The diversity of microorganisms associated with speleological sources has mainly been studied in limestone caves, while studies in silicate caves are still under development. Here, we profiled the microbial diversity of opal speleothems from a silicate cave in Guiana Highlands. Bulk DNAs were extracted from three speleothems of two types, i.e., one soft whitish mushroom-like speleothem and two hard blackish coral-like speleothems. The extracted DNAs were amplified for sequencing the V3–V4 region of the bacterial 16S rRNA gene by MiSeq. A total of 210,309 valid reads were obtained and clustered into 3184 phylotypes or operational taxonomic units (OTUs). The OTUs from the soft whitish speleothem were mostly affiliated with Acidobacteriota, Pseudomonadota (formerly, Proteobacteria), and Chloroflexota, with the OTUs ascribed to Nitrospirota being found specifically in this speleothem. The OTUs from the hard blackish speleothems were similar to each other and were mostly affiliated with Pseudomonadota, Acidobacteriota, and Actinomycetota (formerly, Actinobacteria). These OTU compositions were generally consistent with those reported for limestone and silicate caves. The OTUs were further used to infer metabolic features by using the PICRUSt bioinformatic tool, and membrane transport and amino acid metabolism were noticeably featured. These and other featured metabolisms may influence the pH microenvironment and, consequently, the formation, weathering, and re-deposition of silicate speleothems.