Isolation of anAnaplasmasp. Organism from White-Tailed Deer by Tick Cell Culture

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Abstract
We used tick cell culture to isolate a bacterium previously referred to as the “white-tailed deer (WTD) agent” from two captive fawns inoculated with blood from wild WTD (Odocoileus virginianus). Buffy coat cells were added to ISE6 tick cell cultures and incubated at 34°C, and 8 days later,Anaplasma-like inclusions were demonstrated in Giemsa-stained culture samples. The microbes became established and could be continuously passaged in tick cells. The identity of a culture isolate designated WTD76 was verified as the WTD agent by using specific PCR primers and by DNA sequencing. Comparison with sequences available in GenBank indicated that the isolate was most closely related first toAnaplasma platysand second toAnaplasma phagocytophilum, supporting its placement in the genusAnaplasma. Transmission electron microscopy of thisAnaplasmasp. organism in tick cell cultures revealed large inclusions filled with pleomorphic and rod-shaped bacteria. Tick cells infected with theAnaplasmasp. organism were used to successfully infect a naive deer, thereby proving the infectivity of the isolate for deer.