Dissertation committee:
Μαρία Τσολιά, Καθηγήτρια, Ιατρική Σχολή, ΕΚΠΑ
Βασιλική Παπαευαγγέλου, Καθηγήτρια, Ιατρική Σχολή, ΕΚΠΑ
Ευθυμία Πετεινάκη, Καθηγήτρια, Ιατρική Σχολή, Πανεπιστήμιο Θεσσαλίας
Γεωργία Τζανακάκη, Καθηγήτρια, Σχολή Δημόσιας Υγείας, Πανεπιστήμιο Δυτικής Αττικής
Αθανάσιος Μίχος, Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Βασιλική Σπούλου, Καθηγήτρια, Ιατρική Σχολή, ΕΚΠΑ
Αλεξάνδρα Σολδάτου, Αν. Καθηγήτρια, Ιατρική Σχολή, ΕΚΠΑ
Summary:
Streptococcus pneumoniae (pneumococcus) is one of the most frequent and common pathogens. A total of 1.6 million people worldwide die from pneumococcal diseases each year, of which approximately 700,000-1,000,000 are children under the age of 5. Children under the age of 5, the elderly, and those with an immune deficiency are considered to be at higher risk for pneumococcal infections.
Streptococcus pneumoniae is responsible for a variety of infections: invasive, such as bacteremia, septicemia, meningitis, and the more common and clinically milder non-invasive, acute otitis media, sinusitis, and pharyngotonsillitis.
The virulence of the microorganism is attributed to the antigenic structure of the polysaccharides of its capsule, on the basis of which it is divided into approximately 100 different serotypes.
The increased frequency and the severity of invasive pneumococcal diseases are the main reasons why the epidemiological surveillance of pathogenic serotypes is considered imperative. Vaccination is important not only for prevention, but also due to the ever-increasing resistance to penicillin. The 13-valent pneumococcal conjugate vaccine (PCV-13) includes the serotypes of the 7-valent vaccine (4, 9V, 19F, 14, 23F, 18C, 6B) and additionally serotypes 1, 5, 7, 19A, 3, 6A.
These vaccines cover only a few serotypes and are unable to protect against serotypes not present in the vaccine or strains that do not express capsular antigen. This has led to a rapid increase in non-vaccine serotypes, some of which are resistant to antibiotics. Therefore, there is an urgent need to develop new and effective pneumococcal vaccines that could cover a wide range of serotypes.
Valid and timely diagnosis and characterization of the pneumococcus in terms of it’s serotypes is of critical importance. Tools for identifying the microorganism are classical culture, biochemical methods (optochin sensitivity test and solubility in bile salts, etc.) and serotyping. In recent years molecular techniques have brought about a real revolution as it becomes possible to detect the microorganism. The application of molecular methods (PCR) is crucial in the successful identification of the microorganisms causing infections in a significant proportion of culture-negative clinical samples attributed also to previous antibiotic treatment. Accurate diagnosis and identification of the pathogen is important for patient management and targeted antimicrobial therapy.
The aim of the present study was, to create a technique based on the methods of polymerase chain reaction (PCR) and Sequencing, which has the possibility of detecting the serotypes of the microorganism Streptococcus pneumoniae from isolates as well as from biological materials – where the culture and isolation of the microorganism is not easy, the epidemiological investigation of the Streptococcus pneumoniae serotypes that cause the various infections and whether these serotypes have changed over the years.
A new method - CST, formulated in two reaction steps, was developed and shown to have excellent performance on a wide range of applied biological materials.
With the new method, the percentage of samples that a serotype can be attributed, rose to 92.8% from about 50% before the implementation of the new method. Also, with the completion of the serotyping of the non-typed samples, it became possible to retrospectively study Streptococcus pneumoniae serotypes in Greece for a long period of time, from 2010, after the introduction of the PCV-13 vaccine, until 2020.
Keywords:
Streptococcus pneumoniae, Capsular Sequence Typing, Surveillance, Serotypes, Vaccines
