Bacterial culture broth extracts have been the starting point for the development of numerous therapeutics. However, only a small fraction of bacterial biosynthetic diversity is accessible using this strategy. Here, we apply a discovery approach that bypasses the culturing step entirely by bioinformatically predicting small molecule structures from the primary sequences of the biosynthetic gene clusters. These structures are then chemically synthesized to give synthetic-bioinformatic natural products (syn-BNPs). Using this approach, we screened syn-BNPs inspired by nonribosomal peptide synthetases against microbial pathogens, and discovered an antibiotic for which no resistance could be identified and an antifungal agent with activity against diverse fungal pathogens.

Pigment-based coloration is a common trait found in a variety of organisms across the tree of life. For example, calcareous avian eggs are natural structures that vary greatly in color, yet just a handful of tetrapyrrole pigment compounds are responsible for generating this myriad of colors. To fully understand the diversity and constraints shaping nature’s palette, it is imperative to characterize the similarities and differences in the types of compounds involved in color production across diverse lineages. Pigment composition was investigated in eggshells of eleven paleognath bird taxa, covering several extinct and extant lineages, and shells of four extant species of mollusks. Birds and mollusks are two distantly related, calcareous shell-building groups, thus characterization of pigments in their calcareous structures would provide insights to whether similar compounds are found in different phyla (Chordata and Mollusca). An ethylenediaminetetraacetic acid (EDTA) extraction protocol was used to analyze the presence and concentration of biliverdin and protoporphyrin, two known and ubiquitous tetrapyrrole avian eggshell pigments, in all avian and molluscan samples. Biliverdin was solely detected in birds, including the colorful eggshells of four tinamou species. In contrast, protoporphyrin was detected in both the eggshells of several avian species and in the shells of all mollusks. These findings support previous hypotheses about the ubiquitous deposition of tetrapyrroles in the eggshells of various bird lineages and provide evidence for its presence also across distantly related animal taxa.

The entomopathogenic bacterium Xenorhabdus bovienii exists in a mutualistic relationship with nematodes of the genus Steinernema. Free-living infective juveniles of Steinernema prey on insect larvae and regurgitate X. bovienii within the hemocoel of a host larva. X. bovienii subsequently produces a complex array of specialized metabolites and effector proteins that kill the insect and regulate various aspects of the trilateral symbiosis. While Xenorhabdus species are rich producers of secondary metabolites, many of their biosynthetic gene clusters remain uncharacterized. Here, we describe a nonribosomal peptide synthetase (NRPS) identified through comparative genomics analysis that is widely conserved in Xenorhabdus species. Heterologous expression of this NRPS gene from X. bovienii in E. coli led to the discovery of a family of lipo-tripeptides that chromatographically appear as pairs, containing either a C-terminal carboxylic acid or carboxamide. Coexpression of the NRPS with the leupeptin protease inhibitor pathway enhanced production, facilitating isolation and characterization efforts. The new lipo-tripeptides were also detected in wild-type X. bovienii cultures. These metabolites, termed bovienimides, share an uncommon C-terminal d-citrulline residue. The NRPS lacked a dedicated chain termination domain, resulting in product diversification and release from the assembly line through reactions with ammonia, water, or exogenous alcohols.