Diversity and phylogenetic relationships within the spiral nematodes of Helicotylenchus Steiner, 1945 (Tylenchida: Hoplolaimidae) as inferred from analysis of the D2-D3 expansion segments of 28S rRNA gene sequences

The spiral nematodes of the genus Helicotylenchus are globally distributed and associated with the root system of diverse groups of plants in cultivated and uncultivated areas. Several species are considered serious parasites of crops. The identification of many Helicotylenchus species is not always reliable, in part because many species share very similar diagnostic characters and high intraspecific variation. To verify species identification of geographically distant populations of Helicotylenchus, we tested monophyly of some classical morphospecies and studied their phylogenetic relationships; specifically, we conducted sequence and phylogenetic analysis of 89 sequences of the D2-D3 expansion segments of 28S rRNA gene sequences from 54 Helicotylenchus isolates, including species identified as H. brevis, H. digonicus, H. dihystera, H. labiodiscinus, H. leiocephalus, H. martini, H. multicinctus, H. platyurus, H. pseudorobustus and H. vulgaris, together with three outgroup taxa. Phylogenetic analysis distinguished nine highly or moderately supported major clades within Helicotylenchus. Using the molecular approach we were able to confirm congruence with morphologicalbased identification of samples of H. dihystera and H. multicinctus. However, sequence and phylogenetic analysis using Bayesian inference and maximum parsimony analysis showed that isolates collected in different countries and morphologically identified as H. pseudorobustus, H. digonicus or H. vulgaris were each representative of several different and, sometimes, unrelated lineages. Further detailed comparative morphometrics and morphological studies will help to elucidate if there is some misidentification or if putative species actually comprise a complex of cryptic species. Molecular analysis also revealed that 14 samples were classified as representatives of 11 unidentified species. Molecular characterisation of known Helicotylenchus species especially, using samples collected from type localities, is needed for future reliable identification of species of this genus.