Research note: Evans Blue Dye as an objective quantitative tool for lesion scoring in Eimeria and Histomonas meleagridis infected poultry

Intestinal protozoan infections such as Eimeria spp. and Histomonas meleagridis remain major challenges to turkey production, leading to impaired feed efficiency, intestinal damage, and mortality. Accurate quantification of intestinal lesions is critical for evaluating disease severity and treatment efficacy; however, traditional lesion scoring systems are subjective and limited in detecting subtle or microscopic tissue changes. This research note introduces the use of Evans Blue Dye (EBD) as an objective, quantitative tool to assess intestinal and hepatic lesions in turkeys experimentally challenged with Eimeria or H. meleagridis. A total of one hundred eighty 1-day-old male poults were assigned to non-challenged (NC), Eimeria-challenged (EM), or H. meleagridis-challenged (HM) groups with five replicates per group and reared for 28 days. Evans Blue Dye (10 mg/kg BW) was injected intravenously one hour before necropsy, and dye concentrations were quantified spectrophotometrically in liver, duodenum, jejunum, and ceca tissues. HM poults showed the poorest growth performance and the highest liver and cecal lesion scores (P < 0.001), with EBD concentrations approximately 150% higher than those in controls, indicating severe vascular leakage and tissue damage caused by Histomonas challenge. EM poults exhibited moderate intestinal lesions and corresponding EBD increases in the duodenum and jejunum, consistent with species-specific invasion patterns. Overall, EBD concentration strongly reflected lesion severity and distribution, supporting its use as an objective and reproducible marker for evaluating protozoan-induced intestinal injury in poultry. This method minimizes observer bias and enhances sensitivity in detecting mild or microscopic lesions, offering a valuable measurement tool over conventional lesion scoring systems.