Physiological status and litter size can indeed have a significant impact on ewes' hematological parameters, which are essential indicators of their health. Therefore, this study examined the hematological profiles of ewes during pregnancy with single and twins in the Awassi ewes. The present study involved 232 ewes in good health and at sexual maturity. Among them, 123 ewes had single pregnancies, while 109 ewes had twin pregnancies. The age range of the ewes included in the study was between 3.5 and 4.5 years. Hematological tests were conducted on the sheep's blood samples promptly following collection. The findings demonstrated variations in hematological parameters among pregnant ewes, with differences based on litter size. Ewes carrying twin pregnancies exhibited significantly higher levels of red blood corpuscular, hemoglobin, packed cell volume, and mean corpuscular hemoglobin concentration during pregnancy. In comparison to single-pregnant ewes, Awassi ewes with twin pregnancies displayed elevated counts of white blood cells, lymphocytes, granulocytes, and granulocyte percentage compared to ewes with single pregnancies. Awassi ewes with twin pregnancies also exhibited a strong positive correlation with the leukocytes and erythrocytes constituents. In conclusion, these findings indicate that litter size significantly influences hematological parameters, highlighting the importance of considering the physiological status and litter size as indicators of ewes' health. The findings have practical implications in sheep breeding and reproduction, as they can be utilized to enhance the diagnosis, prognosis, and treatment of related conditions.