Real-time PCR-based detection of Bordetella pertussis and Bordetella parapertussis in an Irish paediatric population

Grogan, J; Logan, Catriona; O’Leary, John; Rush, Rebecca M.; O’Sullivan, Niamh

doi:10.1099/jmm.0.030049-0

Cited by 14 publications

(13 citation statements)

References 27 publications

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“…Thus, when PCR is compared to culture, the sensitivity of PCR is 100%. The sensitivity of culture compared to PCR ranges from 26% to 85% (93,150,(265)(266)(267)(268). It has been shown that the sensitivity of culture with regard to PCR sensitivity decreases with an increasing duration of disease, and thus shows a steeper decline than that of PCR (267).…”

Section: Factors That Influence the Sensitivity Of Diagnostic Methodsmentioning

confidence: 99%

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Laboratory Diagnosis of Pertussis

2015

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SUMMARYThe introduction of vaccination in the 1950s significantly reduced the morbidity and mortality of pertussis. However, since the 1990s, a resurgence of pertussis has been observed in vaccinated populations, and a number of causes have been proposed for this phenomenon, including improved diagnostics, increased awareness, waning immunity, and pathogen adaptation. The resurgence of pertussis highlights the importance of standardized, sensitive, and specific laboratory diagnoses, the lack of which is responsible for the large differences in pertussis notifications between countries. Accurate laboratory diagnosis is also important for distinguishing between the several etiologic agents of pertussis-like diseases, which involve both viruses and bacteria. If pertussis is diagnosed in a timely manner, antibiotic treatment of the patient can mitigate the symptoms and prevent transmission. During an outbreak, timely diagnosis of pertussis allows prophylactic treatment of infants too young to be (fully) vaccinated, for whom pertussis is a severe, sometimes fatal disease. Finally, reliable diagnosis of pertussis is required to reveal trends in the (age-specific) disease incidence, which may point to changes in vaccine efficacy, waning immunity, and the emergence of vaccine-adapted strains. Here we review current approaches to the diagnosis of pertussis and discuss their limitations and strengths. In particular, we emphasize that the optimal diagnostic procedure depends on the stage of the disease, the age of the patient, and the vaccination status of the patient.

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Section: Factors That Influence the Sensitivity Of Diagnostic Methodsmentioning

confidence: 99%

“…The IS elements IS481 and IS1001 are the most used targets for detection of B. pertussis and B. parapertussis, respectively, by PCR (66,67). For simultaneous detection of B. pertussis and B. parapertussis, multiplex PCR assays targeting both IS elements have been developed (90)(91)(92)(93)(94)(95)(96)(97)(98)(99)(100)(101).…”

Section: Pcr Assaysmentioning

confidence: 99%

Laboratory Diagnosis of Pertussis

2015

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“…In addition, two new sets of primers and Taqman probes were designed based on ptx promoter (PT-P) and B pertussis gene for a porin protein (BPTD_0837) (POR). The new ptx promoter assay was designed through modification of the primer and probe sequences described by Grogan et al (12) in an attempt to further improve the specificity of the assay. By aligning the genomes of several different Bordetella species, it was noted that the published probe sequence has only a single nucleotide mismatch with the corresponding sequences of B bronchiseptica and B parapertussis genome.…”

Section: Resultsmentioning

confidence: 99%

“…A single-target PCR, carefully designed to specifically and exclusively amplify B pertussis DNA would, therefore, be preferable to current assays based on IS 481 or multiple targets. Several real-time PCR assays designed to amplify non-IS 481 single-gene targets have been described for the specific detection of B pertussis (12–14). In the present study, we evaluated the performance of these PCR assays in parallel with a novel real-time PCR assay that targets the B pertussis porin gene and the widely used IS 481 -based PCR assay.…”

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confidence: 99%

Evaluation of Amplification Targets for the Specific Detection of Bordetella pertussis Using Real‐Time Polymerase Chain Reaction

Hasan

Tan

Al‐Rawahi

et al. 2014

Canadian Journal of Infectious Diseases and Medical Microbiolog

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BACKGROUND:Bordetella pertussis infections continue to be a major public health challenge in Canada. Polymerase chain reaction (PCR) assays to detect B pertussis are typically based on the multicopy insertion sequence IS481, which offers high sensitivity but lacks species specificity.METHODS:A novel B pertussis real-time PCR assay based on the porin gene was tested in parallel with several previously published assays that target genes such as IS481, ptx-promoter, pertactin and a putative thialase. The assays were evaluated using a reference panel of common respiratory bacteria including different Bordetella species and 107 clinical nasopharyngeal specimens. Discrepant results were confirmed by sequencing the PCR products.RESULTS:Analytical sensitivity was highest for the assay targeting the IS481 element; however, the assay lacked specificity for B pertussis in the reference panel and in the clinical samples. False-positive results were also observed with assays targeting the ptx-promoter and pertactin genes. A PCR assay based on the thialase gene was highly specific but failed to detect all reference strains of B pertussis. However, a novel assay targeting the porin gene demonstrated high specificity for B pertussis both in the reference panel and in clinical samples and, based on sequence-confirmed results, correctly predicted all B pertussis-positive cases in clinical samples. According to Probit regression analysis, the 95% detection limit of the new assay was 4 colony forming units/reaction.CONCLUSION:A novel porin assay for B pertussis demonstrated superior performance and may be useful for improved molecular detection of B pertussis in clinical specimens.

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“…A mortalidade em lactentes é de aproximadamente 1% 10 . Entre os métodos diagnósticos laboratoriais mais utilizados, podemos citar a cultura de material de nasofaringe (método padrão, com alta sensibilidade, porém baixa especificidade), sorologias específicas (técnicas de ELISA, com baixa sensibilidade e especificidade) e reação em cadeia da polimerase (polymerase chain reaction, PCR), método atualmente mais utilizado, com alta especificidade (90%) e maior sensibilidade que a cultura convencional [11][12][13] . Dada a atual elevada incidência de coqueluche e seus variados modos de apresentação, este estudo tem como objetivo descrever as características demográficas e clínicas de pacientes pediátricos menores de 3 anos de idade internados em um hospital universitário terciário com diagnóstico de coqueluche confirmado por PCR no período de julho de 2011 a dezembro de 2012.…”

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