title: Multiplex PCR for molecular screening of Borrelia burgdorferi sensu lato, Anaplasma spp. and Babesia spp.
abstract: BACKGROUND
  Ticks transmit a great variety of pathogenic microorganisms to humans and animals. The detection of tick-borne pathogens (TBP) is mainly by molecular techniques based on polymerase chain reactions (PCR).
  
  
  OBJECTIVE
  To design and evaluate a multiplex PCR for the molecular screening of zoonotic TBP for exploratory studies.
  
  
  METHODS
  Control DNA from reference strains, DNA from experimentally-infected biological specimens, and from Rhipicephalus sanguineus ticks collected from domestic and homeless dogs were used. A multiplex PCR assay to detect the presence of Borrelia burgdorferi sensu lato, Anaplasma spp. and Babesia spp. was designed and optimized using primers previously reported for B. burgdorferi sensu lato and Anaplasma spp., while for Babesia spp. they were designed in silico. The multiplex PCR was evaluated on the DNA from biological samples.
  
  
  RESULTS
  A new set of specific primers for Babesia spp. was designed. Adjustment of the master mix reactive concentrations and amplification conditions for the multiplex PCR allowed the successful amplification of the specific amplicons for each microbial group from the control DNA and experimentally-infected biological specimens. The efficiency of the multiplex PCR amplifying three DNA targets was confirmed. Individual and co-infection of Anaplasma spp. and Babesia spp. were detected in the R. sanguineus ticks from dogs.
  
  
  CONCLUSIONS
  A multiplex PCR assay for the screening of three TBP is available. By using it, B. burgdorferi sensu lato, Anaplasma spp. and Babesia spp. can be detected accurately in one PCR reaction.
authors.affiliation: 'Pedro Kourí', Institute of Tropical Medicine, National Reference Laboratory of Spirochetes, Havana, Cuba.
authors.forename: Islay
authors.initials: I
authors.lastname: Rodríguez
authors.affiliation: Institute of Biology, Laboratory of Eco-Epidemiology of Parasites, University of Neuchâtel, Neuchâtel, Switzerland.
authors.forename: Caroline
authors.initials: C
authors.lastname: Burri
authors.affiliation: 'Pedro Kourí', Institute of Tropical Medicine, National Reference Laboratory of Spirochetes, Havana, Cuba.
authors.forename: Angel A
authors.initials: AA
authors.lastname: Noda
authors.affiliation: Institute of Biology, Laboratory of Eco-Epidemiology of Parasites, University of Neuchâtel, Neuchâtel, Switzerland.
authors.forename: Véronique
authors.initials: V
authors.lastname: Douet
authors.affiliation: Institute of Biology, Laboratory of Eco-Epidemiology of Parasites, University of Neuchâtel, Neuchâtel, Switzerland.
authors.forename: Lise
authors.initials: L
authors.lastname: Gern
mesh_terms: D000709:Anaplasma / Q000145:classification / Q000235:genetics / Q000302:isolation & purification; D000712:Anaplasmosis / Q000175:diagnosis* / Q000382:microbiology; D000818:Animals; D001403:Babesia / Q000145:classification / Q000235:genetics / Q000302:isolation & purification; D001404:Babesiosis / Q000175:diagnosis* / Q000469:parasitology; D015748:Borrelia burgdorferi Group / Q000145:classification / Q000235:genetics / Q000302:isolation & purification; D004283:Dog Diseases / Q000175:diagnosis* / Q000382:microbiology / Q000469:parasitology; D004285:Dogs; D005260:Female; D008193:Lyme Disease / Q000175:diagnosis / Q000382:microbiology / Q000662:veterinary*; D008297:Male; D060885:Multiplex Polymerase Chain Reaction / Q000662:veterinary*; D009758:Nymph / Q000382:microbiology / Q000469:parasitology / Q000502:physiology; D048508:Rhipicephalus sanguineus / Q000254:growth & development / Q000382:microbiology / Q000469:parasitology / Q000502:physiology; D013984:Tick Infestations / Q000469:parasitology / Q000662:veterinary*; D017282:Tick-Borne Diseases / Q000382:microbiology / Q000469:parasitology