Misfolded alpha-synuclein (AS) and other neurodegenerative disorder proteins display prion-like transmission of protein aggregation. Factors responsible for the initiation of AS aggregation are unknown. To evaluate the role of amyloid proteins made by the microbiota we exposed aged rats and transgenic C. elegans to E. coli producing the extracellular bacterial amyloid protein curli. Rats exposed to curli-producing bacteria displayed increased neuronal AS deposition in both gut and brain and enhanced microgliosis and astrogliosis compared to rats exposed to either mutant bacteria unable to synthesize curli, or to vehicle alone. Animals exposed to curli producing bacteria also had more expression of TLR2, IL-6 and TNF in the brain than the other two groups. There were no differences among the rat groups in survival, body weight, inflammation in the mouth, retina, kidneys or gut epithelia, and circulating cytokine levels. AS-expressing C. elegans fed on curli-producing bacteria also had enhanced AS aggregation. These results suggest that bacterial amyloid functions as a trigger to initiate AS aggregation through cross-seeding and also primes responses of the innate immune system.

Porphyromonas gingivalis is an oral/systemic pathogen implicated in chronic conditions, although the mechanism(s) whereby it resists immune defenses and persists in the host is poorly understood. The virulence of this pathogen partially depends upon expression of fimbriae comprising polymerized fimbrillin (FimA) associated with quantitatively minor proteins (FimCDE). In this study, we show that isogenic mutants lacking FimCDE are dramatically less persistent and virulent in a mouse periodontitis model and express shorter fimbriae than the wild type. Strikingly, native fimbriae allowed P. gingivalis to exploit the TLR2/complement receptor 3 pathway for intracellular entry, inhibition of IL-12p70, and persistence in macrophages. This virulence mechanism also required FimCDE; indeed, mutant strains exhibited significantly reduced ability to inhibit IL-12p70, invade, and persist intracellularly, attributable to failure to interact with complement receptor 3, although not with TLR2. These results highlight a hitherto unknown mechanism of immune evasion by P. gingivalis that is surprisingly dependent upon minor constituents of its fimbriae, and support the concept that pathogens evolved to manipulate innate immunity for promoting adaptive fitness and thus their capacity to cause disease.

The leukotoxin of Actinobacillus actinomycetemcomitans has been implicated as a virulence determinant in various human infections and is encoded by a multigene operon consisting of four known genes, designated ltxC, ltxA, ltxB, and ltxD. The ltx operon appears to be present in all A. actinomycetemcomitans strains, but levels of toxin expression vary greatly among strains. Thus, to gain a better understanding of the expression and regulation of the ltx operon, we have analyzed the ltx promoters of a highly toxic (JP2) and a minimally toxic (652) strain of A. actinomycetemcomitans. The nucleotide sequence of the JP2 ltx promoter contains -10 and -35 elements situated 350 bases upstream of ltxC, and primer extension of JP2 RNA confirmed that they are functional in vivo. However, a second primer extension product of 40 bases was present, and analysis of a series of truncated JP2 promoters fused to lacZ suggested that the region immediately upstream of ltxC also promotes transcription in Escherichia coli. These results suggest that two promoters may direct ltx expression in JP2. In addition, a small open reading frame capable of encoding a peptide of 78 amino acids was identified upstream of ltxC. Northern blots showed that this open reading frame is transcribed as part of a 4.2-kb mRNA, a transcript not previously identified as being derived from the ltx operon. In contrast, strain 652 expresses low steady-state levels of ltx mRNA, and its intact ltx promoter was inefficient in transcribing lacZ in E. coli. The nucleotide sequence of the 652 promoter is similar to that of the JP2 promoter but contains a region of 530 bp that is not present in JP2. Of 15 additional strains of A. actinomycetemcomitans that were analyzed, 13 contained promoters resembling the 652 sequence and 2 possessed JP2-like promoters. Both strains possessing the JP2-like promoter expressed 10- to 20-fold-higher levels of leukotoxin than did the strains possessing promoters resembling the 652 promoter. These results suggest that high levels of leukotoxin expression may correlate with the presence of the JP2-like promoter.

Porphyromonas gingivalis, one of the causative agents of adult periodontitis, attaches and forms biofilms on substrata of Streptococcus gordonii. Coadhesion and biofilm development between these organisms requires the interaction of the short fimbriae of P. gingivalis with the SspB streptococcal surface polypeptide. In this study we investigated the structure and binding activities of the short fimbriae of P. gingivalis. Electron microscopy showed that isolated short fimbriae have an average length of 103 nm and exhibit a helical structure with a pitch of ca. 27 nm. Mfa1, the major protein subunit of the short fimbriae, bound to SspB protein, and this reaction was inhibited by purified recombinant Mfa1 and monospecifc anti-Mfa1 serum in a dose-dependent manner. Complementation of a polar Mfa1 mutant with the mfa1 gene restored the coadhesion phenotype of P. gingivalis. Hence, the Mfa1 structural fimbrial subunit does not require accessory proteins for binding to SspB. Furthermore, the interaction of Mfa1 with SspB is necessary for optimal coadhesion between P. gingivalis and S. gordonii.

No abstract

SummaryThe antigen I/II family of cell-surface-anchored polypeptides in oral streptococci are structurally complex multi-functional adhesins, with multiple ligand-binding sites. Discrete regions within these polypeptides bind human salivary glycoproteins, other microbial cells, and calcium. Sequences within the N-terminal region bind preferentially fluid-phase glycoproteins, while the C-terminal half of the polypeptide contains species-specific adhesion-mediating sequences that bind surface-immobilized glycoproteins. These features may assist streptococcal adhesion to oral surface receptors despite the presence of excess fluidphase receptors. Immunological studies reveal an array of T-cell and B-cell epitopes presented by antigen I/II polypeptides and suggest the occurrence of natural suppression of human antibodies to the adhesionmediating sequences. The functional and immunological properties of antigen I/II proteins may account to a major extent for the success of oral streptococci colonizing and surviving within the human host.

Active smokers and those exposed to secondhand smoke are at increased risk of bacterial infection. Tobacco smoke exposure increases susceptibility to respiratory tract infections, including tuberculosis, pneumonia and Legionnaires disease; bacterial vaginosis and sexually transmitted diseases, such as chlamydia and gonorrhoea; Helicobacter pylori infection; periodontitis; meningitis; otitis media; and post-surgical and nosocomial infections. Tobacco smoke compromises the antibacterial function of leukocytes, including neutrophils, monocytes, T cells and B cells, providing a mechanistic explanation for increased infection risk. Further epidemiological, clinical and mechanistic research into this important area is warranted.

Porphyromonas gingivalis is an aggressive periodontal pathogen that persists in the mixed-species plaque biofilm on tooth surfaces. P. gingivalis cells attach to the plaque commensal Streptococcus gordonii and this coadhesion event leads to the development of P. gingivalis biofilms. Binding of these organisms is multimodal, involving both the P. gingivalis major fimbrial FimA protein and the species-specific interaction of the minor fimbrial Mfa1 protein with the streptococcal SspB protein. This study examined the contribution of the Mfa1-SspB interaction to P. gingivalis biofilm formation. P. gingivalis biofilms readily formed on substrata of S. gordonii DL1 but not on Streptococcus mutans cells which lack a coadhesion-mediating homologue of SspB. An insertional inactivation of the mfa1 gene in P. gingivalis resulted in a phenotype deficient in S. gordonii binding and unable to form biofilms. Furthermore, analysis using recombinant streptococci and enterococci showed that P. gingivalis biofilms formed on Enterococcus faecalis strains expressing SspB or translational fusions of SspB with SpaP (the non-adherent SspB homologue in S. mutans) containing the P. gingivalis adherence domain (SspB adherence region, BAR) of SspB. In contrast, an isogenic Ssp null mutant of S. gordonii DL1 was unable to support biofilm growth, even though this strain bound to P. gingivalis FimA at levels similar to wild-type S. gordonii DL1. Finally, site-specific mutation of two functional amino acid residues in BAR resulted in SspB polypeptides that did not promote the development of P. gingivalis biofilms. These results suggest that the induction of P. gingivalis biofilms on a streptococcal substrate requires functional SspB-minor fimbriae interactions.