A multidisciplinary study of silicified volcanoclastic, near-shore deposits from the 3.446 Ga “Kitty's Gap Chert,” Warrawoona Group, Pilbara, reveals that they contain a wealth of carbonaceous microbial fossil remains. The volcanoclastic sediments host predominantly colonies of coccoidal microorganisms that occur in two modal size ranges, 0.4–0.5 μm and 0.75–0.8 μm. These microbial colonies coat the surfaces of the volcanic particles and form either dense, carpetlike associations up to tens of micrometers in diameter comprising hundreds of individuals. They also form less dense concentrations that include many chainlike associations of coccoids. All colonies are associated with a polymer film (extracellular polymeric substances—EPS) that coats both the organisms and their substrate. Multispecies biofilms formed at a boundary representing a short period of nondeposition. They consisted predominantly of coccoids and EPS but also included common, small filaments tens of micrometers in length and 0.25 μm in width and rare, short rods 1 μm in length. Carbon isotopic compositions of about −26‰ to −30‰, measured on individual layers, are compatible with microbial fractionation. The biofilms include possible anoxygenic-photosynthesizing organisms (the filaments), whereas the colonies coating the volcanic clasts probably represent chemolithotrophic organisms. The interaction between the microbes, their colonies and biofilms, and their environment is intimate and complex. The environment provided the substrate and the nutrient, energy, and carbon sources, whereas the metabolic activity of the microbes contributed to the early diagenetic alteration of the volcanic particles, to the binding of the sediment, and to their silicification. The microorganisms were preserved by rapid silicification, with the silica coming partly from hydrothermal sources and partly from pore water enrichment in Si due to the devitrification of the volcanic protoliths (partially mediated by microbial activity). Our multidisciplinary approach to the study of this sample demonstrates the importance of using complementary methods in order to understand the complex microbe/sediment interactions and to be able to relate different types of microbial colonies/biofilms to different microenvironments. The observations and conclusions from this study have important consequences for the methods that need to be used in the search for traces of past life in general and especially in the search for past life on other planets such as Mars.