PCR and Culture Analysis of Streptococcus pneumoniae Nasopharyngeal Carriage in Healthy Children

Abstract

Invasive Streptococcus pneumoniae disease is preceded by asymptomatic nasopharyngeal carriage. Measuring carriage in healthy populations provides data on what serotypes are present in communities, which is of interest in the era of polyvalent pneumococcal conjugate vaccines. Nasopharyngeal swabs from a survey of 682 and 800 healthy children in 2016 and 2018, respectively, were analyzed by culture and Quellung reaction to determine rates of carriage and serotypes. All swabs from 2016 and 300 randomly selected swabs from 2018 were then analyzed using real-time semi-quantitative PCR (qPCR) to detect S. pneumoniae gene targets lytA, piaA, and SP2020 and determine serotype. There were 71 (10.4%) and 68 (8.5%) culture positive samples in 2016 and 2018, respectively. All of these were also positive by qPCR except one that was equivocal. In total, 46.0% of 2016 swabs were positive by qPCR. In 2018, results from the selected sample extrapolated to the complete sample showed 49.0% positive by qPCR. PCV13 serotypes were detected in 29.3% and 21.7% of S. pneumoniae qPCR positive samples from 2016 and 2018, respectively; compared with only 8.4% and 6.0% PCV13 serotypes detected by Quellung reaction in culture positive samples. Compared with culture, qPCR detected S. pneumoniae more frequently. Further, qPCR serotyping detected PCV13 serotypes in a larger proportion of samples than culture and Quellung reaction did, showing that, despite established universal childhood PCV13 immunization, vaccine serotypes can still be detected in a large proportion of young children.

Association of Nonpharmaceutical Interventions During the COVID-19 Pandemic With Invasive Pneumococcal Disease, Pneumococcal Carriage, and Respiratory Viral Infections Among Children in France

“…Sixth, the carriage study used only standard culture methods to identify pneumococci, but PCR tests may be more sensitive. 45 However, we used the same microbiological methods over the study period. Seventh, we cannot exclude the possibility that a fraction of the decrease in IPD incidence may be associated with an avoidance of the pediatric emergency department or a change in criteria for hospital admission.…”

Trends in Asymptomatic Nasopharyngeal Streptococcus pneumoniae Carriage with qPCR and Culture Analysis

“…The detailed methods for nasopharyngeal sample collection, culture detection and Quellung serotyping, as well as qPCR detection, using lytA , piaA , and SP2020 primers (and considered positive if two primers were positive), and qPCR serotyping have been published previously [ 22 ], and results have been reutilized for this study. Of note regarding serotyping, isolates that could not be serotyped by Quellung reaction are referred to as non-typeable, whereas strains that did not match one of the thirty-seven serotypes detected by qPCR are referred to as not-typed due to the limitations of the assay.…”

“…For the not-typed samples, none of the 37 serotypes in the qPCR serotyping assay were identified, but it was not known if other defined serotypes were present. However, although the qPCR assay detects only 37 serotypes, these include all PCV13 vaccine serotypes and almost all common and uncommon serotypes causing pneumococcal disease [ 22 ]. For further analysis, NT and Not-Typed samples were combined as one group.…”

“…For further analysis, NT and Not-Typed samples were combined as one group. As previously reported, there were 682 children who participated in the 2016 survey [ 22 ], of which 679 were included in our study. Two were excluded as they each received at least one dose of the PCV7 vaccine.…”

“…However, molecular analysis with various polymerase chain reaction (qPCR) techniques can more frequently and rapidly detect serotype positive samples compared to standard culture methods [ 15 , 16 ] and these methods consistently detect higher levels of pneumococcal carriage compared to culture methods [ 17 , 18 , 19 , 20 , 21 ]. We have previously reported a higher detection rate for S. pneumoniae through qPCR methods compared with standard culture, as well as a higher proportion of samples carrying vaccine-serotype strains, demonstrating that vaccine serotypes can still be detected in a considerable proportion of children in the late post-PCV13 era [ 22 ].…”