Detection of Burkholderia pseudomallei in blood samples using polymerase chain reaction.
A highly sensitive, specific, rapid and simple method to detect Burkholderia pseudomallei in blood samples was developed. Two 22-base oligonucleotide primers, based on sequences from a specific DNA probe, were used for amplification of bacterial DNA by the polymerase chain reaction (PCR). Amplification with these primers yielded a 178-base pair product in 100 clinical isolates of B. pseudomallei. As little as 0.5 fg of B. pseudomallei DNA was detectable by this method. Experiments involving inoculation of the organism into uninfected blood samples showed that the method could be used to detect as few as 1 bacterial cell ml-1 of whole blood. Non-specific amplification of other bacterial DNAs from 18 samples of bacteria was not observed. Blood samples from seven patients proven to have melioidosis by haemoculture were positive using these primers. The total time required for sample processing, amplification and visualization was approximately 3.5 h. The high sensitivity, rapidity and simplicity of this method should make it valuable for diagnosis, monitoring of drug treatment and for epidemiological studies of the melioidosis.
Rattanathongkom
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A
Department of Oral Biology, Faculty of Dentistry, Khon Kaen University, Thailand.
Sermswan
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RW
Wongratanacheewin
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S
D016470:Bacteremia / Q000382:microbiology*; D016957:Burkholderia pseudomallei / Q000235:genetics / Q000302:isolation & purification*; D017931:DNA Primers; D015342:DNA Probes; D004269:DNA, Bacterial / Q000097:blood* / Q000235:genetics; D006801:Humans; D008554:Melioidosis / Q000097:blood / Q000175:diagnosis*; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA
D017931:DNA Primers; D015342:DNA Probes; D004269:DNA, Bacterial