Production of recombinant EMA-1 protein and its application for the diagnosis of Theileria equi using an enzyme immunoassay in horses from São Paulo State, Brazil.

The researchers produced a recombinant protein known as EMA-1 to diagnose Theileria equi, a horse disease. This diagnostic tool was tested in horses in São Paulo State, Brazil, revealing a high infection rate.

Understanding the Research

• This study focuses on equine piroplasmosis, which is a disease in horses caused by Theileria equi. The team sought to design a reliable diagnostic method for this disease.
• Equi Merozoite Antigen 1 (EMA-1), an erythrocytic-stage surface protein, was chosen to develop this diagnostic. EMA-1 was cloned from the Theileria equi Jaboticabal strain and then expressed in the Escherichia coli bacterium as a histidine-tagged protein (His6-EMA1).

Key Findings

• The expressed EMA-1 reacted with specific antibodies when tested with Western blot, a common method for detecting specific proteins.
• Its molecular mass primarily agreed with the expected value, validating the successful production of EMA-1.
• The EMA-1 gene showed a high degree of homology when compared with published sequences, indicating a strong similarity to EMA-1 genes from various other strains isolated globally.
• The purified His6-EMA1 protein was then used in an enzyme-linked immunosorbent assay (ELISA) to detect anti-T. equi antibodies in horses.
• It was found that the ELISA successfully distinguished between equi-infected horses and Babesia caballi-infected horses or healthy horses, demonstrating the reliability of this diagnostic tool.
• When field serum samples from São Paulo were tested with ELISA, 95.88% (163/170) were found to be positive for T. equi infection, indicating a widespread presence of the disease in this region.

Implications of the Study

• The successful expression of the His6-EMA1 protein in E. coli suggests this method could be used in the future to produce immunodiagnostic antigens reliably.
• The ELISA test developed in the study can effectively differentiate between T. equi and Babesia caballi infections in horses, which could drastically improve diagnostic accuracy for equine piroplasmosis.
• The high prevalence of T. equi infection found in São Paulo underscores the need for effective diagnostic methods and possibly, urgent interventions to manage and control the disease.