ARS | Publication request: In vitro cultivation of a newly recognized Babesia sp. in dogs in North Carolina

Submitted to: Veterinary Parasitology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: October 30, 2007
Publication Date: February 14, 2008
Citation: Lehtinen, L.E., Birkenheuer, A.J., Droleskey, R.E., Holman, P.J. 2008. In vitro cultivation of a newly recognized Babesia sp. in dogs in North Carolina. Veterinary Parasitology. 151:150-157.

Interpretive Summary: Babesia are a type of parasite that live inside red blood cells. They are transmitted to people and animals by the bite of an infected tick. Most of these parasites are found only in animal red blood cells although several new species, or kinds of the parasite, have been found that also infect humans. In fact, the number of these new kinds of Babesia, both in humans and in animals, has been increasing over the past few years. These reports suggest that there are undetected carriers, or natural hosts of these parasites out in the environment. In this report we describe the growth of a new kind of Babesia found in dogs which, although not determined at this time, most probably has a wildlife animal as its natural host. This report illustrates the increasing threat of infection by these parasites to both domesticated animals and man. The ability to grow these parasites under laboratory conditions, as noted here, will aid in developing strategies for detecting them more rapidly and for identifying their natural host.

Technical Abstract: A novel large Babesia sp. from an infected dog was cultivated in vitro by microaerophilous stationary phase culture methodology. uRPMI-1640 medium supplemented with 40% canine serum, Albumax, hypoxanthine and thymidine with antibiotics and buffered with HEPES supported the primary culture of the parasite in a 2% oxygen environment. Subsequent subcultures into HL-1 medium with 20% fetal bovine serum, Albumax, and HB101 supplement with or without hypoxanthine and thymidine also supported parasite propagation. Cultures were successfully introduced to 5% carbon dioxide in air atmosphere at passage 4. To date, the parasites have been continuously cultured through 19 passages, although the parasitemias are low, ranging from 0.2-0.3%. Parasites cultured in RPMI with canine serum were cryopreserved and successfully recovered from liquid nitrogen storage. The small subunit ribosomal rRNA gene sequence was identical in blood-derived and culture-derived parasites, differing in a single base position from the previously reported sequence for this Babesia sp. The fine structure of the parasite was consistent with that of other large Babesia spp., except that the spherical body contained numerous round particles unlike the inclusions previously described in Babesia spp.