Accessing the specialized metabolome of actinobacteria from the bulk soil of Paullinia cupana Mart. on the Brazilian Amazon: a promising source of bioactive compounds against soybean phytopathogens.

The Amazon rainforest, an incredibly biodiverse ecosystem, has been increasingly vulnerable to deforestation. Despite its undeniable importance and potential, the Amazonian microbiome has historically received limited study, particularly in relation to its unique arsenal of specialized metabolites. Therefore, in this study our aim was to assess the metabolic diversity and the antifungal activity of actinobacterial strains isolated from the bulk soil of Paullinia cupana, a native crop, in the Brazilian Amazon Rainforest. Extracts from 24 strains were subjected to UPLC-MS/MS analysis using an integrative approach that relied on the Chemical Structural and Compositional Similarity (CSCS) metric, GNPS molecular networking, and in silico dereplication tools. This procedure allowed the comprehensive understanding of the chemical space encompassed by these actinobacteria, which consists of features belonging to known bioactive metabolite classes and several unannotated molecular families. Among the evaluated strains, five isolates exhibited bioactivity against a panel of soybean fungal phytopathogens (Rhizoctonia solani, Macrophomina phaseolina, and Sclerotinia sclerotiorum). A focused inspection led to the annotation of pepstatins, oligomycins, hydroxamate siderophores and dorrigocins as metabolites produced by these bioactive strains, with potentially unknown compounds also comprising their metabolomes. This study introduces a pragmatic protocol grounded in established and readily available tools for the annotation of metabolites and the prioritization of strains to optimize further isolation of specialized metabolites. Conclusively, we demonstrate the relevance of the Amazonian actinobacteria as sources for bioactive metabolites useful for agriculture. We also emphasize the importance of preserving this biome and conducting more in-depth studies on its microbiota.

Microbial Metabolites Annotation by Mass Spectrometry-Based Metabolomics.

Since the discovery of penicillin, microbial metabolites have been extensively investigated for drug discovery purposes. In the last decades, microbial derived compounds have gained increasing attention in different fields from pharmacognosy to industry and agriculture. Microbial metabolites in microbiomes present specific functions and can be associated with the maintenance of the natural ecosystems. These metabolites may exhibit a broad range of biological activities of great interest to human purposes. Samples from either microbial isolated cultures or microbiomes consist of complex mixtures of metabolites and their analysis are not a simple process. Mass spectrometry-based metabolomics encompass a set of analytical methods that have brought several improvements to the microbial natural products field. This analytical tool allows the comprehensively detection of metabolites, and therefore, the access of the chemical profile from those biological samples. These analyses generate thousands of mass spectra which is challenging to analyse. In this context, bioinformatic metabolomics tools have been successfully employed to accelerate and facilitate the investigation of specialized microbial metabolites. Herein, we describe metabolomics tools used to provide chemical information for the metabolites, and furthermore, we discuss how they can improve investigation of microbial cultures and interactions.

Metabologenomics analysis of Pseudomonas sp. So3.2b, an Antarctic strain with bioactivity against Rhizoctonia solani.

Introduction: Phytopathogenic fungi are a considerable concern for agriculture, as they can threaten the productivity of several crops worldwide. Meanwhile, natural microbial products are acknowledged to play an important role in modern agriculture as they comprehend a safer alternative to synthetic pesticides. Bacterial strains from underexplored environments are a promising source of bioactive metabolites. Methods: We applied the OSMAC (One Strain, Many Compounds) cultivation approach, in vitro bioassays, and metabolo-genomics analyses to investigate the biochemical potential of Pseudomonas sp. So3.2b, a strain isolated from Antarctica. Crude extracts from OSMAC were analyzed through HPLC-QTOF-MS/MS, molecular networking, and annotation. The antifungal potential of the extracts was confirmed against Rhizoctonia solani strains. Moreover, the whole-genome sequence was studied for biosynthetic gene clusters (BGCs) identification and phylogenetic comparison. Results and Discussion: Molecular networking revealed that metabolite synthesis has growth media specificity, and it was reflected in bioassays results against R. solani. Bananamides, rhamnolipids, and butenolides-like molecules were annotated from the metabolome, and chemical novelty was also suggested by several unidentified compounds. Additionally, genome mining confirmed a wide variety of BGCs present in this strain, with low to no similarity with known molecules. An NRPS-encoding BGC was identified as responsible for producing the banamides-like molecules, while phylogenetic analysis demonstrated a close relationship with other rhizosphere bacteria. Therefore, by combining -omics approaches and in vitro bioassays, our study demonstrates that Pseudomonas sp. So3.2b has potential application to agriculture as a source of bioactive metabolites.

Elicitation of Streptomyces lunalinharesii secondary metabolism through co-cultivation with Rhizoctonia solani.

The concern regarding the emergence of phytopathogens strains which are resistant to conventional agrochemicals has given support to the search for alternatives on the use of chemical pesticides in agriculture. In this context, microorganisms are considered as promising sources of useful natural compounds and actinobacteria are particularly relevant since they are known to produce several bioactive metabolites. The objective of this work was to investigate the production of secondary metabolites with antifungal activity by a strain of the actinobacteria Streptomyces lunalinharesii (A54A) under axenic conditions and in co-cultivation with the phytopathogen Rhizoctonia solani. Tests to evaluate antifungal activity of the extracts indicated the presence of diffusable molecules capable of inhibiting the growth of R. solani produced by S. lunalinharesii, especially when in the presence of the fungus during fermentation. Metabolomic analyzes allowed the putative annotation of the bioactive compounds desferrioxamine E and anisomycin, in addition to the evaluation of the metabolic profile of the isolate when grown in axenic mode and in co-cultivation, while statistical analyzes enabled the comparison of such profiles and the identification of metabolites produced in greater relative quantities in the elicitation condition. Such methodologies provided the selection of unknown features with high bioactive potential for dereplication, and several metabolites of S. lunalinharesii possibly represent novel compounds.

Scorpionicidal activity of secondary metabolites from Paecilomyces sp. CMAA1686 against Tityus serrulatus.

INTRODUCTION: Urban pests pose enormous risks to human health. Control initiatives are carried out in regions of high infestation and incidence of accidents caused by scorpions OBJECTIVE: In this study, we aimed to analyze the anti-scorpionic activity of fungal isolates obtained from a cemetery in Brazil. MATERIALS AND METHODS: A total of thirteen fungi were subjected to a bioassay test against Tityus serrulatus, and the two isolates with the highest scorpionicidal activity were selected for molecular identification through sequencing of the ITS DNA hypervariable region and large-scale cultivation on liquid medium for secondary metabolite extraction. The crude extracts were partitioned by solid-phase extraction, and the resulting purified extracts were tested for anti-scorpionic activity. The extracts from one of the isolates presented better results and were submitted to UPLC-MS/MS. The metabolomics data were submitted to GNPS website for Molecular Networking and MASST searches. We also performed a MolNetEnhancer analysis to identify the chemical classes of the molecules found in the samples. RESULTS: The most promising fungal isolate was identified as Paecilomyces sp. CMAA1686 which has 98% of similarity to Paecilomyces formosus. The sub-fractions C and D had the best activity against the scorpions (54 and 32% mortality, respectively). Molecular Networking and MolNetEnhancer revealed a range of molecular classes in our extracts that are known to include bioactive metabolites from Paecilomyces species. CONCLUSIONS: The scorpionicidal activity of Paecilomyces sp. CMAA1686 and its secondary metabolites may provide new alternative compounds for biological and chemical control of scorpions from the species T. serrulatus. Paecilomyces sp. CMAA1686 is an isolate that has great potential for isolation of secondary metabolites.