Technical Abstract: Although the biosynthesis of oxalic acid is known to occur in a number of bacteria, the mechanism(s) regulating its production remains largely unknown. To date, there is no report on the identification of an oxalic acid biosynthetic pathway gene from bacteria. In an attempt to identify such a gene(s), a mutant screen was conducted using the simple oxalic acid producing phytopathogenic bacterium, Burkholderia glumae. Four mutants that failed to produce oxalic acid were isolated from a transposon-mutagenized B. glumae library and named Burkholderia oxalate defective (bod) 1. DNA sequence analysis revealed that each mutant contained an insertion event at a different site in the same open reading frame (ORF), which we referred to as the oxalate biosynthetic component (obc) A locus. Complementation of the bod1 mutant with the obcA gene; however, resulted only in a partial restoration of the oxalic acid producing phenotype. Further complementation studies utilizing a larger DNA fragment encompassing the obcA locus coupled with deletion mutagenesis, resulted in the identification of another ORF, that we named the obcB locus, which was essential for the higher levels of oxalic acid production. Transcript analysis indicated that both the obcA and obcB are co-expressed and encoded on a single polycistron message.