Molecular Detection and Characterization of Ehrlichia ruminantium

Abstract

Ehrlichia ruminantium is an obligate intracellular bacterium that can cause a disease in ruminants known as heartwater. The vertebrate host becomes infected when infected Amblyomma ticks (nymphs or adults) feed on it. When the host survives the infection it becomes a carrier. Ticks become infected with E. ruminantium when they feed on an infected host. The infection is transmitted from one developmental stage to the other except from adult to eggs for as far as is known now. Detection of E. ruminantium is difficult, especially in carrier animals, and is based on either detection of antibodies (MAP1-B ELISA) or detection of bacterial DNA using PCR techniques. The purpose of the research was to molecularly characterize E. ruminantium and to develop new or improved diagnostic tests for the detection of E. ruminantium. For the detection the MAP1-B ELISA was further validated using the TG-ROC method to determine cutoff values with equal sensitivity and specificity, and a new assay based on the reverse line blot (RLB) technique was developed. The developed RLB assays can simultaneously detect and identify 8 Anaplasma and Ehrlichia species. For the characterization several 16S rRNA sequences were determined, which were also used for a reorganization of the taxonomic classification of several rickettsial species. Finally, the transcription of the map1 multigene family was studied in different environments and it was found that map1-1 was transcribed in vitro and in vivo in the vector tick whereas it was not transcribed at a detectable level in vitro in bovine endothelial cells