ISOLATION AND AMPLIFICATION OF Anaplasma marginale IN CHICKEN EMBRYO FIBROBLAST CELL CULTURE

Resumo

 Anaplasma marginale (A. marginale) is a worldwide pathogen that infects a variety of
ruminants, but mostly cattle. The present study aimed to describe an isolation technique for A. marginale, using chicken embryo Þ broblast (CEF) cell culture. Blood and tick samples were collected from 5 calves from 2 to 3 months old, which were considered to be infected with A.marginale due to anemia, jaundiced mucous membranes, and prostration. DNA extraction
and PCR were performed for diagnosis using blood and tick samples. All tick and blood samples tested positive in PCR. Additionally,
ticks were crushed with the aid of a blender for inoculation in CEF cell culture. After inoculation, the cultures were kept at 37ºC and 5%
CO2 for 15 days. The cell supernatant of cell cultures was again analyzed using PCR and Wright stain method to conÞ rm A. marginale
isolation. Cell cultures tested positive in PCR, and the presence of the agent was demonstrated by Wright stain. Therefore, by using CEF
cell culture it was possible to isolate and amplify the A. marginale in a concentration of 1.3
x 107.2 bodies per mL. The CEF cells are undemanding and easy to preserve; they are an
option for isolation and production of A. marginale under laboratory conditions.