I 1 - Structure ,Function and classes of bacteria

What are microorganisms?

• There is a big diversity

• They can be both beneficial, vital for living and can cause disease

• Not all are visible to the naked eye

What is the nomenclature of Bacteria?

• genus/species

• prokaryotic, divides by binary fission

• multiplies, divides, and spreads - growth

• cell wall differentiates into different structures - Blue/Purple ( +), Pink/Red ( - )

What is the general structure of bacteria?

Similarities:
 Plasma membrane
 DNA
 Cytoplasm
 Ribosomes
Differences:
 No nucleus
 Circular DNA
 No membrane-bound organelles
 Complex membrane architecture
 Appendages like fimbriae, pili and flagellum

What are ways to differentiate between types of bacteria?

Ways to classify bacteria:

• shape, size, and aggregation - cocci, rods, clumps, chains, pairs

• cell wall structure - Gram-positive, Gram-negative

• other structures - capsule, spores, flagella, pili

How can you use the structure of the bacterial cell envelope to classify ?

• They can be separated using their cell envelope

• Can be detected using a Gram stain

Gram-positive - monoderm ( Blue/Purple)

- Small periplasmic space

- Thick peptidoglycan layer

- Teichoic & Lipoteichoic acid

-These all help give structure and protect it from the environment

Gram-negative – diderm ( pink)

- Large periplasmic space- peptidoglycan

- Outer membrane - single lipid bilayer lipid polysaccharide, gives protection

What will the Gram stain detection look like ?

What are the functions and structure of peptidoglycan, lipoteichoic acid and LPS?

Peptidoglycan

• Mesh-like polymer made of repeating sugars and amino acids

• Provides cell shape and prevents bursting from osmotic pressure

Lipoteichoic Acid (LTA)

• Polyol phosphate polymers span the cell wall

• Important for stability and mucosal adhesion ( mouth, lungs, etc )

Lipopolysaccharide (LPS)

• Large molecules – Lipid A, inner oligosaccharide and outer polysaccharide

• Lipid A anchors to the cell membrane - reactive in our bodies , causes septic shock

• Endotoxin – causes sepsis

What are other structure that can be found in Bacteria?

Capsule

• Usually polysaccharide – can be protein

• Sits above the cell envelope

• Extra protective coat

• Inhibits white blood cells, sticks to surfaces, protects from dehydration

Fimbriae/Pili

• Hair-like appendages

• Hundreds of them

• Adhesion to surfaces, movement, transfer of genetic material

Flagella

• Whip-like structures

• Motor

• Provides motility

What are the possible effects of bacteria in the body ?

Commensal

• Organism which colonises the host but causes no disease in normal circumstances

Pathogen

• Organism that causes or is capable of causing disease

Opportunist Pathogen

• Microbe that only causes disease if host defences are

compromised

Virulence/Pathogenicity

• Degree to which a given organism is pathogenic

Asymptomatic carriage

• When a pathogen is carried harmlessly at a tissue site but

causes no disease

What is adhesion and its role in bacteria?

• Adhesion is often a critical first step in colonisation or pathogenesis

• Bacteria employ multiple adhesins which can associate with various host molecules such as proteins on the surface, preventing binding of actual ligands

• Host molecules are ‘hijacked’ for association

• Adhesins can associate with just one receptor or multiple receptors

What are exotoxins?

• Soluble peptide that living bacteria secrete into the environment to cause damage to specific host tissues and cells

• they don’t attach to host surfaces but release toxins

What are some exotoxin families ?

Neurotoxin

• Blocks nerve signals, causing flaccid paralysis

Tetanus toxin

• Blocks neurotransmitters, causing severe, rigid muscle spasms

AB toxin

• A causes B, which damages the body

Pore-forming toxins

• Punches holes in cell membrane ,cell lysis ,tissue damage

Superantigens

• Overactivates T cells, systemic inflammation, shock

Streptococcal pyrogenic exotoxins

• toxic shock-like syndrome

Note - Treatment of these conditions often requires killing of the bacteria and neutralisation of the toxin with antibodies

How can oxygen be used to classify bacteria ?

Obligate aerobes

• require oxygen as a terminal electron acceptor

Obligate Anaerobes

• are killed by toxic oxygen radicals; lack protection enzymes like SOD

Facultative Anaerobes

• switch from aerobic respiration to fermentation based on oxygen availability

Microaerophiles

• require reduced oxygen ( 2-10%)

• damaged by atmospheric levels

What is the growth rate of the bacteria?

Lag phase- initial growth phase; growth remains constant as cells adapt to the new environment

Exponential (log) Phase - cells divide rapidly; population doubles at a constant rate

Stationary Phase - plateau: total cells stay constant; rate of cell growth is equal to the rate of cell death

Graph is a sigmoid shape - (S)

What is another way to identify the bacteria?

• Bacteria have very different metabolic processes for different sugars; selective agar can take advantage of this

Examples:

MacConkey - lactose agar

• lactose fermenters - red (pink)

• Acid produced by fermentation turns neutral red dye on the plate RED

Xylose Lysine Deoxycholate ( XLD)

• Lactose fermentation turns phenol red in media yellow

• isolates salmonella and shigella

• Shigella cannot ferment lactose; remains red

• Salmonella cannot ferment lactose but reduces thiosulphate to produce hydrogen sulphide ( Black)

What biochemical tests can help to do distinguish between species?

Catalase test

• Detects the presence of the catalase enzyme

• Converts H2O2 to H2O and O2

• Bacteria cause dilute hydrogen peroxide to bubble

Oxidase test

• Detects cytochrome c oxidase enzyme

• Utilises a reagent called TMPD

• Bacteria oxidises the reagent into a dark blue with the enzyme

Coagulase test

• Detects the coagulase enzyme

• Converts soluble fibrinogen in human plasma to fibrin

• Causes a clot!

Presence of coagulase often suggests the presence of multiple virulence proteins

What are antibiotics?

• Produced by several fungi

• Many groups, split based on chemical structure and mode of action, such as penicillins and tetracyclines

• Can be bactericidal (directly kills bacteria and causes irreversible cell death) or bacteriostatic ( inhibits bacterial growth, reproduction, and multiplication without directly killing cells)

How can the activity of antibiotics be determined?

• Activity is determined by the Minimum Inhibitory Concentration (MIC) – the lower the MIC, the more active the drug

• Disk Diffusion - An agar plate can also be used by measuring the zone of inhibition to see effectiveness

• E-test - strip contains different concentrations of antibiotics - measure the zone of inhibition

What are the antibiotic targets in bacterial structure?

Cell wall synthesis inhibitors:

• penicillins, cephalosporins, vancomycin

Protein synthesis inhibitors:

• 30S subunit (Tetracyclines, aminoglycosides)

• 50S subunit (Macrolides)
  

Nucleic acid synthesis inhibitors

• DNA gyrase (Fluoroquinolones)

• RNA polymerase (Rifamycins)

Metabolic antagonists

• Folic acid synthesis (Sulfonamides, trimethoprim)

What is antibiotic resistance ?

• Developed in human pathogens soon after antibiotics were used in treatment

• Environmental bacteria naturally develop resistance - an ongoing evolutionary battle with some fungi

• Resistance is passed from environmental bacteria or through natural mutation (selection)

• Most common mechanism is via an enzyme – beta-lactamase (ESBL)

• Widespread and often indiscriminate use has led to multi-resistant organisms

• Several bacteria are now very resistant and difficult to treat
   MRSA
   Neisseria gonorrhoeae

• Greater reliance now on infection control and/or vaccines

How does resistance occur ?

4 fundamental mechanisms:

1. Alteration of a DNA sequence changes bacterial proteins that are antimicrobial targets - Target cell wall synthesis, DNA replication, or protein synthesis

2. Reduction in influx of antibiotics into the bacterial cell

3. Increase in efflux of antibiotics from the bacterial cell

4. Enzymatic degradation of antibiotics

-Occurs through mutations within chromosomal DNA OR... acquisition of foreign DNA containing resistance genes

What causes bacterial genetic variation?

Mutation

• Base substitution

• Deletion

• Insertion

Gene transfer

• Transformation- plasmid from the environment

• Transduction - Phage

• Conjugation - Sex pilus

What are the types of vaccines available?

Live attenuated vaccines – BCG tuberculosis)

• Pathogen is live but unable to cause disease

Inactivated whole-cell vaccines - Cholera

• Pathogen killed

Toxoid vaccine - Tetanus

• Toxins inactivated

Polysaccharide conjugate vaccine – Meningitis ACWY

• Capsule sugars bound to a carrier molecule

Subunit vaccine – Meningitis B

• Pathogen proteins extracted and bound to carrier molecules

mRNA – COVID

• Pathogen protein produced by the cell briefly

What are alternative therapeutics?

• Dietary changes

• probiotics

• microbiome replacement

• bacteriocins

• phage therapy

• host- directed therapies - block initial stages of pathogenesis

What is staphylococcus aureus

• Gram-positive: thick cell wall (peptidoglycan)

• Bunch of grapes - can divide in three alternate planes

• Spread through contact

• Methicillin-resistant

• vancomycin resistant

• no vaccine

What is S.aureus lifecycle

Environment

• Food, water, surfaces, stress-resistant (salt/heat)

Carriage

• Nasal carriage or skin survival,30% of the human population

Opportunistic pathogen

• Nosocomial infections

• Biofilm formation

• Antibiotic resistance

What is the S.aureus pathology

• Food poisoning

• Scalded skin syndrome

• Septicaemia

• Osteomyelitis

How can one organism cause so many diseases ?

• Highly adaptable

• Multiple virulence proteins - expressed at different times of growth

• Infection-associated dynamics

1. Interaction with specific target tissue

2. Proliferate (avoid host defences)

3. Local damage

4. Dissemination of pathogen or products (systemic disease)

ʬ All stages require virulence factors

What are some hygiene measures to reduce spread ?

• Clean hands before and after touching patients

• Hands cleaned with soap and water, or alcohol gel or hand rub, reduced from 15.4 cases per 100,000 ̃ 1.9 cases per 100,000

• Wear gloves and aprons when caring for a patient with MRSA

• Isolate MRSA patients from other patients

Summary