title: Direct Detection of Erythromycin-Resistant Bordetella pertussis in Clinical Specimens by PCR.
abstract: Resistance of Bordetella pertussis to erythromycin has been increasingly reported. We developed an allele-specific PCR method for rapid detection of erythromycin-resistant B. pertussis directly from nasopharyngeal (NP) swab samples submitted for diagnostic PCR. Based on the proven association of erythromycin resistance with the A2047G mutation in the 23S rRNA of B. pertussis, four primers, two of which were designed to be specific for either the wild-type or the mutant allele, were used in two different versions of the allele-specific PCR assay. The methods were verified with results obtained by PCR-based sequencing of 16 recent B. pertussis isolates and 100 NP swab samples submitted for diagnostic PCR. The detection limits of the two PCR assays ranged from 10 to 100 fg per reaction for both erythromycin-susceptible and -resistant B. pertussis. Two amplified fragments of each PCR, of 286 and 112 bp, respectively, were obtained from a mutant allele of the isolates and/or NP swab samples containing B. pertussis DNAs. For the wild-type allele, only a 286-bp fragment was visible when the allele-specific PCR assay 1 was performed. No amplification was found when a number of non-Bordetella bacterial pathogens and NP swab samples that did not contain the DNAs of B. pertussis were examined. This assay can serve as an alternative for PCR-based sequencing, especially for local laboratories in resource-poor countries.
authors.affiliation: The Fourth Military Medical University, Xi'an, China Xi'an Center for Disease Prevention and Control, Xi'an, China.
authors.forename: Zengguo
authors.initials: Z
authors.lastname: Wang
authors.affiliation: Xi'an Center for Disease Prevention and Control, Xi'an, China.
authors.forename: Ruijun
authors.initials: R
authors.lastname: Han
authors.affiliation: Xi'an Center for Disease Prevention and Control, Xi'an, China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Xi'an Center for Disease Prevention and Control, Xi'an, China.
authors.forename: Quanli
authors.initials: Q
authors.lastname: Du
authors.affiliation: Xi'an Center for Disease Prevention and Control, Xi'an, China.
authors.forename: Jifeng
authors.initials: J
authors.lastname: Liu
authors.affiliation: Xi'an Center for Disease Prevention and Control, Xi'an, China.
authors.forename: Chaofeng
authors.initials: C
authors.lastname: Ma
authors.affiliation: Xi'an Center for Disease Prevention and Control, Xi'an, China.
authors.forename: Hengxin
authors.initials: H
authors.lastname: Li
authors.affiliation: Department of Medical Microbiology, Capital Medical University, Beijing, China Department of Medical Microbiology and Immunology, University of Turku, Turku, Finland qiushui.he@utu.fi yanyping@fmmu.edu.cn.
authors.forename: Qiushui
authors.initials: Q
authors.lastname: He
authors.affiliation: The Fourth Military Medical University, Xi'an, China qiushui.he@utu.fi yanyping@fmmu.edu.cn.
authors.forename: Yongping
authors.initials: Y
authors.lastname: Yan
mesh_terms: D000900:Anti-Bacterial Agents / Q000627:therapeutic use*; D001483:Base Sequence; D001886:Bordetella pertussis / Q000187:drug effects* / Q000235:genetics / Q000302:isolation & purification; D004269:DNA, Bacterial / Q000235:genetics; D024881:Drug Resistance, Bacterial / Q000235:genetics*; D004917:Erythromycin / Q000627:therapeutic use*; D006801:Humans; D009305:Nasopharynx / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012338:RNA, Ribosomal, 23S / Q000235:genetics*; D017422:Sequence Analysis, DNA; D014917:Whooping Cough / Q000188:drug therapy / Q000382:microbiology
chemical_list: D000900:Anti-Bacterial Agents; D004269:DNA, Bacterial; D012338:RNA, Ribosomal, 23S; D004917:Erythromycin