Myriapoda is no evolutionary sideshow. The subphylum contains more than 18,500 described species, including roughly 14,245 millipedes, 3,300 centipedes, 850 pauropods and 200 symphylans, and it sits as the sister group to Pancrustacea, the clade uniting crustaceans and insects. Together these lineages form Mandibulata, the arthropods equipped with mandibulate mouthparts.

Myriapods also hold the distinction of being the oldest known terrestrial animals: Scottish fossils of archipolypodan millipedes such as Cowiedesmus eroticopodus, Albadesmus almondi and Pneumodesmus newmani date to the mid-Silurian, placing them among the very first creatures recorded on land. Although the monophyly of the four myriapod classes—Chilopoda, Diplopoda, Symphyla and Pauropoda—is now widely accepted, the branching order among them has remained stubbornly unstable across decades of phylogenetic study. Previous genomic attempts had produced a tangle of competing hypotheses.

Early transcriptome-based analyses by Fernandez and colleagues proposed two alternative arrangements, one grouping Diplopoda and Chilopoda with Symphyla as the outgroup, the other placing Symphyla closer to the millipedes, while pauropods were left out entirely. Subsequent transcriptomic studies supported a sister-group relationship between Pauropoda and Symphyla, a clade named Edafopoda, alongside a Chilopoda-Diplopoda pairing called Pectinopoda. Most recently, a reanalysis using quartet-based sampling found support for a monophyletic Progoneata, in which pauropods, symphylans and millipedes descend from a common ancestor, when more stringent character-state filters were employed.

Meanwhile, the phylogenetic identity of rare orders remained contested, most conspicuously the relictual craterostigmomorph centipedes of Tasmania and New Zealand, a two-species lineage whose placement has flip-flopped between morphological and molecular studies, and the minute millipede orders Siphoniulida and Siphonocryptida, which until very recently had never been assessed with molecular data at all. The new study tackles these questions with ultraconserved elements, or UCEs, stretches of DNA so conserved across the animal kingdom that they can be captured and sequenced even in distantly related organisms. The researchers designed a Myriapoda-specific probe set comprising 19,997 baits targeting 1,087 loci, a deliberately dense toolkit intended to work across all four classes, from the subphylum level down to individual species.

In preliminary in silico tests against available genomes and transcriptomes, locus recovery ranged from just 9 loci in the millipede Sigmoria abbotti to 1,051 in the millipede Trigoniulus corallinus, with genome-based samples averaging 1,012 loci compared with 317 for transcriptomes. Notably, the outgroup harvestman Phalangium opilio yielded 925 loci in silico, confirming that many targeted loci are conserved well beyond Myriapoda and across other arthropods. Laboratory testing on freshly sequenced specimens proved even more encouraging.

Across 63 in vitro samples, the average raw locus recovery reached 776, exceeding the 684-locus average for genomes and the 422-locus average for transcriptomes. Scutigeromorph centipedes, the fast-running house centipede relatives, performed exceptionally well, averaging 945 raw loci and topping out at 1,025 in Thereuopoda clunifera. Only two taxa, the millipedes Abacion magnum and Cleidogona sp., fell below 100 loci.

When the team benchmarked their new probe set against a recently published universal Metazoa probe set, the myriapod-specific version generally recovered more loci, particularly from transcriptomic samples, although a handful of species, including the centipede Scolopendra cretica, showed the reverse pattern. The average locus length of 901 base pairs and GC content of 41.76 percent fell squarely within the range of an optimally designed probe set, and the ratio of roughly 20,000 probes capturing about 1,100 loci reflects the exceptional genetic divergence of the taxa used in its design. The phylogenetic results crystallized into two primary topologies, both confirming the monophyly of all four classes but disagreeing on their deeper arrangement.

The favored topology, supported by the 50 and 60 percent occupancy matrices under maximum-likelihood and Bayesian analyses as well as by quartet-based SeaLion analysis, places Chilopoda as the sister group to a clade uniting Diplopoda, Symphyla and Pauropoda, thereby recovering both Progoneata and Edafopoda. The alternative, recovered in the sparsest 70 percent occupancy matrix—which contained only 40 loci—and under CAT-GTR models, pulls Pauropoda out of Myriapoda entirely and groups it with pancrustacean outgroups, a result the authors consider artifactual, likely driven by long-branch attraction and the low number of loci in that matrix. Post-processing of the quartet results strongly rejected the topologies that would separate either Diplopoda or Chilopoda from the pauropod-symphylan pairing, lending further weight to the Progoneata hypothesis.

Within the millipedes, several unexpected relationships emerged. The bristly millipedes of order Polyxenida clustered with Pentazonia, the pill-millipede-bearing group, rather than branching off first as traditionally assumed, a result consistently recovered across all matrices though possibly an artifact of limited polyxenid sampling. Within Pentazonia itself, the analyses placed Glomerida as sister to Glomeridesmida plus Sphaerotheriida, overturning previous arrangements and implying that the ability to roll into a ball, shared by Glomerida and Sphaerotheriida, evolved independently at least twice.

The rare order Siphoniulida, represented here by the recently described Siphoniulus porosus, one of only three known extant species, was robustly resolved as the sister group to Stemmiulida, settling a question that morphology alone had left unresolved. Meanwhile, the siphonocryptid Hirudicryptus canariensis nested firmly within Polyzoniida, supporting the recent proposal to relegate the former order Siphonocryptida to family rank. The clade Nematophora, traditionally united by preanal spinnerets, was not recovered in any analysis, suggesting those structures may have independent origins across millipede lineages or been secondarily lost in the juliform millipedes.

Among the centipedes, the most striking departure from convention concerns Craterostigmomorpha. Rather than Scutigeromorpha, the familiar house centipedes, branching first as most previous morphological and molecular studies concluded, the new trees place Craterostigmomorpha, the least diverse centipede order, as the earliest-splitting lineage, with Scutigeromorpha as sister to the remaining three orders. This arrangement supports the Amalpighiata hypothesis, in which Lithobiomorpha rather than Craterostigmomorpha pairs with the epimorphic centipedes, a topology consistent with recent transcriptomic work.

The authors point to the preferred breakage point of legs during self-amputation as a potential morphological signature: recent work showed that both Craterostigmus and Scutigera share a breakage point between the trochanter and prefemur, a character that, optimized on the new trees, shifts to the coxa-trochanter joint in the ancestor of Amalpighiata. At finer scales, the analyses revealed paraphyly in the lithobiomorph family Henicopidae, confirmed a well-supported basal split within Geophilomorpha between Placodesmata and Adesmata, and recovered expected family-level structure within scutigeromorphs, though the relationships among the three extant scutigeromorph families varied between matrices. The study's authors emphasize that the probe set is designed as a community resource rather than a definitive resolution of every myriapod node.

Sparse sampling of pauropods and symphylans, and the inclusion of only one representative from many polydesmid families, likely contributed to some unstable placements, and denser taxon coverage is expected to clarify contested relationships. Still, the toolkit's ability to recover phylogenetically informative variation even within a single widespread species, Scutigera coleoptrata, demonstrates its utility across taxonomic scales. Because UCE capture works well with the fragmented DNA of historical museum specimens that are unsuitable for PCR amplification or for transcriptomic and genomic sequencing, the probe set opens the door to incorporating such specimens into modern phylogenomic analyses.

The probe set is commercially available, raw sequence data have been deposited in the Sequence Read Archive, and assemblies and matrices are publicly accessible, giving myriapodologists everywhere the means to probe the deep history of one of Earth's oldest and most enigmatic terrestrial lineages.

Subject of Research: Phylogenomics of Myriapoda using a novel ultraconserved-element probe set

Article Title: A myriad of mysteries on many legs: Disentangling the systematics of Myriapoda with UCEs

Article References: Shapiro, L., Rodríguez-Flores, P., Derkarabetian, S., Edgecombe, G. D., Recuero, E., Benavides, L. R., & Giribet, G. (2026). A myriad of mysteries on many legs: Disentangling the systematics of Myriapoda with UCEs. iScience, 29(11), Article 117989. https://doi.org/10.1016/j.isci.2026.117989

Image Credits: AI Generated

DOI: 10.1016/j.isci.2026.117989

Keywords: Myriapoda, millipedes, centipedes, ultraconserved elements, phylogenomics, probe set, Progoneata, Edafopoda, Craterostigmomorpha, Siphoniulida, museum specimens, Silurian fossils