Thelohania solenopsae is a microsporidian parasite that may serve as a biological control agent for the red imported fire ant, Solenopsis invicta. A rapid, filter-based PCR amplification method detecting a portion of the small-subunit ribosomal RNA gene was developed to facilitate field studies detecting the parasite in fire ants. Processing ant homogenates with a commercially available membrane-based system, FTA Classic Card technology, compared favorably with traditional DNA extraction and PCR amplification methods. As few as 100 spores were detected. The FTA membrane system is a simple, extraction-free method for detecting T. solenopsae in fire ants, and allows for easy archival storage of DNA samples.