Nucleic acid amplification for rapid detection of Rhodococcus equi in equine blood and tracheal wash fluids.
OBJECTIVE
To evaluate the ability of nucleic acid amplification techniques to detect Rhodococcus equi in equine buffy coat, blood, and tracheal wash fluid and to differentiate between virulent and avirulent strains of the bacteria.


METHODS
Blood anticoagulated with EDTA and tracheal wash fluid from healthy horses.


METHODS
Logarithmic dilutions of virulent and avirulent strains of R equi were added to equine buffy coat and tracheal wash fluid samples. The DNA was extracted and amplified by polymerase chain reaction (PCR), using primers specific for the 16S ribosomal subunit gene and the virulence plasmid of R equi.


RESULTS
PCR with 16S ribosomal subunit primers amplified a 441-bp segment of DNA from virulent and avirulent strains of R equi, but not from samples containing other species of bacteria. The virulence plasmid primers amplified an 875-bp segment of DNA from virulent strains of R equi, but not from avirulent R equi, or from other species of bacteria. Virulent strains of R equi could be identified by PCR and differentiated from avirulent strains within 12 to 24 hours after sample collection, with as few as 10 to 100 organisms present.


CONCLUSIONS
PCR can be used to rapidly and accurately identify R equi in equine blood and tracheal wash fluid samples and can differentiate between virulent and avirulent strains of the organism.


CONCLUSIONS
Because PCR can confirm a diagnosis of R equi infection in horses more rapidly and specifically than use of standard culture techniques, extrapolation of this assay to soil and fecal samples could be useful in epidemiologic studies and studies of environmental disinfection or decontamination.
Sellon
D C
DC
Department of Food Animal and Equine Medicine, College of Veterinary Medicine, North Carolina State University, Raleigh 27606, USA.
Walker
K
K
Suyemoto
M
M
Altier
C
C
D000193:Actinomycetales Infections / Q000097:blood / Q000175:diagnosis / Q000662:veterinary*; D000818:Animals; D016470:Bacteremia / Q000175:diagnosis / Q000382:microbiology / Q000662:veterinary*; D001483:Base Sequence; D001992:Bronchoalveolar Lavage Fluid / Q000382:microbiology; D017931:DNA Primers / Q000032:analysis / Q000737:chemistry / Q000235:genetics; D004269:DNA, Bacterial / Q000032:analysis / Q000737:chemistry / Q000235:genetics*; D006734:Horse Diseases / Q000097:blood / Q000175:diagnosis* / Q000453:epidemiology; D006736:Horses; D015994:Incidence; D021141:Nucleic Acid Amplification Techniques; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary; D016929:Rhodococcus equi / Q000235:genetics / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D014132:Trachea / Q000378:metabolism / Q000382:microbiology*
D017931:DNA Primers; D004269:DNA, Bacterial