Exceptionally well-preserved pillow lavas and inter-pillow hyaloclastites from the Barberton Greenstone Belt in South Africa contain textural, geochemical, and isotopic biomarkers indicative of microbially mediated alteration of basaltic glass in the Archean. The textures are micrometer-scale tubular structures interpreted to have originally formed during microbial etching of glass along fractures. Textures of similar size, morphology, and distribution have been attributed to microbial activity and are commonly observed in the glassy margins of pillow lavas from in situ oceanic crust and young ophiolites. The tubes from the Barberton Greenstone Belt were preserved by precipitation of fine-grained titanite during greenschist facies metamorphism associated with seafloor hydrothermal alteration. The presence of organic carbon along the margins of the tubes and low δ¹³C values of bulk-rock carbonate in formerly glassy samples support a biogenic origin for the tubes. Overprinting relationships of secondary minerals observed in thin section indicate the tubular structures are pre-metamorphic. Overlapping metamorphic and igneous crystallization ages thus imply the microbes colonized these rocks 3.4–3.5 Ga. Although, the search for traces of early life on Earth has recently intensified, research has largely been confined to sedimentary rocks. Subaqueous volcanic rocks represent a new geological setting in the search for early life that may preserve a largely unexplored Archean biomass.