The existence of polycyclic aromatic hydrocarbons in the various environments has aroused great environmental concerns due to their potential hazards to human health. The presence of polycyclic aromatic hydrocarbons in aquifer is particularly sensitive where groundwater is used as a source of potable water. Anaerobic biodegradation of polycyclic aromatic hydrocarbons is an attractive option for remediation of contaminated aquifer sediment. Bacterial and archaeal community structures of phenanthrene-degrading aquifer sediment under methanogenic condition were investigated using clone library analysis in combination with microcosm study. The bacterial members were all affiliated with γ-Proteobacteria. Phylum Euryarchaeota was the predominant archaeal group, represented by genera Methanosarcina, Methanobacterium and Thermogymnomonas. Both bacteria (genera Citrobacter and Pseudomonas) and archaea (genus Methanosarcina) might have links to phenanthrene degradation process. This work might provide some new insights into developing strategies for the isolation of the putative polycyclic aromatic hydrocarbons degraders under methanogenic condition and bioremediating polycyclic aromatic hydrocarbons in leachate-contaminated aquifer sediment.