Sub-aerial biofilms on urban stone surfaces: community structure and microbial co-occurrence networks

Sub-aerial biofilms (SABs) play a key role in the biodeterioration of stone cultural heritage, yet their microbial composition may vary depending on the colonized lithic substrate. The aim of this study was to analyze the taxonomic composition of SABs in relation to the type of stone substrate. The material consisted of ten SAB samples, including eight collected from a historic cemetery in Kraków and two from retaining walls within the city. Sampling was performed using a low-invasive method directly from stone surfaces. Microbial community composition was assessed using Illumina eDNA metabarcoding targeting fungi, algae, and bacteria. Fungal communities were dominated by representatives of the classes Dothideomycetes and Eurotiomycetes. Among phototrophic organisms, Chlorophyta constituted the majority of algal taxa, while cyanobacteria were also highly represented. Bacterial diversity was largely composed of Proteobacteria and Actinobacteria, which together accounted for approximately 60% of the detected bacterial taxa. The overall taxonomic composition of SAB communities was consistent with results reported previously using culture-dependent approaches. These findings confirm the suitability of using eDNA metabarcoding for comprehensive characterization of SABs and highlight substrate-related patterns in microbial colonization of stone surfaces. Co-occurrence network analysis identified contrasting interaction patterns among microbial domains, with highly integrated fungal communities, more modular bacterial assemblages, and fragmented phototrophic networks, suggesting that the stability of SAB is driven not only by taxonomic composition but also by domain-specific patterns of microbial interactions.