Genome clones and expressed sequence tags (ESTs) from the ascidian Ciona intestinalis and from the larvacean Oikopleura dioica were analysed for the presence of lysozyme-encoding genes. Two genes were found to potentially code for goose-type lysozymes in Oikopleura, while three or possibly more g-type proteins form the lysozyme complement of C. intestinalis, and at least one of these genes from each species is expressed based on EST data. No genes for chicken- or invertebrate-type lysozymes were found in either urochordate species. Consistent with this finding, extracts of Oikopleura animals possessed hydrolysing activity on bacterial cell walls, and this activity was not inhibited in the presence of a known inhibitor of chicken-type lysozyme. A wide range of isoelectric points for the predicted lysozymes from Ciona (pI 4.4, 6.4 and 9.9) and from Oikopleura (pI 5.0 and 8.0) suggests tissue-specific adaptations as well as specific functional roles of the lysozymes. Comparisons of gene structures, encoded sequences, cysteine residue content and their positions in the proteins indicate that the g-type lysozymes of Ciona intestinalis are more closely related to those of vertebrates than are the g-type lysozymes of Oikopleura. Multiple genes from each species may result from separate and lineage-specific duplications followed by functional specialisation.