This study aimed to (1) enrich microbial acetylenotrophs from trichloroethylene (TCE) contaminated groundwater and (2) evaluate whether these enrichments could degrade TCE coupled to acetylene degradation. Acetylenotrophs are microorganisms that use acetylene as their carbon and energy source. TCE contaminated groundwater was collected from wells at the Naval Air Warfare Center (NAWC) in West Trenton, New Jersey. Microbial acetylene uptake in groundwater samples was established by mixing the groundwater with a defined mineral medium to supply nutrients and providing acetylene as the sole electron donor and carbon source. The structure of the microbial community in those enrichments was characterized as shown by 16S rRNA gene sequencing and analysis. The acetylenotrophic groundwater enrichment cultures were then tested to assess whether they could utilize acetylene to drive reduction of TCE and tetrachloroethene (PCE) to vinyl chloride.