Development of a diagnostic test for Yersinia pestis by the polymerase chain reaction.
A 501 bp caf1 gene fragment and a 443 bp of pla gene fragment carried by 100 kb (pFra) and 10 kb (pPst) species-specific extrachromosomal replicons, respectively, were used as targets to study the conditions under which DNA amplification by polymerase chain reaction (PCR) may be applied to detect and identify Yersinia pestis DNA in cell lysates of pure cultures and biological samples. The sensitivity limit of PCR with the crude cell lysates of Y. pestis EV was estimated as 10-50 cfu in reaction mixture. When target Y. pestis EV cells were mixed with fresh blood of white mice, which contained 0.4% potassium citrate, the PCR detection level varied from 400 to 100 cfu ml-1 of blood depending on the method used for preparing the sample. In our tests PCR was effective for the detection of yersinia in the blood of white laboratory mice experimentally infected with virulent Y. pestis KM638 strain. This method can be considered convenient for routine detection and identification of Y. pestis.
Norkina
O V
OV
Antiplague Research Institute Microbe, Saratov, Russia.
Kulichenko
A N
AN
Gintsburg
A L
AL
Tuchkov
I V
IV
Popov YuA
Aksenov
M U
MU
Drosdov
I G
IG
D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D017931:DNA Primers; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification*; D051379:Mice; D008969:Molecular Sequence Data; D010930:Plague / Q000097:blood; D010960:Plasminogen Activators / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D015010:Yersinia pestis / Q000235:genetics / Q000302:isolation & purification*
D001426:Bacterial Proteins; D017931:DNA Primers; D004269:DNA, Bacterial; C086725:caf1 protein, Yersinia pestis; C078140:Pla protease, Yersinia pestis; D010960:Plasminogen Activators