Understanding the mechanisms of surface-associated copper tolerance in marine bacteria

Abstract

Copper can be toxic for bacterial cells when present in excess amounts despite being an essential cellular macromolecule. Copper-based antimicrobial paints are frequently used to inhibit the biofouling of marine vessels. However, some bacterial species can overcome this copper challenge and colonize the surfaces. The early adherent bacterial population of marine vessels plays an important role because of its ability to produce extracellular polymeric substances (EPSs), forming a thin layer of organic matter that traps nutrients from the water and protects other colonizers by blocking the toxic antifouling (AF) coatings. It is of interest to study the factors that drive the initial colonization of copper surfaces. Previous studies have shown Alteromonas species to be one of the early colonizers in the marine biofouling environment. Here, we studied a specific Alteromonas macleodii strain, CUKW, isolated from copper test coupons submerged in tropical oligotrophic coastal seawater. These coupons are used to determine the effects of the coating process on microbial substrate colonization. We demonstrate that CUKW harbors genetic components derived from established copper resistance mechanisms. Transcriptional profiling studies of copper-specific genes showed the induction of multiple plasmid-borne copper genes in response to copper. We also discovered evidence of multiple mobile genetic elements and genomic islands containing copper genes in CUKW, which may be involved in bacterial adaptation to high copper levels. Furthermore, our comparative genomic analysis based on homology-based protein similarity search of copper signal transduction systems across marine bacteria revealed that various potential copper-associated signaling systems are found across marine species. The copper signaling systems varied widely in their abundance and distribution within the same phylum, indicating that the environment from which organisms were isolated might influence their copper-associated signaling systems. We also characterized the microbial diversity and functional potential within the early biofilm that develops on marine copper surfaces. The functional analysis of initial biofilm colonizers on copper surfaces in marine coastal environments provides insights into molecular mechanisms that support biofilm formation on copper surfaces. The taxa associated with copper resistance traits were found to be dominant in the initial microbial communities on copper surfaces, allowing these organisms to survive on copper surfaces despite copper toxicity. Our analysis reveals the dominance of genera Allomuricauda and Ruegeria, which carry several copper genes, as the early colonizers of copper surfaces. Together, our work creates a foundation for subsequent studies to analyze functional signatures unique to various copper-related surfaces, which can guide novel genetic strategies against marine biofouling.