title: First in vitro isolation of Besnoitia besnoiti from chronically infected cattle in Germany.
abstract: Besnoitia besnoiti was in vitro isolated during the first recorded outbreak of bovine besnoitiosis in Germany. Molecular characterization of the new isolate, named Bb-GER1, revealed almost 100% identity with other B. besnoiti isolates obtained in Portugal, Spain, Israel or South Africa, when partial sequences of the 18S ribosomal RNA gene, of the internal transcribed spacer 1 and of the 5.8S RNA gene were compared. Cystozoites obtained from skin tissue of one bull were infectious for gamma-interferon knockout (GKO) mice by intraperitoneal (ip) inoculation. Tachyzoites were detected in the peritoneal cavity, spleen, liver and lung of the mice 5 days post-infection. The parasite could be maintained in GKO mice by ip inoculation for at least 5 passages. Peritoneal washings containing tachyzoites were obtained from infected mice and used to infect five cell lines (Vero, MARC-145, NA42/13, BHK(21), KH-R). The best growth of tachyzoites was observed in BHK(21) cells, but replication occurred to a smaller extent also in MARC-145, NA42/13 and KH-R cells. Subsequent comparative analyses revealed that after direct infection of these cell lines with cystozoites derived from bovine skin, the growth was best in NA42/13 cells. Considerable replication was also observed in the BHK(21) and KH-R cell lines. Our observations on the growth characteristics of Bb-GER1 partially contrast those for other isolates. The preferential growth in particular cell lines may be characteristic for particular B. besnoiti isolates. A potential association between growth properties and differences in virulence remains to be established. This is the first in vitro isolation of B. besnoiti from cattle in Germany.
authors.affiliation: Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Epidemiology, D-16868 Wusterhausen, Germany. gereon.schares@fli.bund.de
authors.forename: G
authors.initials: G
authors.lastname: Schares
authors.forename: W
authors.initials: W
authors.lastname: Basso
authors.forename: M
authors.initials: M
authors.lastname: Majzoub
authors.forename: H C E
authors.initials: HC
authors.lastname: Cortes
authors.forename: A
authors.initials: A
authors.lastname: Rostaher
authors.forename: J
authors.initials: J
authors.lastname: Selmair
authors.forename: W
authors.initials: W
authors.lastname: Hermanns
authors.forename: F J
authors.initials: FJ
authors.lastname: Conraths
authors.forename: N S
authors.initials: NS
authors.lastname: Gollnick
mesh_terms: D000818:Animals; D002417:Cattle; D002418:Cattle Diseases / Q000453:epidemiology / Q000469:parasitology*; D002460:Cell Line; D003048:Coccidiosis / Q000453:epidemiology / Q000469:parasitology / Q000662:veterinary*; D017931:DNA Primers; D016054:DNA, Protozoan / Q000235:genetics*; D021903:DNA, Ribosomal Spacer; D004196:Disease Outbreaks / Q000662:veterinary; D005260:Female; D005858:Germany / Q000453:epidemiology; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D010802:Phylogeny; D016133:Polymerase Chain Reaction / Q000662:veterinary; D012337:RNA, Ribosomal, 18S / Q000235:genetics; D012340:RNA, Ribosomal, 5.8S / Q000235:genetics; D021745:Sarcocystidae / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D017422:Sequence Analysis, DNA; D012867:Skin / Q000469:parasitology
chemical_list: D017931:DNA Primers; D016054:DNA, Protozoan; D021903:DNA, Ribosomal Spacer; D012337:RNA, Ribosomal, 18S; D012340:RNA, Ribosomal, 5.8S