Morphological data and DNA barcoding reveal the presence of the alien freshwater leech Helobdella octatestisaca (Hirudinida: Glossiphoniformes) in North Africa (Tunisia)

Background We hereby report the first occurrence of Helobdella octatestisaca in North Africa, specifically in Tunisia, as a likely introduced species from the Neotropical Region. Historically, leeches bearing a prominent chitinous scute on their dorsal surface were commonly diagnosed as H. stagnalis . Most probably, H. octatestisaca had previously been misidentified as H. stagnalis in Tunisia. Methods and results The identification was primarily based on morphological evidence, supplemented by genetic data obtained from COI DNA barcoding. The morphology of the examined specimens was consistent with the original species description, notably characterized by the presence of four pairs of testisacs. To support our findings, we conducted a phylogenetic analysis using the Maximum Likelihood method based on COI alignment constructed with the newly obtained sequence from Tunisian specimens and complete or nearly complete ‘Folmer fragment’ sequences of congeners sourced from the GenBank database. Conclusions This study highlights the first identification of H. octatestisaca in North Africa and suggests that previous records of H. stagnalis in Tunisia likely misidentified this species.