Morphological and Molecular Insights Into <scp> <i>Oestrus ovis</i> </scp> (Diptera: Oestridae) Infesting Small Ruminants in Pakistan

ABSTRACT Oestrus ovis (sheep nasal botfly) is a cosmopolitan myiasis‐causing parasite responsible for substantial health and productivity losses in small ruminants. Despite its veterinary importance, integrated epidemiological and molecular data from endemic regions such as Pakistan remain limited. This study provides a comprehensive investigation of O. ovis in sheep and goats by combining abattoir‐based epidemiology, detailed larval morphology and mitochondrial cox1 ‐based genetic analysis. A total of 311 small‐ruminant heads (176 goats, 135 sheep) were examined, revealing an overall prevalence of 37.6%, with slightly higher infestation in sheep. In goats, infestation was significantly associated with host sex and nasal pigmentation ( p &lt; 0.05), whereas in sheep only nasal pigmentation showed a significant effect. Morphological and morphometric analyses identified second‐ (L 2 ) and third‐instar (L 3 ) larvae, with L 3 exhibiting greater body dimensions and more developed cephalopharyngeal structures, including robust mouth hooks and dense ventral supination. Molecular characterization based on a 392 bp fragment of the mitochondrial cox1 gene revealed moderate haplotypic diversity, with a dominant haplotype shared among multiple samples. Comparative analysis demonstrated high nucleotide similarity (97%–99%) and low genetic divergence (0.00–0.02) with global reference sequences. Phylogenetic and haplotype network analyses placed Pakistani isolates within a well‐supported, globally distributed O. ovis lineage lacking clear geographic structuring. These findings highlight a high burden of O. ovis infestation in Pakistani small ruminants, confirm distinct larval stage differentiation and demonstrate limited mitochondrial variability consistent with recent common ancestry and host‐mediated dispersal. This integrative approach provides essential baseline data for surveillance and control strategies in endemic regions.