Effects of temperature and aging on viability and infectivity of laboratory-produced actinosporean triactinomyxon spores (infective stage of the organism causing whirling disease) were studied. In vitro staining of triactinomyxon spores with vital fluorescein diacetate correlated with the ability of the spores to infect fry of rainbow trout Oncorhynchus mykiss. The experimentally produced actinosporean stage of Myxobolus cerebralis was short-lived, persisting for only 3–4 d at 12.5°C and for less time at warmer temperatures. The vital staining method has potential for screening therapeutants intended to control myxosporean infection of fish.