Analysis of 16S rRNA and 51-kilodalton antigen gene and transmission in mice of Ehrlichia risticii in virgulate trematodes from Elimia livescens snails in Ohio

Operculate snails (the family Pleuroceridae: Elimia livescens) were collect ed between June and October 1998 from a river in central Ohio where repeate d cases of Potomac horse fever (PHF) have occurred. Of collected snails, co nsistently 50 to 80% carried a combination of cercariae and sporocysts of d igenetic virgulate trematodes. The trematodes obtained from each snail were pooled and tested for Ehrlichia risticii, the agent of PHF, by nested PCR using primers specific to the 16S rRNA gene. Out of a total of 209 trematod e pools, 50 pools were found to be positive by PCR. The DNA sequence of the 16S rRNA gene identified in one trematode pool was identical to that of th e type strain of E. risticii, and the sequence of the gene identified in an other pool differed from that of the type strain by 1 nucleotide. Compariso n of the deduced amino acid sequence of the partial 51-kDa antigen gene fro m various sources revealed that Maryland, Ohio (except Ohio 081), and Kentu cky strains are in a cluster distinct from the sequences obtained from sour ces in California and Oregon. Ohio 081 was shown previously by antigenic co mposition analysis to be distinct from other groups. However, all sequences examined were not segregated according to their sources: horse blood or in fected trematodes. E. risticii was found to be transmittable from trematode s to mice and was subsequently passaged from infected mice to additional mi ce, as determined by PCR analysis. Our findings suggest the evolution off. risticii in the natural reservoir in separate geographic regions and persis tent infection of trematode populations with E. risticii during summer and early fall. The study also suggests that the mouse can be used to isolate E . risticii from the infected trematode.