Gut microbiomes of cycad-feeding insects tolerant to ß-methylamino-L-alanine (BMAA) are rich in siderophore biosynthesis.

Ingestion of the cycad toxins ß-methylamino-L-alanine (BMAA) and azoxyglycosides is harmful to diverse organisms. However, some insects are specialized to feed on toxin-rich cycads with apparent immunity. Some cycad-feeding insects possess a common set of gut bacteria, which might play a role in detoxifying cycad toxins. Here, we investigated the composition of gut microbiota from a worldwide sample of cycadivorous insects and characterized the biosynthetic potential of selected bacteria. Cycadivorous insects shared a core gut microbiome consisting of six bacterial taxa, mainly belonging to the Proteobacteria, which we were able to isolate. To further investigate selected taxa from diverging lineages, we performed shotgun metagenomic sequencing of co-cultured bacterial sub-communities. We characterized the biosynthetic potential of four bacteria from Serratia, Pantoea, and two different Stenotrophomonas lineages, and discovered a suite of biosynthetic gene clusters notably rich in siderophores. Siderophore semi-untargeted metabolomics revealed a broad range of chemically related yet diverse iron-chelating metabolites, including desferrioxamine B, suggesting the occurrence of an unprecedented desferrioxamine-like biosynthetic pathway that remains to be identified. These results provide a foundation for future investigations into how cycadivorous insects tolerate diets rich in azoxyglycosides, BMAA, and other cycad toxins, including a possible role for bacterial siderophores.

MIBiG 3.0: a community-driven effort to annotate experimentally validated biosynthetic gene clusters.

With an ever-increasing amount of (meta)genomic data being deposited in sequence databases, (meta)genome mining for natural product biosynthetic pathways occupies a critical role in the discovery of novel pharmaceutical drugs, crop protection agents and biomaterials. The genes that encode these pathways are often organised into biosynthetic gene clusters (BGCs). In 2015, we defined the Minimum Information about a Biosynthetic Gene cluster (MIBiG): a standardised data format that describes the minimally required information to uniquely characterise a BGC. We simultaneously constructed an accompanying online database of BGCs, which has since been widely used by the community as a reference dataset for BGCs and was expanded to 2021 entries in 2019 (MIBiG 2.0). Here, we describe MIBiG 3.0, a database update comprising large-scale validation and re-annotation of existing entries and 661 new entries. Particular attention was paid to the annotation of compound structures and biological activities, as well as protein domain selectivities. Together, these new features keep the database up-to-date, and will provide new opportunities for the scientific community to use its freely available data, e.g. for the training of new machine learning models to predict sequence-structure-function relationships for diverse natural products. MIBiG 3.0 is accessible online at https://mibig.secondarymetabolites.org/.

Desert Environments Facilitate Unique Evolution of Biosynthetic Potential in Streptomyces.

Searching for new bioactive metabolites from the bacterial genus Streptomyces is a challenging task. Combined genomic tools and metabolomic screening of Streptomyces spp. native to extreme environments could be a promising strategy to discover novel compounds. While Streptomyces of desertic origin have been proposed as a source of new metabolites, their genome mining, phylogenetic analysis, and metabolite profiles to date are scarcely documented. Here, we hypothesized that Streptomyces species of desert environments have evolved with unique biosynthetic potential. To test this, along with an extensive characterization of biosynthetic potential of a desert isolate Streptomyces sp. SAJ15, we profiled phylogenetic relationships among the closest and previously reported Streptomyces of desert origin. Results revealed that Streptomyces strains of desert origin are closer to each other and relatively distinct from Streptomyces of other environments. The draft genome of strain SAJ15 was 8.2 Mb in size, which had 6972 predicted genes including 3097 genes encoding hypothetical proteins. Successive genome mining and phylogenetic analysis revealed the presence of putative novel biosynthetic gene clusters (BGCs) with low incidence in another Streptomyces. In addition, high-resolution metabolite profiling indicated the production of arylpolyene, terpenoid, and macrolide compounds in an optimized medium by strain SAJ15. The relative abundance of different BGCs in arid Streptomyces differed from the non-arid counterparts. Collectively, the results suggested a distinct evolution of desert Streptomyces with a unique biosynthetic potential.

Evolutionary dynamics of natural product biosynthesis in bacteria.

Covering: 2008 up to 2019The forces of biochemical adaptive evolution operate at the level of genes, manifesting in complex phenotypes and the global biodiversity of proteins and metabolites. While evolutionary histories have been deciphered for some other complex traits, the origins of natural product biosynthesis largely remain a mystery. This fundamental knowledge gap is surprising given the many decades of research probing the genetic, chemical, and biophysical mechanisms of bacterial natural product biosynthesis. Recently, evolutionary thinking has begun to permeate this otherwise mechanistically dominated field. Natural products are now sometimes referred to as 'specialized' rather than 'secondary' metabolites, reinforcing the importance of their biological and ecological functions. Here, we review known evolutionary mechanisms underlying the overwhelming chemical diversity of bacterial secondary metabolism, focusing on enzyme promiscuity and the evolution of enzymatic domains that enable metabolic traits. We discuss the mechanisms that drive the assembly of natural product biosynthetic gene clusters and propose formal definitions for 'specialized' and 'secondary' metabolism. We further explore how biosynthetic gene clusters evolve to synthesize related molecular species, and in turn how the biological and ecological roles that emerge from metabolic diversity are acted on by selection. Finally, we reconcile chemical, functional, and genetic data into an evolutionary model, the dynamic chemical matrix evolutionary hypothesis, in which the relationships between chemical distance, biomolecular activity, and relative fitness shape adaptive landscapes.

EvoMining reveals the origin and fate of natural product biosynthetic enzymes.

Natural products (NPs), or specialized metabolites, are important for medicine and agriculture alike, and for the fitness of the organisms that produce them. NP genome-mining aims at extracting biosynthetic information from the genomes of microbes presumed to produce these compounds. Typically, canonical enzyme sequences from known biosynthetic systems are identified after sequence similarity searches. Despite this being an efficient process, the likelihood of identifying truly novel systems by this approach is low. To overcome this limitation, we previously introduced EvoMining, a genome-mining approach that incorporates evolutionary principles. Here, we release and use our latest EvoMining version, which includes novel visualization features and customizable databases, to analyse 42 central metabolic enzyme families (EFs) conserved throughout Actinobacteria, Cyanobacteria, Pseudomonas and Archaea. We found that expansion-and-recruitment profiles of these 42 families are lineage specific, opening the metabolic space related to 'shell' enzymes. These enzymes, which have been overlooked, are EFs with orthologues present in most of the genomes of a taxonomic group, but not in all. As a case study of canonical shell enzymes, we characterized the expansion and recruitment of glutamate dehydrogenase and acetolactate synthase into scytonemin biosynthesis, and into other central metabolic pathways driving Archaea and Bacteria adaptive evolution. By defining the origin and fate of enzymes, EvoMining complements traditional genome-mining approaches as an unbiased strategy and opens the door to gaining insights into the evolution of NP biosynthesis. We anticipate that EvoMining will be broadly used for evolutionary studies, and for generating predictions of unprecedented chemical scaffolds and new antibiotics. This article contains data hosted by Microreact.

Cycad Coralloid Roots Contain Bacterial Communities Including Cyanobacteria and Caulobacter spp. That Encode Niche-Specific Biosynthetic Gene Clusters.

Cycads are the only early seed plants that have evolved a specialized root to host endophytic bacteria that fix nitrogen. To provide evolutionary and functional insights into this million-year old symbiosis, we investigate endophytic bacterial sub-communities isolated from coralloid roots of species from Dioon (Zamiaceae) sampled from their natural habitats. We employed a sub-community co-culture experimental strategy to reveal both predominant and rare bacteria, which were characterized using phylogenomics and detailed metabolic annotation. Diazotrophic plant endophytes, including Bradyrhizobium, Burkholderia, Mesorhizobium, Rhizobium, and Nostoc species, dominated the epiphyte-free sub-communities. Draft genomes of six cyanobacteria species were obtained after shotgun metagenomics of selected sub-communities. These data were used for whole-genome inferences that suggest two Dioon-specific monophyletic groups, and a level of specialization characteristic of co-evolved symbiotic relationships. Furthermore, the genomes of these cyanobacteria were found to encode unique biosynthetic gene clusters, predicted to direct the synthesis of specialized metabolites, mainly involving peptides. After combining genome mining with detection of pigment emissions using multiphoton excitation fluorescence microscopy, we also show that Caulobacter species co-exist with cyanobacteria, and may interact with them by means of a novel indigoidine-like specialized metabolite. We provide an unprecedented view of the composition of the cycad coralloid root, including phylogenetic and functional patterns mediated by specialized metabolites that may be important for the evolution of ancient symbiotic adaptations.

Phylogenomics of 2,4-Diacetylphloroglucinol-Producing Pseudomonas and Novel Antiglycation Endophytes from Piper auritum.

2,4-Diacetylphloroglucinol (DAPG) (1) is a phenolic polyketide produced by some plant-associated Pseudomonas species, with many biological activities and ecological functions. Here, we aimed at reconstructing the natural history of DAPG using phylogenomics focused at its biosynthetic gene cluster or phl genes. In addition to around 1500 publically available genomes, we obtained and analyzed the sequences of nine novel Pseudomonas endophytes isolated from the antidiabetic medicinal plant Piper auritum. We found that 29 organisms belonging to six Pseudomonas species contain the phl genes at different frequencies depending on the species. The evolution of the phl genes was then reconstructed, leading to at least two clades postulated to correlate with the known chemical diversity surrounding DAPG biosynthesis. Moreover, two of the newly obtained Pseudomonas endophytes with high antiglycation activity were shown to exert their inhibitory activity against the formation of advanced glycation end-products via DAPG and related congeners. Its isomer, 5-hydroxyferulic acid (2), detected during bioactivity-guided fractionation, together with other DAPG congeners, were found to enhance the detected inhibitory activity. This report provides evidence of a link between the evolution and chemical diversity of DAPG and congeners.

The neuropeptide Gonadotropin-releasing hormone modifies the spontaneous muscular contraction in the earthworm: Eisenia foetida.

We investigated whether the Gonadotropin-releasing hormone affects the spontaneous muscular contraction in the earthworm Eisenia foetida. In addition, we investigated the presence of Gonadotropin-releasing hormone receptor in ventral nerve cord by immunohistochemistry and polymerase chain reaction. Gonadotropin-releasing hormone induced a significant increase on both amplitude and muscular tone and decrease in the frequency of spontaneous muscular contraction. We found the presence of immunoreactive material to Gonadotropin-releasing hormone receptor in the ventral nerve cord, likewise the Gonadotropin-releasing hormone receptor mRNA expression. In conclusion, the Gonadotropin-releasing hormone modifies the spontaneous muscular contraction in E. foetida and these effects can be due to the activation of the Gonadotropin-releasing hormone receptor.