Social wasp-associated Tsukamurella sp. strains showed promising biosynthetic and bioactive potential for discovery of novel compounds.

In the face of escalating antibiotic resistance, the quest for novel antimicrobial compounds is critical. Actinobacteria is known for producing a substantial fraction of bioactive molecules from microorganisms, nonetheless there is the challenge of metabolic redundancy in bioprospecting. New sources of natural products are needed to overcome these current challenges. Our present work proposes an unexplored potential of Neotropical social wasp-associated microbes as reservoirs of novel bioactive compounds. Using social wasp-associated Tsukamurella sp. strains 8F and 8J, we aimed to determine their biosynthetic potential for producing novel antibiotics and evaluated phylogenetic and genomic traits related to environmental and ecological factors that might be associated with promising bioactivity and evolutionary specialization. These strains were isolated from the cuticle of social wasps and subjected to comprehensive genome sequencing. Our genome mining efforts, employing antiSMASH and ARTS, highlight the presence of BGCs with minimal similarity to known compounds, suggesting the novelty of the molecules they may produce. Previous, bioactivity assays of these strains against bacterial species which harbor known human pathogens, revealed inhibitory potential. Further, our study focuses into the phylogenetic and functional landscape of the Tsukamurella genus, employing a throughout phylogenetic analysis that situates strains 8F and 8J within a distinct evolutionary pathway, matching with the environmental and ecological context of the strains reported for this genus. Our findings emphasize the importance of bioprospecting in uncharted biological territories, such as insect-associated microbes as reservoirs of novel bioactive compounds. As such, we posit that Tsukamurella sp. strains 8F and 8J represent promising candidates for the development of new antimicrobials.

Discovery of antimicrobial peptides clostrisin and cellulosin from Clostridium: insights into their structures, co-localized biosynthetic gene clusters, and antibiotic activity.

Antimicrobial resistance presents a substantial threat to global public health, demanding urgent attention and action. This study focuses on lanthipeptides, ribosomally encoded peptides that display significant structural diversity and hold promising potential as antibiotics. Genome mining was employed to locate biosynthetic gene clusters (BGCs) containing class II lanthipeptide synthetases encoded by lanM genes. A phylogenetic study analyzing homologous sequences of functional LanM sequences revealed a unique evolutionary clade of 17 LanM proteins associated with 12 Clostridium bacterial genomes. In silico exploration identified nine complete BGCs, including one super-cluster containing two co-localized operons from Clostridium cellulovorans 743B, that encode for two new peptides named clostrisin and cellulosin. Each operon was heterologously expressed in Escherichia coli. Molecular weights associated with the expected post-translational modifications of the purified lanthipeptide were confirmed by MS-MS/MS analysis for cellulosin, while clostrisin was not post-translationally modified. Both peptides demonstrated antimicrobial activity against multidrug-resistant bacteria, such as a clinical strain of Staphylococcus epidermidis MIQ43 and Pseudomonas aeruginosa PA14. This is the first report of lanthipeptides from the Clostridium genus produced with its native biosynthetic machinery, as well as chemically and biologically characterized. This study showcases the immense potential of genome mining in identifying new RiPP synthetases and associated bioactive peptides.

Combined effect of SAR-endolysin LysKpV475 with polymyxin B and Salmonella bacteriophage phSE-5.

Endolysins are bacteriophage (or phage)-encoded enzymes that catalyse the peptidoglycan breakdown in the bacterial cell wall. The exogenous action of recombinant phage endolysins against Gram-positive organisms has been extensively studied. However, the outer membrane acts as a physical barrier when considering the use of recombinant endolysins to combat Gram-negative bacteria. This study aimed to evaluate the antimicrobial activity of the SAR-endolysin LysKpV475 against Gram-negative bacteria as single or combined therapies, using an outer membrane permeabilizer (polymyxin B) and a phage, free or immobilized in a pullulan matrix. In the first step, the endolysin LysKpV475 in solution, alone and combined with polymyxin B, was tested in vitro and in vivo against ten Gram-negative bacteria, including highly virulent strains and multidrug-resistant isolates. In the second step, the lyophilized LysKpV475 endolysin was combined with the phage phSE-5 and investigated, free or immobilized in a pullulan matrix, against Salmonella enterica subsp. enterica serovar Typhimurium ATCC 13311. The bacteriostatic action of purified LysKpV475 varied between 8.125 µg ml-1 against Pseudomonas aeruginosa ATCC 27853, 16.25 µg ml-1 against S. enterica Typhimurium ATCC 13311, and 32.50 µg ml-1 against Klebsiella pneumoniae ATCC BAA-2146 and Enterobacter cloacae P2224. LysKpV475 showed bactericidal activity only for P. aeruginosa ATCC 27853 (32.50 µg ml-1) and P. aeruginosa P2307 (65.00 µg ml-1) at the tested concentrations. The effect of the LysKpV475 combined with polymyxin B increased against K. pneumoniae ATCC BAA-2146 [fractional inhibitory concentration index (FICI) 0.34; a value lower than 1.0 indicates an additive/combined effect] and S. enterica Typhimurium ATCC 13311 (FICI 0.93). A synergistic effect against S. enterica Typhimurium was also observed when the lyophilized LysKpV475 at ⅔ MIC was combined with the phage phSE-5 (m.o.i. of 100). The lyophilized LysKpV475 immobilized in a pullulan matrix maintained a significant Salmonella reduction of 2 logs after 6 h of treatment. These results demonstrate the potential of SAR-endolysins, alone or in combination with other treatments, in the free form or immobilized in solid matrices, which paves the way for their application in different areas, such as in biocontrol at the food processing stage, biosanitation of food contact surfaces and biopreservation of processed food in active food packing.

Genome-guided purification of high amounts of the siderophore ornibactin and detection of potentially novel burkholdine derivatives produced by Burkholderia catarinensis 89T.

AIM: The increased availability of genome sequences has enabled the development of valuable tools for the prediction and identification of bacterial natural products. Burkholderia catarinensis 89T produces siderophores and an unknown potent antifungal metabolite. The aim of this work was to identify and purify natural products of B. catarinensis 89T through a genome-guided approach. MATERIALS AND METHODS: The analysis of B. catarinensis 89T genome revealed 16 clusters putatively related to secondary metabolism and antibiotics production. Of particular note was the identification of a nonribosomal peptide synthetase (NRPS) cluster related to the production of the siderophore ornibactin, a hybrid NRPS-polyketide synthase Type 1 cluster for the production of the antifungal glycolipopeptide burkholdine, and a gene cluster encoding homoserine lactones (HSL), probably involved in the regulation of both metabolites. We were able to purify high amounts of the ornibactin derivatives D/C6 and F/C8, while also detecting the derivative B/C4 in mass spectrometry investigations. A group of metabolites with molecular masses ranging from 1188 to 1272 Da could be detected in MS experiments, which we postulate to be new burkholdine analogs produced by B. catarinensis. The comparison of B. catarinensis BGCs with other Bcc members corroborates the hypothesis that this bacterium could produce new derivatives of these metabolites. Moreover, the quorum sensing metabolites C6-HSL, C8-HSL, and 3OH-C8-HSL were observed in LC-MS/MS analysis. CONCLUSION: The new species B. catarinensis is a potential source of new bioactive secondary metabolites. Our results highlight the importance of genome-guided purification and identification of metabolites of biotechnological importance.

Genome analysis of haloalkaline isolates from the soda saline crater lake of Isabel Island; comparative genomics and potential metabolic analysis within the genus Halomonas.

BACKGROUND: Isabel Island is a Mexican volcanic island primarily composed of basaltic stones. It features a maar known as Laguna Fragatas, which is classified as a meromictic thalassohaline lake. The constant deposition of guano in this maar results in increased levels of phosphorus, nitrogen, and carbon. The aim of this study was to utilize high-quality genomes from the genus Halomonas found in specialized databases as a reference for genome mining of moderately halophilic bacteria isolated from Laguna Fragatas. This research involved genomic comparisons employing phylogenetic, pangenomic, and metabolic-inference approaches. RESULTS: The Halomonas genus exhibited a large open pangenome, but several genes associated with salt metabolism and homeostatic regulation (ectABC and betABC), nitrogen intake through nitrate and nitrite transporters (nasA, and narGI), and phosphorus uptake (pstABCS) were shared among the Halomonas isolates. CONCLUSIONS: The isolated bacteria demonstrate consistent adaptation to high salt concentrations, and their nitrogen and phosphorus uptake mechanisms are highly optimized. This optimization is expected in an extremophile environment characterized by minimal disturbances or abrupt seasonal variations. The primary significance of this study lies in the dearth of genomic information available for this saline and low-disturbance environment. This makes it important for ecosystem conservation and enabling an exploration of its biotechnological potential. Additionally, the study presents the first two draft genomes of H. janggokensis.

Complete genome sequence of Pediococcus acidilactici A40, a bacterium with biocontrol and plant growth-promoting properties.

We report the complete genome assembly of Pediococcus acidilactici A40, a bacterium with biocontrol and plant growth-promoting properties, obtained from Colombia.

Whole-genome sequencing-based characterization of Streptomyces sp. 6(4): focus on natural product.

We have sequenced the whole genome of Streptomyces sp. 6(4) isolated from tomato roots that presents antifungal activity against phytopathogenic fungi, mainly Bipolaris sorokiniana. The genome has almost 7 Mb and 3368 hypothetical proteins that were analysed and characterized in Uniprot with the emphasis on biological compounds. Multilocus sequence typing (MLST) analyses were performed in an effort to characterize and identify this isolate, resulting in a new sequence type (ST), classified as ST64. Phenetic and phylogenetic trees were constructed to investigate Streptomyces sp. 6(4) evolution and sequence similarity, and the isolate is a strain closer to Streptomyces prasinus and Streptomyces viridosporus . It is known that the genus Streptomyces possess huge metabolic capacity with the presence of cryptic genes. These genes are usually present in clusters, which are responsible for the production of diverse natural products, mainly antibiotics. In addition, 6(4) showed 11 biosynthetic gene clusters through antiSMASH, including 3 polyketide synthase (PKS) and non-ribosomal peptide synthase (NRPS) type clusters.

In silico characterization of cysteine-stabilized αß defensins from neglected unicellular microeukaryotes.

BACKGROUND: The emergence of multi-resistant pathogens have increased dramatically in recent years, becoming a major public-health concern. Among other promising antimicrobial molecules with potential to assist in this worldwide struggle, cysteine-stabilized αß (CS-αß) defensins are attracting attention due their efficacy, stability, and broad spectrum against viruses, bacteria, fungi, and protists, including many known human pathogens. RESULTS: Here, 23 genomes of ciliated protists were screened and two CS-αß defensins with a likely antifungal activity were identified and characterized, using bioinformatics, from a culturable freshwater species, Laurentiella sp. (LsAMP-1 and LsAMP-2). Although any potential cellular ligand could be predicted for LsAMP-2; evidences from structural, molecular dynamics, and docking analyses suggest that LsAMP-1 may form stably associations with phosphatidylinositol 4,5-bisphosphates (PIP2), a phospholipid found on many eukaryotic cells, which could, in turn, represent an anchorage mechanism within plasma membrane of targeted cells. CONCLUSION: These data stress that more biotechnology-oriented studies should be conducted on neglected protists, such ciliates, which could become valuable sources of novel bioactive molecules for therapeutic uses.

Biosynthetic novelty index reveals the metabolic potential of rare actinobacteria isolated from highly oligotrophic sediments.

Calculations predict that testing of 5 000-10 000 molecules and >1 billion US dollars (£0.8 billion, £1=$1.2) are required for one single drug to come to the market. A solution to this problem is to establish more efficient protocols that reduce the high rate of re-isolation and continuous rediscovery of natural products during early stages of the drug development process. The study of 'rare actinobacteria' has emerged as a possible approach for increasing the discovery rate of drug leads from natural sources. Here, we define a simple genomic metric, defined as biosynthetic novelty index (BiNI), that can be used to rapidly rank strains according to the novelty of the subset of encoding biosynthetic clusters. By comparing a subset of high-quality genomes from strains of different taxonomic and ecological backgrounds, we used the BiNI score to support the notion that rare actinobacteria encode more biosynthetic gene cluster (BGC) novelty. In addition, we present the isolation and genomic characterization, focused on specialized metabolites and phenotypic screening, of two isolates belonging to genera Lentzea and Actinokineospora from a highly oligotrophic environment. Our results show that both strains harbour a unique subset of BGCs compared to other members of the genera Lentzea and Actinokineospora. These BGCs are responsible for potent antimicrobial and cytotoxic bioactivity. The experimental data and analysis presented in this study contribute to the knowledge of genome mining analysis in rare actinobacteria and, most importantly, can serve to direct sampling efforts to accelerate early stages of the drug discovery pipeline.

Antarctic Sphingomonas sp. So64.6b showed evolutive divergence within its genus, including new biosynthetic gene clusters.

Introduction: The antibiotic crisis is a major human health problem. Bioprospecting screenings suggest that proteobacteria and other extremophile microorganisms have biosynthetic potential for the production novel antimicrobial compounds. An Antarctic Sphingomonas strain (So64.6b) previously showed interesting antibiotic activity and elicitation response, then a relationship between environmental adaptations and its biosynthetic potential was hypothesized. We aimed to determine the genomic characteristics in So64.6b strain related to evolutive traits for the adaptation to the Antarctic environment that could lead to its diversity of potentially novel antibiotic metabolites. Methods: The complete genome sequence of the Antarctic strain was obtained and mined for Biosynthetic Gene Clusters (BGCs) and other unique genes related to adaptation to extreme environments. Comparative genome analysis based on multi-locus phylogenomics, BGC phylogeny, and pangenomics were conducted within the closest genus, aiming to determine the taxonomic affiliation and differential characteristics of the Antarctic strain. Results and discussion: The Antarctic strain So64.6b showed a closest identity with Sphingomonas alpina, however containing a significant genomic difference of ortholog cluster related to degradation multiple pollutants. Strain So64.6b had a total of six BGC, which were predicted with low to no similarity with other reported clusters; three were associated with potential novel antibiotic compounds using ARTS tool. Phylogenetic and synteny analysis of a common BGC showed great diversity between Sphingomonas genus but grouping in clades according to similar isolation environments, suggesting an evolution of BGCs that could be linked to the specific ecosystems. Comparative genomic analysis also showed that Sphingomonas species isolated from extreme environments had the greatest number of predicted BGCs and a higher percentage of genetic content devoted to BGCs than the isolates from mesophilic environments. In addition, some extreme-exclusive clusters were found related to oxidative and thermal stress adaptations, while pangenome analysis showed unique resistance genes on the Antarctic strain included in genetic islands. Altogether, our results showed the unique genetic content on Antarctic strain Sphingomonas sp. So64.6, -a probable new species of this genetically divergent genus-, which could have potentially novel antibiotic compounds acquired to cope with Antarctic poly-extreme conditions.

Agronomic efficiency and genome mining analysis of the wheat-biostimulant rhizospheric bacterium Pseudomonas pergaminensis sp. nov. strain 1008T.

Pseudomonas sp. strain 1008 was isolated from the rhizosphere of field grown wheat plants at the tillering stage in an agricultural plot near Pergamino city, Argentina. Based on its in vitro phosphate solubilizing capacity and the production of IAA, strain 1008 was formulated as an inoculant for bacterization of wheat seeds and subjected to multiple field assays within the period 2010-2017. Pseudomonas sp. strain 1008 showed a robust positive impact on the grain yield (+8% on average) across a number of campaigns, soil properties, seed genotypes, and with no significant influence of the simultaneous seed treatment with a fungicide, strongly supporting the use of this biostimulant bacterium as an agricultural input for promoting the yield of wheat. Full genome sequencing revealed that strain 1008 has the capacity to access a number of sources of inorganic and organic phosphorus, to compete for iron scavenging, to produce auxin, 2,3-butanediol and acetoin, and to metabolize GABA. Additionally, the genome of strain 1008 harbors several loci related to rhizosphere competitiveness, but it is devoid of biosynthetic gene clusters for production of typical secondary metabolites of biocontrol representatives of the Pseudomonas genus. Finally, the phylogenomic, phenotypic, and chemotaxonomic comparative analysis of strain 1008 with related taxa strongly suggests that this wheat rhizospheric biostimulant isolate is a representative of a novel species within the genus Pseudomonas, for which the name Pseudomonas pergaminensis sp. nov. (type strain 1008T = DSM 113453T = ATCC TSD-287T) is proposed.

Production of Epoxyketone Peptide-Based Proteasome Inhibitors by Streptomyces sp. BRA-346: Regulation and Biosynthesis.

Streptomyces sp. BRA-346 is an Actinobacteria isolated from the Brazilian endemic tunicate Euherdmania sp. We have reported that this strain produces epoxyketone peptides, as dihydroeponemycin (DHE) and structurally related analogs. This cocktail of epoxyketone peptides inhibits the proteasome chymotrypsin-like activity and shows high cytotoxicity to glioma cells. However, low yields and poor reproducibility of epoxyketone peptides production by BRA-346 under laboratory cultivation have limited the isolation of epoxyketone peptides for additional studies. Here, we evaluated several cultivation methods using different culture media and chemical elicitors to increase the repertoire of peptide epoxyketone production by this bacterium. Furthermore, BRA-346 genome was sequenced, revealing its broad genetic potential, which is mostly hidden under laboratory conditions. By using specific growth conditions, we were able to evidence different classes of secondary metabolites produced by BRA-346. In addition, by combining genome mining with untargeted metabolomics, we could link the metabolites produced by BRA-346 to its genetic capacity and potential regulators. A single biosynthetic gene cluster (BGC) was related to the production of the target epoxyketone peptides by BRA-346. The candidate BGC displays conserved biosynthetic enzymes with the reported eponemycin (EPN) and TMC-86A (TMC) BGCs. The core of the putative epoxyketone peptide BGC (ORFs A-L), in which ORF A is a LuxR-like transcription factor, was cloned into a heterologous host. The recombinant organism was capable to produce TMC and EPN natural products, along with the biosynthetic intermediates DH-TMC and DHE, and additional congeners. A phylogenetic analysis of the epn/tmc BGC revealed related BGCs in public databases. Most of them carry a proteasome beta-subunit, however, lacking an assigned specialized metabolite. The retrieved BGCs also display a diversity of regulatory genes and TTA codons, indicating tight regulation of this BGC at the transcription and translational levels. These results demonstrate the plasticity of the epn/tmc BGC of BRA-346 in producing epoxyketone peptides and the feasibility of their production in a heterologous host. This work also highlights the capacity of BRA-346 to tightly regulate its secondary metabolism and shed light on how to awake silent gene clusters of Streptomyces sp. BRA-346 to allow the production of pharmacologically important biosynthetic products.

Bioprospecting of Ribosomally Synthesized and Post-translationally Modified Peptides Through Genome Characterization of a Novel Probiotic Lactiplantibacillus plantarum UTNGt21A Strain: A Promising Natural Antimicrobials Factory.

The present work describes the genome sequencing and characterization of a novel Lactiplantibacillus plantarum strain assigned UTNGt21A isolated from wild Solanum quitoense (L.) fruits. In silico analysis has led to identifying a wide range of biosynthetic gene clusters (BGCs) and metabolic compounds. The genome had a total of 3,558,611 bp with GC of 43.96%, harboring 3,449 protein-coding genes, among which 3,209 were assigned by the EggNOG database, and 240 hypothetical proteins have no match in the BLASTN database. It also contains 68 tRNAs, 1 23S rRNA, 1 16S rRNA, 6 5S rRNA, and 1 tmRNA. In addition, no acquired resistance genes nor virulence and pathogenic factors were predicted, indicating that UTNGt21A is a safe strain. Three areas of interest (AOI) consisting of multiple genes encoding for bacteriocins and ABC transporters were predicted with BAGEL4, while eight secondary metabolite regions were predicted with the antiSMASH web tool. GutSMASH analysis predicted one metabolic gene cluster (MGC) type pyruvate to acetate-formate, a primary metabolite region essential for anaerobe growth. Several lanthipeptides and non-ribosomal peptide synthetase (NRPS) clusters were detected in the UTNGt21A but not the reference genomes, suggesting that their genome diversity might be linked to its niche-specific lineage and adaptation to a specific environment. Moreover, the application of a targeted genome mining tool (RiPPMiner) uncovered a diverse arsenal of important antimicrobial molecules such as lanthipeptides. Furthermore, in vitro analysis indicated that the crude extract (CE) of UTNGt21A exerted a wide spectrum of inhibition against several pathogens. The results indicated that the possible peptide-protein extract (PC) from UTNGt21A induces morphological and ultrastructural changes of Salmonella enterica subsp. enterica ATCC51741, compatible with its inhibitory potential. Genome characterization is the basis for further in vitro and in vivo studies to explore their use as antimicrobial producers or probiotic strains.

Evolutionary genomics and biosynthetic potential of novel environmental Actinobacteria.

Actinobacteria embroil Gram-positive microbes with high guanine and cytosine contents in their DNA. They are the source of most antimicrobials of bacterial origin utilized in medicine today. Their genomes are among the richest in novel secondary metabolites with high biotechnological potential. Actinobacteria reveal complex patterns of evolution, responses, and adaptations to their environment, which are not yet well understood. We analyzed three novel plant isolates and explored their habitat adaptation, evolutionary patterns, and potential secondary metabolite production. The phylogenomically characterized isolates belonged to Actinoplanes sp. TFC3, Streptomyces sp. L06, and Embleya sp. NF3. Positively selected genes, relevant in strain evolution, encoded enzymes for stress resistance in all strains, including porphyrin, chlorophyll, and ubiquinone biosynthesis in Embleya sp. NF3. Streptomyces sp. L06 encoded for pantothenate and proteins for CoA biosynthesis with evidence of positive selection; furthermore, Actinoplanes sp. TFC3 encoded for a c-di-GMP synthetase, with adaptive mutations. Notably, the genomes harbored many genes involved in the biosynthesis of at least ten novel secondary metabolites, with many avenues for future new bioactive compound characterization-specifically, Streptomyces sp. L06 could make new ribosomally synthesized and post-translationally modified peptides, while Embleya sp. NF3 could produce new non-ribosomal peptide synthetases and ribosomally synthesized and post-translationally modified peptides. At the same time, TFC3 has particularly enriched in terpene and polyketide synthases. All the strains harbored conserved genes in response to diverse environmental stresses, plant growth promotion factors, and degradation of various carbohydrates, which supported their endophytic lifestyle and showed their capacity to colonize other niches. This study aims to provide a comprehensive estimation of the genomic features of novel Actinobacteria. It sets the groundwork for future research into experimental tests with new bioactive metabolites with potential application in medicine, biofertilizers, and plant biomass residue utilization, with potential application in medicine, as biofertilizers and in plant biomass residues utilization. KEY POINTS: • Potential of novel environmental bacteria for secondary metabolites production • Exploring the genomes of three novel endophytes isolated from a medicinal tree • Pan-genome analysis of Actinobacteria genera.

Pan-Genome of the Genus Streptomyces and Prioritization of Biosynthetic Gene Clusters With Potential to Produce Antibiotic Compounds.

Species of the genus Streptomyces are known for their ability to produce multiple secondary metabolites; their genomes have been extensively explored to discover new bioactive compounds. The richness of genomic data currently available allows filtering for high quality genomes, which in turn permits reliable comparative genomics studies and an improved prediction of biosynthetic gene clusters (BGCs) through genome mining approaches. In this work, we used 121 genome sequences of the genus Streptomyces in a comparative genomics study with the aim of estimating the genomic diversity by protein domains content, sequence similarity of proteins and conservation of Intergenic Regions (IGRs). We also searched for BGCs but prioritizing those with potential antibiotic activity. Our analysis revealed that the pan-genome of the genus Streptomyces is clearly open, with a high quantity of unique gene families across the different species and that the IGRs are rarely conserved. We also described the phylogenetic relationships of the analyzed genomes using multiple markers, obtaining a trustworthy tree whose relationships were further validated by Average Nucleotide Identity (ANI) calculations. Finally, 33 biosynthetic gene clusters were detected to have potential antibiotic activity and a predicted mode of action, which might serve up as a guide to formulation of related experimental studies.

Genome Mining and Molecular Networking-Based Metabolomics of the Marine Facultative Aspergillus sp. MEXU 27854.

The marine-facultative Aspergillus sp. MEXU 27854, isolated from the Caleta Bay in Acapulco, Guerrero, Mexico, has provided an interesting diversity of secondary metabolites, including a series of rare dioxomorpholines, peptides, and butyrolactones. Here, we report on the genomic data, which consists of 11 contigs (N50~3.95 Mb) with a ~30.75 Mb total length of assembly. Genome annotation resulted in the prediction of 10,822 putative genes. Functional annotation was accomplished by BLAST searching protein sequences with different public databases. Of the predicted genes, 75% were assigned gene ontology terms. From the 67 BGCs identified, ~60% belong to the NRPS and NRPS-like classes. Putative BGCs for the dioxomorpholines and other metabolites were predicted by extensive genome mining. In addition, metabolomic molecular networking analysis allowed the annotation of all isolated compounds and revealed the biosynthetic potential of this fungus. This work represents the first report of whole-genome sequencing and annotation from a marine-facultative fungal strain isolated from Mexico.

Insights Into the Ecological Role of Pseudomonas spp. in an Ant-plant Symbiosis.

In the myrmecophytic mutualistic relationship between Azteca ants and Cecropia plants both species receive protection and exchange nutrients. The presence of microorganisms in this symbiotic system has been reported, and the symbiotic role of some fungi involved in the myrmecophytic interactions has been described. In this work we focus on bacteria within this mutualism, conducting isolations and screening for antimicrobial activities, genome sequencing, and biochemical characterization. We show that Pantoea, Rhizobium, Methylobacterium, Streptomyces and Pseudomonas are the most common cultivable genera of bacteria. Interestingly, Pseudomonas spp. isolates showed potent activity against 83% of the pathogens tested in our antimicrobial activity assays, including a phytopathogenic fungus isolated from Cecropia samples. Given the predicted nitrogen limitations associated with the fungal patches within this myrmecophyte, we performed nitrogen fixation analyses on the bacterial isolates within the Proteobacteria and show the potential for nitrogen fixation in Pseudomonas strains. The genome of one Pseudomonas strain was sequenced and analyzed. The gene cluster involved in the biosynthesis of cyclic lipodepsipeptides (CLPs) was identified, and we found mutations that may be related to the loss of function in the dual epimerization/condensation domains. The compound was isolated, and its structure was determined, corresponding to the antifungal viscosinamide. Our findings of diazotrophy and production of viscosinamide in multiple Pseudomonas isolates suggests that this bacterial genus may play an important role in the Cecropia-Azteca symbiosis.

Streptomyces sp. M54: an actinobacteria associated with a neotropical social wasp with high potential for antibiotic production.

Streptomyces symbionts in insects have shown to be a valuable source of new antibiotics. Here, we report the genome sequence and the potential for antibiotic production of "Streptomyces sp. M54", an Actinobacteria associated with the eusocial wasp, Polybia plebeja. The Streptomyces sp. M54 genome is composed of a chromosome (7.96 Mb), and a plasmid (1.91 Kb) and harbors 30 biosynthetic gene clusters for secondary metabolites, of which only one third has been previously characterized. Growth inhibition bioassays show that this bacterium produces antimicrobial compounds that are active against Hirsutella citriformis, a natural fungal enemy of its host, and the human pathogens Staphylococcus aureus and Candida albicans. Analyses through TLC-bioautography, LC-MS/MS and NMR allowed the identification of five macrocyclic ionophore antibiotics, with previously reported antibacterial, antitumor and antiviral properties. Phylogenetic analyses placed Streptomyces sp. M54 in a clade of other host-associated strains taxonomically related to Streptomyces griseus. Pangenomic and ANI analyses confirm the identity of one of its closest relatives as Streptomyces sp. LaPpAH-199, a strain isolated from an ant-plant symbiosis in Africa. In summary, our results suggest an insect-microbe association in distant geographic areas and showcase the potential of Streptomyces sp. M54 and related strains for the discovery of novel antibiotics.

Bacillus spp. metabolites are effective in eradicating Aedes aegypti (Diptera: Culicidae) larvae with low toxicity to non-target species.

The growing spread of dengue, chikungunya and Zika viruses demand the development of new and environmentally safe control methods for their vector, the mosquito Aedes aegypti. This study aims to find novel larvicidal agents from mutualistic (endophytic and rhizospheric) or edaphic bacteria that have no action against non-target organisms. Eleven out of the 254 bacterial strains tested were able to kill Ae. aegypti larvae. Larvicidal activity did not depend on presence of cells, since culture supernatants or crude lipopeptide extracts (CLEs) killed the larvae. Bacillus safensis BacI67 and Bacillus paranthracis C21 supernatants were the best performing supernatants, displaying the lowest lethal concentrations (LC50 = 31.11 µL/mL and 45.84 µL/mL, respectively). Bacillus velezensis B64a and Bacillus velezensis B15 produced the best performing CLEs (LC50 = 0.11 mg/mL and 0.12 mg/mL, respectively). Mass spectrometry analysis of CLEs detected a mixture of surfactins, iturins, and fengycins. The samples tested were weakly- or non-toxic to mammalian cells (RAW 264.7 macrophages and VERO cells) and non-target organisms (Caenorhabditis elegans, Galleria mellonella, Scenedesmus obliquus, and Tetrahymena pyriformis) - especially B. velezensis B15 CLE. The biosynthetic gene clusters related to secondary metabolism identified by whole genome sequencing of the four best performing bacteria strains revealed clusters for bacteriocin, beta-lactone, lanthipeptide, non-ribosomal peptide synthetases, polyketide synthases (PKS), siderophores, T3PKS, type 1 PKS-like, terpenes, thiopeptides, and trans-AT-PKS. Purification of lipopeptides may clarify the mechanisms by which these extracts kill Ae. aegypti larvae.

High-quality draft genome sequence of pantanalinema sp. GBBB05, a cyanobacterium from cerrado biome

Cyanobacteria comprise one of the oldest and most diverse phyla in the Bacteria domain and are recognized for their importance in the biosphere evolution. Members of this phylum can be found in a wide variety of environments reflecting their photosynthetic ability, adaptability to various environmental conditions, and diversified metabolism. Such characteristics make cyanobacteria one of the preferred targets for research on bioactive compounds and new enzymes (Schirrmeister et al., 2011; Dittmann et al., 2015). Pantanalinema was described as a new genus of the Leptolyngbyaceae cyanobacterial family by a polyphasic approach, which included morphological characteristics, 16S rRNA gene phylogeny, 16S-23S ITS rRNA secondary structures, and physiological characteristics such as adaptability to pH variations (Vaz et al., 2015). This genus has been described only in Brazilian biomes such as the Pantanal and the Amazon, the first isolates being found in a lake. These Pantanalinema isolates were characterized by their ability to grow over a wide pH range (pH 4 to 11) as well as to modify the culture medium pH around neutrality (pH 6 to 7.4). Due to these characteristics, it is thought that this genus can occupy a variety of ecological niches, such as alkaline or slightly acidic water bodies (Vaz et al., 2015; Genuário et al., 2017). Taxonomic classification of Pantanalinema isolates requires the use of molecular markers as this genus is morphologically very similar to the recently described genus Amazoninema, which, in turn, has comparable morphology to other genera of the Leptolyngbyaceae family (Genuário et al., 2018). In this work, we report the genome sequence of a new Pantanalinema strain, named GBBB05, which was isolated from the Brazilian Cerrado biome. This is the first genome assembly for the Pantanalinema genus, which, along with the analyses provided here, is expected to enhance our understanding of this genus’s metabolic potential