Unusual Sesquiterpenes from Streptomyces olindensis DAUFPE 5622.

In nature, the vast majority of sesquiterpenes are produced by type I mechanisms, and glycosylated sesquiterpenes are rare in actinobacteria. Streptomyces olindensis DAUFPE 5622 produces the sesquiterpenes olindenones A-G, a new class of rearranged drimane sesquiterpenes. Olindenones B-D are oxygenated derivatives of olindenone A, while olindenones E-G are analogs glycosylated with dideoxysugars. 13C-isotope labeling studies demonstrated olindenone A biosynthesis occurs via the methylerythritol phosphate (MEP) pathway and suggested the rearrangement is only partially concerted. Based on the structures, one potential mechanism of olindenone A formation proceeds by cyclization of the linear terpenoid precursor, likely occurring via a terpene cyclase-mediated type II mechanism whereby the terminal alkene of the precursor is protonated, triggering carbocation-driven cyclization followed by rearrangement. Diphosphate hydrolysis may occur either before or after cyclization. Although a biosynthetic route is proposed, the terpene cyclase gene responsible for producing olindenones currently remains unidentified.

An Evaluation of Emergency Medicine Core Content Covered by Podcasts.

INTRODUCTION: Podcasts are used broadly for learning in emergency medicine (EM); however, there is concern about the potential for knowledge gaps for learners who rely on podcasts for their learning. The extent to which EM podcasts cover the core curriculum of EM is not known; thus, we sought to quantify the extent to which podcasts represent the core content of our specialty. METHODS: We performed a retrospective review of all EM podcast episodes published in 2019. All podcasts were given credit for the content they covered as it related to the 2016 American Board of Emergency Medicine (ABEM) Model of Clinical Practice in Emergency Medicine (EM Model). The primary outcome was a description of how podcasts represented the ABEM EM Model content topics compared to the topic representation of the ABEM Qualifying Exam. RESULTS: We included 54 unique EM podcast programs and 1,193 podcast episodes. They covered 2,965 total EM Model core content topics. The topics most covered were "other" (which includes interpersonal skills and professionalism), procedures, and signs and symptoms. Musculoskeletal, hematology, and environmental each accounted for less than 1% of all topics covered. Almost three-quarters of podcast episodes covered other core competencies of the practice of EM. CONCLUSION: Podcasts had a broad yet imbalanced coverage of the ABEM EM Model core content subtopics in 2019, with a notable coverage of other core competencies of the practice of EM. Learners, educators, and scholars should be mindful of these gaps and focus future work on exploring how podcasts should best be used in EM education.

Genome mining of Streptomyces sp. BRB081 reveals the production of the antitumor pyrrolobenzodiazepine sibiromycin.

Employing a genome mining approach, this work aimed to further explore the secondary metabolism associated genes of Streptomyces sp. BRB081, a marine isolate. The genomic DNA of BRB081 was sequenced and assembled in a synteny-based pipeline for biosynthetic gene clusters (BGCs) annotation. A total of 27 BGCs were annotated, including a sibiromycin complete cluster, a bioactive compound with potent antitumor activity. The production of sibiromycin, a pyrrolobenzodiazepine, was confirmed by the analysis of obtained BRB081 extract by HPLC-MS/MS, which showed the presence of the sibiromycin ions themselves, as well as its imine and methoxylated forms. To verify the presence of this cluster in other genomes available in public databases, a genome neighborhood network (GNN) was constructed with the non-ribosomal peptide synthetase (NRPS) gene from Streptomyces sp. BRB081. Although the literature does not report the occurrence of the sibiromycin BGC in any other microorganism than Streptosporangium sibiricum, we have located this BGC in 10 other genomes besides the BRB081 isolate, all of them belonging to the Actinomycetia class. These findings strengthen the importance of uninterrupted research for new producer strains of secondary metabolites with uncommon biological activities. These results reinforced the accuracy and robustness of genomics in the screening of natural products. Furthermore, the unprecedented nature of this discovery confirms the unknown metabolic potential of the Actinobacteria phylum and the importance of continuing screening studies in this taxon. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03305-0.

Mycothiol Peroxidase Activity as a Part of the Self-Resistance Mechanisms against the Antitumor Antibiotic Cosmomycin D.

Antibiotic-producing microorganisms usually require one or more self-resistance determinants to survive antibiotic production. The effectors of these mechanisms are proteins that inactivate the antibiotic, facilitate its transport, or modify the target to render it insensitive to the molecule. Streptomyces bacteria biosynthesize various bioactive natural products and possess resistance systems for most metabolites, which are coregulated with antibiotic biosynthesis genes. Streptomyces olindensis strain DAUFPE 5622 produces the antitumor antibiotic cosmomycin D (COSD), a member of the anthracycline family. In this study, we propose three self-resistance mechanisms, anchored or based in the COSD biosynthetic gene cluster. These include cosIJ (an ABC transporter), cosU (a UvrA class IIa protein), and a new self-resistance mechanism encoded by cosP, which shows response against peroxides by the enzyme mycothiol peroxidase (MPx). Activity-based investigations of MPx and its mutant enzyme confirmed peroxidation during the production of COSD. Overexpression of the ABC transporter, the UvrA class IIa protein, and the MPx led to an effective response against toxic anthracyclines, such as cosmomycins. Our findings help to understand how thiol peroxidases play an antioxidant role in the anthracycline producer S. olindensis DAUFPE 5622, a mechanism which has been reported for neoplastic cells that are resistant to doxorubicin (DOX). IMPORTANCE Anthracycline compounds are DNA intercalating agents widely used in cancer chemotherapeutic protocols. This work focused on the self-resistance mechanisms developed by the cosmomycin-producing bacterium Streptomyces olindensis. Our findings showed that cysteine peroxidases, such as mycothiol peroxidase, encoded by the gene cosP, protected S. olindensis against peroxidation during cosmomycin production. This observation can contribute to much better understanding of resistance both in the producers, eventually enhancing production, and in some tumoral cell lines.

Late hemotoxicity following North American rattlesnake envenomation treated with crotalidae immune F(ab')2 (equine) antivenom and crotalidae immune polyvalent Fab (ovine) antivenom reported to the North American Snakebite Sub-registry.

INTRODUCTION: Late hemotoxicity is common following rattlesnake envenomation treated with crotalidae immune polyvalent Fab (ovine) (FabAV). Initial clinical trials showed crotalidae immune F(ab')2 (equine) (Fab2AV) to be superior to FabAV in preventing late hemotoxicity, but this effect has not been demonstrated in broader populations. This study investigated late hemotoxicity in patients receiving Fab2AV or FabAV after rattlesnake envenomation. METHODS: This is a retrospective analysis of prospectively collected data from patients with snakebite reported to the ToxIC North American Snakebite Registry (NASBR) between January 1, 2019, and December 31, 2020. Inclusion criteria were rattlesnake envenomation and administration of antivenom. Patients were excluded if they received more than one type of antivenom. The primary outcome was occurrence of late hemotoxicity (platelets ≤120 k/mm3 or fibrinogen ≤170 mg/dL) in patients receiving Fab2AV and FabAV. Data collected included demographics, envenomation characteristics, laboratory values, and treatment administered. Statistics including t-test and Fisher's exact test were used. RESULTS: A total of 201 rattlesnake envenomated patients receiving antivenom were reported to the NASBR in the study period; 144 were included. 49 received Fab2AV alone, 45 received FabAV alone and 50 received both antivenoms. Baseline patient and envenomation characteristics were similar between the groups. Late hemotoxicity occurred in 2/49 patients in the Fab2AV group (4% (95% CI 0.7-12.6)) and in 19/45 patients in the FabAV group (42% (95% CI 28.4-59.0); absolute risk reduction 39.1% (95% CI 21.2-46.2) (p = 0.001). On follow up, 0 patients (0%) receiving Fab2AV were retreated with antivenom; 4 patients (9%) receiving FabAV were retreated (p = 0.049). CONCLUSIONS: In the North American Snakebite Registry, late hemotoxicity was less common in rattlesnake envenomated patients treated with Fab2AV compared to FabAV.

Biopolymer production by halotolerant bacteria isolated from Caatinga biome.

Saline environments are extreme habitats with a high diversity of microorganisms source of a myriad of biomolecules. These microorganisms are assigned as extremophiles recognized to be producers of new natural compounds, which can be synthesized by helping to survive under harshness and extreme conditions. In Brazil, in the saline and semi-arid region of Areia Branca (Caatinga biome), halotolerant bacteria (able to growth at high NaCl concentrations) were isolated from rhizosphere of native plants Blutaparon portulacoides and Spergularia sp. and their biopolymer production was studied. A total of 25 bacterial isolates were identified at genus level based on 16S rRNA gene sequence analysis. Isolates were mainly Gram-positive bacteria from Bacillaceae, Staphylococcaceae, Microbacteriaceae, and Bacillales XII incertae sedis families, affiliates to Bacillus, Staphylococcus, Curtobacterium, and Exiguobacterium genera, respectively. One of the Gram-negative isolates was identified as member of the Pseudomonadaceae family, genus Pseudomonas. All the identified strains were halotolerant bacteria with optimum growth at 0.6-2.0 M salt concentrations. Assays for biopolymer production showed that the halotolerant strains are a rich source of compounds as polyhydroxyalkanoates (PHA), biodegradable biopolymer, such as poly(3-hydroxybutyrate) (PHB) produced from low-cost substrates, and exopolysaccharides (EPS), such as hyaluronic acid (HA), metabolite of great interest to the cosmetic and pharmaceutical industry. Also, eight bacterial EPS extracts showed immunostimulatory activity, promising results that can be used in biomedical applications. Overall, our findings demonstrate that these biomolecules can be produced in culture medium with 0.6-2.0 M NaCl concentrations, relevant feature to avoid costly production processes. This is the first report of biopolymer-producing bacteria from a saline region of Caatinga biome that showed important biological activities.

Unveiling Xylanolytic Enzymes Production of Talaromyces wortmannii DR49 on Industrial Agro Wastes

Abstract Xylan degradation is an important step in different industries, such as in biorefinery for biomass hydrolysis. Talaromyces wortmannii is a known fungus due to second metabolite production but only few works showed the xylanolytic potential of this fungus. In this way, the aim of this study was to evaluate the production of xylanolytic enzymes from T. wortmannii DR49 on industrial agro wastes. Cultivation in shake flask showed highest xylanase titration (10.3 U/mL; 9.5 U/mL) for wheat bran (WB) and hydrothermal pretreated sugar cane bagasse (HB); in β-xylosidase production WB and xylose were the best carbon sources (0.57 U/mL; 0.34 U/mL) respectively. STR cultivation revealed that 29°C and pH 6.0 were the best conditions for xylanase (14.5 U/mL) and β-xylosidase (1.7 U/mL) production. T. wortmannii DR49 showed to be a potential candidate for xylanolytic enzymes production using agro wastes in bioreactors, which has never been previously reported in this fungus.

Metabolomic study of marine Streptomyces sp.: Secondary metabolites and the production of potential anticancer compounds.

Resorting to a One Strain Many Compounds (OSMAC) approach, the marine Streptomyces sp. BRB081 strain was grown in six different media settings over 1, 2, 3 or 7 days. Extractions of mycelium and broth were conducted separately for each media and cultivation period by sonication using methanol/acetone 1:1 and agitation with ethyl acetate, respectively. All methanol/acetone and ethyl acetate crude extracts were analysed by HPLC-MS/MS and data treatment was performed through GNPS platform using MZmine 2 software. In parallel, the genome was sequenced, assembled and mined to search for biosynthetic gene clusters (BGC) of secondary metabolites using the AntiSMASH 5.0 software. Spectral library search tool allowed the annotation of desferrioxamines, fatty acid amides, diketopiperazines, xanthurenic acid and, remarkably, the cyclic octapeptides surugamides. Molecular network analysis allowed the observation of the surugamides cluster, where surugamide A and the protonated molecule corresponding to the B-E isomers, as well as two potentially new analogues, were detected. Data treatment through MZmine 2 software allowed to distinguish that the largest amount of surugamides was obtained by cultivating BRB081 in SCB medium during 7 days and extraction of culture broth. Using the same data treatment, a chemical barcode was created for easy visualization and comparison of the metabolites produced overtime in all media. By genome mining of BRB081 four regions of biosynthetic gene clusters of secondary metabolites were detected supporting the metabolic data. Cytotoxic evaluation of all crude extracts using MTT assay revealed the highest bioactivity was also observed for extracts obtained in the optimal conditions as those for surugamides production, suggesting these to be the main active compounds herein. This method allowed the identification of compounds in the crude extracts and guided the selection of best conditions for production of bioactive compounds.

Reciprocal transplantation of the heterotrophic coral Tubastraea coccinea (Scleractinia: Dendrophylliidae) between distinct habitats did not alter its venom toxin composition.

Tubastraea coccinea is an azooxanthellate coral species recorded in the Indian and Atlantic oceans and is presently widespread in the southwestern Atlantic with an alien status for Brazil. T. coccinea outcompete other native coral species by using a varied repertoire of biological traits. For example, T. coccinea has evolved potent venom capable of immobilizing and digesting zooplankton prey. Diversification and modification of venom toxins can provide potential adaptive benefits to individual fitness, yet acquired alteration of venom composition in cnidarians is poorly understood as the adaptive flexibility affecting toxin composition in these ancient lineages has been largely ignored. We used quantitative high-throughput proteomics to detect changes in toxin expression in clonal fragments of specimens collected and interchanged from two environmentally distinct and geographically separate study sites. Unexpectedly, despite global changes in protein expression, there were no changes in the composition and abundance of toxins from coral fragments recovered from either site, and following clonal transplantation between sites. There were also no apparent changes to the cnidome (cnidae) and gross skeletal or soft tissue morphologies of the specimens. These results suggest that the conserved toxin complexity of T. coccinea co-evolved with innovation of the venom delivery system, and its morphological development and phenotypic expression are not modulated by habitat pressures over short periods of time. The adaptive response of the venom trait to specific predatory regimes, however, necessitates further consideration.

Marine Bacteria from Rocas Atoll as a Rich Source of Pharmacologically Active Compounds.

Rocas Atoll is a unique environment in the equatorial Atlantic Ocean, hosting a large number of endemic species, however, studies on the chemical diversity emerging from this biota are rather scarce. Therefore, the present work aims to assess the metabolomic diversity and pharmacological potential of the microbiota from Rocas Atoll. A total of 76 bacteria were isolated and cultured in liquid culture media to obtain crude extracts. About one third (34%) of these extracts were recognized as cytotoxic against human colon adenocarcinoma HCT-116 cell line. 16S rRNA gene sequencing analyses revealed that the bacteria producing cytotoxic extracts were mainly from the Actinobacteria phylum, including Streptomyces, Salinispora, Nocardiopsis, and Brevibacterium genera, and in a smaller proportion from Firmicutes phylum (Bacillus). The search in the spectral library in GNPS (Global Natural Products Social Molecular Networking) unveiled a high chemodiversity being produced by these bacteria, including rifamycins, antimycins, desferrioxamines, ferrioxamines, surfactins, surugamides, staurosporines, and saliniketals, along with several unidentified compounds. Using an original approach, molecular networking successfully highlighted groups of compounds responsible for the cytotoxicity of crude extracts. Application of DEREPLICATOR+ (GNPS) allowed the annotation of macrolide novonestimycin derivatives as the cytotoxic compounds existing in the extracts produced by Streptomyces BRB-298 and BRB-302. Overall, these results highlighted the pharmacological potential of bacteria from this singular atoll.

Venom Composition Does Not Vary Greatly Between Different Nematocyst Types Isolated from the Primary Tentacles of Olindias sambaquiensis (Cnidaria: Hydrozoa).

In this quantitative proteomics study we determined the variety and relative abundance of toxins present in enriched preparations of two nematocyst types isolated from the primary tentacles of the adult medusa stage of the hydrozoan Olindias sambaquiensis. The two nematocyst types were microbasic mastigophores and microbasic euryteles, and these were recovered from the macerated tentacle tissues by using a differential centrifugation approach. Soluble protein extracts from these nematocysts were tagged with tandem mass tag isobaric labels and putative toxins identified using tandem mass spectrometry coupled with a stringent bioinformatics annotation pipeline. Astonishingly, the venom composition of the two capsule types was nearly identical, and there was also little difference in the comparative abundance of toxins between the two nematocyst preparations. This homogeneity suggested that the same toxin complement was present regardless of the penetrative ability of the nematocyst type. Predicted toxin protein families that constituted the venom closely matched those of the toxic proteome of O. sambaquiensis published four years previously, suggesting that venom composition in this species changes little over time. Retaining an array of different nematocyst types to deliver a single venom, rather than sustaining the high metabolic cost necessary to maintain a dynamically evolving venom, may be more advantageous, given the vastly different interspecific interactions that adult medusa encounter in coastal zones.

Genome Sequence of Streptomyces cavourensis 1AS2a, a Rhizobacterium Isolated from the Brazilian Cerrado Biome.

Streptomyces cavourensis strain 1AS2a, isolated from wheat rhizosphere in the Brazilian Neotropical savanna, exhibits strong antimicrobial activities. Its genome comprises 7,600,475 bp with 6,590 open reading frames (ORFs) that reveal 30 biosynthetic gene clusters (BGCs). It provides a genetic basis for further research of the potential of this strain for the production of antimicrobial compounds.

Endophytic Actinomycetes as Potential Producers of Hemicellulases and Related Enzymes for Plant Biomass Degradation

Abstract Tailor made enzymatic preparation must be design to hydrolyze efficiently plant biomass, once that each plant biomass possesses a distinct cell wall composition. Most of actinomycetes used for plant cell wall degradation are focused on the cellulases and xylanases production. However, a wide range of enzymes must be produced for an efficient degradation of lignocellulose materials. During the last decade several unusual environments were studied to obtain strains that produce glycohydrolases with innovator characteristics. In this context, the present work concerned the selection of endophytic actinomycetes as producers of hemicellulases and related enzymes with different enzymatic profiles, for use in the deconstruction of lignocellulosic biomass. A total of 45 Brazilian actinomycetes previously isolated from plants (endophytics) and soil were prospected for hemicellulases and β-glucosidase production. Four strains highlighted for hemicellulase production (DR61, DR63, DR69 and DR66) and were selected for cultivation under other inductors substrates (xylan and pectin). All strains belong to Streptomyces genera and have their extracts tested for degradation of several hemicellulolytic substrates. The strains presented different glicohydrolyse enzymes profiles mainly for xylans and glucans that can be used for specific formulations of enzymes applied on the biomass deconstruction, principally on sugar cane bagasse.

Correction for Ichiwaki et al., "Genome Sequence of Micromonospora sp. NBS 11-29, an Antibiotic and Hydrolytic Enzyme Producer, Isolated from River Sediment in Brazil".

[This corrects the article DOI: 10.1128/genomeA.00552-17.].

The biotechnological potential of Epicoccum spp.: diversity of secondary metabolites.

Epicoccum is a genus of ubiquitous fungi typically found in air, in soil, and on decaying vegetation. They also commonly display an endophytic lifestyle and are isolated from diverse plant tissues. The fungi from the genus Epicoccum are mainly known for their use as biocontrol agents against phytopathogens and for their ability to produce many secondary metabolites with potential biotechnological applications, such as antioxidant, anticancer,r and antimicrobial compounds. Among the bioactive compounds produced by Epicoccum spp., epicocconone is a commercially available fluorophore, D8646-2-6 is a patented telomerase inhibitor, and taxol is an anticancer drug originally isolated from Taxus brevifolia. Epicoccum spp. also produces epicolactone, an antimicrobial compound with a unique and complex structure that has aroused considerable interest in the chemical-synthesis community. The main goal of the present review is to discuss the diversity of secondary metabolites produced by Epicoccum spp., their biotechnological applications, and proposed hypothetical biosynthesis. In addition, the use of Epicoccum spp. as biocontrol agents and the pigments produced by these fungi are also discussed.

Influence of water quality on diversity and composition of fungal communities in a tropical river.

Freshwater fungi are key decomposers of organic material and play important roles in nutrient cycling, bio-remediation and ecosystem functioning. Although aquatic fungal communities respond to pollution, few studies have quantitatively assessed the effect of freshwater contamination on fungal diversity and composition; and knowledge is scarcer for tropical systems. Here we help fill this knowledge gap by studying a heavily-contaminated South American river spanning a biodiversity hotspot. We collected 30 water samples scattered across a quality gradient over two seasons and analyzed them using Terminal Restriction Fragment Length Polymorphisms (T-RFLP) coupled with 454 Pyrosequencing. Using T-RFLP we identified 451 and 442 Operational Taxonomy Units (OTUs) in the dry and rainy seasons respectively, whereas Pyrosequencing revealed 48,553 OTUs from which 11% were shared between seasons. Although 68% of all identified OTUs and 51% of all identified phyla remained unidentified, dominant fungal phyla included the Ascomycota, Basidiomycota, Chytridiomycota, Glomeromycota, Zygomycota and Neocallimastigomycota, while Calcarisporiella, Didymosphaeria, Mycosphaerella (Ascomycota) and Rhodotorula (Basidiomycota) were the most abundant genera. Fungal diversity was affected by pH and dissolved iron, while community composition was influenced by dissolved oxygen, pH, nitrate, biological oxygen demand, total aluminum, total organic carbon, total iron and seasonality. The presence of potentially pathogenic species was associated with high pH. Furthermore, geographic distance was positively associated with community dissimilarity, suggesting that local conditions allowed divergence among fungal communities. Overall, our findings raise potential concerns for human health and the functioning of tropical river ecosystems and they call for improved water sanitation systems.

Corrigendum: Pectins, Endopolygalacturonases, and Bioenergy.

[This corrects the article on p. 1401 in vol. 7, PMID: 27703463.].

Genome Sequence of Micromonospora sp. NBS 11-29, an Antibiotic and Hydrolytic Enzyme Producer, Isolated from River Sediment in Brazil.

The genus Micromonospora comprises actinomycetes with high biotechnological potential, due to their ability to produce secondary metabolites and enzymes. In this study, we report the draft genome sequence of Micromonospora sp. NBS 11-29, which showed antibacterial, cellulolytic, and xylanolytic activities under in vitro conditions.

Comparative proteomics reveals recruitment patterns of some protein families in the venoms of Cnidaria.

Cnidarians are probably the oldest group of animals to be venomous, yet our current picture of cnidarian venom evolution is highly imbalanced due to limited taxon sampling. High-throughput tandem mass spectrometry was used to determine venom composition of the scyphozoan Chrysaora lactea and two cubozoans Tamoya haplonema and Chiropsalmus quadrumanus. Protein recruitment patterns were then compared against 5 other cnidarian venom proteomes taken from the literature. A total of 28 putative toxin protein families were identified, many for the first time in Cnidaria. Character mapping analysis revealed that 17 toxin protein families with predominantly cytolytic biological activities were likely recruited into the cnidarian venom proteome before the lineage split between Anthozoa and Medusozoa. Thereafter, venoms of Medusozoa and Anthozoa differed during subsequent divergence of cnidarian classes. Recruitment and loss of toxin protein families did not correlate with accepted phylogenetic patterns of Cnidaria. Selective pressures that drive toxin diversification independent of taxonomic positioning have yet to be identified in Cnidaria and now warrant experimental consideration.