Epitope analyses of pneumococcal surface protein A: a combination of two monoclonal antibodies detects 94% of clinical isolates

Abstract

Immunisation of BALB/c mice with seven heat-treated Norwegian clinical isolates of Streptococcus pneumoniae of different serotypes elicited mainly monoclonal antibodies (mAbs) to pneumococcal surface protein A (PspA). It was remarkable that the fusions resulted only in a few mAbs directed against other protein antigens. Dot blot analysis with 16 mAbs using clinical isolates representing 23 different capsular types and the uncapsulated reference strain R36A showed that some of the mAbs bound to PspA epitopes expressed by a low number of strains whereas others bound to broadly distributed epitopes. On the basis of their reactivities, seven of these mAbs could be divided into two groups recognising different subsets of pneumococci. The three mAbs in the narrow reacting group bound to epitopes found in 21-25% of the strains whereas the four mAbs in the broad reacting group detected more than 57% of the analysed strains. The epitopes for these seven antibodies were surface exposed on live exponential phase grown pneumococci as shown by flow cytometry. The finding that a combination of mAb 180,C-1 (IgG2a) from the first group and mAb 170,E-11 (IgG2a) from the second group detected 94% of the examined strains is interesting because PspA has been reported by others to be a serological highly variable protein.

Epitope mapping of pneumococcal surface protein A of strain Rx1 using monoclonal antibodies and molecular structure modelling

“…None of the six PspA/Rx1 reactive mAbs were dot blot positive against the recently genomic sequenced Norwegian clinical isolate TIGR4, which belongs to family 2 [33]. Epitope differences between PspA families 1 and 2 were further supported by the finding that the three PspA specific mAbs (143,F‐2; 149,B‐3; 180,C‐1) which showed positive flow cytometric reactions with strain 7/87 (TIGR4) [16] were non‐reactive with R6. We have previously reported that a combination of two PspA specific mAbs detected 94% of the examined Norwegian strains [16].…”

“…The mAbs were tested for binding to live pneumococci using flow cytometry as described previously [16]. Briefly, the bacteria were grown on horse blood agar overnight, then in Todd–Hewitt broth for the next night.…”

“…We have recently reported the properties of 16 PspA specific mAbs prepared against seven different clinical Norwegian isolates [16]. Also, the structural properties of the N‐terminal, helical, coiled‐coil, and functional module of PspA were investigated by us for the Rx1 and other strains [5,10].…”

Towards Identifying Protective B-Cell Epitopes: The PspA Story

“…The active immunization of PspA from family 2 containing α-helical domain and proline-rich region provided significant cross protection when challenged with the strains from PspA families 1 and 2, while immunization with the α-helical domain of family 2 PspA alone was found involved in providing family specific protection (Moreno et al, 2010; Kothari et al, 2015). Using monoclonal antibodies, Kolberg et al (2001) found that individual mAb cross-reacted with 20–50% strains and in combination could recognize about 94% of the strains analyzed (Kolberg et al, 2001). With higher cross reactivity, antibodies generated against PspA held greater possibility of providing cross-protection against varied pneumococcal strains (Tart et al, 1996; Briles et al, 2000; Andre et al, 2015; Kristian et al, 2016).…”

“…It was found that all epitopes recognized by these mAbs were surface accessible (Kolberg et al, 2001). Surprisingly, a combination of the two anti-PspA mAbs detected 94% of the 77 strains analyzed.…”