Detection and Quantification of Pathogenic Bacteria and Fungi

[Guest guest]

Vol. 128 No. 5, May 2010

Detection and Quantification of Pathogenic Bacteria and Fungi

Using Real-Time Polymerase Chain Reaction by Cycling Probe in Patients With

Corneal Ulcer

Motoki Itahashi, MD, PhD; Shiro Higaki, MD, PhD; Masahiko Fukuda, MD, PhD;

Yoshikazu Shimomura, MD, PhD

http://archopht.ama-assn.org/cgi/content/short/128/5/535

Arch Ophthalmol. 2010;128(5):535-540.

Objective To detect and quantitate the causative pathogens in patients with

corneal ulcer using real-time polymerase chain reaction (PCR) by cycling probe.

Design Clinical and laboratory study of 40 eyes of 40 patients diagnosed with

corneal ulcer. Two methods were used for pathogen detection: bacterial culture

and real-time PCR with the patient's corneal scrapings. Probes and primers of

real-time PCR were designed to be pathogen specific for simultaneous detection

of Staphylococcus aureus, Staphylococcus pneumoniae, Pseudomonas aeruginosa,

methicillin-resistant S aureus, Candida species, and Fusarium species. Results

by both methods were evaluated and compared.

Results Of 40 eyes, 20 eyes had the same pathogens detected by both methods and

those were S aureus (3 eyes; mean [sE], 3.8 [1.3] x 101 copies/sample), S

pneumoniae (5 eyes; mean [sE], 5.6 [5.1] x 103 copies/sample), P aeruginosa (8

eyes; 5.1 [4.0] x 103 copies/sample), methicillin-resistant S aureus (1 eye; 1.0

x 102 copies/sample), and Candida species (3 eyes; mean [sE], 8.8 [4.9] x 103

copies/sample). Six eyes showed negative results by both methods. Results of

both methods disagreed in 14 eyes; specifically, 11 had positive PCR results

only, 2 had positive culture results only, and 1 eye had positive results for

different pathogens.

Conclusions The real-time PCR assay can simultaneously detect and quantitate

bacterial and fungal pathogens in patients with corneal ulcer. Real-time PCR can

be a fast diagnostic tool and may be useful as an adjunct to identify potential

pathogens.

Author Affiliations: Department of Ophthalmology, Kinki University School of

Medicine, Osaka-Sayama, Japan.