Pneumococcal Serotype Identification by Capsular Sequence Typing (CST): A Modified Novel Approach for Serotyping Directly in Clinical Samples.

Nektarios Marmaras,Efthymia Petinaki,Anastasia Papandreou,Georgina Tzanakaki,Maria Tsolia,Vana Papaevangelou,Athanasia Xirogianni

doi:10.3390/diagnostics11122353

Abstract

As almost 60–70% of Invasive Pneumococcal Disease (IPD) is identified by nonculture methods in Greece, serotyping is of high importance for the better monitoring of pneumococcal serotypes due to the availability of conjugate vaccines. The aim of the study was the modification and direct application of the Capsular Sequence Typing (CST) assay in clinical samples in order to serotype Streptococcus pneumoniae culture-negative, Polymerase Chain Reaction (PCR_-positive samples, followed by CST group specific single-tube PCR assays. A two-step PCR modified assay was applied on a total of 306 samples (such as CSF, blood, pleural and middle ear fluids, isolates) obtained from 283 patients with IPD. The overall performance permits a rapid, accurate and cost-effective method for nonculture pneumococcal serotyping. As the management of IPD is closely related to the continuous monitoring of pneumococcal serotypes, the proposed approach proved to be a valuable tool for the typing and epidemiological monitoring of S. pneumoniae, for the evaluation of the overall impact of vaccination programs in the era of pneumococcal conjugate vaccines, in order to initiate the appropriate vaccination strategy.

Highlights

The present study describes a modification of the Capsular Sequence Typing (CST) assay in a two-step Polymerase Chain Reaction (PCR) protocol and its direct application on S. pneumoniae culture-negative/PCR-positive clinical samples (CSF, whole blood, pleural fluid, middle ear fluid), which were obtained from patients with
The CST was shown to be well-performed in all clinical samples (n = 239) and bacterial isolates (n = 67) identified previously by PCR as S. pneumoniae
The specificity was estimated at 100% and evaluated from the 67 S. pneumoniae isolates for which the serotype was identified by the Quellung reaction (PPV 100% and NPV 100%)

Summary

Introduction

Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. Streptococcus pneumoniae is a major cause of serious bacterial infections including meningitis, septicemia and pneumonia and is associated with significant morbidity and mortality worldwide. The ability of S. pneumoniae to cause disease is directly related to the capsule production, a polysaccharide structure external to the cell wall that provides resistance to phagocytosis and promotes the invasion of the host immune system by the bacteria [2]. Approximately 100 different pneumococcal serotypes have been identified, based on the unique antigen structure of the capsular polysaccharide, each with their own characteristics, adaptability for nasopharyngeal carriage and potential for invasive disease [3]

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