Preliminary indication of the role of AHL-dependent quorum sensing systems in calcium carbonate precipitation in Gram-negative bacteria

Abstract

<abstract> <p>Numerous microbial species participate in precipitation of carbonates in various natural environments, including soils, geological formations, freshwater biofilms and oceans. Despite the geochemical interest of such a biomineralization process, its molecular mechanisms and adaptive aspects remain poorly known. Many Gram-negative bacteria use cell-to-cell communication systems relying on N-acylhomoserine lactone (AHLs) signal molecules to express certain phenotypic traits in a density-dependent manner, a phenomenon referred as to quorum-sensing (QS). In this work, bacterial isolates collected from cave and rhizosphere soil were analyzed to study the occurrence of the AHL-mediated QS in bacterial calcium carbonate (CaCO₃) precipitation. To test the production of AHLs signal molecules, we cross-streaked Gram-negative calcifying strains, selected among the environmental strains studied, with the AHL-negative mutant <italic>Chromobacterium subtsugae</italic> strain CV026. Only <italic>Burkholderia ambifaria</italic> LMG 11351 was able to restore violacein production in CV026 among the tested strains. The constructed AHL-negative mutant of <italic>B. ambifaria</italic> LMG 11351 could not precipitate CaCO₃ on B-4 agar. Scanning Electron Microscopy (SEM) analysis on CaCO₃ crystals obtained <italic>in vitro</italic> shows crystals of different morphologies, calcified biofilms and bacteria in close contact with the precipitated crystals. In the inner layers of the bioliths deposited by <italic>B. ambifaria</italic> LMG 11351, a stream-like organization of the <italic>Burkholderia</italic> imprints was not detected by SEM. Our data provide preliminary evidence that the activation of AHL-regulated genes may be a prerequisite for <italic>in vitro</italic> bacterial carbonatogenesis, in some cases, confirming the specific role of bacteria as CaCO₃ precipitating agents. We enhance the understanding of bacterial CaCO₃ biomineralization and its potential biotechnology implications for QS-based strategies to enhance or decrease CaCO₃ precipitation through specific bacterial processes. The AHL-negative mutant of <italic>B. ambifaria</italic> LMG 11351 (a well-known plant growth-promoting bacterium) could also be used to study plant-bacteria interactions. The adaptive role of bacterial CaCO₃ biomineralization was also discussed.</p> </abstract>