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This study investigated the genetic variability within fish louse Argulus japonicus (Crustacea: Branchiura) from Africa, Middle East, and Asia by polymerase chain reaction in three mitochondrial DNA (mtDNA) regions, namely, cytochrome c oxidase subunit 1 ( cox1) and NADH dehydrogenase subunits 1 and 4 ( nad1 and nad4). Six different sequences from a portion of the cox1 gene (p cox1) and a portion of the nad1 and nad4 genes (p nad1 and p nad4) for ten adult specimens from infected fish in China, Egypt, and Syria were amplified separately from individual and the amplicons were subjected to direct sequencing. A + T percentages were 68.8–69% for p cox1, 77.1–77.6% for p nad1, and 60.4–60.9% for p nad4. Among all the collected parasites, A. japonicus sequence variations were 0.0–1.9% for cox1, 0.0–2.3% for nad1, and 0.0–0.8% for nad4. In rivers, sequence variations among all individuals were 0.4–0.8% for cox1, 1.0–2.3% for nad1, and 0.4–0.8% for nad4, while sequence variations among all the collected parasites in fish farms were 0.6–1.9% for cox1, 0.0–1.7% for nad1, and 0.2–0.6% for nad4. The nad1 was the most variable gene among selected markers, while nad4 was a more conserved gene than cox1. All isolates of A. japonicus were sister to Argulus americanus in phylogenetic tree and they grouped together in one sub-clade, while isolates from China and Egypt fish farms were closely clustered together. However, moderate genetic drift and slight mutation could be observed among A. japonicus individuals. These findings demonstrated the convenience and attributes of the three selected mtDNA sequences for population genetic studies of A. japonicus where nad1 is a new and reliable marker to detect the sequence variation among A. japonicus individuals. [ABSTRACT FROM AUTHOR]