A549 cell lines. The predicted absorption, distribution, metabolism and excretion (ADME) properties and drug-likeness properties of B15, B16, B19, and B20 indicate that these compounds can be used as lead compounds for further study to develop excellent MTAs.

Biodiversity and antimicrobial potential of bacterial endophytes from halophyte Salicornia brachiata.

Extreme natural habitats like halophytes, marsh land, and marine environment are suitable arena for chemical ecology between plants and microbes having environmental impact. Endophytes are an ecofriendly option for the promotion of plant growth and to serve as sustainable resource of novel bioactive natural products. The present study, focusing on biodiversity of bacterial endophytes from Salicornia brachiata, led to isolation of around 336 bacterial endophytes. Phylogenetic analysis of 63 endophytes revealed 13 genera with 27 different species, belonging to 3 major groups: Firmicutes, Proteobacteria, and Actinobacteria. 30% endophytic isolates belonging to various genera demonstrated broad-spectrum antibacterial and antifungal activities against a panel of human, plant, and aquatic infectious agents. An endophytic isolate Bacillus amyloliquefaciens 5NPA-1, exhibited strong in-vitro antibacterial activity against human pathogen Staphylococcus aureus and phytopathogen Xanthomonas campestris. Investigation through LC-MS/MS-based molecular networking and bioactivity-guided purification led to the identification of three bioactive compounds belonging to lipopeptide class based on 1H-, 13C-NMR and MS analysis. To our knowledge, this is the first report studying bacterial endophytic biodiversity of Salicornia brachiata and the isolation of bioactive compounds from its endophyte. Overall, the present study provides insights into the diversity of endophytes associated with the plants from the extreme environment as a rich source of metabolites with remarkable agricultural applications and therapeutic properties.

Genetic Characterization of Influenza A (H1N1) Pandemic 2009 Virus Isolates from Mumbai.

Pandemic influenza A (H1N1) 2009 virus was first detected in India in May 2009 which subsequently became endemic in many parts of the country. Influenza A viruses have the ability to evade the immune response through its ability of antigenic variations. The study aims to characterize influenza A (H1N1) pdm 09 viruses circulating in Mumbai during the pandemic and post-pandemic period. Nasopharyngeal swabs positive for influenza A (H1N1) pdm 09 viruses were inoculated on Madin-Darby canine kidney cell line for virus isolation. Molecular and phylogenetic analysis of influenza A (H1N1) pdm 09 isolates was conducted to understand the evolution and genetic diversity of the strains. Nucleotide and amino acid sequences of the HA gene of Mumbai isolates when compared to A/California/07/2009-vaccine strain revealed 14 specific amino acid differences located at the antigenic sites. Amino acid variations in HA and NA gene resulted in changes in the N-linked glycosylation motif which may lead to immune evasion. Phylogenetic analysis of the isolates revealed their evolutionary position with vaccine strain A/California/07/2009 but had undergone changes gradually. The findings in the present study confirm genetic variability of influenza viruses and highlight the importance of continuous surveillance during influenza outbreaks.

Targeted Isolation of Antibiofilm Compounds from Halophytic Endophyte Bacillus velezensis 7NPB-3B Using LC-HR-MS-Based Metabolomics.

The discovery of new natural products has become more challenging because of the re-isolation of compounds and the lack of new sources. Microbes dwelling in extreme conditions of high salinity and temperature are huge prospects for interesting natural metabolites. In this study, the endophytic bacteria Bacillus velezensis 7NPB-3B isolated from the halophyte Salicornia brachiata was screened for its biofilm inhibition against methicillin-resistant Staphylococcus aureus (MRSA). The fractionation of the crude extract was guided by bioassay and LC-HRMS-based metabolomics using multivariate analysis. The 37 fractions obtained by high-throughput chromatography were dereplicated using an in-house MS-Excel macro coupled with the Dictionary of Natural Products database. Successive bioactivity-guided separation yielded one novel compound (1), a diketopiperazine (m/z 469.258 [M - H]-) with an attached saturated decanoic acid chain, and four known compounds (2-5). The compounds were identified based on 1D- and 2D-NMR and mass spectrometry. Compounds 1 and 5 exhibited excellent biofilm inhibition properties of >90% against the MRSA pathogen at minimum inhibition concentrations of 25 and 35 µg/mL, respectively. The investigation resulted in the isolation of a novel diketopiperazine from a bacterial endophyte of an untapped plant using an omics approach.

Th1 polarization in Bordetella pertussis vaccine responses is maintained through a positive feedback loop.

Outbreaks of Bordetella pertussis (BP), the causative agent of whooping cough, continue despite broad vaccination coverage and have been increasing since vaccination switched from whole-BP (wP) to acellular BP (aP) vaccines. wP vaccination has been associated with more durable protective immunity and an induced Th1 polarized memory T cell response. Here, a multi-omics approach was applied to profile the immune response of 30 wP and 31 aP-primed individuals and identify correlates of T cell polarization before and after Tdap booster vaccination. We found that transcriptional changes indicating an interferon response on day 1 post-booster along with elevated plasma concentrations of IFN-γ and interferon-induced chemokines that peaked at day 1-3 post-booster correlated best with the Th1 polarization of the vaccine-induced memory T cell response on day 28. Our studies suggest that wP-primed individuals maintain their Th1 polarization through this early memory interferon response. This suggests that stimulating the interferon pathway during vaccination could be an effective strategy to elicit a predominant Th1 response in aP-primed individuals that protects better against infection.

Glaucarubinone from Simarouba glauca DC. as a potential biocontrol agent for mosquito vector management.

The study explored Simarouba glauca DC. for mosquito larvicidal potential by performing bioactivity-guided chemical investigation of its root extract resulting in isolation of the known bioactive metabolite glaucarubinone (1). Mosquito larvicidal activity of glaucarubinone (1) against the three vector species viz. Anopheles stephensi, Aedes aegypti, and Culex quinquefasciatus was determined using a modified WHO 2005 protocol. It was observed that Culex quinquefasciatus larvae were the most susceptible species with LC50 13.88 ppm and LC90 70.01 ppm followed by Aedes aegypti and Anopheles stephensi at 24 h of exposure. The mode of action as observed microscopically is the lysis of midgut and thorax cells of the third instar larvae. The crystal structure of the glaucarubinone (1) is reported for the first time using X-ray crystallography. This phytochemical product has the potential to act as a green alternative to existing chemical-based insecticides for integrated vector management.

A multi-omics systems vaccinology resource to develop and test computational models of immunity.

Systems vaccinology studies have identified factors affecting individual vaccine responses, but comparing these findings is challenging due to varying study designs. To address this lack of reproducibility, we established a community resource for comparing Bordetella pertussis booster responses and to host annual contests for predicting patients' vaccination outcomes. We report here on our experiences with the "dry-run" prediction contest. We found that, among 20+ models adopted from the literature, the most successful model predicting vaccination outcome was based on age alone. This confirms our concerns about the reproducibility of conclusions between different vaccinology studies. Further, we found that, for newly trained models, handling of baseline information on the target variables was crucial. Overall, multiple co-inertia analysis gave the best results of the tested modeling approaches. Our goal is to engage community in these prediction challenges by making data and models available and opening a public contest in August 2024.

Putting computational models of immunity to the test - an invited challenge to predict B. pertussis vaccination outcomes.

Systems vaccinology studies have been used to build computational models that predict individual vaccine responses and identify the factors contributing to differences in outcome. Comparing such models is challenging due to variability in study designs. To address this, we established a community resource to compare models predicting B. pertussis booster responses and generate experimental data for the explicit purpose of model evaluation. We here describe our second computational prediction challenge using this resource, where we benchmarked 49 algorithms from 53 scientists. We found that the most successful models stood out in their handling of nonlinearities, reducing large feature sets to representative subsets, and advanced data preprocessing. In contrast, we found that models adopted from literature that were developed to predict vaccine antibody responses in other settings performed poorly, reinforcing the need for purpose-built models. Overall, this demonstrates the value of purpose-generated datasets for rigorous and open model evaluations to identify features that improve the reliability and applicability of computational models in vaccine response prediction.

Integrated longitudinal multiomics study identifies immune programs associated with acute COVID-19 severity and mortality.

BACKGROUNDPatients hospitalized for COVID-19 exhibit diverse clinical outcomes, with outcomes for some individuals diverging over time even though their initial disease severity appears similar to that of other patients. A systematic evaluation of molecular and cellular profiles over the full disease course can link immune programs and their coordination with progression heterogeneity.METHODSWe performed deep immunophenotyping and conducted longitudinal multiomics modeling, integrating 10 assays for 1,152 Immunophenotyping Assessment in a COVID-19 Cohort (IMPACC) study participants and identifying several immune cascades that were significant drivers of differential clinical outcomes.RESULTSIncreasing disease severity was driven by a temporal pattern that began with the early upregulation of immunosuppressive metabolites and then elevated levels of inflammatory cytokines, signatures of coagulation, formation of neutrophil extracellular traps, and T cell functional dysregulation. A second immune cascade, predictive of 28-day mortality among critically ill patients, was characterized by reduced total plasma Igs and B cells and dysregulated IFN responsiveness. We demonstrated that the balance disruption between IFN-stimulated genes and IFN inhibitors is a crucial biomarker of COVID-19 mortality, potentially contributing to failure of viral clearance in patients with fatal illness.CONCLUSIONOur longitudinal multiomics profiling study revealed temporal coordination across diverse omics that potentially explain the disease progression, providing insights that can inform the targeted development of therapies for patients hospitalized with COVID-19, especially those who are critically ill.TRIAL REGISTRATIONClinicalTrials.gov NCT04378777.FUNDINGNIH (5R01AI135803-03, 5U19AI118608-04, 5U19AI128910-04, 4U19AI090023-11, 4U19AI118610-06, R01AI145835-01A1S1, 5U19AI062629-17, 5U19AI057229-17, 5U19AI125357-05, 5U19AI128913-03, 3U19AI077439-13, 5U54AI142766-03, 5R01AI104870-07, 3U19AI089992-09, 3U19AI128913-03, and 5T32DA018926-18); NIAID, NIH (3U19AI1289130, U19AI128913-04S1, and R01AI122220); and National Science Foundation (DMS2310836).

Characterization of FK506 biosynthetic intermediates involved in post-PKS elaboration.

The post-PKS modification steps of FK506 biosynthesis include C9-oxidation and 31-O-methylation, but the sequence of these reactions and the exact route have remained unclear. This study details the post-PKS modification pathways in FK506 biosynthesis through the identification of all intermediates and in vitro enzymatic reactions of the cytochrome P450 hydroxylase FkbD and the methyltransferase FkbM. These results complete our understanding of post-PKS modification steps to FK506 showing the substrate flexibility of two enzymes involved and the existence of two parallel biosynthetic routes to FK506.

Dactylides A-C, three new bioactive 22-membered macrolides produced by Dactylosporangium aurantiacum.

Three new 22-membered polyol macrolides, dactylides A-C (1-3), were isolated from Dactylosporangium aurantiacum ATCC 23491 employing repeated chromatographic separations, and their structures were established based on detailed analysis of NMR and MS data. The relative configurations at the stereocenters were established via vicinal 1H-1H coupling constants, NOE correlations, and by application of Kishi's universal NMR database. In order to get insights into the biosynthetic pathway of 1-3, the genome sequence of the producer strain D. aurantiacum was obtained and the putative biosynthetic gene cluster encoding their biosynthesis was identified through bioinformatic analysis using antiSMASH. Compounds 1-3 showed significant in-vitro antimycobacterial and cytotoxic activity.

A systems vaccinology resource to develop and test computational models of immunity.

Computational models that predict an individual's response to a vaccine offer the potential for mechanistic insights and personalized vaccination strategies. These models are increasingly derived from systems vaccinology studies that generate immune profiles from human cohorts pre- and post-vaccination. Most of these studies involve relatively small cohorts and profile the response to a single vaccine. The ability to assess the performance of the resulting models would be improved by comparing their performance on independent datasets, as has been done with great success in other areas of biology such as protein structure predictions. To transfer this approach to system vaccinology studies, we established a prototype platform that focuses on the evaluation of Computational Models of Immunity to Pertussis Booster vaccinations (CMI-PB). A community resource, CMI-PB generates experimental data for the explicit purpose of model evaluation, which is performed through a series of annual data releases and associated contests. We here report on our experience with the first such 'dry run' for a contest where the goal was to predict individual immune responses based on pre-vaccination multi-omic profiles. Over 30 models adopted from the literature were tested, but only one was predictive, and was based on age alone. The performance of new models built using CMI-PB training data was much better, but varied significantly based on the choice of pre-vaccination features used and the model building strategy. This suggests that previously published models developed for other vaccines do not generalize well to Pertussis Booster vaccination. Overall, these results reinforced the need for comparative analysis across models and datasets that CMI-PB aims to achieve. We are seeking wider community engagement for our first public prediction contest, which will open in early 2024.

Integrated longitudinal multi-omics study identifies immune programs associated with COVID-19 severity and mortality in 1152 hospitalized participants.

Hospitalized COVID-19 patients exhibit diverse clinical outcomes, with some individuals diverging over time even though their initial disease severity appears similar. A systematic evaluation of molecular and cellular profiles over the full disease course can link immune programs and their coordination with progression heterogeneity. In this study, we carried out deep immunophenotyping and conducted longitudinal multi-omics modeling integrating ten distinct assays on a total of 1,152 IMPACC participants and identified several immune cascades that were significant drivers of differential clinical outcomes. Increasing disease severity was driven by a temporal pattern that began with the early upregulation of immunosuppressive metabolites and then elevated levels of inflammatory cytokines, signatures of coagulation, NETosis, and T-cell functional dysregulation. A second immune cascade, predictive of 28-day mortality among critically ill patients, was characterized by reduced total plasma immunoglobulins and B cells, as well as dysregulated IFN responsiveness. We demonstrated that the balance disruption between IFN-stimulated genes and IFN inhibitors is a crucial biomarker of COVID-19 mortality, potentially contributing to the failure of viral clearance in patients with fatal illness. Our longitudinal multi-omics profiling study revealed novel temporal coordination across diverse omics that potentially explain disease progression, providing insights that inform the targeted development of therapies for hospitalized COVID-19 patients, especially those critically ill.

Engineered biosynthesis of glycosylated derivatives of narbomycin and evaluation of their antibacterial activities.

A 14-membered macrolide antibiotic narbomycin produced from Streptomyces venezuelae ATCC 15439 is composed of polyketide macrolactone ring and D-desosamine as a deoxysugar moiety, which acts as an important determinant of its antibacterial activity. In order to generate diverse glycosylated derivatives of narbomycin, expression plasmids carrying different deoxysugar biosynthetic gene cassettes and the gene encoding a substrate-flexible glycosyltransferase DesVII were constructed and introduced into S. venezuelae YJ003 mutant strain bearing a deletion of thymidine-5'-diphospho-D-desosamine biosynthetic gene cluster. The resulting recombinants of S. venezuelae produced a range of new analogs of narbomycin, which possess unnatural sugar moieties instead of native deoxysugar D-desosamine. The structures of narbomycin derivatives were determined through nuclear magnetic resonance spectroscopy and mass spectrometry analyses and their antibacterial activities were evaluated in vitro against erythromycin-susceptible and -resistant Enterococcus faecium and Staphylococcus aureus. Substitution with L-rhamnose or 3-O-demethyl-D-chalcose was demonstrated to exhibit greater antibacterial activity than narbomycin and the clinically relevant erythromycin. This work provides new insight into the functions of deoxysugar biosynthetic enzymes and structure-activity relationships of the sugar moieties attached to the macrolides and demonstrate the potential of combinatorial biosynthesis for the generation of new macrolides carrying diverse sugars with increased antibacterial activities.

Nucleotide-based genetic networks: Methods and applications.

Genomic variations have been acclaimed as among the key players in understanding the biological mechanisms behind migration, evolution, and adaptation to extreme conditions. Due to stochastic evolutionary forces, the frequency of polymorphisms is affected by changes in the frequency of nearby polymorphisms in the same DNA sample, making them connected in terms of evolution. This article presents all the ingredients to understand the cumulative effects and complex behaviors of genetic variations in the human mitochondrial genome by analyzing co-occurrence networks of nucleotides, and shows key results obtained from such analyses. The article emphasizes recent investigations of these co-occurrence networks, describing the role of interactions between nucleotides in fundamental processes of human migration and viral evolution. The corresponding co-mutation-based genetic networks revealed genetic signatures of human adaptation in extreme environments. This article provides the methods of constructing such networks in detail, along with their graph-theoretical properties, and applications of the genomic networks in understanding the role of nucleotide co-evolution in evolution of the whole genome.

Selective Partition of Lipopeptides from Fermentation Broth: A Green and Sustainable Approach.

Lipopeptide (LP) biosurfactants from microbes have the potential to gradually replace chemical synthetic surfactants and fit the contemporary green and sustainable industrial production concept. However, their active participation is comparatively low in the global market pertaining to their low yield in microbial broth and costly downstream processes arising due to tedious isolation and purification methods. Herein, an efficient extraction method is developed that utilizes an aqueous biphasic system (ABS) comprising ionic liquids and polypropylene glycol 400 (PPG) to selectively extract a mixture of cyclic lipopeptides, namely, surfactin and fengycin from the culture broth of Bacillus amyloliquefaciens 5NPA-1, isolated from the halophyte Salicornia brachiata Roxb. Out of four different ABSs, the ABS composed of 2-hydroxyethyl ammonium formate and PPG displayed a maximum extraction efficiency of 82.30%. PPG-rich phase containing lipopeptides exhibited excellent antimicrobial and mosquito larvicidal properties with no toxic effect on plants. The developed method is simple, novel and accelerates the application of cyclic lipopeptides produced by the microbial source.

Impact of modular mitochondrial epistatic interactions on the evolution of human subpopulations.

Investigation of human mitochondrial (mt) genome variation has been shown to provide insights to the human history and natural selection. By analyzing 24,167 human mt-genome samples, collected for five continents, we have developed a co-mutation network model to investigate characteristic human evolutionary patterns. The analysis highlighted richer co-mutating regions of the mt-genome, suggesting the presence of epistasis. Specifically, a large portion of COX genes was found to co-mutate in Asian and American populations, whereas, in African, European, and Oceanic populations, there was greater co-mutation bias in hypervariable regions. Interestingly, this study demonstrated hierarchical modularity as a crucial agent for these co-mutation networks. More profoundly, our ancestry-based co-mutation module analyses showed that mutations cluster preferentially in known mitochondrial haplogroups. Contemporary human mt-genome nucleotides most closely resembled the ancestral state, and very few of them were found to be ancestral-variants. Overall, these results demonstrated that subpopulation-based biases may favor mitochondrial gene specific epistasis.