Prevalence of Bordetella pertussis and Bordetella parapertussis in infants presenting to the emergency department with bronchiolitis.

BACKGROUND: The clinical presentation of Bordetella pertussis can overlap with that of respiratory syncytial virus (RSV), and coinfection does occur, but management differs. HYPOTHESIS: The prevalence of B. pertussis is < 2% among Emergency Department (ED) patients with bronchiolitis. Our secondary hypothesis was that the prevalence of Bordetella parapertussis is also < 2% among these patients. METHODS: Nasal washings were obtained from children up to 18 months of age (inclusive) who presented to a county hospital ED with a clinical diagnosis of bronchiolitis. These washings were frozen to -70°C before testing for B. pertussis and B. parapertussis using species-specific real-time polymerase chain reaction (PCR) assays. The assays were optimized to target conserved regions within a complement gene and the CarB gene, respectively. A Bordetella spp. genus-specific real-time PCR assay was designed to detect the Bhur gene of B. pertussis, B. parapertussis, and B. bronchiseptica. RSV antigen detection was also performed. RESULTS: There were 227 patients enrolled. After exclusions, 204 remained in the analysis. RSV antigen testing was positive in 109/186 (59%) of the patients in whom it was performed. All samples were tested for B. pertussis. B. parapertussis testing could not be completed on 23 samples. No cases (0/204; 95% confidence interval [CI] 0-1.8%) tested positive for B. pertussis or B. parapertussis (0/181; 95% CI 0-2%). CONCLUSION: The prevalence of B. pertussis in children presenting to the ED with bronchiolitis was < 2%.

Comparison of respiratory virus detection rates for infants and toddlers by use of flocked swabs, saline aspirates, and saline aspirates mixed in universal transport medium for room temperature storage and shipping.

A nylon flocked swab/universal transport medium collection method developed for bacterial sexually transmitted infections was adapted to detect respiratory viruses in infants and toddlers. This method significantly outperformed the traditional use of nasal aspirates in terms of PCR-based virus detection (P = 0.016), and the samples were easier for clinicians to evaluate, store, and transport.

Prevalence of Bordetella pertussis and Bordetella parapertussis in Samples Submitted for RSV Screening.

BACKGROUND: The clinical presentation of Bordetella pertussis can overlap with that of respiratory syncytial virus (RSV); however, management differs. HYPOTHESIS: First, the prevalence of B. pertussis is less than 2% among patients screened for RSV, and second the prevalence of B. parapertussis is also less than 2% among these patients. METHODS: Nasal washings submitted to a clinical laboratory for RSV screening were tested for B. pertussis and B. parapertussis, using species-specific real-time polymerase chain reaction (PCR) assays. These were optimized to target conserved regions within a complement gene and the CarB gene, respectively. A Bordetella spp. genus-specific real-time PCR assay was designed to detect the Bhur gene of B. pertussis, B. parapertussis, and B. bronchiseptica. RSV A and B subtypes were tested by reverse transcription-PCR. RESULTS: Four hundred and eighty-nine clinical samples were tested. There was insufficient material to complete testing for one B. pertussis, 10 RSV subtype A, and four RSV subtype B assays. Bordetella pertussis was detected in 3/488 (0.6%) (95% CI 0.1% to 1.8%), while B. parapertussis was detected in 5/489 (1.0%) (95% CI 0.3% to 2.4%). Dual infection of B. pertussis with RSV and of B. parapertussis with RSV occurred in two and in three cases respectively. RSV was detected by PCR in 127 (26.5%). CONCLUSION: The prevalence of B. pertussis in nasal washings submitted for RSV screening was less than 2%. The prevalence of parapertussis may be higher than 2%. RSV with B. pertussis and RSV with B. parapertussis coinfection do occur.

Simultaneous detection of herpes simplex virus types 1 and 2 by real-time PCR and Pyrosequencing.

BACKGROUND: Up to 80% of the US adult population has been exposed to herpes simplex virus (HSV) type 1, primarily during childhood. Also, approximately 20% of the US population has contracted genital herpes from HSV-2 infections. Clinical symptoms can present as fever, headache, malaise, myalgia, and cold sores/lesions that cause pain, itching, dysuria, and vaginal or urethral discharge. A recurrence of infection is common. HSV culturing is characterized by low sensitivity with variable success rates due to shipping conditions. OBJECTIVE: To design and validate a real-time PCR assay capable of simultaneously detecting each HSV subtype. STUDY DESIGN: ATCC-purchased HSV-1 and HSV-2 positive samples and HSV-1 and HSV-2 infected clinical specimens were assayed simultaneously with shared amplification primers and subtype-specific probes against the HSV glycoprotein B gene on a Rotor-Gene 3000 platform. Separately, two PCR reactions were performed in which one primer contained a 5' biotin modification. Single-stranded DNA from the amplicon was purified and Pyrosequenced. RESULTS: The quantitative range of the assay extended from 10(8) through 10(0) copies of each virus (r(2) > 0.991) and specificity was determined by non-amplification of 37 different human pathogens, including other herpesviruses such as VZV, CMV, and EBV. Sensitivity and specificity values of 100% were calculated by concordance analysis between the real-time PCR and the DNA Pyrosequencing results (HSV-1: n = 119, HSV-2: n = 120). Application of this assay to 4581 cervical swab specimens collected from women visiting physicians primarily in six states provided detection rates of 3.1% for HSV-1 and 7.6% for HSV-2. The average age of women infected with HSV-1 was 29.5 versus 35.6 for HSV-2. CONCLUSIONS: This procedure was demonstrated as both highly sensitive and specific for the detection of HSV-1 and HSV-2 in a single reaction. Also, the integration of Pyrosequencing analysis permitted an innovative and rapid verification for each subtype.