4.2 - 4.3 Bacterial growth, metabolism and taxonomy Flashcards
What physical properties are used to classify bacteria?
- Shape
- Staining
- Specialised features
What are the metabolic properties that are used to classify bacteria?
- Relationship with oxygen
- Biochemical activity
How is pathogenicity used to classify bacteria?
- Surface antigens
- Susceptibility to killing by specific viruses
- Capacity to cause disease
How is the genetic material in the bacteria arranged?
- Bacterial genomes are smaller than eukaryotes
- Bacteria don’t have a membrane bound nucleus
- The bacterial nucleoid comprises the genetic information (DNA) and the associated machinery (RNA, proteins) of the cell
- single chromosome is a closed circle of dsDNA, looped and supercoiled
- Variable in size
What are plasmids?
- Bacteria may also carry one or more plasmids
- Physically separate from chromosome and replicate independently
- Commonly occur as small circular, double stranded (supercoiled) DNA
- Variable size from 1 Kbp to several Mbp and copy number (1-40)
How much of bacterial DNA is functional genes?
- Majority of bacterial DNA encodes functional genes a bacterial strain
- Bacterial DNA does also contain pseudogenes and sRNAs, but no introns
- Bacterial lifestyle (free-living vs. pathogenic) tends to correlate with genome size
What is the difference between the core genome and pan genome?
- Core genome comprises the conserved genes of a species
- Pan genome is core genome and the accessory genome (variable content within a species)
What proportion does the core genome and the pan genome comprise?
Where do bacteria acquire genetic diversity from?
- Bacteria can evolve mia modification of existing genetic information (mutations)
- Bacteria can calso acquire genetic diversity via horizontal gene transer
- Mechanisms include plasmids
- Transposons (mobile sequence of DNA)
- Bacteriophages (virus that infects bacteria)
- Pathogenicity islands (series of genes that includes one or more virulence determinants)
- Integrons (genetic elements that allow efficient capture and expression of exogenous genes)
How were bacteria classified in the early days?
- Morphological, biochemical and metabolic classification defined prokaryotic taxonomic systems until the late 20th century
- Classification based on microorganism cell walls, cell shapes and the substances they consume
What is the common characteristic in prokaryotes used for phylogenetics?
- 16S ribosomal RNA (or 16S rRNA) is the component of the 30S small subunit of a prokaryotic ribosome
- 16S rRNA is highly conserved between different species of bacteria and archaea
- 16S rRNA gene can be used as a molecular clock for mapping evolution
What does 16s RNA do?
- It has structural roles where it acts as a scaffold defining positions of the ribosomal proteins
- Interacts with 23s rRNA aiding in the binding od the two ribosomal subunits
- 30S subunit binds to the Shine-Dalgarno sequence
- The 3’ of end 16S contains an anti-Shine-Dalgarno sequence
- The 3’-end of 16S RNA binds to the proteins S1 and S21 known to be involved in initiation of protein synthesis
How can 16S rRNA be used for phylogeny?
- Bacterial 16S rRNA gene contains highly conserved and hypervariable regions
- Nine hypervariable regions (V1-V9) of ~30-100 base pairs long
- Involved in the secondary structure of the 30S ribosomal subunit
How is the varibale regions of 16s rRNA used for taxonomy?
- Highly conserved sequences between V regions facilitates identification (via universal primers) by sequencing the same sections of the 16S sequence across different taxa
- 16S rRNA to measure relatedness: species-level >= 97%; genera >= 95%
What are the five bacterial shapes?
Is cell shape of bacteria fixed?
Cell shape is fixed due to the cell wall and peptidoglycan
What stain distinguishes cells by cell wall type?
How does gram staining separate bacteria into distinct groups?
How does gram staining work?
- Exploits differences in cell wall composition:
- Gram-positive bacteria have more peptidoglycan and retain the crystal violet stain
- Gram-negative bacteria have a less peptidoglycan and do not retain the crystal violet stain when washed and so are counter-stained by safranin.
What is the composition of gram positive cells?
- Peptidoglycan is a major component of the cell wall (~15- 80 nm)
- Cell wall also contains teichoic acid and lipoteichoic acid
- Only one membrane (cell membrane), which is functionally similar to the inner membrane of Gram-negative organisms
What is the structure of gram negative cells?
- Major feature is the outer membrane
- Small molecules, e.g. glucose, pass through protein channels (porins)
- Larger molecules require specific carriers and/or secretion pathways
- Lipopolysaccharide (LPS) stabilises the outer membrane and acts as endotoxin (a PAMP = pathogen-associated molecular pattern)
What is acid fast stain used to identify?
- Exploits the abundance of mycolic acids in the cell walls of certain bacteria, which resist other staining methods
- Acid-fast bacteria – stain bright red
- Non-acid-fast bacteria – counterstain blue
- Useful in the identification of Mycobacterium tuberculosis, the pathogen responsible for tuberculosis.
What are bacterial capsules?
- Polysaccharide-based material extending from cell surface sometimes called glycocalyx
- Visualised by negative staining
- Makes colonies large and shiny
How do capsules contribute to virulence?
- Facilitate adherence to host cells and surfaces
- Protection against phagocytosis and immune
surveillance
- Protection against dehydration
What does the flagella consist of?
- Consists of basal body, hook and filament
- Helical with a sharp bend just outside the outer membrane
- Hook allows the axis of the helix to point directly away from the cell.
- Shaft between hook and basal body via protein rings in the membrane, which act as bearings
How is flagella identified?
- Filament composed of flagellin protein (H-antigen)
- Visible by light microscopy with special stain
- Presence inferred by bacterial motility
How can the antigens of flagella be used for serotyping?
- Surface feature are bacterial antigens
- Surface antigens can be used for serotyping
- Serotyping of E.coli and Salmonella enterica species
What happens in the lag phase of bacterial metabolism?
Lag phase – bacteria adapt to the growth conditions, but are not yet (able to) divide. The bacterial cells are not dormant, as they are remodelling their metabolism for the available growth conditions.
What happens in the stationary phase of bacterial metabolism?
- Stationary phase – is due to the depletion of a crucial growth factor (nutrient) and/or the formation of an inhibitory (waste) product(s). Appears as a flattening/horizontal linear stage of the curve during the stationary phase.
- Mutations can occur.
What happens in the death phase of bacterial metabolism?
Death phase – typically arises from catastrophic nutrient depletion. Can also be caused by environmental or injurious factors.
How can you classify metabolic pathways according to the final electron acceptor in bacteria?
- Oxygen – respiration
- Inorganic compound (not O2) – anaerobic respiration
- Organic compound – fermentation
What is the difference between catalase positive and catalase negative?
- Staphylococcus aureus is catalase positive
- Streptococcus pneumoniae is catalase negative
- It secretes hydrogen peroxide to kill other bacteria, e.g. Haemophilus influenzae, Neisseria meningitidis and Staphylococcus aureus
What happens if a bacteria is alpha haemolytic?
- Streptococcus pneumoniae is a-haemolytic
- Hydrogen peroxide that oxidizes haemoglobin to green methaemoglobin (haem iron is oxidised to ferric [Fe3+] ion)
- Lacks catalase so h2o2 production results in secretion into culture medium. Incomplete oxidation of hb to mb which has a green colour. Haem not broken down further
What happens if a bacteria is beta haemolytic?
- Streptococcus pyogenes is b-haemolytic
- Streptolysin, is an exotoxin (enzyme) produced by the bacteria that causes lysis of red blood cells
- Complete haemolysis of red cells in the media appears lightened (red to yellow) and transparent around the colonies
- B haemolytic is complete haemolysis. Destruction of RBC and heme
What groups are separated based on using MacConkey agar?
- Lactose fermenters (e.g., E. coli) produce acid and are pink
- Lactose non-fermenters (e.g., Salmonella) cannot use lactose, so don’t produce acid – not pink
What are bacteriophages?
Bacteriophages (phages) are viruses that infect bacteria and archaea
How does phage typing work?
- Phage typing is a diagnostic approach that exploits the restricted host range of certain bacteriophages
- Phage can have very narrow host ranges even within a bacterial species
- Can be used to rapidly detect (hrs vs. days) susceptible species of bacteria associated with specific traits, such as drug resistance
