Molecular and morphological insights from the Ha’il region, King Salman Bin Abdulaziz Royal Natural Reserve

This study documents the first confirmed record of the Buthid scorpion Leiurus nigellus from Jabal Arnan in the Ha’il region, located within the King Salman Bin Abdulaziz Royal Natural Reserve (KSRNR) in the north-western Kingdom of Saudi Arabia (KSA). This species was originally described by Abu Afifeh, Aloufi & Al-Saraireh (2023). This locality extends the known distribution area for L. nigellus at over 200 km southeast of the type locality in Al-Ula, Al Madinah Province. A total of six copies of L. nigellus were collected during fieldwork carried out between June 2024 and April 2025, including two adult males, one adult female and three juveniles. The aim of this study was to confirm the taxonomic identity of Leiurus nigellus from a newly discovered locality using morphological examination and mitochondrial DNA analysis and documentation of its known geographic distribution. Adult specimens (one male and one female) were examined using comparative morphometric analysis following standard taxonomic protocols for scorpions, which confirmed diagnostic features consistent with the original species description. Meanwhile, habitat assessments indicated adaptation to semi-arid rock and gravel substrates. Molecular analysis was performed on an adult male using targeted mitochondria 16S rRNA gene sequencing (Sanger method). Phylogenetic relationships were inferred using neighbor-joining and maximum parsimony analyses, placement L. nigellus within Arabic Leiurus clade with bootstrap-supported affinity to Arab congeners and limited intraspecific divergence. It generated 16S rRNA sequence represents the first molecular record for L. nigellus and has been deposited in GenBank. Sexual dimorphism was evident in morphometric traits, but these differences reflect normal biological variation rather than taxonomic differentiation. The discovery of L. nigellus in northern Saudi Arabia emphasizes the importance of continued faunistic and genetic investigations in underexplored regions, both to delineate species distributions and to inform the conservation management of specialized desert arachnofauna.