Abstract
BackgroundNasopharyngeal colonization is considered a necessary step in the initiation of pneumococcal diseases. Real time PCR (RT-PCR) is an alternative approach for the identification and quantification of pneumococci directly from samples.ObjectivesTo compare pneumococcal detection rates using culture-based method versus RT-PCR direct detection and to quantify pneumococcal colonization in two study cohorts (healthy children and hospitalized children with respiratory symptoms) using quantitation through RT-PCR.MethodologyA total of 101 nasopharyngeal swabs (NPS) from healthy children and 183 NPSs from hospitalized children with respiratory symptoms were included in the study. None of the children were vaccinated. All children were between 2 months to 2 years. In parallel to routine culture and identification, a RT-PCR assay targeting the lytA gene was done.ResultsConsidering all 284 samples tested, colonization rate by conventional culture was 41.2% (n = 117) while positive colonization using RT-PCR was 43.7% (n = 124). The colonization rate detected by RT-PCR in the healthy cohort was 33.7% (n = 34) and it was 49.2% (n = 90) in the hospitalized cohort. It was 37.6% (n = 38) and 43.2% (n = 79) for the two cohorts by culture. The mean Cq value for the healthy cohort is 29.61 (SD 2.85) and 28.93 (SD 3.62) for the hospitalized cohort.With the standard curve obtained from amplifying a dilution series of control DNA, the mean amount of genomic DNA copy numbers detected in children with respiratory symptoms was log10 7.49 (SD 1.07) while it was log10 7.30 (SD 0.23) in healthy children and the difference was not statistically significant.ConclusionsThe overall colonization rate was higher when detected using RT-PCR compared to culture. However, it was lower in the healthy group when detected with RT-PCR compared to culture. Even though there was a higher detection of pneumococcal colonization density in children with respiratory symptoms, this was not significantly higher unlike many previous studies. Therefore, the use of RT-PCR to detect pneumococcal colonization needs further evaluation with careful analysis of interpretation and confounders.
Highlights
Streptococcus pneumoniae or pneumococcus is a pathogenic bacterium which can cause a variety of diseases ranging from non-invasive diseases such as otitis media, sinusitis to invasive diseases including pneumonia, meningitis, and sepsis
With the standard curve obtained from amplifying a dilution series of control DNA, the mean amount of genomic DNA copy numbers detected in children with respiratory symptoms was log10 7.49 (SD 1.07) while it was log10 7.30 (SD 0.23) in healthy children and the difference was not statistically significant
The overall colonization rate was higher when detected using Real Time Polymerase Chain Reaction (RT-PCR) compared to culture
Summary
Streptococcus pneumoniae or pneumococcus is a pathogenic bacterium which can cause a variety of diseases ranging from non-invasive diseases such as otitis media, sinusitis to invasive diseases including pneumonia, meningitis, and sepsis. According to the World Health Organization (WHO), pneumococcal diseases were responsible for an estimated 317,300 pediatric deaths in 2015 [1]. Nasopharyngeal colonization of pneumococci has been identified as a prerequisite for the initiation pneumococcal disease and transmission of pneumococci [2]. Pneumococcal colonization rates differ globally according with socioeconomic factors and age [3]. It is highest among children below the age of 5 years [4]. In this age group too, it is found to vary between countries [4,5,6,7,8]. Nasopharyngeal colonization is considered a necessary step in the initiation of pneumococcal diseases. Real time PCR (RT-PCR) is an alternative approach for the identification and quantification of pneumococci directly from samples
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