A multiplex-PCR for the diagnosis of contagious agalactia of sheep and goats

A multiplex polymerase chain reaction (m-PCR) assay was optimized for the simultaneous detection of several species of small ruminant mycoplasmas. Two sets of oligonucleotide primers specific for Mycoplasma agalactiae (Ma) and Mycoplasma 'mycoides' cluster (M.m. cluster) were used in the test. The m-PCR was able to amplify a 375-bp fragment of Ma chromosomal DNA and a 257-260-bp fragment of M.m. cluster chromosomal DNA. Four reference strains (M. agalactiae, M. mycoides subsp. mycoides LC, M. capricolum subsp. capricolum, M. mycoides subsp, capri) and 56 samples (44 milk samples, two nasal swabs, six ocular swabs, three vaginal swabs and one sample of fibrinous exudate from carpal joint), from sheep and gears with clinical signs of contagious agalactia (CA), were examined. The m-PCR confirmed the identification of reference strains and allowed to identify, of the 43 positive samples examined, 35 Ma strains, 12 M. 'mycoides' cluster strains; in four samples both Ma and mycoplasmas of M. 'mycoides' cluster were revealed. The m-PCR was able to detect 7 pg of mycoplasma DNA. The specificity and sensitivity of the m-PCR suggest its use for the 'routine' diagnosis of CA. (C) 2001 Academic Press.