Discriminative PCR of Bordetella pertussis from closely related Bordetella species using 16S rDNA Gene.
- Author:
Sang Oun JUNG
1
;
Yu Mi MOON
;
Hwa Young SUNG
;
Yeon Ho KANG
;
Jae Yon YU
Author Information
- Publication Type:Original Article
- Keywords: Bordetella pertussis; Bordetella holmesii; 16S rDNA; species-specific PCR primer
- MeSH: Bordetella; Bordetella bronchiseptica; Bordetella parapertussis; Bordetella pertussis; Discrimination (Psychology); DNA; DNA, Ribosomal; Polymerase Chain Reaction; Sensitivity and Specificity; Whooping Cough
- From: Infection and Chemotherapy 2008;40(1):24-31
- CountryRepublic of Korea
- Language:English
- Abstract: BACKGROUND: Polymerase-chain reaction (PCR) detection is useful to diagnosis of pertussis at initial stage because the growth rate of Bordetella pertussis (B. pertussis) is relatively slow. Currently, the primer set for the insertion sequence IS481 (BP primer) is used widely for PCR detection of B. pertussis. However, the cross-reactivity of BP primer set with Bordetella holmesii (B. holmesii) was reported recently. Therefore, discrimination of B. pertussis and B. holmesii is needed in PCR step. For this reason, we developed new primer sets based on 16S rDNA sequence for diagnostic use and estimated the efficiency of these new primer sets. MATERIALS AND METHODS: The specific PCR primers were designed from the aligned sequence matrix of 16S rDNA genes of various Bordetella species. The specificity of designed primers were estimated using clinically important 4 Bordetella species, B. pertussis, B. holmesii, Bordetella parapertussis (B. parapertussis) and Bordetella bronchiseptica (B. bronchiseptica). The sensitivity to B. pertussis of designed primers was also estimated and compared with BP primer set. RESULTS: As the results, the developed new primer set successfully distinguished B. pertussis and other Bordetella species containing B. holmesii. In the sensitivity assay, the detectable limits of 16S-F2/16S-R1 primer set for B. pertussis were revealed as 5 pg of genomic DNA and 105 cells/mL of cell suspension. In addition to these, identical results between BP with primer and new primer were obtained in clinical samples. CONCLUSION: In this study, the specific primer set for B. pertussis was developed based on 16S rDNA sequence and this primer set did not show cross-reactivity to B. holmesii. In addition to these, the applicability of this primer set to the clinical specimens was also confirmed.
