Antimicrobial resistance (AMR) is a growing global health crisis, with mortality already in the millions and projections of up to 10 million deaths annually by 2050. Traditional discovery strategies, such as one strain many compounds-based screening of large strain collections, have yielded most of our current antibiotics but are slow, resource-intensive, and highly prone to rediscovery. In parallel, high-throughput sequencing has uncovered an enormous reservoir of biosynthetic gene clusters (BGCs), of which only a small fraction has been experimentally characterized, and many show extensive overlap in gene content. This combination of hidden diversity and functional redundancy demands new tools to prioritize, monitor, and rationally activate BGCs. In this review, we discuss how biosensors can be integrated with genome mining to accelerate antimicrobial discovery and BGC dereplication. We summarize the chemical scaffolds, biosynthetic logic and diagnostic enzymatic signatures of major antimicrobial classes, including β-lactams, tetracyclines, macrolides, aminoglycosides, glycopeptides, lincosamides, polyenes, and azoles, and highlighting representative biosensors that target each scaffold. The biosensors discussed use transcription factors, enzymes, aptamers, CRISPR systems and stress-response modules to generate specific and sensitive signals in complex matrices. We argue that translating BGC architecture into biosensor design creates a practical framework to rapidly discriminate from novel activities, but most importantly, to guide the exploration of chemical space around clinically important scaffolds in the era of escalating AMR.

Sclerotium cepivorum Berk is the etiological agent of white rot disease that affects plants of the genus Allium. This fungus produces resistance structures called sclerotia that are formed by a rolled mycelium with a thick layer of melanin and it can remain dormant for many years in the soil. Current interest in S. cepivorum has arisen from economic losses in Allium crops in the agricultural sector. Melanin is a component that protects the sclerotia from adverse environmental conditions In many organisms, it plays an important role in the infectious process; in S. cepivorum, the pathway by which this component is synthetized is not fully described. By using infrared spectrophotometric assays applied direct to the sclerotia and a melanin extract followed by an NMR analysis and a tricyclazole melanin inhibition experiment, it allowed us to determine the dihydroxynaphthalene (DHN)-melanin pathway by which S. cepivorum performs its melanin synthesis. Moreover, we focused on studying scytalone dehydratase (SDH) as a key enzyme of the DHN-melanin synthesis. We obtained the recombinant SDH enzyme and tested its activity by a zymogram assay. Thereby, the S. cepivorum melanogenic route was established as a DHN pathway.