TABLE 1.
Determination of K. kingae in animal blood via detection of rtxA and rtxB by qPCR analysisa
| Inoculum and animal | CP value |
Bacteremia | Lesion | Relative wt gain | |
|---|---|---|---|---|---|
| rtxA | rtxB | ||||
| Buffer | |||||
| 1 | − | − | − | − | 1.22 |
| 2 | − | − | − | − | 1.33 |
| KKNB100 | |||||
| 1 | − | − | − | − | 1.23 |
| 2 | − | − | − | − | 1.29 |
| PYKK081 | |||||
| 1 | 20.63 | 25.78 | + | + | 1.04 |
| 2 | 20.61 | 25.89 | + | + | 1.15 |
| 3 | 18.71 | 25.29 | + | + | 0.88 |
The table lists CP values and the major clinical symptoms for representative animals of each group. CP values are presented as the cycle at which fluorescence is deemed to be detectable above the background during the exponential phase of amplification as described in Materials and Methods. DNA purified from PYKK081 was used to determine the minimum detectable CFU, which was equivalent to 8 to 75 CFU. qPCRs were performed from three PYKK081-, two KKNB100-, and two PBS-injected animals, each in technical duplicate. Bacteremia was defined as a positive call from both the rtxA and rtxB primer sets. The threshold for specific amplification (positive qualitative identification) was set at 26 cycles, below that of the negative control. The major clinical presentations, lesion formation, and weight changes are shown. Mortality is not reflected in the table, since the animals were sacrificed at 48 h postinfection; however, formation of a lesion always correlated with a lethal outcome. The weight gain ratio was determined by dividing the PN 9 (preinjection) by the PN 7 (preinjection) rat weight.