Pine bark, a low-cost industrial residue, has been suggested as a promising substitute for granular activated carbon in the on-site treatment of water contaminated with 2,4,6-trinitrotoluene (TNT). However, the complex organic structure and indigenous microbial community of pine bark have thus far not been thoroughly described in the context of TNT-contaminated water treatment. This two-week batch study examined the removal efficiency ofTNT from water by (1) adsorption on pine bark and (2) simultaneous adsorption on pine bark and biotransformation by specialized TNT-biotransforming microbial inocula. The bacterial community composition of experimental batches, inocula and pine bark, was profiled by Illumina sequencing of the V6 region of the 16S rRNA gene. The results revealed that the inocula and experimental batches were dominated by phylotypes belonging to the Enterobacteriaceae family and that the tested inocula had good potential for TNT biotransformation. The type of applied inocula had the most profound effect on the TNT-transforming bacterial community structure in the experimental batches. The indigenous microbial community of pine bark harboured phylotypes that also have a potential to degrade TNT. Altogether, the combination of a specialized inoculum and pine bark proved to be the most efficient treatment option for TNT-contaminated water.