Detection of naturally occuring mixed infection of Theileria lestoquardi and Theileria annulata in buffalo, in Pakistan.

Theileriosis caused by Theileria lestoquardi (malignant ovine theileriosis) in sheep and Theileria annulata (topical theileriosis) in cattle, is a tick-borne hemoprotozoal disease that causes major economic losses in animal production caused by a tick-transmitted haemoprotozoan Theileria species, infecting various domestic and wild animals. It is a threat to the buffalo population in Pakistan due to high mortality rates and economic losses. Extensive mixed farming in Pakistan poses threats due to mixed infections and increased mortality rates. The current case report describes the mixed infection and characterization of Theileria lestoquardi with Theileria annulata in a buffalo raised in a mixed farming setup, in Pakistan. A 6.5-year-old female buffalo presented at the Outdoor Hospital exhibiting swollen sub-scapular lymph nodes, pale mucous membranes, a high fever (41.5°C), dullness, depression, shortness of breath, and a decrease in milk production. Investigation through microscopy, hematology, and PCR confirmed that the buffalo was suffering from theileriosis. A Giemsa-stained thin blood smear showed piroplasms indicating theileriosis. Hematological analysis of the blood revealed lower values for hemoglobin and red blood cells (RBCs), while higher counts of lymphocytes, neutrophils, and monocytes were observed during the infection. The PCR performed on the blood sample was positive for Theileria spp. using RLB universal primers and T. annulata-specific primers. The sequence revealed similarity with T. lestoquardi reported in Pakistan, Iran, Tunisia, and India. The buffalo was treated with Bupralex® and Oxtra LA® at a dosage of 1ml/20kg of body weight I/M, respectively, along with supportive treatment. Despite these interventions, the animal died probably due to damage to cardiac muscles. This study reports the first case of T. annulata mixed infection with T. lestoquardi in a buffalo in Punjab, Pakistan. It can be concluded that mixed farming exacerbates the epidemiology and spread of ovine malignant theileriosis and mortalities possibly due to cardiac arrest in mixed infections.