The availability of completely sequenced genomes from eight species of nematodes has provided an opportunity to identify novel cis-regulatory elements in the promoter regions of Caenorhabditis elegans transcripts using comparative genomics. We determined orthologues for C. elegans transcripts in C. briggsae, C. remanei, C. brenneri, C. japonica, Pristionchus pacificus, Brugia malayi and Trichinella spiralis using the WABA alignment algorithm. We pooled the upstream region of each transcript in C. elegans with the upstream regions of its orthologues and identified conserved DNA sequence elements by de novo motif discovery. In total, we discovered 158 017 novel conserved motifs upstream of 3847 C. elegans transcripts for which three or more orthologues were available, and identified 82% of 44 experimentally proven regulatory elements from ORegAnno. We annotated 26% of the motifs as similar to known binding sequences of transcription factors from ORegAnno, TRANSFAC and JASPAR. This is the first catalogue of annotated conserved upstream elements for nematodes and can be used to find putative regulatory elements, improve gene models, discover novel RNA genes, and understand the evolution of transcription factors and their binding sites in phylum Nematoda. The annotated motifs provide novel binding site candidates for both characterized transcription factors and orthologues of characterized mammalian transcription factors.