title: A simplified PCR assay for fast and easy mycoplasma mastitis screening in dairy cattle.
abstract: A simplified polymerase chain reaction (PCR) assay was developed for fast and easy screening of mycoplasma mastitis in dairy cattle. Species of major mycoplasma strains [Mycoplasma (M.) bovis, M. arginini, M. bovigenitalium, M. californicum, M. bovirhinis, M. alkalescens and M. canadense] in cultured milk samples were detected by this simplified PCR-based method as well as a standard PCR technique. The minimum concentration limit for detecting mycoplasma by the simplified PCR was estimated to be about 2.5 × 10(3) cfu/mL and was similar to that of the standard PCR. We compared the specificity and sensitivity of the simplified PCR to those of a culture method. Out of 1,685 milk samples cultured in mycoplasma broth, the simplified PCR detected Mycoplasma DNA in 152 that were also positive according to the culture assay. The sensitivity and specificity of the simplified PCR were 98.7% and 99.7%, respectively, for detecting mycoplasma in those cultures. The results obtained by the simplified PCR were consistent with ones from standard PCR. This newly developed simplified PCR, which does not require DNA purification, can analyze about 300 cultured samples within 3 h. The results from our study suggest that the simplified PCR can be used for mycoplasma mastitis screening in large-scale dairy farms.
authors.affiliation: Animal Health Laboratory, Faculty of Veterinary Medicine, Rakuno Gakuen University, Hokkaido 069-8501, Japan. higuchi@rakuno.ac.jp
authors.forename: Hidetoshi
authors.initials: H
authors.lastname: Higuchi
authors.forename: Hidetomo
authors.initials: H
authors.lastname: Iwano
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Kawai
authors.forename: Takehiro
authors.initials: T
authors.lastname: Ohta
authors.forename: Tetsu
authors.initials: T
authors.lastname: Obayashi
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Hirose
authors.forename: Nobuhiko
authors.initials: N
authors.lastname: Ito
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Yokota
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Tamura
authors.forename: Hajime
authors.initials: H
authors.lastname: Nagahata
mesh_terms: D000818:Animals; D002417:Cattle; D015169:Colony Count, Microbial / Q000662:veterinary; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004196:Disease Outbreaks / Q000517:prevention & control / Q000662:veterinary; D005260:Female; D008414:Mastitis, Bovine / Q000175:diagnosis / Q000382:microbiology*; D008892:Milk / Q000166:cytology / Q000382:microbiology*; D009174:Mycoplasma / Q000235:genetics / Q000302:isolation & purification*; D009175:Mycoplasma Infections / Q000175:diagnosis / Q000382:microbiology / Q000662:veterinary*; D016133:Polymerase Chain Reaction / Q000662:veterinary
chemical_list: D004269:DNA, Bacterial