Diagnosis of community-acquired pertussis infection: comparison of both culture and fluorescent-antibody assays with PCR detection using electrophoresis or dot blot hybridization.
Diagnosis of Bordetella pertussis infection has been difficult due to the low sensitivity of culture. PCR tests have been shown to be more sensitive than culture, but the reported sensitivity of PCR is variable. We evaluated PCR product detection by using either agarose gel electrophoresis (PCR-gel) or dot blot hybridization with (32)P-labeled oligonucleotide probes, and we compared these methods to both culture and direct fluorescent-antibody (DFA) assays with microscopy for the detection of pertussis. This was done with 225 nasopharyngeal swab specimens collected in community clinic settings. The multiplexed PCR amplified the multiply repeated IS481 B. pertussis sequence and a sequence from the human globin gene as a positive control for specimen adequacy. Of 225 specimens, 179 were judged to be adequate for PCR analysis. Among the adequate specimens, 9, 4, and 10 were culture, DFA, and PCR-gel positive, respectively. The sensitivity of PCR-gel versus culture was 89% while the sensitivity of culture versus PCR-gel was 80%. DFA had the lowest sensitivity. Thirty specimens were positive by PCR with dot blot hybridization; no negative control specimens showed a signal above the background. Among the 79 (44%) adequate specimens with clinical data available, the rates of reported cough or persistent cough were similar for persons who were pertussis positive by each assay. The IS481 PCR, with either electrophoresis or dot blot hybridization, is a sensitive assay; however, at this time it cannot completely replace culture without an overall loss in sensitivity for the detection of pertussis. Further study is required to understand the clinical significance of B. pertussis PCR products detected by dot blot hybridization alone.
Lingappa
Jairam R
JR
Department of Laboratory Medicine, University of Washington, Seattle, USA. JWL8@cdc.gov
Lawrence
William
W
West-Keefe
Sheyla
S
Gautom
Romesh
R
Cookson
Brad T
BT
D001431:Bacteriological Techniques; D001886:Bordetella pertussis / Q000235:genetics / Q000254:growth & development / Q000276:immunology / Q000302:isolation & purification*; D017714:Community-Acquired Infections / Q000175:diagnosis* / Q000382:microbiology; D003470:Culture Media; D019085:Fluorescent Antibody Technique, Direct; D006801:Humans; D015151:Immunoblotting; D009305:Nasopharynx / Q000382:microbiology; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction / Q000379:methods; D014917:Whooping Cough / Q000175:diagnosis* / Q000382:microbiology
D003470:Culture Media