It Takes Two to Tango: Combining Conventional Culture With Molecular Diagnostics Enhances Accuracy of Streptococcus pneumoniae Detection and Pneumococcal Serogroup/Serotype Determination in Carriage
BackgroundThe 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 sampl...
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2022-04-01
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| Series: | Frontiers in Microbiology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fmicb.2022.859736/full |
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| author | Willem R. Miellet Willem R. Miellet Janieke van Veldhuizen David Litt Rob Mariman Alienke J. Wijmenga-Monsuur Paul Badoux Tessa Nieuwenhuijsen Rebecca Thombre Sanaa Mayet Seyi Eletu Carmen Sheppard Marianne Alice van Houten Nynke Y. Rots Elizabeth Miller Norman K. Fry Norman K. Fry Elisabeth A. M. Sanders Elisabeth A. M. Sanders Krzysztof Trzciński |
| author_facet | Willem R. Miellet Willem R. Miellet Janieke van Veldhuizen David Litt Rob Mariman Alienke J. Wijmenga-Monsuur Paul Badoux Tessa Nieuwenhuijsen Rebecca Thombre Sanaa Mayet Seyi Eletu Carmen Sheppard Marianne Alice van Houten Nynke Y. Rots Elizabeth Miller Norman K. Fry Norman K. Fry Elisabeth A. M. Sanders Elisabeth A. M. Sanders Krzysztof Trzciński |
| author_sort | Willem R. Miellet |
| collection | DOAJ |
| description | BackgroundThe 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.MethodsCulture 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 Cq 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.ResultsDetection 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 < 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 < 0.05).ConclusionThe 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. |
| first_indexed | 2024-04-14T07:22:41Z |
| format | Article |
| id | doaj.art-87b6911c6e95436e94f1182f4ddfd734 |
| institution | Directory Open Access Journal |
| issn | 1664-302X |
| language | English |
| last_indexed | 2024-04-14T07:22:41Z |
| publishDate | 2022-04-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Microbiology |
| spelling | doaj.art-87b6911c6e95436e94f1182f4ddfd7342022-12-22T02:06:06ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2022-04-011310.3389/fmicb.2022.859736859736It Takes Two to Tango: Combining Conventional Culture With Molecular Diagnostics Enhances Accuracy of Streptococcus pneumoniae Detection and Pneumococcal Serogroup/Serotype Determination in CarriageWillem R. Miellet0Willem R. Miellet1Janieke van Veldhuizen2David Litt3Rob Mariman4Alienke J. Wijmenga-Monsuur5Paul Badoux6Tessa Nieuwenhuijsen7Rebecca Thombre8Sanaa Mayet9Seyi Eletu10Carmen Sheppard11Marianne Alice van Houten12Nynke Y. Rots13Elizabeth Miller14Norman K. Fry15Norman K. Fry16Elisabeth A. M. Sanders17Elisabeth A. M. Sanders18Krzysztof Trzciński19Department of Pediatric Immunology and Infectious Diseases, Wilhelmina Children’s Hospital, University Medical Center Utrecht (UMCU), Utrecht, NetherlandsCentre for Infectious Disease Control, National Institute for Public Health and the Environment (RIVM), Bilthoven, NetherlandsCentre for Infectious Disease Control, National Institute for Public Health and the Environment (RIVM), Bilthoven, NetherlandsRespiratory and Vaccine Preventable Bacterial Reference Unit (RVPBRU), Public Health England – National Infection Service, London, United KingdomCentre for Infectious Disease Control, National Institute for Public Health and the Environment (RIVM), Bilthoven, NetherlandsCentre for Infectious Disease Control, National Institute for Public Health and the Environment (RIVM), Bilthoven, NetherlandsRegional Laboratory of Public Health (Streeklab) Haarlem, Haarlem, NetherlandsDepartment of Pediatric Immunology and Infectious Diseases, Wilhelmina Children’s Hospital, University Medical Center Utrecht (UMCU), Utrecht, NetherlandsRespiratory and Vaccine Preventable Bacterial Reference Unit (RVPBRU), Public Health England – National Infection Service, London, United KingdomRespiratory and Vaccine Preventable Bacterial Reference Unit (RVPBRU), Public Health England – National Infection Service, London, United KingdomRespiratory and Vaccine Preventable Bacterial Reference Unit (RVPBRU), Public Health England – National Infection Service, London, United KingdomRespiratory and Vaccine Preventable Bacterial Reference Unit (RVPBRU), Public Health England – National Infection Service, London, United KingdomSpaarne Gasthuis, Haarlem, NetherlandsCentre for Infectious Disease Control, National Institute for Public Health and the Environment (RIVM), Bilthoven, NetherlandsImmunisation and Countermeasures Division, Public Health England (PHE) – National Infection Service, London, United KingdomRespiratory and Vaccine Preventable Bacterial Reference Unit (RVPBRU), Public Health England – National Infection Service, London, United KingdomImmunisation and Countermeasures Division, Public Health England (PHE) – National Infection Service, London, United KingdomDepartment of Pediatric Immunology and Infectious Diseases, Wilhelmina Children’s Hospital, University Medical Center Utrecht (UMCU), Utrecht, NetherlandsCentre for Infectious Disease Control, National Institute for Public Health and the Environment (RIVM), Bilthoven, NetherlandsDepartment of Pediatric Immunology and Infectious Diseases, Wilhelmina Children’s Hospital, University Medical Center Utrecht (UMCU), Utrecht, NetherlandsBackgroundThe 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.MethodsCulture 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 Cq 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.ResultsDetection 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 < 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 < 0.05).ConclusionThe 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.https://www.frontiersin.org/articles/10.3389/fmicb.2022.859736/fullStreptococcus pneumoniae (pneumococcus)carriagediagnostic accuracyqPCR (quantitative PCR)conventional culture |
| spellingShingle | Willem R. Miellet Willem R. Miellet Janieke van Veldhuizen David Litt Rob Mariman Alienke J. Wijmenga-Monsuur Paul Badoux Tessa Nieuwenhuijsen Rebecca Thombre Sanaa Mayet Seyi Eletu Carmen Sheppard Marianne Alice van Houten Nynke Y. Rots Elizabeth Miller Norman K. Fry Norman K. Fry Elisabeth A. M. Sanders Elisabeth A. M. Sanders Krzysztof Trzciński It Takes Two to Tango: Combining Conventional Culture With Molecular Diagnostics Enhances Accuracy of Streptococcus pneumoniae Detection and Pneumococcal Serogroup/Serotype Determination in Carriage Frontiers in Microbiology Streptococcus pneumoniae (pneumococcus) carriage diagnostic accuracy qPCR (quantitative PCR) conventional culture |
| title | It Takes Two to Tango: Combining Conventional Culture With Molecular Diagnostics Enhances Accuracy of Streptococcus pneumoniae Detection and Pneumococcal Serogroup/Serotype Determination in Carriage |
| title_full | It Takes Two to Tango: Combining Conventional Culture With Molecular Diagnostics Enhances Accuracy of Streptococcus pneumoniae Detection and Pneumococcal Serogroup/Serotype Determination in Carriage |
| title_fullStr | It Takes Two to Tango: Combining Conventional Culture With Molecular Diagnostics Enhances Accuracy of Streptococcus pneumoniae Detection and Pneumococcal Serogroup/Serotype Determination in Carriage |
| title_full_unstemmed | It Takes Two to Tango: Combining Conventional Culture With Molecular Diagnostics Enhances Accuracy of Streptococcus pneumoniae Detection and Pneumococcal Serogroup/Serotype Determination in Carriage |
| title_short | It Takes Two to Tango: Combining Conventional Culture With Molecular Diagnostics Enhances Accuracy of Streptococcus pneumoniae Detection and Pneumococcal Serogroup/Serotype Determination in Carriage |
| title_sort | it takes two to tango combining conventional culture with molecular diagnostics enhances accuracy of streptococcus pneumoniae detection and pneumococcal serogroup serotype determination in carriage |
| topic | Streptococcus pneumoniae (pneumococcus) carriage diagnostic accuracy qPCR (quantitative PCR) conventional culture |
| url | https://www.frontiersin.org/articles/10.3389/fmicb.2022.859736/full |
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