A polymerase chain reaction enzyme immunoassay for diagnosing infection caused by Aspergillus fumigatus

Aim-To develop reaction enzyme immunoassay (PCR-EIA) to measure levels of c irculating aspergillus DNA in invasive aspergillosis caused by Aspergillus fumigatus. Methods-The PCR reaction was based on primers from the 18s rRNA gene. Bindi ng of the product to a streptavidin coated microtitration plate was mediate d by a biotinylated capture probe. The product was digoxigenylated during P CR and this was the tag to which antibody was bound in the subsequent EIA. Results-The optical density (OD) endpoint was < 0.1 in 10 sera from neutrop enic patients with no evidence of invasive aspergillosis, and in 10 sera fr om nonneutropenic patients with bacterial pneumonia (group 1). The OD from five of 12 patients with allergic bronchopulmonary aspergillosis (ABPA) (gr oup 2), three with an aspergilloma (group 3), and five with possible invasi ve aspergillosis (group 4) was greater than or equal to 0.1. In 63 sera fro m 33 cases of proven invasive aspergillosis (group 5) an OD greater than or equal to 0.1 was achieved in 48 sera from 30 patients. The maximum OD was 0.510. The level fell in survivors and gradually rose in fatal cases. Conclusions-This assay validated the concept of diagnosing invasive aspergi llosis by measuring levels of circulating fungal DNA in serum.