Multiplex polymerase chain reaction for diagnosis of viral and chlamydial keratoconjunctivitis

PURPOSE. TO develop a multiplex polymerase chain reaction (PCR) for the det ection of adenovirus, herpes simplex virus, and Chlamydia trachomatis in co njunctival swabs. METHODS. Oligonucleotide primers for detection of the 3 agents were combine d in one reaction and evaluated for optimal performance using control DNAs of adenovirus type 2, herpes simplex virus, and C. trachomatis plasmid. The multiplex PCR was evaluated prospectively against its corresponding uniple x PCRs, virus isolation, Chlamydia Amplicor PCR, and an immunoassay techniq ue (immune dot blot test) in a total of 805 conjunctival swabs from patient s with suspected viral and chlamydial keratoconjunctivitis. RESULTS. The multiplex PCR was as sensitive as uniplex PCRs for the detecti on of the agents in clinical specimens. In the prospective study, 48 of 49 (98%) clinical specimens were positive for adenovirus by the multiplex PCR compared with 26 of 49 (53%) by adenovirus isolation. For herpes simplex vi rus detection, the multiplex PCR had a sensitivity of 92% (34/37) compared with 94.5% (35/37) by cell culture. The multiplex PCR produced identical re sults to the Amplicor PCR (21/21; 100%) compared with 71% (15/21) by the im mune dot blot test. CONCLUSIONS. With clinical specimens the multiplex PCR was as sensitive as ifs respective uniplex PCRs but more sensitive than adenovirus isolation an d as sensitive as herpes simplex virus isolation or C. trachomatis Amplicor PCR. It has the potential to replace several diagnostic tests with consequ ent savings in cost. The test also reduces the risk of misdiagnosis by the clinicians.