Integrative taxonomy of the genus Longidorus (Nematoda: Longidoridae) reveals two new species in the Mediterranean Basin

The needle nematode genus Longidorus comprises approximately 194 species of polyphagous plant ectoparasites distributed worldwide, some of which serve as vectors for plant viruses. However, the high species diversity and conserved morphology of these nematodes pose significant challenges for accurate identification of species. To address this issue, we conducted an integrative taxonomic study across 264 sites in major olive-growing regions (Greece, Morocco, Italy, Portugal, and Spain) of the Mediterranean Basin, including nearby patches of natural vegetation. Herein, we describe two new species, Longidorus olearum sp. nov. and Longidorus morocciensis sp. nov., and report Longidorus oakgracilis in Portugal for the first time. We performed a comprehensive study that integrates morphological and morphometric traits with molecular data from nuclear ribosomal RNA (rRNA) genes (D2-D3 expansion segments of 28 S, ITS1, and partial 18 S) and a mitochondrial DNA (mtDNA) marker, specifically the Cytochrome c oxidase subunit I (COI). The results of our phylogenetic analyses provided robust support for the delimitation of the newly described species, L. olearum sp. nov. and L. morocciensis sp. nov., and further clarified of three previously recognized species within the genus: L. magnus, L. oakgracilis, and L. vineacola. Phylogenetic relationships inferred from ribosomal and mitochondrial markers revealed that the majority of Longidorus species from the Mediterranean Basin clustered within subclades of Clade I. The phylogenetic placement of these species demonstrated strong congruence across lineages, corroborating previous studies on the genus. These findings contribute to a broader understanding of Longidorus biodiversity in the Mediterranean region and highlight the need for further intensive and wide-ranging nematological surveys.