Evaluation of PCR Sensitivity
To determine the lowest detection limit of our nested
PCR, seven 10-fold dilutions were prepared from
simulated positive blood specimen. Then, 180 ll of
each dilution containing 2.07 9 105–2.07 9 10-1
Four amplicons were detected in all 34 P. insidiosum
isolates. Only one genus-specific product (512 bp)
was found in five non-insidiosum Pythium spp. as
demonstrated in Fig. 2. No product was found after
amplification of DNA from all the remaining 29 fungal
and 10 bacterial isolates. Therefore, our single-tube
nested PCR had 100 % specificity within the group of
clinical microorganisms tested in this study.