Viability Assessment of Cryptosporidium parvum Oocysts by Vital Dyes: Dry Mounts Overestimate the Number of “Ghost” Oocysts

Viability assessment of Cryptosporidium parvum oocysts is crucial for evaluation of the public health significance of this important zoonotic protozoon. Viability is commonly assessed in wet mounts after acid pretreatmentand staining with fluorogenic vital dyes. However, in some studies, oocyst viability is evaluated in dry mounts after staining in suspension.

Here, we evaluate the effect of acid pretreatment in nine replicate samples and compare the assessment of oocyst viability after evaluation in wet and dry mounts, respectively. Although acid pretreatment had no significant effect on the viability scores, data obtained by scoring oocysts in drymounts resulted in∼25% underestimation of the proportion of viable oocyst (82.5% –+ 0.9% [wet mount + acid],57.7% –+ 2.3% [dry mount - Oacid], 76.0% –+ 1.7% [wet mount, - Oacid]), while the proportions of nonviable oocysts (DAPI+/PI+) were comparable for wet and dry mounts (9.7% –+ 0.4% [wet mount +acid], 12.1 –+ 1.5%[dry mount, - acid], 15.5% –+ 1.1% [wet mount, - acid]).