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J. Christopher Post, MD; Robert A. Preston, PhD; Jerome J. Aul; Margaret Larkins-Pettigrew, MD; Jill Rydquist-White; Kenneth W. Anderson; Robert M. Wadowsky, ScD; David R. Reagan, MD, PhD; Elaine S. Walker, PhD; Lawrence A. Kingsley, DrPH; Anthony E. Magit, MD; Garth D. Ehrlich, PhD

JAMA. 1995;273(20):1598-1604.

Abstract
Objective.
—To determine if the polymerase chain reaction (PCR) can detect bacterial DNA in pediatric middle ear effusions that are sterile by standard cultural methods.

Design.
—Single-center, blinded, comparative study of diagnostic assays. The PCR-based detection systems for Moraxella catarrhalis, Haemophilus influenzae, and Streptococcus pneumoniae were designed and validated using a battery of DNAs obtained from cultured bacteria. Chronic middle ear effusion specimens were collected and comparatively analyzed by culture and the PCR.

Setting.
—Tertiary care pediatric hospital.

Patients.
—A total of 97 middle ear effusions were collected from pediatric outpatients at Children's Hospital of Pittsburgh (Pa) during myringotomy and tube placement for chronic otitis media with effusion (duration >3 months). All patients had failed multiple courses of antimicrobial therapy and were diagnosed by a combination of validated otoscopy and tympanograms.

Main Outcome Measure.
—Differences in the percentage of positive test results between PCR-based assays and culture for M catarrhalis, H influenzae, and S pneumoniae.

Results.
—Of the 97 specimens of otitis media with effusion, 28 (28.9%) tested positive by both culture and PCR for M catarrhalis, H influenzae, or S pneumoniae. An additional 47 specimens (48%) were PCR positive/culture negative for these three bacterial species. Thus, 75 (77.3%) of the 97 specimens tested PCR positive for one or more of the three test organisms. The minimum number of bacterial genomic equivalents present in the average culture-negative ear was estimated to be greater than 10⁴ based on dilutional experiments.

Conclusions.
—The PCR-based assay systems can detect the presence of bacterial DNA in a significant percentage of culturally sterile middle ear effusions. While this finding is not proof of an active bacterial infectious process, the large number of bacterial genomic equivalents present in the ears is suggestive of an active process.

(JAMA. 1995;273:1598-1604)

Author Affiliations
From the Departments of Otolaryngology (Drs Post, Larkins-Pettigrew, Magit, and Ehrlich), Pathology (Drs Preston, Larkins-Pettigrew, Wadowsky, and Ehrlich, Messrs Aul and Anderson, and Ms Pydquist-White), and Infectious Diseases and Microbiology (Drs Wadowsky, Kingsley, and Ehrlich), University of Pittsburgh (Pa); Departments of Pediatric Otolaryngology (Drs Post, Larkins-Pettigrew, Magit, and Ehrlich) and Pathology (Dr Wadowsky), Children's Hospital of Pittsburgh (Pa); and Department of Internal Medicine, James H. Quillen College of Medicine, East Tennessee State University, and Mountain Home Veterans Administration Medical Center, Johnson City, Tenn (Drs Reagan and Walker). Dr Post was a recipient of the Academy Research and Training Award, American Academy of Otolaryngology-Head and Neck Surgery.

Footnotes
Reprint requests to Department of Pathology, University of Pittsburgh, 737 Scaife Hall, Pittsburgh, PA 15261 (Dr Ehrlich).

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