Advancements in understanding of the microbial ecology in soils containing light non-aqueous phase liquids (LNAPLs) are needed to drive development of optimized bioremediation technologies. In this study, depth-resolved characterization of geochemical parameters and microbial communities was conducted for a shallow hydrocarbon-impacted aquifer. Four distinct biogeochemical zones were identified: (I) an aerobic, low-contaminant mass zone at the top of the vadose zone, (II) a moderate to high-contaminant mass, low-oxygen to anaerobic transition zone in the middle of the vadose zone, (III) an anaerobic, high-contaminant mass zone spanning the bottom of the vadose zone and saturated zone, and (IV) a sulfate-reducing, low-contaminant mass zone below the LNAPL body. Evidence suggested hydrocarbon degradation is mediated by syntrophic fermenters and methanogens in zones II and III. Upward flux of methane likely contributes to promoting anaerobic conditions in zone II by limiting downward flux of oxygen as methane and oxygen fronts converge at the top of this zone. Observed sulfate gradients and microbial communities suggested that sulfate reduction and methanogenesis both contribute to hydrocarbon degradation in zone IV. Pyrosequencing revealed that Syntrophus- and Methanosaeta-related species dominate bacterial and archaeal communities, respectively, thus linking these genera with in situ hydrocarbon degradation in the LNAPL body.