The overall sensitivity of PCR for diagnosing pertussis was 65% in the present study, where serology was used to confirm the great majority of cases
The overall sensitivity of PCR for diagnosing pertussis was 65% in the present study, where serology was used to confirm the great majority of cases. to 100%, but the sensitivity is only about 60% if the culture is taken early during the disease and even lower the longer the disease Golgicide A persists (11, 14). Demonstrations of significant increases in serum antibodies against pertussis toxin (PT) or filamentous hemagglutinin (FHA) are also well-documented methods for diagnosing pertussis with high specificity and sensitivity (3, 26, 27). Serology requires paired sera taken about 1 month apart. Hence, results are not available until the patient is recovering. Thus, there is a Golgicide A need for a specific and sensitive diagnostic method for pertussis that can be used in the early stage Golgicide A of the disease. Detection of and infections by comparison of PCR with cultures, serology, and clinical diagnosis. MATERIALS AND METHODS Study design. All individuals in the present study were participants or family members of participants in a double-blind, placebo-controlled efficacy trial of a monocomponent pertussis toxoid vaccine performed in the G?teborg area of western Sweden (21, 22). In Sweden, there was no licensed pertussis vaccine between 1979 and 1996. Because a Swedish-made whole-cell vaccine was ineffective, pertussis experienced already recurred during the early 1970s, with a yearly incidence rate in preschool children of about 10% (9, 10). A total of 3,450 healthy infants were randomized to vaccination with diphtheria-tetanus toxoids with or without pertussis toxoid at 3, 5, and 12 months of age. There were 10,200 family members: 6,900 parents and 3,300 siblings. Families were enrolled in the study between September 1991 and June 1992. The Golgicide A study was completed in January 1995. The vaccine trial was approved by the institutional evaluate board of the National Institute of Child Health and Human Development; the Food and Drug Administration; the Medical Products Agency, Uppsala, Sweden; and the Ethics Committee, G?teborg University or college. All parents gave their written consent after receiving oral and written information. Follow-up of coughing episodes. Parents were instructed to contact the study nurse if anyone in the family coughed for 7 days. In addition a study nurse called Golgicide A each family once a month. Clinical and laboratory investigations were performed in the same way for study children and family members. A nasopharyngeal sample for culture and PCR and a serum sample for antibody determination were obtained. A convalescent serum sample was taken 4 weeks later. Information about exposure to pertussis within and outside the family, duration of the cough, paroxysms, whooping, vomiting, fever, rhinitis, and use of antibiotics was recorded. Laboratory assays. (i) Nasopharyngeal secretions. Nasopharyngeal secretions were collected by swabbing with a rayon swab (PurFybr Inc., Munster, Ind.) and transported to the laboratory in a altered Stuart medium. The same samples were utilized for culture and PCR. (ii) Culture. The swab was inoculated on Regan-Lowe medium (15) and then inserted into a tube containing enrichment medium. The enrichment medium experienced the same composition as the culture medium but with only half the concentration of charcoal agar (5, 6). The plates and the enrichment tubes were inoculated at 35 to 37C. A subculture from your enrichment medium was made after 72 h of incubation. The primary plates were inspected from the third through the seventh days after inoculation. The subcultures from your enrichment medium were incubated at 35 to 37C for at least 4 days. Colonies of and were verified by Gram staining, agglutination with specific antisera (Difco, Detroit, Mich.), and biochemical assessments (6). (iii) Pretreatment of the clinical samples for PCR. While the initial swab was put into the enrichment tube for culture, a new sterile swab (OTE Sjukv?rdsprodukter, Billdal, Sweden) was put into the transport tube to obtain material for PCR. The swab was tested to ensure that it did not inhibit the PCR (data not shown). The swab was kept in the transport medium for about 10 min before being inserted into an Eppendorf tube made up of 100 l of sterile, double-distilled water. After 5 min of incubation at space temperature, the pipe was shaken for 60 s as well as the swab was eliminated. The pipes were incubated inside a temperature stop at 95 to 100C for 10 min. The examples were kept at 4 to 8C until these were useful for PCR. If the examples needed to be kept for a CACN2 lot more than 48 h these were held frozen at.