Microsporidia are a group of obligate intracellular eukaryotic parasites with small genomes. They infect animals from a wide variety of phyla, including humans. Two manganese superoxide dismutase (MnSOD) genes, designated NbMnSOD1 and NbMnSOD2, were found to be organized in a tandem array within the Nosema bombycis genome. The genes, both 678 bp in length, were found to be more similar to each other than they are to homologous genes of other Microsporidia, suggesting that the tandem duplication occurred subsequent to the development of this lineage. Reverse transcript PCR shows that mRNA for both genes is present in the spores. Analysis of the primary structure, hydrophobic cluster analysis, target signal analysis, and phylogenetic analysis all indicate that NbMnSOD1 is dimeric and targeted to the cytosol. NbMnSOD2 seems to have changed more rapidly and is under less evolutionary constraint than NbMnSOD1 suggesting that NbMnSOD2 may function under different conditions or in different tissues of its host rather than simply resulting in an increase in expression. A phylogenetic analysis of MnSOD sequences from eukaryotes, Archaea, and bacteria shows the microsporidial MnSODs to be grouped with the bacteria suggesting a possible horizontal gene transfer.