Ethical considerations

All experimental procedures were approved by the Institutional Animal Care and Use Committee (Protocol no. 15-017), United States Department of Agriculture, Beltsville, Maryland. Outbred SW and KO mice were purchased from the Jackson Laboratory (Bar Harbor, ME, USA) in compliance with the Institutional Animal Ethics Committee guidelines. The WTD were euthanized in the field, often in remote locations and tissues were shipped overnight by the collector with ice packs. Consequently, by the time tissues were received at the USDA laboratory, they often were contaminated with bacteria and not suited for cell culture to isolate T. gondii. The number of T. gondii in tissues of naturally infected large animal tissues is low (estimated one tissue cyst per 50–100 g) and the probability of isolation of T. gondii is very low unless large numbers of mice (10 or more) are used (Dubey et al., 1995; Dubey, 2010). To balance the possibility of isolating parasites and using the minimum number of mice, we decided to use 3–5 mice for the bioassay of each deer sample in the current study.

All mice used in the current study were treated humanely and examined twice daily for any signs of illness. A veterinarian was assigned exclusively to the toxoplasmosis project. Any sick mice were euthanized because our objective was isolation of T. gondii and not testing for mortality. We wanted to collect mouse tissues aseptically for cultivation in cell culture or subpassage to other mice.