Please enable JavaScript.
Coggle requires JavaScript to display documents.
Human Infection & Immunity - Coggle Diagram
Human Infection & Immunity
Bacteriology
Bacteria
Free-living prokaryotes
largest group of medically significant microbes
Many thousands of species
Most non-pathogenic
Bacteria energy requirements
Synthesis of macromolecules
Repair and maintenance
Synthesis of structural components
Active transport
Movement (flagella)
Storage of nutrients
Removal of waste products
Catabolic (Energy-producing) Pathways
Many bacteria can breakdown polysaccharides to yield glucose
Glucose catabolism commences with glycolysis to generate pyruvic acid and small amount of energy
Pyruvic acid further metabolised by respiration (aerobic) or fermentation (anaerobic)
Respiration (aerobic)
Involves the conversion of pyruvic acid into acetyl CoA
Acetyl CoA enters the TCA cycle
TCA cycle and oxidative phosphorylation through the electron transport chain generate ATP
Most bacteria possess all or most of the enzymes of the TCA cycle
Electron transport chains different in prokaryotes
Fermentation (anaerobic)
Of pyruvic acid occurs through several pathways
Energy yield relatively low
Products of fermentation may be of value in indentification of bacteria
Anabolic pathways
Energy stored in ATP used:
To synthesise carbohydrates, lipids, nucleic acids, proteins (enzymes)
Cell wall
Cell membrane (phospholipids)
Movement of flagella
Active transport
Bacteria physical factors
Temperature
pH
Oxygen
Water and osmotic pressure
Temperature
pH
pH optimum for most bacteria is close to neutral
Most do not grow at all above pH 8 or below pH 6
Exception are the genus Lactobacillus (tolerate acid to approx pH 4)
Oxygen
Aerobes: use oxygen
Obligate aerobes: require oxygen (Mycobacterium tuberculosis)
Anaerobes: do not require oxygen
Obligate anaerobes: destroyed by oxygen (Clostridium)
Facultative anaerobes - grow in the presence or absence of oxygen (adaptable)
Water & osmotic pressure
Bacteria grow optimally in an environment that is water saturated (liquid environment)
Bacteria obtain all nutrients from their surroundings and the solute content of those surroundings will exert osmotic pressure
Optimal growth if surroundings isotonic
Hypotonic environment: leads to turgidity and possibly lysis
Hypertonic environment: leads to membrane shrinkage and inhibits growth
Bacteria nutritional factors
Carbon
Nitrogen
Phosphorus and sulphur
Cations (iron, potassium, calcium and magnesium)
Trace elements
Organic factors
Growth of Bacteria
Implies an increase in cell number
Bacteria reproduce by binary fission when the bacterium approx. doubles in size
Time taken to reproduce is the generation time (varies from organism to organism and the environment)
Generation time may be as little as 20 minutes (E.coli) or several hours (Mycobacteria)
Growth of Bacteria in the Lab
Bacteria may be grown in a wide variety of different media in the lab
Liquid media contain salts and nutrients in water adjusted to the correct pH and sterilised
Solid media are generated through the addition of agar (complex polysaccharide extracted from seaweed)
Agar is poured and allowed to set in a petri dish
Agar plates are usually used to isolate a bacterial species from a clinical sample for identification
Clinical samples invariably contain a mixture of the 'pathogenic' bacteria and the normal flora
Various methods are used to identify bacteria
Culture Media
Defined medium: all constituents known
Complex media: some unknown constituents such as yeast extract (MacConkey agar, tryptic soy broth)
Enriched media: presence of special growth factors for fastidious microorganisms (Streptococcus pyogenes requires red blood cells)
Selective Media
Encourage the growth of some microorganisms while inhibiting the growth of others (Sabouraud dextrose medium promotes fungal growth and inhibits bacterial growth)
Differential Media
Distinguishes between bacteria by the nature of their growth (blood agar between haemolytic and non-haemolytic bacteria)
Phases of Bacterial Growth
In liquid media bacterial numbers can be sampled over time and a growth curve constructed
Lag phase
Exponential/log phase
Stationary phase
Terminal phase
Lag phase
Adjusting: bacteria is adjusting to the new environment upregulating genes that can take advantage of environment and down regulating genes that are no longer required
Log phase
Replicating: bactiera is at its maximal or most efficient rate that can take advantage of the nutrients available
Stationary phase
Switching off: bacteria starts to switch itself off, switching off enzymes required for replication etc. because there are no nutrients remaining to allow further replication and waste is also starting to accumulate which slows bacterial growth.
Bacterial death
Wastes start to accumulate more and more which causes bacterial death. This occurs if you're not replenishing the media and moving the culture to fresh media
Identification/Study of Bacteria
Patient with a possible Infectious Disease
Patient history (onset, symptoms, contacts, travel)
Clinical examination (focus of infection)
Investigations (pathology, radiology)
Provisional diagnosis or differential diagnoses
Plan for treatment
Clincial Specimens
Normally sterile fluids (blood, urine, CSF, peritoneal fluid, pleural fluid, pericardial fluid)
Tissue/organ biopsy
Sputum
Faeces
Pus
Foreign body (intravascular catheter tip)
Processing of Clinical Specimens
Microscopy (wet and fixed/stained preparations)
Establish a pure culture
Various techniques to identify pathogenic bacteria
Antibiotic suscepribility testing
Specimen Collection
Specimen selection: collect from a site where organism likley to be found
Timing of collection: knowledge of clinical course of disease and patient history, prior to antibiotic therapy
Proper collection protocols: sterile containers, labelling, patient care, quantity of organism, contamination, potential hazard
Transport: fragile organisms, accurate representation of organisms present at time of collection, special conditions for some organisms
Microscopy
Wet preparations
Observation of living microbes
Principally used for fungi and protozoa
Useful for direct observation of urine and CSF (prescence of other cell types)
Often use of weak stains to enhance contrast (methylene blue)
Fixed and Stained
Bacteria killed and fixed on microscope slide
Many different stains but most useful and important is the Gram stain
Most bacteria stain Gram-POS (stained purple) or Gram-NEG (decolourised)
May be performed immediately on clinical specimens, which in some cases can result in a presumptive diagnosis
Microorganisms that do not Gram stain
Mycobacteria (waxy cell wall)
Treponemes, Borrelia, Leptospira (very slender, spiral)
Mycoplasma (no cell wall)
Rickettsia, Coxiella, Chlamydia (intracellular parasites and very small)
Other stains
Ziehl-Neelsen stain or acid fast stain (Mycobacteria)
Flagella stains
Capsule stains
Pure Cultures
Most clinical specimens contain mixtures of microorganisms (normal flora)
Liquid media may be used initally to increase the number of bacteria in the sample (blood, anaerobes)
Follow with culture on solid media (agar)
Some bacteria cannot be grown in media (Mycobacterium leprae, Treponema pallidum, Chlamydia)
Preparation of pure cultures
Spread plate: drop of liquid containing 100-200 cells spread over agar plate
Streak plate: mix of cells in a 3-4 way streak on agar
Pour plate: microorganisms mixed with liquid agar at 45 degrees and poured into Petri-dish
Colony Morphology on Agar Plates
Use of defined, complex, enriched, selective, differential media
Colonies appear the same for a given species but differ berween species
Differ in cololur, size, form, elevation, margin, texture
Some bacteria show haemolytic activity in media containing red blood cells (blood agar)
Presumptive Identification
Usually based on:
Gram stain
Cell and colony morphology
Aerobic, anaerobic growth (or both)
Specialised requirements for growth
Further tests to identify bacteria
Serological Tests
Molecular techniques
Biochemical tests
Serological Tests
Implies the detection of microorganisms by specific antibodies
An infected patient will produce antibodies specific for the antigens present on the microorganism causing the infection
Antigens whole organisms or components of them (flagella, toxins)
Antibodies detected principally in the serum (blood of the patient)
Advantages & Methods
Most useful if a bacteria is impossible to grow (Treponema pallidum) or grows very slowly (Legionella)
Usually based on the notions of seroconversion and antibody titre (concentration)
Various methods to determine antibody titre (agglutination, neutralisation, complement fixation, fluorescent antibody test, ELISA)
Related notion of antigen detection with specific antibodies (commercial kits)
Often used for microorganisms with impossible, difficult or time consuming growth characteristics
May provide a rapid presumptive diagnosis (Meningitis)
Available for Legionella, Neisseria meningitidis, Clostridium difficile toxin
Disadvantages
Time lag between the infection and appearance of serum antibody and hence retrospective
May develop specific IgM assays to indicate active disease as opposed to past infection (not achieved for many infections at present)
Molecular Techniques
Bacteria Morphology
Most range in size from 1-10um in length & 0.1-1um in diameter
3 principle shapes:
rods (bacillus)
spheres (cocci)
spirals (3 variants - spirochaetes (corkscrews), spirilla (less tightly coiled), and vibrios (incomplete spirals or comma shaped)
May grow in chains or clusters
May have one or more flagella
Bacteria Structure
Cell membrane
Cell wall
External structure
Internal structure
Endospores
Cell membrane
Phospholipid bilayer and similar cell membranes of other cells
Absence of sterols (subgroup of steroids) but they contain similar compounds 'hopanoids' (lipids) found in many prokaryotic membranes
Cell membrane function
Prinicipal role is to act as a semi-permeable barrier
Osmosis
Passive and active transport
Specialised functions which include:
ATP synthesis and cellular respiration as bacteria don't contain mitochondria this occurs on the underside of the cell membrane where all metabolic functions occur
Enzyme and protein secretion and synthesis of cell wall components
Cell wall
Semi-rigid structure present in most bacteria
Lies outside the cell membrane
Principal component is peptidoglycan
Most bacteria can be classified into 2 major groups: Gram-POS & Gram-NEG
Gram-positive cell wall
Thick peptidoglycan layer
Contain complex polysaccharides called teichoic acids
Teichoic acids are negatively charged and influence the passage of materials in and out of the bacterial cell and play a role in antigenicity
Gram-Negative cell wall
More complex structure than Gram-POS
Thin peptidoglycan layer bounded by an outer membrane
Outer membrane contains lipopolysaccharides (LPS), lipoproteins and phospholipids
Outer membrane provides a barrier to various substances (antibiotics, lysozyme) but is more permeable than the cell membrane (porins)
Lipopolysaccharide (LPS)
Composed of 3 components:
Lipid A: or endotoxin causes toxic effects in Gram-Neg infections
Core polysaccharide
O side-chain (antigen) distinctive structure in bacterial strains (serology)
Cell wall function
Determines cell shape
Protects against osmotic lysis
Determines some staining properties
Offers protection from some toxic substances
Contains components that contribute to pathogenicity
Damage to the cell wall
Lysozome breaks the bonds in the peptidoglycan later
Some antibiotics target the cell wall (penicillins)
Cell wall compromise exposes bacteria (protoplast or spheroplast) to increased sensitivity to osmotic lysis
External structure
Glycocalyx
Capsule
Slime layer
Flagella
Pili
Fimbriae
Glycocalyx
Sticky polysaccharide layer secreted onto the surface of some bacteria
Depending on structure and function referred to as a capsule or a slime layer
Protect from desiccation
Capsule
Polysaccharide and/or polypeptide molecules
Usually synthesised inside the cell and secreted to form an organised gelatinous layer attached to the cell wall
Involved in adherence to host cells and in evasion of phagocytosis (virulence factors)
Slime layer
Polysaccharide and glycoprotein molecules
Less organised than capsule and loosely attached
Involved in attachment to host cells
May contribute to motility
May contribute to biofilm formation on medical instruments
Flagella (flagellum)
Thin, rigid filaments of protein flagellin arranged as a hollow cylinder
Attached to bacterial cell by the basal body
Many different patterns of flagellation useful in the indentification of bacteria
Pili (pilus)
Occur as hairlike appendages 1-10 per cell
Required for bacterial mating (conjugation)
Determined genetically by sex factors or conjugative plasmids
Transfer of antibiotic resistance
Fimbriae (fimbria)
Hairlike appendages thinner than Pili and more numerous (thousands per cell)
Involved with attachment of bacteria to host cells
Virulence factor
Internal structure
Cytoplasm
Bacterial chromosome
Plasmids
Ribosomes
Inclusion bodies
Bacteria don't contain subcellular organelles but they do contain structures that serve like organelles
Cytoplasm
Approx. 80% water
Contains the macromolecules essential for cell activity
Some bacteria possess internal membrane systems (nitrifying and photosynthetic bacteria)
Mesosomes may be present
Proteins positioned at certain sites (cytoskeleton-like arrangement)
Bacterial chromosome
Located in an ill-defined cytoplasmic region called the nucleoid
Usually circular double stranded DNA but may be linear and some bacteria possess more than one chromosome
Carries the genetic information required for the structure and function of the cell
Plasmids
One or more small double stranded circular DNA molecules
Replicate independently of the bacterial chromosome or may be integrated with it
Are not mandatory for bacterial growth and reproduction but carry genes that may confer selective advantage (drug resistance, pathogenicity)
Carry gene for the production of sex pili
May move between bacteria
Plasmids carry many genes that can provide a selective advantage
Ribosomes
Small granular particles present in the cytoplasm composed of 2 subunits
Subunits contain both protein and RNA
Sites of protein synthesis
Designated 70S ribosomes (sedimentation rate in ultracentrifuge)
Eukaryotic ribosomes are 80S (slightly larger and more dense than prokaryotic ribosomes)
Inclusion bodies
Granules of organic or inorganic materials utilised as storage sites and contribute to reducing osmotic pressure
Organic inclusion bodies usually contain glycogen or poly-B-hydroxybutyrate
Inorganic inclusion bodies often represent storage of phosphate (polyphosphate or volutin granules)
May be free in cytoplasm or membrane bound
Some bacteria also contain gas vacuoles or vesicles, which confer buoyancy on the cell
Endospores
A specialised type of resting cell formed inside the bacterial cell membrane of some Gram-POS bacteria
Surrounded by a spore coat that is composed of peptidoglycan and protein
Occurs under conditions of adversity
Endospores survive boiling, freezing, desiccation and exposure to radiation and chemicals
Possess low water content and do not perform metabolic functions
Nucleic acid, enzymes and metabolites present for successful germination to transpire in the advent of favourable conditions
May survive for years
Many endospore forming bacteria are important pathogens (genus Clostridium)
-
Virology
Immunology
