Comparison of three DNA extraction methods for detection of Erysipelothrix rhusiopathiae in chicken blood by polymerase chain reaction.
A previously reported Erysipelothrix-specific polymerase chain reaction (PCR) was used to detect Erysipelothrix bacteremia in chickens. The sensitivity of PCR using 3 DNA extraction methods (boiling method, commercial gene matrix, and DNA extractor kit) was compared by using a serial 10-fold dilution of a chicken isolate of Erysipelothrix rhusiopathiae strain in chicken blood. Of the techniques used, the DNA extractor kit, followed by PCR, provided the most sensitive method for the detection of the E. rhusiopathiae strain in chicken blood (approximately 10(0) CFU/0.1 ml of blood). Two E. rhusiopathiae infection experiments were then attempted. In a total of 10 inoculated chickens, bacteremia developed in 9 chickens, consisting of all 5 chickens used in the first trial (ranging from 5.1 x 10(1) to 2.0 x 10(3) CFU/0.1 ml of blood) and 4 of the 5 chickens used in the second trial (ranging from 1.0 x 10(0) to 3.3 x 10(2) CFU/0.1 ml of blood). In the second trial, the 3 detection techniques were applied to the chickens with bacteremia, and the organism could be detected by using the DNA extractor kit in blood specimens from the 3 chickens exhibiting bacteremia of > or =4.2 x 10(1) CFU/0.1 ml of blood. This observation suggests that most E. rhusiopathiae-infected chickens develop more critical bacteremia than the detectable level by PCR with the DNA extractor kit, and the PCR detection method can be used as a first-line screening of avian erysipelas.
Harada
Kazuki
K
National Veterinary Assay Laboratory, Ministry of Agriculture, Forestry and Fisheries, 1-15-1 Tokura, Kokubunji, Tokyo 185-8511, Japan. harada@nval.go.jp
Uchiyama
Mariko
M
Hoshi
Teruyuki
T
Takahashi
Toshio
T
D000818:Animals; D002645:Chickens; D004269:DNA, Bacterial / Q000302:isolation & purification*; D004888:Erysipelothrix; D004889:Erysipelothrix Infections / Q000097:blood* / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D011201:Poultry Diseases / Q000175:diagnosis; D012680:Sensitivity and Specificity
D004269:DNA, Bacterial