Discovery and Biosynthesis of Persiathiacins: Unusual Polyglycosylated Thiopeptides Active Against Multidrug Resistant Tuberculosis.

Thiopeptides are ribosomally biosynthesized and post-translationally modified peptides (RiPPs) that potently inhibit the growth of Gram-positive bacteria by targeting multiple steps in protein biosynthesis. The poor pharmacological properties of thiopeptides, particularly their low aqueous solubility, has hindered their development into clinically useful antibiotics. Antimicrobial activity screens of a library of Actinomycetota extracts led to discovery of the novel polyglycosylated thiopeptides persiathiacins A and B from Actinokineospora sp. UTMC 2448. Persiathiacin A is active against methicillin-resistant Staphylococcus aureus and several Mycobacterium tuberculosis strains, including drug-resistant and multidrug-resistant clinical isolates, and does not significantly affect the growth of ovarian cancer cells at concentrations up to 400 µM. Polyglycosylated thiopeptides are extremely rare and nothing is known about their biosynthesis. Sequencing and analysis of the Actinokineospora sp. UTMC 2448 genome enabled identification of the putative persiathiacin biosynthetic gene cluster (BGC). A cytochrome P450 encoded by this gene cluster catalyzes the hydroxylation of nosiheptide in vitro and in vivo, consistent with the proposal that the cluster directs persiathiacin biosynthesis. Several genes in the cluster encode homologues of enzymes known to catalyze the assembly and attachment of deoxysugars during the biosynthesis of other classes of glycosylated natural products. One of these encodes a glycosyl transferase that was shown to catalyze attachment of a D-glucose residue to nosiheptide in vitro. The discovery of the persiathiacins and their BGC thus provides the basis for the development of biosynthetic engineering approaches to the creation of novel (poly)glycosylated thiopeptide derivatives with enhanced pharmacological properties.

Hibiscus acid and hydroxycitric acid dimethyl esters from Hibiscus flowers induce production of dithiolopyrrolone antibiotics by Streptomyces Strain MBN2-2.

Plants and microbes are closely associated with each other in their ecological niches. Much has been studied about plant-microbe interactions, but little is known about the effect of phytochemicals on microbes at the molecular level. To access the products of cryptic biosynthetic gene clusters in bacteria, we incorporated an organic extract of hibiscus flowers into the culture media of different Actinobacteria isolated from plant rhizospheres. This approach led to the production of broad-spectrum dithiolopyrrolone (DTP) antibiotics, thiolutin (1) and aureothricin (2), by Streptomyces sp. MBN2-2. The compounds from the hibiscus extract responsible for triggering the production of these two DTPs were found to be hibiscus acid dimethyl ester (3) and hydroxycitric acid 1,3-dimethyl ester (4). It was subsequently found that the addition of either Fe2+ or Fe3+ to culture media induced the production of 1 and 2. The Chrome Azurol S (CAS) assay revealed that 3 and 4 can chelate iron, and therefore, the mechanism leading to the production of thiolutin and aureothricin appears to be related to changes in iron concentration levels. This work supports the idea that phytochemicals can be used to activate the production of cryptic microbial biosynthetic gene clusters and further understand plant-microbe interactions.

Chemistry and biology of specialized metabolites produced by Actinomadura.

Covering: up to the end of 2022In recent years rare Actinobacteria have become increasingly recognised as a rich source of novel bioactive metabolites. Actinomadura are Gram-positive bacteria that occupy a wide range of ecological niches. This review highlights about 230 secondary metabolites produced by Actinomadura spp., reported until the end of 2022, including their bioactivities and selected biosynthetic pathways. Notably, the bioactive compounds produced by Actinomadura spp. demonstrate a wide range of activities, including antimicrobial, antitumor and anticoccidial effects, highlighting their potential in various fields.

Autochthonous case of Trichophyton indotineae in Kuwait.

Trichophyton indotineae is an emerging dermatophyte that has remarkable impact on public health worldwide. In addition to producing severe extensive skin lesions, this species is frequently resistant to terbinafine, used as a first line agent. As a result, the infection is often refractory, making treatment very challenging. The current report describes the first case of Trichophyton indotineae infection in Kuwait. The infected woman had no recent travel history. She failed to respond to several courses of antifungals, but finally responded to voriconazole. The report suggests that T. indotineae is under recognised, hence, active surveillance of dermatophytes is warranted.

On the mechanisms of lysis triggered by perturbations of bacterial cell wall biosynthesis.

Inhibition of bacterial cell wall synthesis by antibiotics such as ß-lactams is thought to cause explosive lysis through loss of cell wall integrity. However, recent studies on a wide range of bacteria have suggested that these antibiotics also perturb central carbon metabolism, contributing to death via oxidative damage. Here, we genetically dissect this connection in Bacillus subtilis perturbed for cell wall synthesis, and identify key enzymatic steps in upstream and downstream pathways that stimulate the generation of reactive oxygen species through cellular respiration. Our results also reveal the critical role of iron homeostasis for the oxidative damage-mediated lethal effects. We show that protection of cells from oxygen radicals via a recently discovered siderophore-like compound uncouples changes in cell morphology normally associated with cell death, from lysis as usually judged by a phase pale microscopic appearance. Phase paling appears to be closely associated with lipid peroxidation.

Enemy in disguise: A case report of solitary trichoepithelioma initially diagnosed as BCC.

INTRODUCTION: Solitary trichoepitheliomas (TE) are benign tumors that are strikingly similar to their malignant counterpart, basal cell carcinoma (BCC). PRESENTATION OF CASE: An 83-year-old man presented with a 10-year history of a right lower lid skin mass initially diagnosed as BCC. Intraoperatively, an excisional biopsy was performed with primary reconstruction of the skin defect and the specimen was submitted for histopathology processing. Eventually, histopathology findings suggested the diagnosis of benign hair follicle tumor. The postoperative results were aesthetically pleasing and the integrity of the lower lid was preserved. DISCUSSION: Despite being rare, benign solitary TE are frequently misdiagnosed as malignant BCC, and vice versa. Oculoplastic surgeons face considerable difficulty distinguishing the two pathologies due to their similar clinical and histological pictures. Hence, excisional biopsy should be considered whenever such discrepancy is confronted to avoid the possibility of recurrence or malignant transformation. Furthermore, immunohistochemical staining could increase the accuracy of diagnosis in such unequivocal findings. CONCLUSION: Correlation of clinical, dermoscopic and histopathological findings are essential to establish an accurate diagnosis and select the appropriate management. In-depth understanding of eyelid reconstruction principles is mandatory to achieve desirable goals.

Mirubactin C rescues the lethal effect of cell wall biosynthesis mutations in Bacillus subtilis.

Growth of most rod-shaped bacteria is accompanied by the insertion of new peptidoglycan into the cylindrical cell wall. This insertion, which helps maintain and determine the shape of the cell, is guided by a protein machine called the rod complex or elongasome. Although most of the proteins in this complex are essential under normal growth conditions, cell viability can be rescued, for reasons that are not understood, by the presence of a high (mM) Mg2+ concentration. We screened for natural product compounds that could rescue the growth of mutants affected in rod-complex function. By screening > 2,000 extracts from a diverse collection of actinobacteria, we identified a compound, mirubactin C, related to the known iron siderophore mirubactin A, which rescued growth in the low micromolar range, and this activity was confirmed using synthetic mirubactin C. The compound also displayed toxicity at higher concentrations, and this effect appears related to iron homeostasis. However, several lines of evidence suggest that the mirubactin C rescuing activity is not due simply to iron sequestration. The results support an emerging view that the functions of bacterial siderophores extend well beyond simply iron binding and uptake.

Discovery of Demurilactone A: A Specific Growth Inhibitor of L-Form Bacillus subtilis.

Metabolic profiling of the extracts from a library of actinobacteria led to the identification of a novel polyketide, demurilactone A, produced by Streptomyces strain DEM21308. The structure of the compound was assigned based on a detailed investigation of 1D/2D NMR spectra and HR-MS. Whole genome DNA sequencing, followed by bioinformatics analysis and insertional mutagenesis, identified type I polyketide synthases encoded by the dml gene cluster to direct the biosynthesis of this polyene macrolide. While the number of modules is consistent with the carbon backbone of the assigned structure, some discrepancies were identified in the domain organization of five modules. Close investigation of the amino acid sequences identified several mutations in the conserved motifs of nonfunctional domains. Furthermore, the absolute configuration of hydroxy-bearing stereocenters was proposed based on analyses of the ketoreductase domains. Remarkably, although demurilactone A has little detectable activity against normal-walled bacteria, it specifically inhibits the growth of cell wall-deficient "L-form" Bacillus subtilis at a minimum inhibitory concentration value of 16 µg/mL. Time-lapse microscopy analyses revealed that demurilactone affects membrane dynamics, probably by reducing membrane fluidity. This compound could be a powerful reagent for studying long-standing questions about the involvement of L-forms in recurrent infection.

Discovery, isolation, heterologous expression and mode-of-action studies of the antibiotic polyketide tatiomicin from Amycolatopsis sp. DEM30355.

A genomic and bioactivity informed analysis of the metabolome of the extremophile Amycolatopsis sp. DEM30355 has allowed for the discovery and isolation of the polyketide antibiotic tatiomicin. Identification of the biosynthetic gene cluster was confirmed by heterologous expression in Streptomyces coelicolor M1152. Structural elucidation, including absolute stereochemical assignment, was performed using complementary crystallographic, spectroscopic and computational methods. Tatiomicin shows antibiotic activity against Gram-positive bacteria, including methicillin-resistant Staphylococcus aureus (MRSA). Cytological profiling experiments suggest a putative antibiotic mode-of-action, involving membrane depolarisation and chromosomal decondensation of the target bacteria.

Adolescent's self-reported weight and its association with media impact on decision to lose weight and body thinness perception.

This study investigated the accuracy of self-reported weight among Kuwaiti adolescents and the associations of self-reported weight and calculated BMI with the impact of media use on adolescent's decision to lose weight and body thinness perception. A total of 706 Kuwaiti adolescents (363 females) 15-18 year-olds were randomly selected from schools. Weight and height were self-reported by the adolescents and measured by the researchers. A specifically designed questionnaire reported the impact of media use on adolescent's decision to lose weight and body thinness perception. There were significant (p < 0.001) relationships between measured and reported weight (r = 0.947), height (r = 0.777), and calculated BMI (r = 0.623). Intraclass correlation coefficients (95% CI) between self-reported and measured weight and height were 0.973 (0.968-0.977) and 0.867 (0.839-0.891), respectively. The mean differences between self-reported and measured weight (1.26 kg) and BMI (0.478 kg/m2) were relatively small. Females under-reported their weight and the calculated BMI from estimated weight and height was underestimated by adolescents with overweight/obesity. The impact of media use on the adolescent's perception of being thin relative to the accuracy of the calculated BMI was significant (p = 0.043-0.001). The age-adjusted odds ratio of the calculated (underestimated) BMI in adolescents without overweight/obesity was 0.437 (95% CI = 0.257-0.741; p = 0.002). It was concluded that the validity of self-reported weight was high. Adolescents with overweight/obesity were more likely to underestimate their weight and calculated BMI. Educating adolescents about proper lifestyles and weight loss through media appears warranted.

Prevalence of overweight and obesity among Kuwaiti adolescents and the perception of body weight by parents or friends.

OBJECTIVE: Recently, the State of Kuwait has witnessed a steady rise in the prevalence of obesity among children and adolescents. The present study aims to provide an update on the rate of overweight or obesity among Kuwaiti adolescents and examines the associations between adolescents' overweight/obesity levels and their perception of body weight as seen by parents or friends. METHODS: A cross-sectional study was conducted in Kuwaiti secondary schools and included adolescents between the ages of 15 and 18 years, using a multistage stratified random sampling method. Body weight and height were measured. A specifically designed self-report questionnaire was used to assess parents' and friends' perceptions of an adolescent's body weight. RESULTS: A total of 706 adolescents were included the study. The prevalence of overweight or obesity among Kuwaiti adolescents reached nearly 50%, with males (54.3%) having a significantly higher overweight or obesity percentage than females (44.6%). No significant difference in the prevalence of obesity relative to age, from 15 to 18 years, was found. In addition, logistic regression analysis, adjusted for age and gender, revealed that adolescents perceived their parents (p = 0.011 and p < 0.001) or friends (p = 0.002 and p < 0.001) as more likely to classify their weight as overweight or obese, respectively. CONCLUSION: Overweight or obesity levels appear to be high among Kuwaiti adolescents, and appears to have reached a plateau recently. Efforts to combat obesity and promote physical activity and healthy nutrition are needed. Future studies should seek to identify important moderators of parental and social underestimation/overestimation of children's overweight or obesity.

Date Palm Fruit (Phoenix dactylifera): Effects on Vascular Health and Future Research Directions.

Cardiovascular disease is a leading cause of death globally, presenting an immense public and economic burden. Studies on cardioprotective foods and their bioactive components are needed to address both personal and public health needs. Date fruit is rich in polyphenols, particularly flavonoids, certain micronutrients, and dietary fiber, which can impact vascular health, and have the potential to attenuate vascular disease in humans. Data from in vitro and animal studies report that consumption of date fruit or extracts can modulate select markers of vascular health, particularly plasma lipid levels including triglycerides and cholesterol, indices of oxidative stress and inflammation, but human data is scant. More investigation is needed to better characterize date polyphenols and unique bioactive compounds or fractions, establish safe and effective levels of intake, and delineate underlying mechanisms of action. Implementing scientific rigor in clinical trials and assessment of functional markers of vascular disease, such as flow-mediated dilation and peripheral arterial tonometry, along with gut microbiome profiles would provide useful information with respect to human health. Emerging data supports the notion that intake of date fruit and extracts can be a useful component of a healthy lifestyle for those seeking beneficial effects on vascular health.

Kill and cure: genomic phylogeny and bioactivity of Burkholderia gladioli bacteria capable of pathogenic and beneficial lifestyles.

Burkholderia gladioli is a bacterium with a broad ecology spanning disease in humans, animals and plants, but also encompassing multiple beneficial interactions. It is a plant pathogen, a toxin-producing food-poisoning agent, and causes lung infections in people with cystic fibrosis (CF). Contrasting beneficial traits include antifungal production exploited by insects to protect their eggs, plant protective abilities and antibiotic biosynthesis. We explored the genomic diversity and specialized metabolic potential of 206 B. gladioli strains, phylogenomically defining 5 clades. Historical disease pathovars (pv.) B. gladioli pv. allicola and B. gladioli pv. cocovenenans were distinct, while B. gladioli pv. gladioli and B. gladioli pv. agaricicola were indistinguishable; soft-rot disease and CF infection were conserved across all pathovars. Biosynthetic gene clusters (BGCs) for toxoflavin, caryoynencin and enacyloxin were dispersed across B. gladioli, but bongkrekic acid and gladiolin production were clade-specific. Strikingly, 13 % of CF infection strains characterized were bongkrekic acid-positive, uniquely linking this food-poisoning toxin to this aspect of B. gladioli disease. Mapping the population biology and metabolite production of B. gladioli has shed light on its diverse ecology, and by demonstrating that the antibiotic trimethoprim suppresses bongkrekic acid production, a potential therapeutic strategy to minimize poisoning risk in CF has been identified.

Genomics-Driven Discovery of a Novel Glutarimide Antibiotic from Burkholderia gladioli Reveals an Unusual Polyketide Synthase Chain Release Mechanism.

A gene cluster encoding a cryptic trans-acyl transferase polyketide synthase (PKS) was identified in the genomes of Burkholderia gladioli BCC0238 and BCC1622, both isolated from the lungs of cystic fibrosis patients. Bioinfomatics analyses indicated the PKS assembles a novel member of the glutarimide class of antibiotics, hitherto only isolated from Streptomyces species. Screening of a range of growth parameters led to the identification of gladiostatin, the metabolic product of the PKS. NMR spectroscopic analysis revealed that gladiostatin, which has promising activity against several human cancer cell lines and inhibits tumor cell migration, contains an unusual 2-acyl-4-hydroxy-3-methylbutenolide in addition to the glutarimide pharmacophore. An AfsA-like domain at the C-terminus of the PKS was shown to catalyze condensation of 3-ketothioesters with dihydroxyacetone phosphate, thus indicating it plays a key role in polyketide chain release and butenolide formation.

Discovery and Biosynthesis of Bolagladins: Unusual Lipodepsipeptides from Burkholderia gladioli Clinical Isolates*.

Two Burkholderia gladioli strains isolated from the lungs of cystic fibrosis patients were found to produce unusual lipodepsipeptides containing a unique citrate-derived fatty acid and a rare dehydro-ß-alanine residue. The gene cluster responsible for their biosynthesis was identified by bioinformatics and insertional mutagenesis. In-frame deletions and enzyme activity assays were used to investigate the functions of several proteins encoded by the biosynthetic gene cluster, which was found in the genomes of about 45 % of B. gladioli isolates, suggesting that its metabolic products play an important role in the growth and/or survival of the species. The Chrome Azurol S assay indicated that these metabolites bind ferric iron, which suppresses their production when added to the growth medium. Moreover, a gene encoding a TonB-dependent ferric-siderophore receptor is adjacent to the biosynthetic genes, suggesting that these metabolites may function as siderophores in B. gladioli.

Heterologous reconstitution of the biosynthesis pathway for 4-demethyl-premithramycinone, the aglycon of antitumor polyketide mithramycin.

BACKGROUND: Mithramycin is an anti-tumor compound of the aureolic acid family produced by Streptomyces argillaceus. Its biosynthesis gene cluster has been cloned and characterized, and several new analogs with improved pharmacological properties have been generated through combinatorial biosynthesis. To further study these compounds as potential new anticancer drugs requires their production yields to be improved significantly. The biosynthesis of mithramycin proceeds through the formation of the key intermediate 4-demethyl-premithramycinone. Extensive studies have characterized the biosynthesis pathway from this intermediate to mithramycin. However, the biosynthesis pathway for 4-demethyl-premithramycinone remains unclear. RESULTS: Expression of cosmid cosAR7, containing a set of mithramycin biosynthesis genes, in Streptomyces albus resulted in the production of 4-demethyl-premithramycinone, delimiting genes required for its biosynthesis. Inactivation of mtmL, encoding an ATP-dependent acyl-CoA ligase, led to the accumulation of the tricyclic intermediate 2-hydroxy-nogalonic acid, proving its essential role in the formation of the fourth ring of 4-demethyl-premithramycinone. Expression of different sets of mithramycin biosynthesis genes as cassettes in S. albus and analysis of the resulting metabolites, allowed the reconstitution of the biosynthesis pathway for 4-demethyl-premithramycinone, assigning gene functions and establishing the order of biosynthetic steps. CONCLUSIONS: We established the biosynthesis pathway for 4-demethyl-premithramycinone, and identified the minimal set of genes required for its assembly. We propose that the biosynthesis starts with the formation of a linear decaketide by the minimal polyketide synthase MtmPKS. Then, the cyclase/aromatase MtmQ catalyzes the cyclization of the first ring (C7-C12), followed by formation of the second and third rings (C5-C14; C3-C16) catalyzed by the cyclase MtmY. Formation of the fourth ring (C1-C18) requires MtmL and MtmX. Finally, further oxygenation and reduction is catalyzed by MtmOII and MtmTI/MtmTII respectively, to generate the final stable tetracyclic intermediate 4-demethyl-premithramycinone. Understanding the biosynthesis of this compound affords enhanced possibilities to generate new mithramycin analogs and improve their production titers for bioactivity investigation.

Effects of short-term consumption of strawberry powder on select parameters of vascular health in adolescent males.

Cardiovascular disease is a leading cause of death in the United States and much of the developed world, costing billions of dollars in lost work time, lower productivity and high health care expenditures. Research on foods and bioactive food components that have cardioprotective benefits may provide new insights as to how modest changes in one's diet may result in a reduced risk of vascular disease. In intervention trials, the consumption of strawberries, either fresh or freeze-dried, has been reported to improve select markers of cardiovascular health, including improved lipid profiles, microvascular function, and platelet reactivity. Consistent with the above, epidemiological studies suggest beneficial effects of strawberries on vascular function. Preliminary studies on the effects of freeze-dried strawberry powder on vascular health are reviewed in the current paper.

An unusual Burkholderia gladioli double chain-initiating nonribosomal peptide synthetase assembles 'fungal' icosalide antibiotics.

Burkholderia is a multi-talented genus of Gram-negative bacteria, which in recent years has become increasingly recognised as a promising source of bioactive natural products. Metabolite profiling of Burkholderia gladioli BCC0238 showed that it produces the asymmetric lipopeptidiolide antibiotic icosalide A1, originally isolated from a fungus. Comparative bioinformatics analysis of several genome-sequenced B. gladioli isolates identified a gene encoding a nonribosomal peptide synthase (NRPS) with an unusual architecture that was predicted to be responsible for icosalide biosynthesis. Inactivation of this gene in B. gladioli BCC0238 abolished icosalide production. PCR analysis and sequencing of total DNA from the original fungal icosalide A1 producer revealed it has a B. gladioli strain associated with it that harbours an NRPS with an identical architecture to that responsible for icosalide A1 assembly in B. gladioli BCC0238. Sequence analysis of the icosalide NRPS indicated that it contains two chain-initiating condensation (CI) domains. One of these is appended to the N-terminus of module 1 - a common architecture for NRPSs involved in lipopeptide assembly. The other is embedded in module 3, immediately downstream of a putative chain-elongating condensation domain. Analysis of the reactions catalysed by a tridomain construct from module 3 of the NRPS using intact protein mass spectrometry showed that the embedded CI domain initiates assembly of a second lipopeptide chain, providing key insights into the mechanism for asymmetric diolide assembly.

Effects of Short-Term Dried Plum (Prune) Intake on Markers of Bone Resorption and Vascular Function in Healthy Postmenopausal Women: A Randomized Crossover Trial.

Osteoporosis and cardiovascular disease are global health burdens, with postmenopausal women being at great risk. Dried plums/prunes (DPs) have been reported to provide bone health benefits in animal models, which is consistent with in vitro models. Data from human studies suggest that DP intake can enhance lipid metabolism, anti-inflammatory, and oxidant defense systems, which can impact cardiovascular health. We tested the hypothesis that short-term consumption of low and reasonable levels of DPs augments bone resorption and vascular function. Twenty-seven healthy, postmenopausal women were randomly assigned to consume six DPs (∼42 g) or two DPs (∼14 g) per day for 2 weeks, then a 2-week washout period and then crossed over. Serum C-telopeptide, beta-crosslinked (CTX) was used as a measure of bone resorption. Peripheral artery tonometry (PAT) was used to assess microvascular function. The pattern of changes in CTX in the second 2-week period (no change or decline) differed significantly from the pattern in the first 2 weeks (increases in both groups; F = 9.26, P = .006), suggesting a trend in CTX reduction (i.e., a decrease in bone resorption) in those consuming six DPs per day in the second phase. No effects on vascular function were noted. A significant interaction was observed for the augmentation index, a measure of arterial stiffness, between treatment and years after menopause (P = .045). The results suggest a potentially favorable impact of DPs on bone health when assessed with a short-term, crossover study design in postmenopausal women. Given the novel assessments used in this study, follow-up studies are warranted.