[Bordetella pertussis detection by real-time PCR, immunofluorescence and culture: prospective evaluation and molecular epidemiology].
BACKGROUND
An increase in the incidence of pertussis has been observed in recent years. The aim of this study was to determine the usefulness of several procedures, including real-time PCR, for the laboratory diagnosis of pertussis, and to investigate clonal relationships among clinical isolates of Bordetella pertussis.


METHODS
During the period of August 2002 to October 2003, nasopharyngeal swabs were collected from pediatric and adult patients with symptoms of pertussis, and contact cases. The samples were processed by culture, direct fluorescence assay (DFA), and real-time PCR. Most of the isolates were further characterized by pulsed-field gel electrophoresis (PFGE).


RESULTS
Among 121 clinical samples corresponding to 117 patients, B. pertussis was detected in 17 samples by culture (14.1%), 30 samples (24.8%) by DFA and 41 samples (33.9%) by real-time PCR. Real-time PCR diagnosed 26 and 24 more cases than culture and DFA, respectively. Seventeen clinical isolates were available for PFGE analysis, 14 collected during the study period and three in 1997, 2000 and 2001. PFGE identified 5 different genotypes, 2 of which included 8 (genotype C) and 6 (genotype E) isolates. Two of the older isolates (1997 and 2001) were identified as genotype C.


CONCLUSIONS
Real-time PCR applied to the diagnosis of pertussis provided more positive results than DFA and culture, but the true diagnostic value of these results should be clarified. Two bacterial clones were dominant, and one of them has circulated at least since 1997.
García-Martínez
Jesús
J
Servicio de Microbiología. Hospital Universitario 12 de Octubre. Madrid. España.
Chaves
Fernando
F
Salto
Efrén
E
Otero
Joaquín R
JR
D000293:Adolescent; D000328:Adult; D001431:Bacteriological Techniques; D001886:Bordetella pertussis / Q000254:growth & development* / Q000302:isolation & purification*; D002648:Child; D002675:Child, Preschool; D019085:Fluorescent Antibody Technique, Direct; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D008875:Middle Aged; D016133:Polymerase Chain Reaction; D011446:Prospective Studies; D014917:Whooping Cough / Q000175:diagnosis* / Q000453:epidemiology* / Q000382:microbiology