Comparison of Serological and Real-Time PCR Assays To Diagnose Bordetella pertussis Infection in 2007

Abstract

Bacterial culture for diagnosing pertussis infection has high specificity but poor sensitivity and is slow. Highly sensitive real-time PCR assays and single-serum pertussis serology have been developed to overcome these limitations, but there are few data available on the relative sensitivities and specificities of such assays for pertussis diagnosis. Using data on 195 participants (>7 years old) from an epidemiological study, we assessed the sensitivity, specificity, and performance (Youden index) for pertussis diagnosis of the pertussis toxin enzyme-linked immunosorbent assay (using single and paired serology) and of real-time PCR assays (using the IS481 and ptxA-Pr targets). All available diagnostic information (clinical and laboratory) was pooled to serve as the gold standard. Single serology was the most efficient diagnostic test (Youden index, 0.57 to 0.58), with relatively high sensitivity (>64%) and high specificity (>90%), independent of the cutoff level. IS481 PCR performance was superior to that of ptxA-Pr PCR, and it was the second-most-efficient tool (Youden index, 0.30). Performing both ptxA-Pr and IS481 PCRs did not improve diagnostic performance. The greatest test efficiency (Youden index, 0.69 to 0.74) was achieved when single-serum serology was used in combination with IS481 or ptxA-Pr PCR or paired serology. Combining single serology with one PCR or paired serology increased the sensitivity with an associated limited decrease in specificity. The most specific tests for diagnosis of pertussis were single serology and ptxA-Pr PCR, and the most sensitive diagnostic tool was the combination of IS481 PCR with single serology.Currently there is no satisfactory gold standard technique for laboratory confirmation of a pertussis infection. Although culture is highly specific, sensitivity is low and declines with the duration of illness, and the method may take up to 7 days to provide a result. Using culture alone to diagnose pertussis is likely to lead to underreporting of pertussis cases (8). Serological assays, using one or two serum samples, have been developed to improve the sensitivity of the pertussis diagnosis, but they have a lower specificity than culture (3,9,20,25). PCR techniques were developed to overcome these limitations, but despite a consensus meeting in 2005 (24), these techniques still require further standardization and optimization. To date there are few reports (8) in which the sensitivities and specificities of these different techniques are compared using a population exposed to pertussis.The objective of this study was to compare the sensitivities and specificities of the current most widely used techniques to diagnose pertussis in exposed populations, namely, two PCR methods (IS481 and ptxA-Pr targets) and single and paired serology for detection of antipertussis toxin (PT) antibodies in the serum of the patients using enzyme-linked immunosorbent assay (ELISA) with two different cutoffs (two-or fourfold change for paired serology and Ն100 or Ն125 ELISA units [EU]/ml for single...

Performance of Commercial Enzyme-Linked Immunosorbent Assays for Detection of Antibodies to Bordetella pertussis

“…In clinical practice, however, pertussis diagnosis is made mostly by single-sample serology using a single or a dual cutoff. Single-sample serology together with PCR was recently found to be the most sensitive method for diagnosing pertussis (1).…”

Significant Finding of Bordetella holmesii DNA in Nasopharyngeal Samples from French Patients with Suspected Pertussis

“…To identify the species, we used previously developed specific "in-house" real-time PCR. These real-time PCRs are based on the amplification of the promoter of the pertussis toxin operon (ptxAPr-based PCR) specific for B. pertussis (1) and of the recA gene (RecA-based PCR) specific for B. holmesii (2). We also used a real-time PCR based on the BP3385 gene (BP3385-based PCR), which was initially described as specific for B. pertussis (5) but was subsequently shown to score positive for some B. bronchiseptica isolates (8).…”

Proficiency Program for Real-Time PCR Diagnosis of Bordetella pertussis Infections in French Hospital Laboratories and at the French National Reference Center for Whooping Cough and other Bordetelloses

“…They demonstrated that real-time PCR could be used to diagnose whooping cough in young children for up to three weeks after the start of treatment. A study comparing current widely used techniques for whooping cough diagnosis in exposed populations showed a combination of real-time PCR for IS481 and single serological tests to be the most sensitive diagnostic approach (1).…”

“…The insertion sequence element IS481, found in several hundred copies in the B. pertussis genome, is frequently used as a target for B. pertussis detection and has a much greater analytical sensitivity than assays with single-copy target sequences, such as that of the pertussis toxin promoter (1). The European consensus group EUPertStrain has issued several recommendations for the standardization of real-time PCR diagnosis to increase the quality of the results (20).…”

“…B. pertussis culture remains the gold standard method for diagnosis but has low sensitivity, takes a long time, and may be difficult in practical terms. The detection of anti-PT antibodies by reference enzyme-linked immunosorbent assay is highly specific (1), but there is a lack of validated commercial tests (22). Furthermore, the increasing use of adult vaccination renders this approach to diagnosis much less useful (2).…”