Determination of avidity of IgG against protein antigens from Streptococcus pneumoniae: assay development and preliminary application in clinical settings
The measurement of antibody levels is a common test for the diagnosis of Streptococcus pneumoniae infection in research. However, the quality of antibody response, reflected by avidity, has not been adequately evaluated. We aimed to evaluate the role of avidity of IgG against eight pneumococcal proteins in etiologic diagnosis. Eight pneumococcal proteins (Ply, CbpA, PspA1 and 2, PcpA, PhtD, StkP-C, and PcsB-N) were used to develop a multiplex bead-based avidity immunoassay. The assay was tested for effects of the chaotropic agent, multiplexing, and repeatability. The developed assay was applied to paired samples from children with or without pneumococcal disease (n = 38 for each group), determined by either serology, polymerase chain reaction (PCR), or blood culture. We found a good correlation between singleplex and multiplex assays, with r ≥ 0.94.The assay was reproducible, with mean inter-assay variation ≤ 9% and intra-assay variation < 6%. Children with pneumococcal disease had lower median avidity indexes in the acute phase of disease for PspA1 and 2 (p = 0.042), PcpA (p = 0.002), PhtD (p = 0.014), and StkP-C (p < 0.001). When the use of IgG avidity as a diagnostic tool for pneumococcal infection was evaluated, the highest discriminative power was found for StkP-C, followed by PcpA (area under the curve [95% confidence interval, CI]: 0.868 [0.759–0.977] and 0.743 [0.607–879], respectively). The developed assay was robust and had no deleterious influence from multiplexing. Children with pneumococcal disease had lower median avidity against five pneumococcal proteins in the acute phase of disease compared to children without disease.
We thank Sanofi Pasteur (Lyon, France) for supplying PcpA and PhtD; Prof. Elaine Tuomanen at St. Jude Children’s Research Hospital (Memphis, TN, USA) for supplying Ply, CbpA, and PspA1; Profs. Susan Hollingshead, David Briles, and Pat Coan at University of Alabama (Birmingham, AL, USA) for supplying PspA2; and Valneva Austria GmbH (Vienna, Austria) for supplying StkP-C and PcsB-N.
Data availability statement
The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.
This work was supported by: Bahia State Agency for Research Funding (FAPESB), Brazil; Brazilian Council for Scientific and Technological Development (CNPq), Brazil; Turku University Hospital Research Foundation, Finland; Rauno and Anne Puolimatka Foundation, Finland; Sohlberg Foundation, Finland.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
The use of the samples was approved by the Ethics Committee of the Federal University of Bahia in Brazil, the Ethics Committee of the National Institute for Health and Welfare in Finland (formerly National Public Health Institute), and the Ethics Committee of Satakunta Central Hospital, Pori, Finland.
Written informed consent was obtained from legal guardians before recruitment.
- 1.O’Brien KL, Wolfson LJ, Watt JP, Henkle E, Deloria-Knoll M, McCall N, Lee E, Mulholland K, Levine OS, Cherian T; Hib and Pneumococcal Global Burden of Disease Study Team (2009) Burden of disease caused by Streptococcus pneumoniae in children younger than 5 years: global estimates. Lancet 374:893–902CrossRefPubMedGoogle Scholar
- 3.Ekström N, Ahman H, Palmu A, Grönholm S, Kilpi T, Käyhty H; FinOM Study Group (2013) Concentration and high avidity of pneumococcal antibodies persist at least 4 years after immunization with pneumococcal conjugate vaccine in infancy. Clin Vaccine Immunol 20:1034–1040CrossRefPubMedPubMedCentralGoogle Scholar
- 13.Adamou JE, Heinrichs JH, Erwin AL, Walsh W, Gayle T, Dormitzer M, Dagan R, Brewah YA, Barren P, Lathigra R, Langermann S, Koenig S, Johnson S (2001) Identification and characterization of a novel family of pneumococcal proteins that are protective against sepsis. Infect Immun 69:949–958CrossRefPubMedPubMedCentralGoogle Scholar
- 14.Giefing C, Meinke AL, Hanner M, Henics T, Bui MD, Gelbmann D, Lundberg U, Senn BM, Schunn M, Habel A, Henriques-Normark B, Ortqvist A, Kalin M, von Gabain A, Nagy E (2008) Discovery of a novel class of highly conserved vaccine antigens using genomic scale antigenic fingerprinting of pneumococcus with human antibodies. J Exp Med 205:117–131CrossRefPubMedPubMedCentralGoogle Scholar
- 17.Chen A, Mann B, Gao G, Heath R, King J, Maissoneuve J, Alderson M, Tate A, Hollingshead SK, Tweten RK, Briles DE, Tuomanen EI, Paton JC (2015) Multivalent pneumococcal protein vaccines comprising pneumolysoid with epitopes/fragments of CbpA and/or PspA elicit strong and broad protection. Clin Vaccine Immunol 22:1079–1089CrossRefPubMedPubMedCentralGoogle Scholar
- 20.Andrade DC, Borges IC, Ivaska L, Peltola V, Meinke A, Barral A, Käyhty H, Ruuskanen O, Nascimento-Carvalho CM (2016) Serological diagnosis of pneumococcal infection in children with pneumonia using protein antigens: a study of cut-offs with positive and negative controls. J Immunol Methods 433:31–37CrossRefPubMedGoogle Scholar
- 21.Andrade DC, Borges IC, Laitinen H, Ekström N, Adrian PV, Meinke A, Barral A, Nascimento-Carvalho CM, Käyhty H (2014) A fluorescent multiplexed bead-based immunoassay (FMIA) for quantitation of IgG against Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis protein antigens. J Immunol Methods 405:130–143CrossRefPubMedGoogle Scholar
- 23.Meriluoto M, Hedman L, Tanner L, Simell V, Mäkinen M, Simell S, Mykkänen J, Korpelainen J, Ruuskanen O, Ilonen J, Knip M, Simell O, Hedman K, Söderlund-Venermo M (2012) Association of human bocavirus 1 infection with respiratory disease in childhood follow-up study, Finland. Emerg Infect Dis 18:264–271CrossRefPubMedPubMedCentralGoogle Scholar
- 24.Chen T, Tanner L, Simell V, Hedman L, Mäkinen M, Sadeghi M, Veijola R, Hyöty H, Ilonen J, Knip M, Toppari J, Simell O, Söderlund-Venermo M, Hedman K (2014) Diagnostic methods for and clinical pictures of polyomavirus primary infections in children, Finland. Emerg Infect Dis 20:689–692CrossRefPubMedPubMedCentralGoogle Scholar
- 25.Ota MO, Oluwalana C, Howie SR, Gomez M, Ogunniyi AD, Mendy-Gomez AL, Owolabi O, Mureithi MW, Townend J, Secka O, Antonio M, Sutherland JS, Adegbola RA (2011) Antibody and T-cell responses during acute and convalescent stages of invasive pneumococcal disease. Int J Infect Dis 15:e282–e288CrossRefPubMedGoogle Scholar
- 26.Posfay-Barbe KM, Galetto-Lacour A, Grillet S, Ochs MM, Brookes RH, Kraehenbuhl JD, Cevey-Macherel M, Gehri M, Gervaix A, Siegrist CA (2011) Immunity to pneumococcal surface proteins in children with community-acquired pneumonia: a distinct pattern of responses to pneumococcal choline-binding protein A. Clin Microbiol Infect 17:1232–1238CrossRefPubMedGoogle Scholar
- 29.Orihuela CJ, Mahdavi J, Thornton J, Mann B, Wooldridge KG, Abouseada N, Oldfield NJ, Self T, Ala’Aldeen DA, Tuomanen EI (2009) Laminin receptor initiates bacterial contact with the blood brain barrier in experimental meningitis models. J Clin Invest 119:1638–1646CrossRefPubMedPubMedCentralGoogle Scholar
- 30.Borges IC, Andrade DC, Cardoso MR, Toppari J, Vähä-Mäkilä M, Ilonen J, Knip M, Hyöty H, Veijola R, Simell O, Jartti T, Käyhty H, Ruuskanen O, Nascimento-Carvalho CM (2016) Natural development of antibodies against Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis protein antigens during the first 13 years of life. Clin Vaccine Immunol 23:878–883CrossRefPubMedPubMedCentralGoogle Scholar
- 38.Andrade DC, Borges IC, Adrian PV, Meinke A, Barral A, Ruuskanen O, Käyhty H, Nascimento-Carvalho CM (2016) Effect of pneumococcal conjugate vaccine on the natural antibodies and antibody responses against protein antigens from Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis in children with community-acquired pneumonia. Pediatr Infect Dis J 35:683–689CrossRefPubMedGoogle Scholar