Abstract
Bordetella pertussis is the causative agent of whooping cough, commonly referred to as pertussis. Although the incidence of pertussis was reduced through vaccination, during the last thirty years it has returned to high levels in a number of countries. This resurgence has been linked to the switch from the use of whole-cell to acellular vaccines. Protection afforded by acellular vaccines appears to be short-lived compared to that afforded by whole cell vaccines. In order to inform future vaccine improvement by identifying immune correlates of protection, a human challenge model of B. pertussis colonisation has been developed. Accurate measurement of colonisation status in this model has required development of a qPCR-based assay to enumerate B. pertussis in samples that distinguishes between viable and dead bacteria. Here we report the development of this assay and its performance in the quantification of B. pertussis from human challenge model samples. This assay has future utility in diagnostic labs and in research where a quantitative measure of both B. pertussis number and viability is required.
Highlights
Whooping cough, or pertussis, is a highly contagious respiratory tract infection of humans caused by the gram-negative coccobacillus Bordetella pertussis
IS481 is often used as the target for qPCR detection of B. pertussis as it is present at ~250 copies per cell in B. pertussis, providing great sensitivity
To develop a propidium monoazide (PMA)-qPCR assay, the sensitivity of qPCR for detection of B. pertussis was tested over a range of template genomic DNA (gDNA) concentrations
Summary
Pertussis, is a highly contagious respiratory tract infection of humans caused by the gram-negative coccobacillus Bordetella pertussis. Clinical manifestations of pertussis depend on age and immune status of the host and include a low-grade fever, cyanosis, and paroxysmal cough accompanied by a high-pitched “whoop” [1]. Infants aged less than 1 year old present the highest incidence of pertussis and are at the greatest risk of severe disease and death [2]. The introduction of vaccination in the early 1950s significantly reduced the incidence of pertussis in developed nations, the number of reports of pertussis has been. Enumerating viable B. pertussis using PMA-qPCR design, data collection and analysis, decision to publish, or preparation of the manuscript
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