An improved method for quantifying hematozoa by digital microscopy

Intensity of hematozoan infection is infrequently quantified because accurate calculations require visual counts of parasites relative to a large number of erythrocytes. Manual quantification of erythrocytes can be circumvented by using ImageJ software (developed by the National Institutes of Health) to count erythrocyte nuclei from digital images. Here we use the ratio of microscope erythrocyte counts to digital image erythrocyte counts (field:image ratio) to extrapolate erythrocyte counts from smaller digital images to the microscope's larger field of view. Field:image ratios were consistently calculated from 10 slides (resampling P = 0.049) and used to rapidly estimate intensity of infection within 50,000 or more erythrocytes. Intensity of hematozoan infection calculated from manual quantification of 2,000 erythrocytes was significantly lower (0.46 times) than intensity calculated from digital quantification of 50,000 erythrocytes (bootstrap P = 0.02). We contend that digital quantification of hematozoan infection offers a rapid and precise method to quantify infections of low to moderate intensity.