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  • 1
    Online Resource
    Online Resource
    Table_1.docx
    Frontiers Media SA ; 2023
    In:  Frontiers in Microbiology Vol. 14 ( 2023-4-17)
    Details
    In: Frontiers in Microbiology, Frontiers Media SA, Vol. 14 ( 2023-4-17)
    Abstract: Despite strong historical records on the accuracy of saliva testing, oral fluids are considered poorly suited for pneumococcal carriage detection. We evaluated an approach for carriage surveillance and vaccine studies that increases the sensitivity and specificity of pneumococcus and pneumococcal serotype detection in saliva samples. Methods Quantitative PCR (qPCR)-based methods were applied to detect pneumococcus and pneumococcal serotypes in 971 saliva samples collected from 653 toddlers and 318 adults. Results were compared with culture-based and qPCR-based detection in nasopharyngeal samples collected from children and in nasopharyngeal and oropharyngeal samples collected from adults. Optimal C q cut-offs for positivity in qPCRs were determined via receiver operating characteristic curve analysis and accuracy of different approaches was assessed using a composite reference for pneumococcal and for serotype carriage based on isolation of live pneumococcus from the person or positivity of saliva samples determined with qPCR. To evaluate the inter-laboratory reproducibility of the method, 229 culture-enriched samples were tested independently in the second center. Results In total, 51.5% of saliva samples from children and 31.8% of saliva samples from adults were positive for pneumococcus. Detection of pneumococcus by qPCR in culture-enriched saliva exhibited enhanced sensitivity and higher agreement with a composite reference compared to diagnostic culture of nasopharyngeal samples in children (Cohen’s κ: 0.69–0.79 vs. 0.61–0.73) and in adults (κ: 0.84–0.95 vs. 0.04–0.33) and culture of oropharyngeal samples in adults (κ: 0.84–0.95 vs. −0.12–0.19). Similarly, detection of serotypes with qPCR in culture-enriched saliva exhibited enhanced sensitivity and higher agreement with a composite reference compared to nasopharyngeal culture in children (κ: 0.73–0.82 vs. 0.61–0.73) and adults (κ: 0.90–0.96 vs. 0.00–0.30) and oropharyngeal culture in adults (κ: 0.90–0.96 vs. −0.13 to 0.30). However, results of qPCRs targeting serotype 4, 5, and 17F and serogroups 9, 12, and 35 were excluded due to assays’ lack of specificity. We observed excellent quantitative agreement for qPCR-based detection of pneumococcus between laboratories. After exclusion of serotype/serogroup-specific assays with insufficient specificity, moderate agreement (κ 0.68, 95% CI 0.58–0.77) was observed. Conclusion Molecular testing of culture-enriched saliva samples improves the sensitivity of overall surveillance of pneumococcal carriage in children and adults, but limitations of qPCR-based approaches for pneumococcal serotypes carriage detection should be considered.
    Type of Medium: Online Resource
    ISSN: 1664-302X
    URL: Article
    URL: Article
    URL: Article
    URL: Article
    DOI: 10.3389/fmicb.2023.1156695
    DOI: 10.3389/fmicb.2023.1156695.s001
    DOI: 10.3389/fmicb.2023.1156695.s002
    DOI: 10.3389/fmicb.2023.1156695.s003
    Language: Unknown
    Publisher: Frontiers Media SA
    Publication Date: 2023
    detail.hit.zdb_id: 2587354-4
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  • 2
    Online Resource
    Online Resource
    Table_7.docx
    Frontiers Media SA ; 2022
    In:  Frontiers in Microbiology Vol. 13 ( 2022-4-18)
    Details
    In: Frontiers in Microbiology, Frontiers Media SA, Vol. 13 ( 2022-4-18)
    Abstract: The specificity of molecular methods for the detection of Streptococcus pneumoniae carriage is under debate. We propose a procedure for carriage surveillance and vaccine impact studies that increases the accuracy of molecular detection of live pneumococci in polymicrobial respiratory samples. Methods Culture and qPCR methods were applied to detect pneumococcus and pneumococcal serotypes in 1,549 nasopharyngeal samples collected in the Netherlands ( n = 972) and England ( n = 577) from 946 toddlers and 603 adults, and in paired oropharyngeal samples collected exclusively from 319 Dutch adults. Samples with no live pneumococci isolated at primary diagnostic culture yet generating signal specific for pneumococcus in qPCRs were re-examined with a second, qPCR-guided culture. Optimal C q cut-offs for positivity in qPCRs were determined via receiver operating characteristic (ROC) curve analysis using isolation of live pneumococci from the primary and qPCR-guided cultures as reference. Results Detection of pneumococcus and pneumococcal serotypes with qPCRs in cultured (culture-enriched) nasopharyngeal samples exhibited near-perfect agreement with conventional culture (Cohen’s kappa: 0.95). Molecular methods displayed increased sensitivity of detection for multiple serotype carriage, and implementation of qPCR-guided culturing significantly increased the proportion of nasopharyngeal and oropharyngeal samples from which live pneumococcus was recovered ( p & lt; 0.0001). For paired nasopharyngeal and oropharyngeal samples from adults none of the methods applied to a single sample type exhibited good agreement with results for primary and qPCR-guided nasopharyngeal and oropharyngeal cultures combined (Cohens kappa; 0.13–0.55). However, molecular detection of pneumococcus displayed increased sensitivity with culture-enriched oropharyngeal samples when compared with either nasopharyngeal or oropharyngeal primary cultures ( p & lt; 0.05). Conclusion The accuracy of pneumococcal carriage surveillance can be greatly improved by complementing conventional culture with qPCR and vice versa , by using results of conventional and qPCR-guided cultures to interpret qPCR data. The specificity of molecular methods for the detection of live pneumococci can be enhanced by incorporating statistical procedures based on ROC curve analysis. The procedure we propose for future carriage surveillance and vaccine impact studies improves detection of pneumococcal carriage in adults in particular and enhances the specificity of serotype carriage detection.
    Type of Medium: Online Resource
    ISSN: 1664-302X
    URL: Article
    URL: Article
    URL: Article
    URL: Article
    URL: Article
    URL: Article
    URL: Article
    URL: Article
    URL: Article
    URL: Article
    DOI: 10.3389/fmicb.2022.859736
    DOI: 10.3389/fmicb.2022.859736.s001
    DOI: 10.3389/fmicb.2022.859736.s002
    DOI: 10.3389/fmicb.2022.859736.s003
    DOI: 10.3389/fmicb.2022.859736.s004
    DOI: 10.3389/fmicb.2022.859736.s005
    DOI: 10.3389/fmicb.2022.859736.s006
    DOI: 10.3389/fmicb.2022.859736.s007
    DOI: 10.3389/fmicb.2022.859736.s008
    DOI: 10.3389/fmicb.2022.859736.s009
    Language: Unknown
    Publisher: Frontiers Media SA
    Publication Date: 2022
    detail.hit.zdb_id: 2587354-4
    Location Call Number Limitation Availability
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