We examined the presence of TTAGG telomeric repeats in 22 species from 20 insect orders with no or inconclusive information on the telomere composition by single-primer polymerase chain reaction with (TTAGG)6 primers, Southern hybridization of genomic DNAs, and fluorescence in situ hybridization of chromosomes with (TTAGG)n probes. The (TTAGG)n sequence was present in 15 species and absent in 7 species. In a compilation of new and published data, we combined the distribution of (TTAGG)n telomere motif with the insect phylogenetic tree. The pattern of phylogenetic distribution of the TTAGG repeats clearly supported a hypothesis that the sequence was an ancestral motif of insect telomeres but was lost repeatedly during insect evolution. The motif was conserved in the "primitive" apterous insect orders, the Archaeognatha and Zygentoma, in the "lower" Neoptera (Plecoptera, Phasmida, Orthoptera, Blattaria, Mantodea, and Isoptera) with the exception of Dermaptera, and in Paraneoptera (Psocoptera, Thysanoptera, Auchenorrhyncha, and Sternorrhyncha) with the exception of Heteroptera. Surprisingly, the (TTAGG)n motif was not found in the "primitive" pterygotes, the Palaeoptera (Ephemeroptera and Odonata). The Endopterygota were heterogeneous for the occurrence of TTAGG repeats. The motif was conserved in Hymenoptera, Lepidoptera, and Trichoptera but was lost in one clade formed by Diptera, Siphonaptera, and Mecoptera. It was also lost in Raphidioptera, whereas it was present in Megaloptera. In contrast with previous authors, we did not find the motif in Neuroptera. Finally, both TTAGG-positive and TTAGG-negative species were reported in Coleoptera. The repeated losses of TTAGG in different branches of the insect phylogenetic tree and, in particular, in the most successful lineage of insect evolution, the Endopterygota, suggest a backup mechanism in the genome of insects that enabled them frequent evolutionary changes in telomere composition.

The (TTAGG)n sequence is supposed to be an ancestral DNA motif of telomeres in insects. Here we examined the occurrence of TTAGG telomeric repeats in other arthropods and their close relatives by Southern hybridization of genomic DNAs and fluorescence in-situ hybridization (FISH) of chromosomes with (TTAGG)n probes or, alternatively, with the 'vertebrate' telomeric probe, (TTAGGG)n. Our results show that the (TTAGG)n motif is conserved in entognathous hexapods (Diplura and Collembola), crustaceans (Malacostraca, Branchiura, Pentastomida, and Branchiopoda), myriapods (Diplopoda and Chilopoda), pycnogonids, and most chelicerates (Palpigradi, Amblypygi, Acari, Opiliones, Scorpiones, Pseudoscorpiones, and Solifugae) but not in spiders (Araneae). The presence of TTAGG repeats in these groups suggests that the sequence is an ancestral motif of telomeres not only in insects but in Arthropoda. We failed, however, to detect the TTAGG repeats in close relatives of the arthropods, Tardigrada and Onychophora. But while Onychophora had the 'vertebrate' (TTAGGG)n motif instead, the Tardigrada did not. The (TTAGG)n motif probably evolved from the (TTAGGG)n motif. Based on our and compiled data, we presume that the 'vertebrate' motif (TTAGGG)n is an ancestral motif of telomeres in bilaterian animals and possibly also in the superclade including animals, fungi and amoebozoans.

In most eukaryotes the telomeres consist of short DNA tandem repeats and associated proteins. Telomeric repeats are added to the chromosome ends by telomerase, a specialized reverse transcriptase. We examined telomerase activity and telomere repeat sequences in representatives of basal metazoan groups. Our results show that the 'vertebrate' telomere motif (TTAGGG)( n ) is present in all basal metazoan groups, i.e. sponges, Cnidaria, Ctenophora, and Placozoa, and also in the unicellular metazoan sister group, the Choanozoa. Thus it can be considered the ancestral telomere repeat motif of Metazoa. It has been conserved from the metazoan radiation in most animal phylogenetic lineages, and replaced by other motifs-according to our present knowledge-only in two major lineages, Arthropoda and Nematoda.