Report of three species of Belondiroidea Thorne, 1939 (Nematoda: Dorylaimida) from Taf region of Khorramabad County, Iran

The Taf region encompasses several distinct local climate zones and harbors rich biodiversity, yet its nematode fauna remains unstudied. The distinguishing characteristics of this superfamily include a relatively short odontostyle, a slender but weakly muscular anterior section of the esophagus, an enlarged basal expansion of the esophagus, which varies in length and is encased by a discernible sheath composed of spiral muscles, and a potentially similar or dissimilar tail morphology in males and females. Presently, the superfamily comprises 36 genera. To investigate the nematode biodiversity of the Taf region in Khorramabad County, 50 soil samples were collected. Among the extracted nematodes, three species belonging to Belondiroidea—Oxydirus oxycephalus, Metaxonchium bihariense, and Syncheilaxonchium nairi—were identified. The morphometric, morphological, and molecular characteristics of these species were analyzed using the 28S rDNA gene region.

Soil samples were collected from the rhizosphere of forest trees in the Taf region. Nematodes were extracted utilizing a modified Brown & Boag (1988) method. Following extraction, specimens were fixed and preserved in anhydrous glycerin according to the protocol outlined by De Grisse (1969) and mounted on permanent glass slides. Species identification was conducted using a light microscope equipped with a Dino-Eye Eyepiece Camera, relying on morphological and morphometric characteristics alongside established taxonomic keys. Photomicrographs were captured using the same microscope outfitted with a DP50 digital camera. Nematode DNA was extracted from individual specimens following the protocol described by Archidona-Yuste et al. (2016). The D2-D3 segments were amplified utilizing the D2A and D3B forward and reverse primers, respectively. Subsequently, the generated sequences were aligned with other 28S rDNA gene sequences available in GenBank using MEGA6 software. Phylogenetic analysis of the sequence datasets was conducted via Bayesian inference (BI) employing MrBayes 3.1.2 under the GTR + I + G model. The resulting tree was visualized using FigTree v1.4.3.

In this study, three species from the Belondiroidea superfamily were identified. Through morphological, morphometric, and D2-D3 expansion analysis of the 28S rDNA, these species were determined to be Oxydirus oxycephalus, Metaxonchium bihariense, and Syncheilaxonchium nairi. Notably, O. oxycephalus was found for the first time within Iran's nematode fauna. In females, O. oxycephalus displays a cylindroid body shape, measuring 1.5-1.8 mm in length. The lip region is slightly asymmetrical, while the spear is fusiform, measuring 6.5-8.0 µm in length. The oesophagus assumes a bottle-shaped configuration, with the basal bulb being cylindrical and encased by a prominent muscular spiral sheath. The vulva is observed as a transverse slit with non-sclerotized lips, and the posterior gonad is normal (opisthodelphic) with a reduced anterior gonad. The tail is long and filiform, typically presenting a swelling. Male specimens share a similar morphology to females, with spicules measuring 34.0-35.5 µm in length and 9-11 ventromedian supplements. The phylogenetic Bayesian consensus tree, derived from D2-D3 28S rDNA analysis, reveals that O. oxycephalus, along with O. oxycephalus (AY593012), forms a subclade with 100% posterior probability (PP) support, situated within a larger clade containing Opisthodorylaimus sylphoides (AY593010 and AY593008) from the Thornenematinae subfamily. The morphological and morphometric characteristics of the S. nairi population studied closely resemble those of the type specimens in terms of general morphology and measurements, indicating a high degree of similarity. A single D2-D3 28S rDNA gene sequence (OR594282) was obtained for S. nairi, which forms a highly supported (100% PP) clade with another representative of Syncheilaxonchium sp. (KP792285 and KP792284).
The Iranian population of M. bihariense closely resembles the type population, with only minor differences observed in body length and pharyngeal length. Notably, a sequence (OR600367) was obtained for the first time for this population.

The placement of O. oxycephalus sequences in the phylogenetic tree, segregated from other taxa within this superfamily and grouped within the same clade (96% posterior probability), alongside O. sylphoides from the Thornenematinae, suggests that Belondiroidea constitutes a polyphyletic group. Additionally, as noted by Peña-Santiago and Abolafia (2023), the presence of a spiral sheath surrounding the expanded part of the pharynx is considered an apomorphic feature.