Genomic Survey of Bordetella pertussis Diversity, United States, 2000–2013

We characterized 170 complete genome assemblies from clinical Bordetella pertussis isolates representing geographic and temporal diversity in the United States. These data capture genotypic shifts, including increased pertactin deficiency, occurring amid the current pertussis disease resurgence and provide a foundation for needed research to direct future public health control strategies

Analysis of Toxigenic Corynebacterium ulcerans Strains Revealing Potential for False-Negative Real-Time PCR Results▿

Diphtheria surveillance depends on the rapid and reliable recognition of the toxin gene in Corynebacterium diphtheriae. Real-time PCR is a rapid tool to confirm the presence of the diphtheria toxin gene (tox) in an isolate or specimen. We report that some toxigenic Corynebacterium ulcerans strains show atypical results in a real-time PCR for tox

Development and Analytical Validation of an Immunoassay for Quantifying Serum Anti-Pertussis Toxin Antibodies Resulting from Bordetella pertussis Infection ▿

Adequately sensitive and specific methods to diagnose pertussis in adolescents and adults are not widely available. Currently, no Food and Drug Administration-approved diagnostic assays are available for the serodiagnosis of Bordetella pertussis. Since concentrations of B. pertussis-specific antibodies tend to be high during the later phases of disease, a simple, rapid, easily transferable serodiagnostic test was developed. This article describes test development, initial evaluation of a prototype kit enzyme-linked immunosorbent assay (ELISA) in an interlaboratory collaborative study, and analytical validation. The data presented here demonstrate that the kit met all prespecified criteria for precision, linearity, and accuracy for samples with anti-pertussis toxin (PT) immunoglobulin G (IgG) antibody concentrations in the range of 50 to 150 ELISA units (EU)/ml, the range believed to be most relevant for serodiagnosis. The assay met the precision and linearity criteria for a wider range, namely, from 50 to 200 EU/ml; however, the accuracy criterion was not met at 200 EU/ml. When the newly adopted World Health Organization International Standard for pertussis antiserum (human) reference reagent was used to evaluate accuracy, the accuracy criteria were met from 50 to 200 international units/ml. In conclusion, the IgG anti-PT ELISA met all assay validation parameters within the range considered most relevant for serodiagnosis. This ELISA was developed and analytically validated as a user-friendly kit that can be used in both qualitative and quantitative formats. The technology for producing the kit is transferable to public health laboratories

Clinical evaluation and validation of laboratory methods for the diagnosis of <i>Bordetella pertussis</i> infection: Culture, polymerase chain reaction (PCR) and anti-pertussis toxin IgG serology (IgG-PT)

<div><p>Introduction</p><p>The appropriate use of clinically accurate diagnostic tests is essential for the detection of pertussis, a poorly controlled vaccine-preventable disease. The purpose of this study was to estimate the sensitivity and specificity of different diagnostic criteria including culture, multi-target polymerase chain reaction (PCR), anti-pertussis toxin IgG (IgG-PT) serology, and the use of a clinical case definition. An additional objective was to describe the optimal timing of specimen collection for the various tests.</p><p>Methods</p><p>Clinical specimens were collected from patients with cough illness at seven locations across the United States between 2007 and 2011. Nasopharyngeal and blood specimens were collected from each patient during the enrollment visit. Patients who had been coughing for ≤ 2 weeks were asked to return in 2–4 weeks for collection of a second, convalescent blood specimen. Sensitivity and specificity of each diagnostic test were estimated using three methods—pertussis culture as the “gold standard,” composite reference standard analysis (CRS), and latent class analysis (LCA).</p><p>Results</p><p>Overall, 868 patients were enrolled and 13.6% were <i>B</i>. <i>pertussis</i> positive by at least one diagnostic test. In a sample of 545 participants with non-missing data on all four diagnostic criteria, culture was 64.0% sensitive, PCR was 90.6% sensitive, and both were 100% specific by LCA. CRS and LCA methods increased the sensitivity estimates for convalescent serology and the clinical case definition over the culture-based estimates. Culture and PCR were most sensitive when performed during the first two weeks of cough; serology was optimally sensitive after the second week of cough.</p><p>Conclusions</p><p>Timing of specimen collection in relation to onset of illness should be considered when ordering diagnostic tests for pertussis. Consideration should be given to including IgG-PT serology as a confirmatory test in the Council of State and Territorial Epidemiologists (CSTE) case definition for pertussis.</p></div

Laboratory diagnostic test results for all participants enrolled in the clinical validation study (N = 868).

<p>Laboratory diagnostic test results for all participants enrolled in the clinical validation study (N = 868).</p

Sensitivity and specificity estimates of <i>B</i>. <i>pertussis</i> culture, PCR, serology, and the clinical case definition.

<p>Sensitivity and specificity estimates of <i>B</i>. <i>pertussis</i> culture, PCR, serology, and the clinical case definition.</p

<i>B</i>. <i>pertussis</i>-positive laboratory test results by time of specimen collection.

<p><i>B</i>. <i>pertussis</i>-positive laboratory test results by time of specimen collection.</p

<i>B</i>. <i>pertussis</i>-positive laboratory test results by age group (N = 868).

<p>Acute sera are collected ≤ 2 weeks after cough onset, and convalescent sera are collected > 2 weeks after cough onset.</p

Enrollment criteria for participation in the clinical validation study, 2007–2011.

<p>Enrollment criteria for participation in the clinical validation study, 2007–2011.</p