Streptococcus Adhesion and Pathogenesis Mechanisms
Current funding: The Wellcome Trust, Health Research Board of Ireland.
Press Release: Safer streptococcus vaccines
Participants: HF Jenkinson, ME Barbour, S Maddocks, L Franklin, C Wright, C Keane.
Collaborators: MA Jepson (Department of Biochemistry, Bristol), M Miles (Department of Physics, Bristol), A Corfield (Mucin Research Group, Bristol), A Kadioglu (University of Leicester), D Cox (Royal College of Surgeons in Ireland, Dublin), S Kerrigan (RCSI, Dublin), C Kelly (GKT, London), S Matthews (Imperial College, London), N Strömberg (Umeå University, Sweden).
Introduction
Over 100 species of streptococci are now recognized. The mitis group streptococci, including the species gordonii, mitis, oralis, parasanguinis and sanguinis (45) colonize the mucosal surfaces of the mouth and nasopharynx as well as dental surfaces (16). The mutans group bacteria, including the species mutans and sobrinus (45), colonize mainly the teeth (38). Dental caries, manifested as the demineralization of tooth tissues by bacterial acids, is strongly associated with the mutans group streptococci, in conjunction with other predisposing factors such as frequency of sugar intake, oral hygiene, use of fluoride and host susceptibility. Since the early 1970s there has been much interest in developing an effective immunization method for S. mutans. A potential vaccine candidate, protective against caries in animal models, is the major surface adhesin AgI/II protein, also designated antigen B (42), SpaP or P1 (2,10) or PAc (27). A monoclonal antibody to AgI/II, and a synthetic peptide adhesion epitope, have each been shown to be effective in preventing re-colonization of human volunteers with S. mutans (28,34). Thus, topical application of adhesion inhibitors may be effective for controlling colonization by some species of oral bacteria.
The mitis group streptococci are amongst the primary colonizers of oral cavity surfaces (24). Their patterns of adhesion and accumulation, in conjunction with other bacteria, e.g. Actinomyces, facilitate the development of multi-species biofilm communities (29,31). These communities can initiate enamel or root caries, or various forms of periodontal disease (43), and provide sources of bacteria for invasive infections of the root and pulp (33), systemic spread, and development of endocarditis and abscesses (31). Colonization of infants by S. sanguinis correlates with time of first emergence of primary teeth (6). A window of infectivity for colonization by S. sanguinis is suggested at a median age of 9 months (8), while acquisition of mutans group streptococci occurs at a median age of 26 months (7). Earlier colonization by S. sanguinis, and elevated levels of these organisms, are correlated with a significant delay in colonization by S. mutans (8). These findings are important because they demonstrate that bacterial colonization patterns can strongly influence disease outcome, thus emphasizing the necessity to understand better the molecular mechanisms by which streptococci recognize host ligands to effectively colonize humans.
Streptococcal adhesion and the AgI/II family of polypeptides.
Adhesion of bacteria to peptide or carbohydrate receptors on host surfaces is a key event in colonization and infection. A wide range of salivary components support streptococcal adhesion (24). The proline rich proteins (PRPs) and high molecular mass parotid salivary agglutinins (SAGs) (14) provide salivary pellicle receptors (17,41). SAGs also mediate aggregation and clearance of streptococci from the oral cavity, so are important general modulators of oral microbial ecology (41). Allelic variants of PRPs and isomorphs of SAG (now designated gp-340) (40) display large variations in their binding efficiencies to streptococci (4,5). SAG (gp-340), PRPs, collagen type I, other oral micro-organisms (3,19) and host cells are all bound by the AgI/II family polypeptides, which are produced by most indigenous species of oral streptococci (reviewed in ref. 23). The AgI/II proteins are cell wall anchored adhesins of about 1500 aa residues, with highest sequence identities (62%) within their C-terminal regions. S. mutans serotypes c and f, and S. sobrinus each express a single AgI/II (SpaP, Sr and SpaA, respectively) (27,30,35), while S. gordonii produces two polypeptides designated SspA (171 kDa) and SspB (160 kDa) (12,13). The overall structure of the AgI/II protein is conserved across the family, with major substrate-binding sites being localized to within the alanine-rich (A) region, and C- terminal to the proline-rich (P) region.
AgI/II structure and proposed substrate-binding regions
The mechanisms by which AgI/II polypeptides bind their substrates are not fully understood, but there is evidence that the various polypeptides display different ligand binding specificities. The S. gordonii SspA and SspB proteins, when expressed on the cell surface of non-pathogenic Lactococcus lactis, have different binding affinities for gp-340 and for collagen (20). These specificities may be determined, at least in part, by sequences within the C-terminal region of AgI/II. A synthetic 20 aa residue peptide (SpaP residues 1024-1045) binds gp-340 (26,28) so the ligand-binding site within native AgI/II may form a flexible loop. Recent studies utilizing surface plasmon resonance (BiaCore) to measure adhesion-inhibition kinetics by synthetic 20-mer peptides suggest a role for Glu1037 in binding of S. mutans SpaP to SAG (28). The corresponding residue in S. gordonii SspB is Lys, which might be related to different ligand-binding specificity of SspB (21).
Deletion of genes sspA and sspB encoding AgI/II proteins in S. gordonii inhibits interactions of bacteria with collagen (see ref 18).
In addition to mediating adhesion of streptococci to saliva-coated surfaces, the AgI/II proteins promote biofilm formation (39), type I collagen-dependent bacterial invasion of dentine (18,32), bind to monocytes via lectin interactions (9) inducing IL-1beta, TNF-alpha and IL-6, and activate endothelial cells inducing IL-8 and IL-6 and up-regulation of ICAM-1 and VCAM-1 expression (1). Recently we have demonstrated that SspA and SspB mediate adhesion of S. gordonii to human epithelial cells (36), stimulating host signalling pathways and secretion of IL-8. Also, S. gordonii wild-type cells, but not isogenic AgI/II polypeptide mutants, block integrin (alpha5 beta1)-mediated adhesion and internalization of pathogenic group A streptococci suggesting that competition for host receptor binding may influence dissemination of streptococci from oral sites. Therefore the AgI/II polypeptides appear to be globally important molecules for streptococcal adhesion, oral microbial community formation, host tissue invasion, and in the pathogenesis of inflammatory disorders.
Adhesion specificity and host susceptibility.
Allelic variations in salivary receptors (15) may modulate oral microbial ecology and biofilm formation (32,44) and recent evidence suggests that saliva adhesion levels due to gp-340 and PRP polymorphisms rank among the strongest factors underlying host susceptibility to dental caries (5,25). The interplay of adhesin affinities and ligand specificities with host receptor availability may then determine the outcome of competition between different streptococci for host receptors. Thus, defining the molecular basis of oral streptococcal AgI/II protein interactions with their ligands may provide a key to understanding colonization properties of streptococci.
Deletion of sspA and sspB genes encoding AgI/II proteins in S. gordonii abrogates bacterial invasion of deninal tubules (see ref 32).
Mechanisms of receptor recognition by AgI/II polypeptides.
Evidence suggests that AgI/II proteins from different streptococci preferentially recognize different host ligands. However, the mechanisms by which these occur are not understood. Competition between streptococci for binding these polymorphic host receptors may determine colonization patterns (37), influence the composition and biological activities of oral bacterial communities (11), and impact on disease outcome.
More recently we have characterized further the molecular basis for the interactions of AgI/II polypeptides from different streptococcal species with purified forms of gp-340 (a member of the scavenger receptor cysteine rich family of polypeptides), other oral bacteria, and with epithelial cell receptors. The relative binding levels of 6 different AgI/II polypeptides from 3 species of oral streptococci for bacterial and host ligands were determined. The S. gordonii polypeptides SspA and SspB were most efficient at binding gp-340, while SspB was unique in binding a carbohydrate receptor on Actinomyces naeslundii T14V cells (21). AgI/II polypeptides were shown, for the first time, to interact directly with the polypeptide backbone of gp-340 as well as with sugar residues that decorate the gp-340 glycoprotein. In conjunction with analyses of adherence properties of purified AgI/II polypeptides by surface plasmon resonance we have shown that at least three regions of AgI/II proteins were necessary for interactions with host ligands. The N-terminally located A and V regions carried major binding sites while the presence of sequences within the C-terminal half were necessary for presentation of the N-terminal region binding functions. It was shown that AgI/II polypeptides interact with beta1 integrins via a fibronectin-independent mechanism that may be significant in blocking integrin-mediated adhesion and invasion by pathogenic streptococci. The AgI/II polypeptides were also demonstrated themselves to bind fibronectin and to interplay with a major sialic acid binding adhesin (Hsa) in promoting interactions of streptococci with extracellular matrix components, epithelial cell receptors, and platelets (22).
Future prospects
Results of our research have enhanced understanding of the molecular basis by which interactions of commensal and pathogenic streptococci with host ligands relate to oral infectious disease. This information will be broadly relevant in studies of mechanisms underlying general bacterial infections of host surfaces and may assist development of novel strategies involving peptide or carbohydrate inhibitors to combat infectious diseases.
References1. Al-Okla S, Chatenay-Rivauday C, Klein J-P, Wachsmann D. 1999. Involvement of alpha5 beta1 integrins in interleukin 8 production induced by oral viridans streptococcal protein I/IIf in cultured endothelial cells. Cell Microbiol 1: 157-168.
2. Brady LJ, Piacentini DA, Crowley PJ, Oyston PCF, Bleiweis AS. 1992. Differentiation of salivary agglutinin-mediated adherence and aggregation of mutans streptococci by use of monoclonal antibodies against the major surface adhesin P1. Infect Immun 60: 1008-1017.
3. Brooks W, Demuth DR, Gil S, Lamont RJ. 1997. Identification of a Streptococcus gordonii SspB domain that mediates adhesion to Porphyromonas gingivalis. Infect Immun 65: 3753-3758.
4. Carlen A, Olsson J. 1995. Monoclonal antibodies against a high-molecular-weight agglutinin block adherence to experimental pellicles on hydroxyapatite and aggregation of Streptococcus mutans. J Dent Res 74: 1040-1047.
5. Carlen A, Bratt P, Stenudd C, Olsson J, Stromberg N. 1998. Agglutinin and acidic proline rich protein receptor patterns may modulate bacterial adherence and colonization on tooth surfaces. J Dent Res 77: 81-90.
6. Carlsson J, Grahnen H, Jonsson G, Wikner S. 1970. Establishment of Streptococcus sanguis in the mouths of infants. Arch Oral Biol 15: 1143-1148.
7. Caufield PW, Cutter GR, Dasanayake AP. 1993. Initial acquisition of mutans streptococci infections in infants: evidence for a discrete window of infectivity. J Dent Res 72: 37-45.
8. Caufield PW, Dasanayake AP, Yihong LI, Pan Y, Hsu J, Hardin JM. 2000. Natural history of Streptococcus sanguinis in the oral cavity of infants: evidence for a discrete window of infectivity. Infect Immun 68: 4018-4023.
9. Chatenay-Rivauday C, Yamodo I, Sciotti M-A, Ogier JA, Klein J-P. 1998. The A and extended V N-terminal regions of streptococcal antigen I/IIf mediate the production of tumour necrosis factor in the monocyte cell line THP-1. Mol Microbiol 29: 39-48.
10. Crowley PJ, Brady LJ, Piacentini DA, Bleiweis AS. 1993. Identification of salivary agglutinin-binding domain within cell surface adhesin P1 of Streptococcus mutans. Infect Immun 61: 1547-1552.
11. Daep CA, James DM, Lamont RJ, Demuth DR. 2006. Structural characterization of peptide-mediated inhibition of Porphyromonas gingivalis biofilm formation. Infect Immun 74: 5756-5762/
12. Demuth DR, Lammey MS, Huck M, Lally ET, Malamud D. 1990. Comparison of Streptococcus mutans and Streptococcus sanguis receptors for human salivary agglutinin. Microb Patho 9: 199-211.
13. Demuth DR, Duan Y, Brooks W, Holmes AR, McNab R, Jenkinson HF. 1996. Tandem genes encode cell-surface polypeptides SspA and SspB which mediate adhesion of the oral bacterium Streptococcus gordonii to human and bacterial receptors. Mol Microbiol 20: 403-413.
14. Ericson T, Rundegren J. 1983. Characterization of a salivary agglutinin reacting with a serotype c strain of Streptococcus mutans. Eur J Biochem 133: 255-261.
15. Eriksson C, Frangsmyr L, Danielsson Niemi L, Loimaranta V, Holmskov U, Bergman T, Leffler H, Jenkinson HF, Stromberg N. 2007. Variant size- and glycoforms of the scavenger receptor cysteine-rich protein gp-340 with differential bacterial aggregation. Glycoconj J 24: 131-142.
16. Frandsen EVG, Pedrazzoli V, Kilian M. 1991. Ecology of viridans streptococci in the oral cavity and pharynx. Oral Microbiol Immunol 6: 129-133.
17. Gibbons RJ, Hay DI, Schlesinger DH. 1991. Delineation of a segment of adsorbed salivary acidic proline-rich proteins which promotes adhesion of Streptococcus gordonii to apatitic surfaces. Infect Immun 59: 2948-2954.
18. Heddle C, Nobbs AH, Jakubovics NS, Gal M, Mansell JP, Dymock D, Jenkinson HF. 2003. Host collagen signal induces antigen I/II adhesin and invasin gene expression in oral Streptococcus gordonii. Mol Microbiol 50: 597-607.
19. Holmes AR, McNab R, Jenkinson HF. 1996. Candida albicans binding to the oral bacterium Streptococcus gordonii involves multiple adhesin-receptor interactions. Infect Immun 64: 4680-4685.
20. Holmes AR, Gilbert C, Wells JM, Jenkinson HF. 1998. Binding properties of Streptococcus gordonii SspA and SspB (antigen I/II family) polypeptides expressed on the cell surface of Lactococcus lactis MG1363. Infect Immun 66: 4633-4639.
21. Jakubovics NS, Stromberg N, van Dolleweerd CJ, Kelly CG, Jenkinson HF. 2005. Differential binding specificities of oral streptococcal antigen I/II family adhesins for human or bacterial ligands. Mol Microbiol 55: 1591-1605.
22. Jakubovics NS, Kerrigan SW, Nobbs AH, Strömberg N, van Dolleweerd CJ, Cox DM, Kelly CG, Jenkinson HF. 2005. Functions of cell surface anchored antigen I/II family and Hsa polypeptides in interactions of Streptococcus gordonii with host receptors. Infect Immun 73: 6629-6638.
23. Jenkinson HF, Demuth DR. 1997. Structure, function and immunogenicity of streptococcal antigen I/II polypeptides. Mol Microbiol 23: 183-190.
24. Jenkinson HF, Lamont RJ. 1997. Streptococcal adhesion and colonization. Crit Rev Oral Biol Med 8: 175-200.
25. Jonasson A, Eriksson C, Jenkinson HF, Kallestal C, Johansson I, Stromberg N. 2007. Innate immunity glycoprotein gp-340 variants may modulate human susceptibility to dental caries. BMC Infect Dis 7: 57
26. Kelly CG, Todryk S, Kendal HL, Munro GH, Lehner T. 1995. T-cell, adhesion, and B-cell epitopes of the cell surface Streptococcus mutans protein antigen I/II. Infect Immun 63: 3649-3658.
27. Kelly CG, Evans P, Bergmeier L, Lee SF, Progulske-Fox A, Harris AC, Aitken A, Bleiweis AS, Lehner T. 1989. Sequence analysis of the cloned streptococcal cell surface antigen I/II. FEBS Letts 258: 127-132.
28. Kelly CG, Younson JS, Hikmat BY, Todryk SM, Czisch M, Haris PI, Flindall IR, Newby C, Mallet AI, Ma JK-C, Lehner T. 1999. A synthetic peptide adhesion epitope as a novel antimicrobial agent. Nature Biotechnol 17: 42-47.
29. Kolenbrander PE. 2000. Oral microbial communities: biofilms, interactions, and genetic systems. Annu Rev Microbiol 54: 413-437.
30. LaPolla RJ, Haron JA, Kelly CG, Taylor WR, Bohart C, Hendricks M, Pyati J, Graff RT, Ma JK-C, Lehner T. 1991. Sequence and structural analysis of surface protein antigen I/II (SpaA) of Streptococcus sobrinus. Infect Immun 59: 2677-2685.
31. Lamont RJ, Jenkinson HF. 2000. Adhesion as an ecological determinant in the oral cavity. In: Kuramitsu HK, Ellen RP, eds Oral Bacterial Ecology: The Molecular Basis, Wymondham U.K., Horizon Scientific Press, pp 131-168.
32. Li T, Johansson I, Hay DI, Stromberg N. 1999. Strains of Actinomyces naeslundii and Actinomyces viscosus exhibit structurally variant fimbrial subunit proteins and bind to different peptide motifs in salivary proteins. Infect Immun 67: 2053-2059.
33. Love RM, McMillan MD, Park Y, Jenkinson HF. 2000. Coinvasion of dentinal tubules by Porphyromonas gingivalis and Streptococcus gordonii depends upon binding specificity of streptococcal antigen I/II adhesin. Infect Immun 68: 1359-1365.
34. Ma JK-C, Hunjan M, Smith R, Kelly C, Lehner T. 1990. An investigation into the mechanism of protection by local passive immunization with monoclonal antibodies against Streptococcus mutans. Infect Immun 58: 3407-3414.
35. Moisset A, Schatz N, Lepoivre Y, Amadio S, Wachsmann D, Scholler M, Klein J-P. 1994. Conservation of salivary glycoprotein-interacting and human immunoglobulin G-cross-reactive domains of antigen I/II in oral streptococci. Infect Immun 62: 184-193.
36. Nobbs AH, Shearer BH, Drobni M, Jepson MA, Jenkinson HF. 2007. Adherence and internalization of Streptococcus gordonii by epithelial cells involves beta1 integrin recognition by SspA and SspB (antigen I/II family) polypeptides. Cell Microbiol 9: 65-83
37. Nobbs AH, Zhang Y, Khammanivong A, Herzberg MC. 2007. Streptococcus gordonii Hsa environmentally constrains competitive binding by Streptococcus sanguinis to saliva-coated hydroxylapatite. J Bacteriol 189: 3106-3114.
38. Nyvad B, Kilian M. 1987. Microbiology of the early colonization of human enamel and root surfaces in vivo. Scand J Dent Res 95: 369-380.
39. Pecharki D, Petersen FC, Assev S, Scheie AA. 2005. Involvement of antigen I/II surface proteins in Streptococcus mutans and Streptococcus intermedius biofilm formation. Oral Microbiol Immunol 20: 366-371.
40. Prakobphol A, Xu F, Hoang VM, Larsson T, Bergstrom J, Johansson I, Frangsmyr L, Holmskov U, leffler H, Nilsson C, Boren T, Wright JR, Stromberg N, Fisher SJ. 2000. Salivary agglutinin, which binds Streptococcus mutans and Helicobacter pylori, is the lung scavenger receptor cystein-rich protein gp-340. J Biol Chem 275: 39860-39866.
41. Rosan B, Appelbaum B, Golub E, Malamud D, Mandel ID. 1982. Enhanced saliva-mediated bacterial aggregation and decreased bacterial adhesion in caries-resistant versus caries-susceptible individuals. Infect Immun 38: 1056-1059.
42. Russell RRB. 1979. Wall associated antigens of Streptococcus mutans. J Gen Microbiol 114: 109-115.
43. Socransky SS, Haffajee AD, Cugini MA, Smith C, Kent Jr RL. 1998. Microbial complexes in subgingival plaque. J Clin Periodontol 25: 134-144.
44. Stromberg N, Boren T. 1992. Actinomyces tissue specificity may depend on differences in receptor specificity for GalNAcβ-containing glycoconjugates. Infect Immun 60: 3268-3277.
45. Whiley RA, Beighton D. 1998. Current classification of the oral streptococci. Oral Microbiol Immunol 13: 195-216.