Evaluation of multiple methods for the diagnosis of equine Hemomycoplasmosis in Misan, Iraq.

Overview

• This study evaluates various diagnostic methods for detecting equine Hemomycoplasmosis, aiming to identify the most accurate approach for diagnosing this infection in horses in Misan, Iraq.
• The research compares cytological techniques, molecular diagnosis via qPCR, assessment of acute phase response, and postmortem examinations to determine their effectiveness and agreement in diagnosis.

Introduction and Background

• Hemomycoplasmosis is caused by hemotropic mycoplasmas (Hemoplasma), formerly classified as Eperythrozoon spp., which are parasites that infect the surface of erythrocytes (red blood cells).
• These parasites affect various mammals, including horses, cattle, sheep, goats, dogs, cats, buffalo, camels, and can even transmit to humans.
• Hemoplasmas are uncultivable, meaning they cannot be grown in laboratory culture media, making diagnosis challenging.
• The study focuses on diagnosing equine hemomycoplasmosis accurately using multiple available diagnostic methods.

Methods Examined

• Cytological examinations: Includes Giemsa-stained blood smears and fluorescent microscopic diagnosis through acridine orange staining to detect parasites on erythrocytes.
• Molecular diagnosis: Quantitative polymerase chain reaction (qPCR) was used to detect and quantify Hemoplasma DNA in blood samples.
• Acute phase response evaluation: Measurement of inflammatory markers in infected horses to observe immune system activation.
• Postmortem examination: Inspection of dead horses for gross pathological changes, and histopathological analysis of tissues to understand organ damage caused by the infection.

Results

• Infection rates: Cytological testing identified 88% of sampled horses as infected (88 out of 100), while qPCR confirmed infection in 79% (79 out of 100).
• Agreement between cytology and qPCR:
  • The overall diagnostic agreement was 91% (91 out of 100 samples).
  • Cohen’s Kappa coefficient, a statistical measure of agreement beyond chance, was calculated at 0.678, which represents substantial agreement between the two methods based on Landis and Koch benchmarks.
• Acute phase response: Infected horses showed significant changes in inflammatory markers, indicating an active immune response to infection.
• Postmortem findings:
  • Dead horses exhibited general emaciation and paleness of internal organs.
  • There was noticeable enlargement of spleen (splenomegaly), liver, kidneys, and heart; some organs showed white patches indicating tissue changes.
  • Histopathology indicated:
    • Degeneration and necrosis in cardiac muscles.
    • Kidney damage including atrophic glomeruli and dilated renal tubules.
    • Marked reduction in white pulp of the spleen, suggestive of immune tissue alteration.
    • Degenerative changes in hepatocytes with nuclear abnormalities (loss of chromatin).

Conclusions and Recommendations

• Evaluating multiple diagnostic tests is crucial to identify the most accurate method for detecting equine Hemomycoplasmosis effectively.
• Each diagnostic test has specific advantages and limitations, and the choice of method should be informed by the context of use.
• Cytological diagnosis is useful and recommended for field screening due to its relative simplicity and cost-effectiveness.
• Quantitative PCR (qPCR) provides confirmatory diagnosis with high sensitivity, making it valuable for precise diagnosis where laboratory facilities are available.
• A combined approach using cytology for initial screening followed by qPCR for confirmation can optimize diagnostic accuracy in managing this disease.