Telomeres are made up of tandem arrays of telomeric-repeat motifs (TRMs) and telomere-binding proteins (TBPs), which address the difficulties of end-protection and end-replication. TRMs are usually highly preserved due to the sequence specificity of TBPs, although significant TRM alterations have been observed in several taxa but not in Nematoda. We used public whole-genome sequencing data to analyse putative TRMs of 100 nematode species to investigate TRM evolution in Nematoda. We discovered that six distinct branches included specific novel TRMs, suggesting frequent TRM evolution in Nematoda. We concentrated on one of the six branches, the Panagrolaimidae family, to validate TRM evolution by collecting nematode species and obtaining whole-genome sequencing data. We also de novo assembled five high-quality draft genomes of Panagrolaimidae species and these genomes showed densely clustered arrays of the novel TRM. With reference to the telomere evolution in Caenorhabditis elegans, we comprehensively analysed subtelomeric regions in the genomes to determine how the unique TRM evolved. We found that the novel TRM was used to preserve telomere integrity by alternative lengthening of telomeres even in species that employ the canonical TRM. We propose a hypothetical scenario in which some pre-existing TBPs may be capable of binding both canonical and novel TRMs, resulting in pre-adaptation of the novel TRM and aiding its evolution in the genus Panagrellus.
