Much is known about microbes originally identified in caves, but little is known about the entrapment of microbes (bacteria) in stalactites and their possible environmental origins. This study presents data regarding the significant environmental distribution of prokaryotic bacterial taxa of a Greek stalactite core. We investigated the involvement of those bacteria communities in stalactites using a metataxonomic analysis approach of partial 16S rRNA genes. The metataxonomic analysis of stalactite core material revealed an exceptionally broad ecological spectrum of bacteria classified as members of Proteobacteria, Actinobacteria, Firmicutes, Verrucomicrobia, and other unclassified bacteria. We concluded that (i) the bacterial transport process is possible through water movement from the upper ground cave environment, forming cave speleothems such as stalactites, (ii) bacterial genera such as Polaromonas, Thioprofundum, and phylum Verrucomicrobia trapped inside the stalactite support the paleoecology, paleomicrobiology, and paleoclimate variations, (iii) the entrapment of certain bacteria taxa associated with water, soil, animals, and plants such as Micrococcales, Propionibacteriales, Acidimicrobiales, Pseudonocardiales, and α-, β-, and γ-Proteobacteria.

Speleothems found in caves worldwide are considered the natural libraries of paleontology. Bacteria found in these ecosystems are generally limited to Proteobacteria and Actinomycetota, but rare microbiome and “Dark Matter” is generally under-investigated and often neglected. This research article discusses, for the first time to our knowledge, the diachronic diversity of Actinomycetota entrapped inside a cave stalactite. The planet’s environmental microbial community profile of different eras can be stored in these refugia (speleothems). These speleothems could be an environmental “Microbial Ark” storing rare microbiome and “Dark Matter” bacterial communities evermore.