Examples of a new class of phytase related to protein tyrosine phosphatases (PTP) were recently isolated from several anaerobic bacteria from the rumen of cattle. In this study, the diversity of PTP-like phytase gene sequences in the rumen was surveyed by using the polymerase chain reaction (PCR). Two sets of degenerate primers were used to amplify sequences from rumen fluid total community DNA and genomic DNA from nine bacterial isolates. Four novel PTP-like phytase sequences were retrieved from rumen fluid, whereas all nine of the anaerobic bacterial isolates investigated in this work contained PTP-like phytase sequences. One isolate, Selenomonas lacticifex, contained two distinct PTP-like phytase sequences, suggesting that multiple phytate hydrolyzing enzymes are present in this bacterium. The degenerate primer and PCR conditions described here, as well as novel sequences obtained in this study, will provide a valuable resource for future studies on this new class of phytase. The observed diversity of microbial phytases in the rumen may account for the ability of ruminants to derive a significant proportion of their phosphorus requirements from phytate.