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Development of a multiplex real-time PCR for contagious agalactia diagnosis in small ruminants
- Source :
- Journal of Microbiological Methods. 90:73-79
- Publication Year :
- 2012
- Publisher :
- Elsevier BV, 2012.
-
Abstract
- Contagious agalactia is an important disease worldwide that affects small ruminants. Clinical manifestations vary from mastitis, pneumonia, arthritis and keratoconjunctivitis to septicemia. Four mycoplasmal etiological agents have been identified: Mycoplasma (M.) agalactiae, M. mycoides subsp. capri, M. capricolum subsp. capricolum and M. putrefaciens. The current procedure for direct diagnosis, recommended by the World Organization for Animal Health, involves the isolation of one or several causative agents from clinical specimens and further time-consuming identification steps. The present study reports the development of a new multiplex real-time PCR (including an internal positive control) that detects all four pathogens simultaneously and distinguishes M. agalactiae from the others. First, intra- and inter-species polymorphisms of the two target house-keeping genes, namely polC and fusA, were analyzed to design primers and probes adapted to the diversity of currently circulating strains. The specificity and the sensitivity of the assay were then challenged and the limit of detection was found to be as low as 6 to 12 copies of the target genes. The assay requires further assessment on clinical specimens but its performances (notably low intra- and inter-assay variability) are already very promising for use in large-scale diagnosis and prophylactic surveys of contagious agalactia.
- Subjects :
- DNA, Bacterial
Veterinary Medicine
Microbiology (medical)
Isolation (health care)
Sheep Diseases
DNA-Directed DNA Polymerase
Disease
Biology
Real-Time Polymerase Chain Reaction
medicine.disease_cause
Sensitivity and Specificity
Microbiology
Mycoplasma
Bacterial Proteins
medicine
Animals
Mycoplasma Infections
Multiplex
Molecular Biology
Keratoconjunctivitis
DNA Primers
Bacteriological Techniques
Goat Diseases
Polymorphism, Genetic
Sheep
Goats
Peptide Elongation Factor G
medicine.disease
Virology
Mastitis
Real-time polymerase chain reaction
Molecular Diagnostic Techniques
Multiplex Polymerase Chain Reaction
Pneumonia (non-human)
Subjects
Details
- ISSN :
- 01677012
- Volume :
- 90
- Database :
- OpenAIRE
- Journal :
- Journal of Microbiological Methods
- Accession number :
- edsair.doi.dedup.....b7b4bbd5f8404d76c2eb4398f42f4ea6