FIG. 4.

The PCR-based assay can detect as few as four bacteria/ml. The minimum number of bacteria that can be detected by this method was determined by diluting a culture which contains 1.5 × 10⁸ bacteria and adjusting it to an OD₆₀₀ of 0.3 with various dilutions in PBS and sterile blood. The diluted culture samples were then subjected to one round of PCR amplification for 35 cycles; the products were then visualized by electrophoresis on a 0.8% agarose gel. The corresponding number of bacteria/ml is given at the top of each lane. (A) Different numbers of S. Typhi CT18 diluted in PBS, showing that one bacterium/ml can produce a visible band. (B) Different numbers of S. Typhi CT18 diluted in blood, showing that the procedure can detect as few as four bacteria/ml of blood. (C) Different numbers of S. Paratyphi A diluted in PBS, showing a sensitivity of detection of four bacteria/ml. (D) Different numbers of S. Paratyphi A diluted in blood, showing a lower detection limit of one bacterium/ml of blood. M lanes contain molecular size markers.