Fimbriae & Pili

Fimbriae

Present in many Gram negative and some Gram positive bacteria, fimbriae (singular fimbria) are thin, short hair like appendages on the surface of bacteria. They are thinner than flagella and are not involved in motility. They help in the attachment of bacteria to host tissues and some solid surfaces like rock in streams.  They are made up of helically arranged protein subunits and are about 3 to 10 nm in diameter and several micrometers in length. They can either occur at the pole of bacterial cell or over the entire surface of bacterial cell and also they vary in number from a few to several hundred per cell.

Pili

As fimbriae, pili (singular pilus) are thin, hairlike appendages on the surface of bacteria. Pili are 9 to 10 nm in diameter; slightly larger than fimbriae. They are present mostly in Gram negative bacteria with a very few Gram positive one like Corynbacterium renale. They are made up of oligomeric protein called pilins. They helps in the process of conjugation (transfer of genetic material) and hence they are also called sex pili. Pili also acts as a receptor for donor specific phages and provides a site for virus attachment at the start of their reproductive cycle. 

Note: Fimbria is basically a short pilus that helps in the attachment of bacteria to a surface. Hence, fimbriae are sometime known as attachment pili.

Protoplast & Spheroplast

Protoplast

Protoplast are generally defined as plasma membrane and everything within i.e. a cell without a cell wall is protoplast. They are generally derived from Gram positive bacteria. When Gram positive bacteria are incubated in an isotonic solution containing antibiotic penicillin, protoplast formation takes place. Penicillin basically inhibits the peptidoglycan synthesis and cell wall of Gram positive bacteria contains peptidoglycan mainly. So, it inhibits cell wall formation. Due to the absence of cell wall, protoplast are spherical in shape and are sensitive to osmotic stress. They are used for DNA transformation as presence of cell wall will interfere with the uptake of DNA into the cell.

Spheroplast 

Spheroplast are formed by the partial removal of the cell wall component. When Gram negative bacteria is treated with penicillin, they retain their outer membrane and forms spheroplast. They are spherical in shape and are osmotically sensitive.

L-form bacteria

Some strains of bacteria lack cell wall. They are called L-form bacteria or L-phase bacteria or cell wall deficient bacteria. A German microbiologist Emmy Klieneberger-Nobel, working at Lister institute in London, had first isolated them and named them L-form, after her institute name. They are generated from the bacterial species that usually have the cell walls like Escherichia coli and Bacillus subtilis. L-form can be generated from the cell wall containing bacteria by treating them with lysozyme (an enzyme that digests cell wall) or by inhibiting the peptidoglycan synthesis with antibiotics. The morphology of L-form differs from the strain from which they are derived and they are generally spheres or spheroids in shape. For example, the L-form of Bacillus subtilis appears round in shape. 

Note: Mycoplasma differs from L-form. Both of them lacks the cell wall but later are derived from the cell wall containing bacteria.

Cell Wall

Bacterial cells are surrounded by a complex cell wall that protects against osmotic lysis. Cell wall is basically made up of peptidoglycan.

Cell wall of Gram positive bacteria

Gram positive bacterial cell wall is mainly composed of peptidoglycan; 20 to 80 nm thick homogenous layer of peptidoglycan. Peptidoglycan often contains peptide interbridge of short amino acid chain. Cell wall also contains large amount of an acidic substance called teichoic acid, a polymer of ribitol or glycerol joined by phosphate group. Teichoic acids are negatively charged and responsible for providing negative charge to the cell wall. Teichoic acid are either linked directly to N-acetylmuramic acid (NAM) of peptidoglycan or to the lipids of the plasma membrane. Teichoic acids linked to the lipids are known as lipoteichoic acid. Teichoic acid provides rigidity to the cell wall thereby maintaining the cell wall structure. Although the lipid content of Gram positive bacteria is very less but some like Mycobacteria and Cornyebacterium sp are rich in lipids known as Mycolic acid. The outer membrane and lipopolysaccharide is absent in Gram positive bacteria.

Cell wall of Gram negative bacteria

As compare to Gram positive, Gram negative bacteria has much more complex cell wall. They have a thin peptidoglycan layer about 2-7 nm (in E.coli, it is 2 nm) and contain a 7-8 nm thick outer membrane. The space between the two i.e. between plasma membrane and outer membrane is called periplasmic space and it contains periplasm. The outer membrane and peptidoglycan are linked by Braun’s lipoprotein.

The other characteristics component of Gram negative bacterial cell wall is lipopolysaccharides. They are the complex of lipids and polysaccharides and are present in outer membrane. They provide chemical protection against the action of lipophilic antibiotics and bile salts and also contribute to the structural integrity of bacteria.

The outer membrane acts as a protective barrier and allows the entry of only few molecules. But the outer membrane is more permeable as compare to the plasma membrane due to the presence of special porin proteins that permits small molecules like glucose and other monosaccharides.

LPS (Lipopolysaccharide)

LPS are large complex molecules containing both carbohydrate and lipid and are generally found in the outer membrane of Gram negative bacteria. It comprises of three parts: 1) lipid A, (2) the core polysaccharide, and (3) the O side chain.

Lipid A: It consists of two phosphorylated glucosamine sugar derivatives, each linked with three fatty acids. LPS projects from the cell surface except the hydrophobic fatty acid chain which is buried in the outer membrane. The toxicity of the Gram negative bacteria is mainly due to lipid A, making LPS to act as an endotoxin. When fragments of lipid A containing membrane are released into circulation due to the cell lysis by immune system they causes fever, diarrhea and other symptoms.

Core polysaccharide: It is linked to lipid A. It commonly contains sugar like heptose and keto-deoxyoctulosonate (KDO or 3-Deoxy-D-manno-oct-2-ulosonic acid). Non-carbohydrate components like amino acids and phosphate are found in the core of many bacteria. The presence of charged sugar and phosphate in the core makes LPS to contribute to the negative charge on the bacterial surface.

O side chain: It is also termed as O-polysaccharide or O-antigen. It is a polysaccharide chain, attached to the core polysaccharide and extends outward. It also contains some unusual sugar molecules. Its composition varies between different strains of Gram negative bacteria. Presence of the full length O-antigen makes LPS smooth whereas absence or incomplete O-antigen makes LPS rough. They functions as an antigen thereby acting as a target for host antibodies. 

Note: Mycoplasma lacks cell wall.

Peptidoglycan

Peptidoglycan Structure

Peptidoglycan or murein is an enormous polymer of two sugar derivatives- N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM) joined by β-1,4 glycosidic bond. A tetrapeptide i.e. a peptide chain of four alternating D- and L- amino acids is connected to the carboxyl group of NAM. The amino acid present in tetrapeptide includes L- alanine, D- glutamic acid, D- alanine and either L- lysine or meso-diaminopimelic acid (DAP). The three amino acids i.e. D- glutamic acid, D- alanine and DAP are not found in proteins and these D- amino acids provides protection against most of the peptidases. The two tetrapeptide chain lying side by side may be linked directly to each other (as seen in many Gram negative bacteria) or indirectly by a peptide interbridge of short chain amino acids (such as polyglycine). Peptide interbridge are generally observed in Gram positive bacteria and they helps to retain their shape and intergrity. This reaction of peptide cross link formation during peptidoglycan synthesis is known as transpeptidation.

Peptidoglycan Synthesis

The biosynthesis of peptidoglycan starts in the cytoplasm as monomer and then with the help of membrane carrier bactoprenol they get transported across the cell membrane and joins the existing peptidoglycan. Firstly, the synthesis of nucleotide sugar-linked precursors UDP-NAM (Uridine diphosphate- N-acetylmuramic acid) and UDP-NAG (UDP- N-acetylglucosamine) takes place. NAG with a lactic acid attached by ether linkage is NAM. Next, amino acids are added to UDP-NAM and NAM-pentapeptide is formed. Third step takes place in cytoplasmic membrane. Bactoprenol, a lipid carrier, is present in the cytoplasmic membrane. This bactoprenol attacks UDP-NAM-pentapeptide and forms lipid I which is PP-NAM-pentapeptide. This is followed by the addition of NAG from UDP-NAG to lipid I, resulting in lipid II (PP-NAM-pentapeptide-NAG, a disaccharide and a precursor to peptidoglycan). The last step, which takes place outside the cytoplasmic membrane, is basically the polymerization of newly synthesized disaccharide with the growing glycan chain. This reaction is known as transglycosylation and enzyme involved is called transglycosylase. Hydroxyl group of the NAG will attach to the NAM in the growing chain during transglycosylation and thereby displacing the lipid-PP from the glycan chain.