Rapid differential diagnosis of Mycoplasma agalactiae and Mycoplasma bovis based on a multiplex-PCR and a PCR-RFLP.
The membrane-protein 81 gene (mb-mp81) of Mycoplasma bovis was cloned, sequenced and compared to membrane-protein 81 gene (ma-mp81) of Mycoplasma agalactiae. After alignment of both sequences, specific primers pairs were designed from variable or unchanging nucleotide segments. In this study, we describe the development and optimization of a multiplex-PCR (MPCR) for the rapid detection of M. agalactiae and M. bovis strains. In addition, a simple and rapid PCR-restriction fragment length polymorphism (RFLP) assay, using the restriction enzymes AluI, DraI, RsaI and XbaI, is described to distinguish between both species. The results suggest that MPCR and PCR-RFLP assays could be used as an alternative method in routine diagnosis for rapid and specific simultaneous detection of M. agalactiae and M. bovis.
Foddai
A
A
Istituto Zooprofilattico Sperimentale della Sardegna G.Pegreffi, Via Duca degli Abruzzi 8, 07100 Sassari, Italy.
Idini
G
G
Fusco
M
M
Rosa
N
N
de la Fe
C
C
Zinellu
S
S
Corona
L
L
Tola
S
S
D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics; D003001:Cloning, Molecular; D003937:Diagnosis, Differential; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics; D008970:Molecular Weight; D045644:Mycoplasma agalactiae / Q000235:genetics* / Q000302:isolation & purification*; D045682:Mycoplasma bovis / Q000235:genetics* / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length
D001426:Bacterial Proteins; D008565:Membrane Proteins