title: Optimization of a species-specific polymerase chain reaction assay for identification of Pentatrichomonas hominis in canine fecal specimens.
abstract: OBJECTIVE
  To determine the optimum reaction conditions and detection limits of PCR assay for identification of Pentatrichomonas hominis in DNA extracted from canine feces.
  
  
  METHODS
  DNA extracted from feces of 4 dogs with diarrhea from which trichomonads were observed, 81 dogs that had feces submitted to a diagnostic laboratory, and 19 dogs residing in a laboratory animal facility.
  
  
  METHODS
  Optimum reaction conditions and absolute and practical detection limits of 2 P hominis 18S species-specific primer pairs were determined by use of an in vitro cultivated canine isolate of P hominis in the presence and absence of canine feces. The optimized PCR assay was applied to amplification of P hominis 18S rRNA genes from DNA extracted from the feces of dogs.
  
  
  RESULTS
  Under optimized conditions, a primer pair was identified as able to detect as few as 1 P hominis organism/180-mg fecal sample. The PCR assay identified P hominis in diarrheic feces of 4 dogs in which trichomonads were seen by light microscopy. The P hominis genes were not amplified from other fecal samples examined.
  
  
  CONCLUSIONS
  Molecular identification of P hominis in feces of 4 dogs with trichomonosis and diarrhea reported here validates the identity of this species in such infections. Sensitive and specific PCR amplification of P hominis 18S rRNA genes from DNA extracted from feces will directly facilitate studies examining pathogenicity of this trichomonad and enable differentiation of P hominis from other known or novel species of trichomonads that may infect the gastrointestinal tract of dogs.
authors.affiliation: Department of Molecular Biomedical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27606, USA.
authors.forename: Jody L
authors.initials: JL
authors.lastname: Gookin
authors.forename: Stephen H
authors.initials: SH
authors.lastname: Stauffer
authors.forename: Maria R
authors.initials: MR
authors.lastname: Coccaro
authors.forename: Miriam J
authors.initials: MJ
authors.lastname: Marcotte
authors.forename: Michael G
authors.initials: MG
authors.lastname: Levy
mesh_terms: D000818:Animals; D016054:DNA, Protozoan / Q000737:chemistry / Q000235:genetics; D003967:Diarrhea / Q000175:diagnosis / Q000469:parasitology*; D004283:Dog Diseases / Q000175:diagnosis / Q000469:parasitology*; D004285:Dogs; D005243:Feces / Q000469:parasitology*; D005260:Female; D008297:Male; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D011529:Protozoan Infections, Animal / Q000175:diagnosis / Q000469:parasitology*; D013045:Species Specificity; D016834:Trichomonadida / Q000235:genetics* / Q000302:isolation & purification*
chemical_list: D016054:DNA, Protozoan