The mitochondrial DNA (mtDNA) molecules of the nematode worms, Caenorhabditis elegans and Ascaris suum contain 22 putative genes for non-standard forms of tRNAs. The inferred transcripts can be folded into 20 separate structures each resembling a tRNA whose T psi C arm and variable loop are replaced with a simple loop of 6-12 nucleotides. In two further structures [that resemble tRNAs for ser(UCN) and ser(AGN)], the dihydrouridine arm is replaced by a loop of 5-8 nucleotides. By hybridizing mt-tRNA gene-specific oligonucleotide probes to nematode RNAs, we have obtained evidence for transcription of at least nine C.elegans and three A.suum mt-tRNA genes. Each transcript (tRNA) is the exact size predicted from the respective DNA sequence, to which three nucleotides, presumably CCA, have been added following transcription. An exception was C.elegans mt-tRNAasn, most molecules of which had one nucleotide (plus CCA) more than predicted from the gene. The data presented strongly support the conclusion that the functional mt-tRNAs of nematode worms are direct transcripts (with only CCA addition) of the structurally unusual mt-tRNA genes. There is no evidence of trans-splicing or RNA editing to add the sequences missing from these nonstandard tRNAs. We presume, therefore, that the non-standard forms are active in mitochondrial protein synthesis.