Analysis of Streptococcus pneumoniae using Fourier-transformed infrared spectroscopy allows prediction of capsular serotype

Abstract

Determination of the capsule type of clinical isolates of Streptococcus pneumoniae is a prerequisite for epidemiological studies and further vaccine development. The Quellung reaction for serotyping is expensive and mostly done in reference centres. We wanted to evaluate whether Fourier-transformed infrared (FT-IR) spectroscopy is suitable for capsular type analysis and prediction of pneumococcal serotypes. We used the IR-Biotyper™ (Bruker) to create a database containing the spectra of 120 strains from invasive disease. The strains covered the 24 vaccine serotypes contained in the 13-valent conjugate vaccine (PCV13) and the 23-valent polysaccharide vaccine (PSV23). Hierarchical clustering analysis was performed. Finally, two different classification sets were created (PCV13 and PSV23). They were used to predict the serotype of 168 different challenge strains (invasive and non-invasive disease) covering 48 different serotypes (vaccine and non-vaccine types). FT-IR spectra from pneumococci (1300-800 cm^-1) clustered along their serotype as determined by the Quellung reaction (120 strains, 24 different serotypes). Strains with unknown serotype fell within the cluster of the correct serotype, as long as the latter was represented in the database (168 strains, 48 different serotypes). Concordance between the Quellung reaction and FT-IR spectroscopy was excellent (kappa ≥ 0.75). FT-IR spectroscopy is a fast and cost-effective method to predict the capsular serotype of pneumococci.

Keywords: Capsular typing; FT-IRS; PCV13; PSV23; Pneumococcus; Vaccine serotypes.

Fig. 1

Classification sets for typing/clustering of a 13 different serotypes (PCV13 classification set) and b 24 different serotypes (PSV23 classification set); strains cluster according to their serotype with the exception of serotype 6A and B; to improve discernibility within the tree, spaces between strains clustering together with a cut-off of less than 0.2 are filled with very light grey; individual strains are colour-coded according to their serotype: serotype 14: black, serotype 5: red, serotype 9V: ochre, serotype 7F: blue, serotype 4: light green, serotype 18C: dark brown, serotype 23F: magenta, serotype 6B: light orange, serotype 6A: yellow, serotype 19F: grey, serotype 19A: light purple, serotype 1: dark green, serotype 3: light blue, serotype 15B: white, serotype 10A: cobalt blue, serotype 33F: light grey, serotype 17F: light brown, serotype 9N: rose, serotype 11A: light blue, serotype 22F: light rose, serotype 20: neon green, serotype 12F: dark green, serotype 2: dark blue, serotype 8: orange

Fig. 2

Classification sets for typing/clustering (PCV13, PSV23 classification set) with challenge strains: challenge strains are pink; cut-off for match or no match: 0.2; serotype colour-codes: see legend Fig. 1. Shown are the decisive sections of the respective classification sets: a challenge strain (AL612616): serotype 1, matches with the serotype 1 cluster of the PCV13 classification set; b challenge strain (AL218499): serotype 11A, no match within the PCV13 classification set; c challenge strain (AL218499): serotype 11A, matches with the serotype 11A cluster of the PSV23 classification set; d challenge strain (AL211459): serotype 21, no match within the PSV23 classification set; e challenge strain (BL202254): serotype 15B, matches with the serotype 14 cluster of the PCV13 classification set; f challenge strain (BL202254): serotype 15B, matches with the serotype 15B (sub-)cluster of the PSV23 classification set

Fig. 3

Typing results of 168 challenge strains with PCV13 and PSV23 classification sets; pattern codes: according to serotypes of challenge strains: solid: serotypes of challenge strains are part of the respective classification set; checkerboard: serotypes of challenge strains are not part of the respective classification set; colour codes: according to level of typing agreement between the Quellung reaction and FT-IRS: green: typing result concordant to serotype level between methods; yellow: result concordant to serogroup level; red: discordant results