title: Development of a TaqMan Array Card for Pneumococcal Serotyping on Isolates and Nasopharyngeal Samples.
abstract: Streptococcus pneumoniae is both a commensal and a major pathogen that causes invasive disease in people of all ages. The introduction of serotype-specific pneumococcal vaccines has reduced the burden of disease but has also led to replacement with new strains; thus, serotyping remains important for vaccine-related disease surveillance. Conventional serotyping methods are laborious and expensive. We developed an easy-to-perform genotypic TaqMan array card (TAC) to identify S. pneumoniae strains, including lytA-based sequences, and 53 sequence-specific PCRs to identify 74 serotypes/serogroups covering all current vaccine types as well as prevalent nonvaccine types. The TAC method was evaluated on 146 clinical S. pneumoniae isolates and 13 nonpneumococcal species that naturally inhabit the upper respiratory tract and yielded 97% (142/146) sensitivity and 100% (13/13) specificity versus results of standard Quellung serotyping. The calculated limit of detection was 20 to 200 fg (∼8 to 84 genome equivalents) per reaction. On 23 blinded nasopharyngeal specimens that were pneumococcus culture positive, the TAC pan-pneumococcus lytA assay was positive in 21 (91% sensitivity versus culture). On TAC lytA-positive specimens, a serotype result was obtained on 86%, and the result was 95% accurate versus the subsequent culture's Quellung result. TAC also detected mixed serotypes in two specimens where Quellung detected only the predominant serotype. This TAC method yields fast and comprehensive serotyping compared to the standard method and may be useful on direct specimens.
authors.affiliation: Division of Infectious Diseases and International Health, Department of Medicine, University of Virginia, Charlottesville, Virginia, USA.
authors.forename: Suporn
authors.initials: S
authors.lastname: Pholwat
authors.affiliation: Hubert Department of Global Health, Rollins School of Public Health, Emory University, Atlanta, Georgia, USA.
authors.forename: Fuminori
authors.initials: F
authors.lastname: Sakai
authors.affiliation: Cambodia Oxford Medical Research Unit, Angkor Hospital for Children, Siem Reap, Cambodia.|Centre for Tropical Medicine and Global Health, Nuffield Department of Medicine, University of Oxford, Oxford, United Kingdom.
authors.forename: Paul
authors.initials: P
authors.lastname: Turner
authors.affiliation: Hubert Department of Global Health, Rollins School of Public Health, Emory University, Atlanta, Georgia, USA.
authors.forename: Jorge E
authors.initials: JE
authors.lastname: Vidal
authors.affiliation: Division of Infectious Diseases and International Health, Department of Medicine, University of Virginia, Charlottesville, Virginia, USA erh6k@virginia.edu.
authors.forename: Eric R
authors.initials: ER
authors.lastname: Houpt
mesh_terms: D002353:Carrier State / Q000382:microbiology*; D002648:Child; D002675:Child, Preschool; D006801:Humans; D046228:Microarray Analysis / Q000379:methods; D058889:Molecular Typing / Q000379:methods*; D009305:Nasopharynx / Q000382:microbiology*; D011008:Pneumococcal Infections / Q000382:microbiology*; D012680:Sensitivity and Specificity; D012703:Serotyping / Q000379:methods*; D013296:Streptococcus pneumoniae / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification