Bacterial Cell Structure Basic Bacteriology Part-1-2 Bacterial Cell Structure (First Semester 2017-2018)

The bacterial genome: The bacterial genome consists of a single chromosome (in most cases) , plasmid (s), mobile genetic elements and viral DNA (prophage). A- Nucleoid: The nucleoid represents the bacterial chromosome (folded /super-coiled). Most bacteria have a single chromosome, which is circular when it is unfolded. The bacterial chromosome may contain up to 2000 genes. The bacterial nucleoid is not separated from the cytoplasm by a membrane. The bacterial nucleoid has no nucleolus, no histons and no mitotic spindles The HU protein (not histons as in eukaryotes) mediates super-coiling/folding of the bacterial chromosome. The bacterial DNA has no introns (unlike eukaryotic DNA , which contains introns and exons)

B- Plasmids: Plasmids are small extra-chromosomal, double-stranded, circular DNA that are capable of replicating independently of the bacterial chromosome. Some times, bacterial plasmid integrates with the bacterial chromosome . In this case, the plasmid is called Episom. Although plasmids are not essential for bacterial survival, they may carry genes that can provide the bacterial cells with additional features that may increase their pathogenesis and antibiotic resistance Plasmids can be classified into: 1- Transmissible plasmids (Conjugative plasmids): these plasmids harbor genes that can control their transfer from one bacterial cell to another by a process known as conjugation. In addition, they may also have other genes such as antibiotic resistance genes. 2- Non-transmissible plasmids: these are usually small plasmids. Although they may have different types of genes, these plasmids do not have genes that are responsible for the conjugation process.

Plasmids may be named according to the type of genes they have such as: R plasmids: these plasmids have genes that are involved in antibiotic resistance. ( in addition, they may or may not have conjugative genes) UV resistance plasmids: such plasmids carry genes that encode for proteins or enzymes involved in DNA damage that may result because of UV radiation. ( in addition, they may or may not have conjugative genes) 3. Virulence plasmids: carry genes that are involved in bacterial pathogenesis such as toxin genes. (in addition, they may or may not have conjugative genes)

C- Mobile genetic elements (Transmissible genetic elements /Jumping genes) These are pieces or regions of DNA that are capable of moving them selves from one position to another on the chromosome or plasmid, or between the chromosome and the plasmid. Their moving is just like a cut and a paste processes that is mediated by an enzyme known as Transposase. This enzyme is encoded by the transposase gene that is carried by these mobile genetic elements. Unlike plasmids, mobile genetic elements are not capable of independent replication. As they are always found integrated with the plasmid or the chromosome, they can only get replicated along the replication of the plasmid or the chromosome. The main two types of these mobile genetic elements are: Insertion Sequence ( IS element): it contains only the transposase gene Transposon (Tn): it contains the transposase gene and other additional gene (s), such as antibiotic resistance genes.

Exception: Replicative transposon: This type of transposons is capable of making a copy of it self that can move to another positing on the chromosome or the plasmid or between the plasmid and the chromosome. This type of transposons has a special additional gene known as the resolvase gene. This gene encodes for the resolvase enzyme that mediates the release of the newly formed copy of the replicative transposon, which moves to another position.

D- Viral DNA (prophage / provirus) These DNA regions represent the DNA of temperate phages that has been integrated a long time ago with the bacterial chromosome. However, because of certain mutations, these temperate phages were unable to leave the bacterial chromosome and became part of it. The integrated viral DNA is known as a prophage or provirus. In some cases, this viral DNA may contain a toxin gene that is involved in bacterial pathogenesis. The bacterium which has a prophage integrated with its chromosome is known as a lysogenic bacterium (Example: lysogenic Corynebacterium diphtheriae)

2- The Cytoplasmic Membrane (Cell membrane): It represents the physical boundary of bacterial cells. It consists of a phospholipid bilayer and integral (structural) and non-structure proteins that have different functions (transporters, enzymes, sensors/receptors) Unlike eukaryotic cell membranes, which contain sterols, bacterial cell membranes do not contain sterols (except the bacterium, Mycoplasma). The bacterial cytoplasmic membrane has four important functions: It contains transporters ( some may utilize energy) It contains enzymes involved in energy generation (ATP) by oxidative phosphorylation It contains enzymes that are involved in biosynthesis such as the synthesis of cell wall It contains sensors that monitor and detect changes in extracellular environmental conditions such as pH, nutrient concentration…… It contains translocation and secretion systems

3- The Bacterial Cell Wall: The bacterial cell wall is a multi-layered structure located external to the cytoplasmic membrane. The main component of the bacterial cell wall that is responsible for its rigidity is peptidoglycan. All bacteria (except Mycoplasma ) have cell walls In Gram-positive bacteria: it is mainly consists of a thick layer of peptidoglycan (see Peptidoglycan) In Gram-negative bacteria: it consists of a thin layer of peptidoglycan and an outer membrane

Peptidoglycan: Peptidoglycan is a polymer that is made of a single subunit (monomer). This monomer is a disaccharide that is consist of two amino sugars. These are : N-acetyl glucose amine (NAG) and N-acetyl muramic acid (NAM) A tetra-peptide side chain is linked to NAM. The tetra-peptide side chain consists of 4 amino acids, which are: 1- L-alanine 2- D-glutamate 3- L-lysine ( in Gram-positive bacteria) or (Diaminopimelic acid (DAP) in Gram- negative bacteria) 4- D-alanine Note: Murein and mucopeptide are old names of peptidoglycan.

A single chain of peptidoglycan is composed of alternating N-acetylmuramic acid and N-acetylglucosamine molecules

The chains of peptidoglycan are cross-linked together resulting in the formation of a strong network. The cross-linking is mediated by the tetra- peptide side chains in such a way that the third amino acid of one side chain is linked to the forth amino acid of the tetra-peptide side chain that is closed to it.

In Gram-positive bacteria, a pentaglycine bridge mediates the cross linking between L-lysine of one tetra-peptide side chain to D-alanine of the other tetra-peptide side chain. In Gram-negative bacteria, a peptide bond mediates the cross-linking between DAP of one tetra-peptide side chain to D-Alanine of the other tetra-peptide side chain.

The peptidoglycan layer is very thick in Gram-positive The peptidoglycan layer is very thick in Gram-positive. It consists of about 40-50 layer (network) of peptidoglycan. ( Note: Some Gram-positive bacteria have Teichoic acid or Lipoteichpoic acid with its peptidoglycan) The peptidoglycan layer is thin in Gram-negative bacteria. It consists of 3-5 layers (networks) of peptidoglycan.

Functions of peptidoglycan: It protects bacterial cells from their high cytoplasmic osmotic pressure specially if the bacterial cell is found in an environment of lower osmotic pressure It plays a role in the attachment of Gram-positive bacteria to environmental surfaces and to human cell and tissues It plays an important role in determining the morphology of bacterial cells It may play role in pathogenesis (some bacteria have peptidoglycan that has some toxicity)

The Periplasmic space: The Periplasmic space is the area between the cell membrane (cytoplasmic membrane) and the outer-membrane of Gram-negative bacteria. This periplasmic space contains many types of proteins such as: Braun’s lipoproteins (which connects the outer membrane with peptidoglycan which locates underneath it) Carrier proteins involved in nutrient transport Enzymes involved in nutrient degradation Enzymes involved in antibiotic resistance Note: in Gram-negative bacteria, the peptidoglycan is found within the periplasmic space.

The Outer membrane of Gram-negative bacteria It is a phospholipids bilayer. However, it contains special components such as: 1- Lipopolysaccharide 2- Porins: these are proteins that form small channels through the outer membrane of Gram-negative bacteria . Porins play a role in facilitating the passage of small, hydrophilic molecules such as sugars, amino acids, vitamins, and metals as well as many antimicrobial drugs.

Lipopolysaccharide (Found only in the outer membrane of Gram-negative bacteria) The lipopolysaccharides (LPS) is only found in the outer membrane of Gram-negative bacteria The LPS is composed of: 1- A lipid portion that is known as lipid A, which is very toxic 2- A polysaccharide portion that is consists of two parts: a- A core polysaccharide of five sugars linked to lipid A b- An outer polysaccharide that is a polymer of about 25 repeated subunit. Each subunit consists of about 3 or 5 sugars. This outer polymer is known as the somatic, or O, antigen.

LPS is know as Endotoxin (found only in Gram-negative bacteria) The toxic part of LPS is lipid A Lipid A is responsible for many of the features of disease upon infection with Gram-negative bacteria such as inflammation and fever. In case of septicemia caused by Gram-negative bacteria, Lipid A may result in toxic shock (septic shock) that may be fatal. Antibodies raised against the O antigen can be used in bacterial diagnosis Note: LPS of Gram-negative bacteria is called Endotoxin because it is an integral part of the cell wall of Gram-negative bacteria .

Some important functions of LPS: It may play role in attachment (adherence of bacterial cells to human cells and tissues) It interferes with phagocytosis It plays an important role in the pathogenesis of Gram-negative bacteria

Teichoic Acid and Lipoteichoic acid (Found only in the cell wall of Gram-positive bacteria): Teichoic Acid and Lipoteichoic acid are made of glycerol phosphate or ribitol phosphate They are located in the outer layer of peptidoglycan of only Gram-positive bacteria. Importance of Teichoic Acid and Lipoteichoic acid: They play role in attachment of bacterial to human cells and tissues In case of septicemia, Teichoic Acid and Lipoteichoic acid of certain types of Gram-positive bacteria can induce septic shock Note: Gram-negative bacteria do not have Teichoic Acid and Lipoteichoic acids

Specialized Structures Outside the Cell Wall: Glycocalyx: A layer of polysaccharides (in most cases) that lactates just next the external surface of the bacterial wall (NOT ALWAYS PRESENT) -In Gram-positive bacteria, it will locates next to the external surface of peptidoglycan -In Gram-negative bacteria, it will locates next to the external surface of outer membrane Structurally, Glycocalyx can be classified into capsule and slime layer: A- Capsule: The bacterial capsule is dense and thick layer that covers the bacterial cell wall and it is well attached to it. In most cases, it is composed of polysaccharides. However, in some bacteria, it could be made of amino acids such as in the case of the capsule of Bacillus anthracis, which is made of D-Gultamate. Importance of bacterial capsule: 1- It is highly hydrated (water storage) and can be used as a nutrient by the bacterial cell in case of starvation. 2- It plays an important role in attachment (adherence) of bacterial cell to environmental surfaces as well as to human tissues and cells during infection. 3- It interferes with phagocytosis ( In general, the bacterial capsule is anti-phagocytic)

4- In some cases, the bacterial may be made of polysaccharides similar to those found in human body. In this case, the bacterial cell can hardly be recognized by immune cells (antigenic disguise) 5- The capsules of certain bacteria can be used as vaccines. 6- Antibodies raised against capsules can be used in bacterial diagnosis.

B- Slime Layer: The slime layer is a loose polysaccharide layer that is produced by many bacteria. It can cover the surface of bacterial cell, however, unlike the capsule, it is not dense and can be easily detached off the surface of the bacterial cells. Importance of slime layer: 1- It plays an important role in attachment (adherence) of bacterial cell to environmental surfaces as well as to human tissues and cells during infection. 2- It interferes with phagocytosis ( similar to capsule, it is anti-phagocytic) 3- It facilitates gliding motility 4- Biofilm formation: the material of the slime layer can be secreted into the surrounding environment to contribute in Biofilm formation. Biofilm formation on human tissues or prosthetic medical implants (joints, heart valves) is major clinical problem and can be a source of recurrent infections. In addition, Biofilm formation on teeth (plaque) is the first step in dental caries .

Biofilms: When a slime-producing bacteria attaches to environmental surfaces, human tissues, teeth, prosthetic implants, it starts producing and secreting slime material into the surrounding environment. The attached bacteria produce slime in large quantities , so that ultimately, the slime material along with other substances (proteins) cover the bacterial cells ( in other words, the bacterial cells become embedded within a layer that is composed of slime material and other substances (matrix of the Biofilm). Biofilm= Matrix + bacterial cells that live within the matrix Biofilms represent a dynamic and a natural community of bacteria (may be of different types). Within this community, bacteria can survive and replicate. Biofilms protect bacterial cells: Within biofilms, bacteria are protected (shielded) from: A- Disinfectants and antibiotics B- Immune defenses such as antibodies and phagocyte In addition, it has been found that bacteria in biofilms become more virulent.

Bacterial Biofilms

Flagella: Flagella are long, hollow, whip-like appendages that mediates the swimming motility of bacteria by acting like a propeller. Bacterial flagellum can be considered as a polymer that is made of a protein known as flagellin. Bacterial flagellum is composed of three parts: A- Basal body (motor) B- Hook (connect the filament to basal body) C- Filament Energy is needed for the rotation of the bacterial flagellum.

Some bacteria have only one flagellum Some bacteria have only one flagellum. However, some other flagellated bacteria may have 2 or more flagella. Distribution of flagella on bacterial surface: Forward motility (swimming) of the bacterial cells requires that the flagellum rotates counter clockwise. Tumbling motility of the bacterial cell (local), cells requires that the flagellum rotates clockwise

A flagellated bacterial call can swim toward nutrients and other attractants or a way from harmful substances. The bacterial cell can detect these chemicals by surface receptors (sensors) Swimming of a flagellated bacterial cells toward a nutrient or a way from harmful chemicals is called chemotaxis Importance of bacterial flagellum in pathogenesis: Some species of motile bacteria such as E. coli and Proteus, are common causes of urinary tract infections. Flagella may play a role in pathogenesis by propelling the bacteria up the urethra into the bladder. It can play role in attachment Note: Bacteria that do not have flagella are non-motile bacteria such as Shigella Klebsiella , Staphylococci, Streptococci Note: bacterial cell can detect and respond (by using sensors/receptors) to many other stimuli, such as alteration in temperature, pH, light intensity…..

Pili (Fimbriae): Pili are hair-like filaments that extend from the surface of many types of bacteria (mostly, Gram-negative ///About 1000/ cell/// They are shorter than flagella and are composed of a protein subunit known as of pilin. There are two types of pili: 1- Common pili: mediate attachment (adherence) of bacteria to human cells and tissues, which is a necessary step for infection initiation Example: mutants of Neisseria gonorrhoeae that do not form pili are not pathogenic.

Sex pili: is a specialized kind of pili that mediates attachment between bacterial cells during the conjugation process. There about 1-10 sex pili/bacterial cell ( which has conjugative plasmid)

Bacterial Endospores (Spores): These are highly resistant structures are formed by certain types of bacteria in response to adverse conditions, such as lack of nutrients and water. The main aim of sporulation is bacteria is to survive adverse conditions (unlike fungi that make spores for reproduction) Although there are several types of bacteria that can make spores, the only two medically important genera are the genus Bacillus and the genus Clostridium (Both of which, are Gram-positive bacilli). Infection with these bacteria is initiated upon entry into bacterial endospores of these bacteria into our bodies through wounds, digestive tracts, or respiratory tract

Bacterial endospore can be defined as is a dormant bacterial cell that is metabolically inactive but not dead. Bacterial endospore is highly resistant to severe conditions such as high and low temperatures, chemicals, antibiotics radiations. The ability of the bacterial endospore to resist severe conditions is mainly attributed to its structure Bacterial endospores can remain viable for long period of time (hundreds or thousands of years) Bacterial endospores can be killed by autoclaving (but not by boiling). :

The structure of the bacterial endospore include: 1- Core: which is the inner most part of the endospore. It contains bacterial DNA, a small amount of partially dehydrated cytoplasm, calcium salts, DNA repair enzymes, small acidic soluble proteins (SASP) that bind to DNA . Lack of water, calcium salt , SASP and DNA repair enzymes maintains bacterial DNA integrity and stability. The core is surrounded by a membrane known as the inner membrane. 2-Cortex: comes next to the inner membrane and it composed of unusual peptidoglycan 3-Coat: comes next to the cortex. It consists of a strong keratin-like proteins. . 4- Exosporium: is the outer most layer that consists of a phospholipids membrane Notes: The Cortex and the coat are responsible for giving the bacterial endospore its highly resistant nature to severe conditions The Cortex, the coat and the exosporium are known together as the spore wall.

Upon exposure to water and the appropriate nutrients, specific enzymes degrade the coat, water and nutrients enter, and germination into a metabolically active bacterial cell occurs (vegetative cell).