title: Isolation and propagation of the Ap-Variant 1 strain of Anaplasma phagocytophilum in a tick cell line.
abstract: The first tissue culture isolates of the unique Anaplasma phagocytophilum strain, Ap-Variant 1, were obtained in the Ixodes scapularis tick-derived cell line ISE6. Two isolates were from goat blood samples: one from a goat infected with I. scapularis ticks from Rhode Island and a second from a goat infected by serial passage of blood from the first infected goat. Eight isolates were made directly from I. scapularis ticks collected from white-tailed deer in Minnesota and represent the first isolations of an Anaplasma species directly from ticks. Each of the 10 isolates had a 16S rRNA gene sequence identical to that previously described for Ap-Variant 1, but differences within the ank gene were found that suggest natural variation. Prevalence of Anaplasma in the Minnesota ticks was 63.9%; 23 of 36 ticks tested by PCR were positive. Six of the tick-derived isolates were obtained from a set of 18 PCR-positive ticks, for a 33.3% isolation success rate. The conservation of host tropism among the Rhode Island and Minnesota isolates of Ap-Variant 1 was examined by use of experimental infections of mice and a goat. A Minnesota tick-derived isolate (MN-61-2) was used to inoculate naïve animals, and this isolate was able to infect a goat but unable to infect each of five mice, confirming that the Minnesota isolates have the same host tropism as Ap-Variant 1 from the northeastern United States. Light and electron microscopy of the Ap-Variant 1 isolate MN-61-2 in ISE6 cells showed cytoplasmic inclusions characteristic of A. phagocytophilum with pleomorphic bacteria in membrane-bound vacuoles and both electron-dense and electron-lucent forms.
authors.affiliation: Viral and Rickettsial Zoonoses Branch, Centers for Disease Control and Prevention, 1600 Clifton Rd., MS G-13, Atlanta, GA 30333, USA. rfm2@cdc.gov
authors.forename: Robert F
authors.initials: RF
authors.lastname: Massung
authors.forename: Michael L
authors.initials: ML
authors.lastname: Levin
authors.forename: Ulrike G
authors.initials: UG
authors.lastname: Munderloh
authors.forename: David J
authors.initials: DJ
authors.lastname: Silverman
authors.forename: Meghan J
authors.initials: MJ
authors.lastname: Lynch
authors.forename: Jariyanart K
authors.initials: JK
authors.lastname: Gaywee
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Kurtti
mesh_terms: D041081:Anaplasma phagocytophilum / Q000145:classification* / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure; D000818:Animals; D002460:Cell Line; D016873:Ehrlichiosis / Q000382:microbiology / Q000662:veterinary; D005260:Female; D015511:Goat Diseases / Q000382:microbiology; D006041:Goats; D051379:Mice; D010802:Phylogeny; D012336:RNA, Ribosomal, 16S; D013987:Ticks / Q000166:cytology* / Q000382:microbiology*
chemical_list: D012336:RNA, Ribosomal, 16S