Drug Resistance in Eimeria tenella. VII. Acriflavine-Mediated Loss of Resistance to Amprolium

An amprolium-resistant strain of Eimeria tenella was propagated serially in chickens fed mash containing 0.05% acriflavine hydrochloride. Increased sensitivity to amprolium was evident after the first passage. After six passages in the presence of acriflavine, the response of the experimental strain was approximately the same as that of an amprolium-sensitive control strain. Exposure to acriflavine, in vitro or in vivo, induces dyskinetoplasy in a number of genera of the order Kinetoplastida. The literature on this phenomenon has been summarized by Trager and Rudzinska (1964) and by Stuart and Hanson (1967). Mukherjea and Ray (1956) found cytochemical aberrations including alteration in nuclear DNA in Tritrichomonas foetus that had been cultured in the presence of acriflavine. Moreover, Mitsuhashi, Harada, and Kameda (1961) reported that transmissible (F factor) drug resistance could be eliminated in vitro by culturing multiple resistant strains of Shigella flexneri and Escherichia coli in medium containing acriflavine. The cells became sensitive to all drugs to which they previously had been resistant. The present report describes the response of an amprolium-resistant strain of Eimeria tenella to propagation in chickens fed mash containing acriflavine. MATERIALS AND METHODS Three-week-old chicks were used. They were grouped by weight as suggested by Gardiner and Wehr (1950) into lots of 10, and each lot was started on the appropriate mash 24 hr before the infected groups were inoculated with approximately 100,000 sporulated Eimeria tenella oocysts. In the initial passage, groups of birds fed unmedicated mash, mash containing 0.0125% amprolium, or mash containing 0.05% acriflavine hydrochloride were given oocysts from an amproliumsensitive strain. In subsequent passages of this strain, oocysts recovered from the unmedicated group were used to inoculate the three groups of the succeeding passage. Similar groups of birds initially were given oocysts of a strain that had been serially passed through 64 groups of birds fed mash containing amprolium. In subsequent passages, oocysts recovered from the unmedicated and from the acriflavine-medicated birds were used to inoculate the respective groups of the next Received for publication 22 March 1968. passage. This protocol is presented as a flow sheet in Figure 1. An uninfected, unmedicated group was carried at each passage to allow comparison of weight gains. On the 8th day after inoculation, the surviving birds were weighed and necropsied. The ceca were removed and cecal lesions evaluated. Oocyst cultures were made from the ceca and cecal contents of the respective infected groups, and the number of oocysts produced per surviving bird was estimated for each group. Weight gains, oocyst production, mortality rates, and cecal lesion scores obtained at each passage were used to calculate the Anticoccidial Indices for the respective groups as suggested in the Merck Bulletin on Amprolium (1960). These Indices form the basis for the comparisons made.