Unveiling the Pool of Metallophores in Native Environments and Correlation with Their Potential Producers

For many organisms, metallophores are essential biogenic ligands that ensure metal scavenging and acquisition from their environment. Their identification is challenging in highly organic matter rich environments like peatlands due to low solubilization and metal scarcity and high matrix complexity. In contrast to common approaches based on sample modification by spiking of metal isotope tags, we have developed a two-dimensional (2D) Solid-phase extraction-Liquid chromatography-mass spectrometry (SPE-LC-MS) approach for the highly sensitive (LOD 40 fmol per g of soil), high-resolution direct detection and identification of metallophores in both their noncomplexed (apo) and metal-complexed forms in native environments. The characterization of peat collected in the Bernadouze (France) peatland resulted in the identification of 53 metallophores by a database mass-based search, 36 among which are bacterial. Furthermore, the detection of the characteristic (natural) metal isotope patterns in MS resulted in the detection of both Fe and Cu potential complexes. A taxonomic-based inference method was implemented based on literature and public database (antiSMASH database version 3.0) searches, enabling to associate over 40% of the identified bacterial metallophores with potential producers. In some cases, low completeness with the MIBiG reference BCG might be indicative of alternative producers in the ecosystem. Thus, coupling of metallophore detection and producers' inference could pave a new way to investigate poorly documented environment searching for new metallophores and their producers yet unknown.

Automated summary

Metallophores play a crucial role in metal scavenging and acquisition for many organisms. However, their identification in highly organic matter rich environments like peatlands is challenging due to low solubilization and metal scarcity. In this study, a novel two-dimensional solid-phase extraction-liquid chromatography-mass spectrometry (SPE-LC-MS) approach was developed for sensitive detection and identification of metallophores in native environments. By applying this method, 53 metallophores, including 36 bacterial ones, were identified in peat collected from a French peatland. The detection of characteristic metal isotope patterns also revealed potential complexes of iron and copper. Furthermore, a taxonomic-based inference method was implemented to associate the identified bacterial metallophores with potential producers, suggesting a new way to explore poorly documented environments for new metallophores and their producers.