Development of a Dry Medium for Isolation of Histomonas meleagridis in the Field

Abstract

Blackhead disease is caused by Histomonas meleagridis, an anaerobic protozoan parasite, and results in mortality rates of up to 100% in turkeys and 30% in chickens. Outbreaks of blackhead disease are unpredictable, and the harvesting of H. meleagridis strains from the field would be a great resource for researchers to study its epidemiology. Therefore, the objective of this study was to develop a dry medium that would allow storage at ambient temperatures until needed. Fifty milliliters of horse serum was dried and then mixed with dry medium M199 with Hanks balanced salts (10.6 g), sodium bicarbonate (0.35 g), and rice powder (0.8 g). To test the ability of reconstituted medium to support growth of H. meleagridis, groups of 10 flasks containing 0.2 g of dry medium were stored for 24 hr at 25 and 60 C before testing. Other groups of flasks containing dry medium were stored at 25, 37, and 42 C for 1, 3, or 6 mo. At each test period, the flasks were reconstituted with 10 ml of water, inoculated with 100 000 H. meleagridis cells, and incubated at 40 C for 48 hr. Fresh liquid medium was used as a control. There were no differences in cell counts in medium stored at 25 or 60 C for 24 hr. After 1 mo, cell counts in reconstituted medium were about half that of fresh liquid medium after 48 hr of incubation. But after 3 and 6 mo, the cell counts were not significantly different in all groups (P < 0.05) after 72 hr of incubation. These results show that dried Dwyer medium can be stored at ambient temperatures for extended times and would be an effective tool for obtaining isolates of H. meleagridis from the field.