PCR-based detection of bacterial DNA after antimicrobial treatment is indicative of persistent, viable bacteria in the chinchilla model of otitis media.
OBJECTIVE
Bacterial deoxyribonucleic acid (DNA) has been previously detected by polymerase chain reactions (PCR) in a significant percentage of culturally-sterile pediatric middle-ear effusions. The current study was designed to determine whether this represents the existence of viable bacteria or the persistence of residual DNA in the middle-ear cleft.


METHODS
The middle-ear cavities of two sets of chinchillas were inoculated with either: 1) 100 colony-forming units (CFU) of live Haemophilus influenzae, 2.2 x 10(6) CFU of pasteurized Moraxella catarrhalis, and 1000 ng of DNA (>10(8) genomic equivalents) from Streptococcus pneumoniae; or 2) 100 CFU of live S pneumoniae, 2.2 x 10(6) CFU of pasteurized M catarrhalis and 1000 ng of purified DNA from H influenzae. Animals were treated with ampicillin for 5 days beginning on day 3. A single-point longitudinal study design was used for sampling to eliminate the possibility of contamination.


RESULTS
No DNA was detectable from the heat-killed bacteria or the purified DNA after day 3. However, DNA from the live bacteria persisted through day 21, even though all specimens were culture-negative following the initiation of antimicrobial therapy.


CONCLUSIONS
These findings indicate that purified DNA and DNA from intact but nonviable bacteria do not persist in the middle-ear cleft in the presence of an effusion, even following high copy inoculation. In contrast, antibiotic-treated bacteria persist in some viable state for weeks as evidenced by the differential ability of the PCR-based assay systems to detect the live bacteria, but not detect the heat-killed organisms.
Post
J C
JC
Department of Pathology, School of Medicine, University of Pittsburgh Medical Center, Pennsylvania 15261, USA.
Aul
J J
JJ
White
G J
GJ
Wadowsky
R M
RM
Zavoral
T
T
Tabari
R
R
Kerber
B
B
Doyle
W J
WJ
Ehrlich
G D
GD
D000667:Ampicillin / Q000627:therapeutic use*; D000818:Animals; D000900:Anti-Bacterial Agents / Q000627:therapeutic use*; D002682:Chinchilla; D004269:DNA, Bacterial; D006193:Haemophilus influenzae / Q000472:pathogenicity*; D001936:Moraxella catarrhalis / Q000472:pathogenicity*; D010033:Otitis Media / Q000188:drug therapy / Q000209:etiology / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D013296:Streptococcus pneumoniae / Q000472:pathogenicity*
D000900:Anti-Bacterial Agents; D004269:DNA, Bacterial; D000667:Ampicillin