Rapid identification of Klebsiella pneumoniae, Corynebacterium kutscheri, and Streptococcus pneumoniae using triplex polymerase chain reaction in rodents.
Klebsiella pneumoniae, Corynebacterium kutscheri, and Streptococcus pneumoniae are important pathogens that cause respiratory infections in laboratory rodents. In this study, we used species-specific triplex PCR analysis to directly detect three common bacterial pathogens associated with respiratory diseases. Specific targets were amplified with conventional PCR using the tyrB gene from K. pneumoniae, gyrB gene from C. kutscheri, and ply gene from S. pneumoniae. Our primers were tested against purified DNA from another eleven murine bacteria to determine primer specificity. Under optimal PCR conditions, the triplex assay simultaneously yielded a 931 bp product from K. pneumoniae, a 540 bp product from C. kutscheri, and a 354 bp product from S. pneumoniae. The triplex assay detection thresholds for pure cultures were 10 pg for K. pneumoniae and S. pneumoniae, and 100 pg for C. kutscheri. All three bacteria were successfully identified in the trachea and lung of experimentally infected mice at the same time. Our triplex PCR method can be used as a useful method for detecting pathogenic bacterial infections in laboratory rodents.
Department of Laboratory Animal Medicine, College of Veterinary Medicine, Konkuk University, 1 Hwayang-dong, Gwangjin-gu, Seoul 143-701, Republic of Korea.
Eui-Suk
ES
Jeong
Kyoung-Sun
KS
Lee
Seung-Ho
SH
Heo
Jin-Hee
JH
Seo
Yang-Kyu
YK
Choi
D000818:Animals; D000830:Animals, Laboratory / Q000382:microbiology*; D003352:Corynebacterium / Q000235:genetics / Q000302:isolation & purification*; D004269:DNA, Bacterial / Q000302:isolation & purification; D007711:Klebsiella pneumoniae / Q000235:genetics / Q000302:isolation & purification*; D008168:Lung / Q000382:microbiology; D051379:Mice; D008807:Mice, Inbred BALB C / Q000382:microbiology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012141:Respiratory Tract Infections / Q000382:microbiology* / Q000662:veterinary*; D012376:Rodent Diseases / Q000382:microbiology*; D013296:Streptococcus pneumoniae / Q000235:genetics / Q000302:isolation & purification*; D014132:Trachea / Q000382:microbiology
D004269:DNA, Bacterial