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BY2012 Microbiology Bacterial Fimbriae & Adherence Bacterial Fimbriae Fimbriae Flagella Colourised electron micrograph of an Escherichia coli cell bearing type 1 fimbriae Escherichia coli with Type 1 Fimbriae Fimbriae Transmission electron micrograph of E. coli cell bearing type 1 fimbriae – negative staining Red Blood Cells (Erythrocytes) - Model System to Demonstrate Fimbrial Adhesion Colourised electron micrograph of red blood cells Fimbrial Adherence to Red Blood Cells Adhesion Schematic of a bacterium with cell-surface fimbriae or adhesins (represented by the black tips) binding to host cell receptors (sugars of glycoproteins or glycolipids) (represented by the open circles) on the host cell surface Fimbrial Adherence to Red Blood Cells Mannose RBC Adhesion Blocking Schematic of a bacterium with cell-surface fimbriae or adhesins (represented by the black tips) binding to host cell receptors (sugars of glycoproteins or glycolipids) (represented by the open circles) on the host cell surface. Addition of mannose in the case of type 1 fimbriae blocks adherence. Fimbrial Adherence Haemagglutination Assay Fimbriate bacteria Fimbriate bacteria causing cross-linking of red blood cells Escherichia coli Type 1 Fimbriae Cross-Linking Red Blood Cells RBC 0.1 μm RBC NCTC 8623 Type 1 Fimbriae RBC Transmission electron micrograph of red blood cells agglutinated by E. coli NCTC 8623. Type 1 fimbriae are attached (arrows) to the surfaces of three RBCs, cross-linking these to give rise to haemagglutination. Escherichia coli NCTC 8623 Mannose-Sensitive Haemagglutination by Type 1 Fimbriae No Mannose Plus Mannose → Inhibition of fimbrial haemagglutination Escherichia coli NCTC 8623 Not Expressing Type 1 Fimbriae No Mannose Plus Mannose Bacteria not expressing Type 1 fimbriae do not cause haemagglutination Escherichia coli with K88 Fimbriae K88 fimbriae differ from Type 1 fimbriae. They are very fine, flexible fibrillar structures rather than rod-like structures like type 1 fimbriae. Shadowing technique – Transmission electron micrograph Escherichia coli K88 Fimbriae K88 fimbriae are very fine fibrillar structures rather than rod-like structures. Transmission electron micrograph – negative staining Escherichia coli K88 Fimbriae Mannose-Resistant Haemagglutination No Mannose Plus Mannose → No inhibition of fimbrial haemagglutination The receptor for K88 fimbriae is not mannose but β-linked N-acetylhexosamine (HexNAc), either GlcNAc or GalNAc Escherichia coli K88 Fimbriae Mannose-Resistant Haemagglutination With continued incubation No Mannose increased clumping of RBCs Plus Mannose → No inhibition of fimbrial haemagglutination The receptor for K88 fimbriae is not mannose but β-linked N-acetylhexosamine (HexNAc), either GlcNAc or GalNAc Saccharomyces cerevisiae - Model System to Demonstrate Type 1 Fimbrial Adhesion Yeast cell suspension viewed by phase contrast microscopy A Schematic of the Structure of the Yeast Cell Wall SS Mannoprotein 40% GPI Anchor β1-3 glucan 50% Transmembrane Protein β1-6 glucan 8% Chitin 2% Saccharomyces – Cell Surface Features Protease treated 100 nm The outer cell wall layer is composed mainly of mannoproteins (dense zone arrowed), covalently bound to ß-1,6-glucan (light zone), can be removed by protease treatment (right hand panel) Saccharomyces Mannoproteins [α1–6-Mannose] Module GPI = Glycosyl phosphatidyl inositol 50-200 Wall Lattice Agglutination of Yeast Cells by E. coli Bearing Type 1 Fimbriae Agglutination Broth-grown cells expressing type 1 fimbriae No Agglutination Agar-grown cells not expressing type 1 fimbriae Agglutination of Yeast Cells by E. coli Bearing Type 1 Fimbriae Agglutination Broth-grown cells No Mannose No Agglutination Broth-grown cells with Mannose
