title: Development of loop-mediated isothermal amplification (LAMP) assays for rapid detection of Ehrlichia ruminantium.
abstract: BACKGROUND
  The rickettsial bacterium Ehrlichia ruminantium is the causative agent of heartwater, a potential zoonotic disease of ruminants transmitted by ticks of the genus Amblyomma. The disease is distributed in nearly all of sub-Saharan Africa and some islands of the Caribbean, from where it threatens the American mainland. This report describes the development of two different loop-mediated isothermal amplification (LAMP) assays for sensitive and specific detection of E. ruminantium.
  
  
  RESULTS
  Two sets of LAMP primers were designed from the pCS20 and sodB genes. The detection limits for each assay were 10 copies for pCS20 and 5 copies for sodB, which is at least 10 times higher than that of the conventional pCS20 PCR assay. DNA amplification was completed within 60 min. The assays detected 16 different isolates of E. ruminantium from geographically distinct countries as well as two attenuated vaccine isolates. No cross-reaction was observed with genetically related Rickettsiales, including zoonotic Ehrlichia species from the USA. LAMP detected more positive samples than conventional PCR but less than real-time PCR, when tested with field samples collected in sub-Saharan countries.
  
  
  CONCLUSIONS
  Due to its simplicity and specificity, LAMP has the potential for use in resource-poor settings and also for active screening of E. ruminantium in both heartwater-endemic areas and regions that are at risk of contracting the disease.
authors.affiliation: Department of Collaboration and Education, Research Center for Zoonosis Control, Hokkaido University, Kita 20, Nishi 10, Kita-ku, Sapporo, Hokkaido 001-0020, Japan.
authors.forename: Ryo
authors.initials: R
authors.lastname: Nakao
authors.forename: Ellen Y
authors.initials: EY
authors.lastname: Stromdahl
authors.forename: Joseph W
authors.initials: JW
authors.lastname: Magona
authors.forename: Bonto
authors.initials: B
authors.lastname: Faburay
authors.forename: Boniface
authors.initials: B
authors.lastname: Namangala
authors.forename: Imna
authors.initials: I
authors.lastname: Malele
authors.forename: Noboru
authors.initials: N
authors.lastname: Inoue
authors.forename: Dirk
authors.initials: D
authors.lastname: Geysen
authors.forename: Kiichi
authors.initials: K
authors.lastname: Kajino
authors.forename: Frans
authors.initials: F
authors.lastname: Jongejan
authors.forename: Chihiro
authors.initials: C
authors.lastname: Sugimoto
mesh_terms: D000818:Animals; D001096:Arachnid Vectors / Q000382:microbiology; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D002417:Cattle; D002418:Cattle Diseases / Q000382:microbiology*; D017931:DNA Primers / Q000235:genetics; D016995:Ehrlichia ruminantium / Q000235:genetics / Q000302:isolation & purification*; D005260:Female; D006357:Heartwater Disease / Q000382:microbiology*; D008297:Male; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D013987:Ticks / Q000382:microbiology
chemical_list: D001426:Bacterial Proteins; D017931:DNA Primers