A cross-sectional study was conducted in River Nile State, Sudan, between June to August 2018 to determine the prevalence of tick infestation on sheep and the potential risk factors associated with the infestation. A total of 135 sheep from five different localities (Shendi, Al Matamah, Ad-Damer, Atbara, and Berber) were examined and, of these, 90 were tick-infested (66.7%). A total of 340 ticks (male 185, female 155) were collected and identified using zoological taxonomic keys. The most dominant tick species collected in this survey were Rhipicephalus eversti Neumann, 1897 (38%), followed by Hyalomma anatolicum Koch, 1844 (23.8%), Rhipicephalus sanguineus Latreille, 1806 (20.6%), Rhipicephalus praetextatus Gerstacker, 1873 (16.4%), and Hyalomma dromedarii Koch, 1844 (1.2%). The chi-square analysis showed that there was a significant association (p ≤ 0.05) between tick infestation and localities, housing type, sex, control of ticks, and removal of manure. The highest prevalence rate was recorded in Atbara, in the open housing type, in females, in farms that did not use acaricides and did not remove the manure frequently. On the other hand, there was no significant association (p ˃ 0.05) between tick infestation and herd size, rearing system, breed, age and colour of coat (p=0.846). This study expanded the knowledge on tick fauna and associated risk factors in the River Nile State, and it demonstrated that multiple tick species are infesting sheep with the potential to transmit several tick-borne diseases.