Bacterial Identification, growth, gene exchange & Isolation Flashcards
4 main environmental factors on bacterial growth
- Temp
- pH
- Availability of water
- Air/Oxygen
Nutrients & microbial growth
- As nutrient conc. increases, bacterial growth rate increases exponentially until eventually reach a plateau
- growth can’t increase further because transport mechanisms for nutrient uptake is saturated
Temperature & microbial growth
-3 types of organisms
- Thermophiles - like it hot
- Mesophiles - include moth pathogens (optimum = 37)
- Psychrophiles - grow at low temps
*slow increase in growth rate w/ temp, then sudden drop off as temp denatures enzymes & melts membrane proteins
pH and microbial growth
- All bacteria need aqueous phase, therefore pH will always be important
- many have optima around pH 7.0
Salts/Osmotic pressure & Microbial growth
e.g. of halophile
-Halphiles - i.e. Staph aureus can tolerate saline solutions
Gaseous environment & microbial growth
-5 types & examples of organisms
- Obligate aerobes - little or no fermentation (as lack enzymes)
- e.g. Pseudomonas sp., mycobacterium tuberculosis
- Facultative anaerobes - can grow w/out air, but shift to aerobic respiration in presence of air
- e.g. Enterobacteriaceae
- Aerotolerant anaerobes: survive in air but don’t shift to aerobic respiration
- e.g. streptococcus
- Microaerophilic: prefer reduced oxygen tensions (and extra CO2) such as found in respiratory systems
- e.g. Brucella sp.
- Obligate Anaerobes: cannot tolerate oxygen - lacks enzymes to deal w/ very oxidative molecule that reacts rapidly (aerobes have enzymes to deal with it)
- e.g. Clostridia sp.
Features of Culture media
Need to supply;
- Energy
- carbon source
- nitrogen source
- inorganic radicals
- Some require accessory growth factors (i.e. vitamins)
Types of media to grow pathogens (4)
Basal media: Contains meat extracts; support most micro-organisms
-e.g. nutrient agar/broth
Enriched media: basal media + supplements
-e.g. blood, serum
Solid media: Used for isolation of single cells: these cells grow into discrete colonies
-agar (long chain polysaccharide from algae)
Liquid Media: used for subsequent growth of pure cultures (e.g. peptone water = meat digest + salt + water)
Selective Media
-To enhance isolation/growth of specific bacterial species can use selective media (agar) or enrichment media (broth)
Selective media: agar
-used to select for growth of part. groups of bacteria - contain inhibitors to certain organisms
e.g. Maccokey agar, brilliant green agar (Salmonella), Mannitol salt agar (staphs), Edwards media (for Streps)
Enrichment Media: Browths
-contains factors that favour growth of desired organism
e.g. selenite broth for salmonella
3 reasons we would need to identify bacteria
- Know what we are working with
- Describe roles in the environment
- Diagnose and treat diseases
Techniques for identifying bacteria
- colony morphology
- growth on different media
- Biochemical tests - can test metabolic pathways
- protein analysis
- different staining
- serological methods
- phage typing (bacterial viruses - highly specific on what they infect)
- nucleotide analysis
Shapes of cells (3)
- Coccus (round)
- Rod
- Spiral
API 20E test strip
- Rapid identification of Enterobacteriaceae and other Gram -tive bacteria)
- different substrates in vials - inoculate w/ bacteria and incubate overnight
- indicates whether bacteria has grown or not
Genotype vs Phenotype
Genotype: genetic potential of an individual
Phenotype: Observed characteristics - not all genes are expressed
Mutation: 2 types
- Mutation: cause inheritable change in the genes
1. Spontaneous mutation: due to v. rapid transcription in bacterial cells - random errors occur
i) Point mutation (change in one base)
ii) Frame shift mutation (either deletion or insertion)
2. Induced mutations: due to mutagens such as UV light, gamma irradiation, alkylating agents. Can either have;- no effect
- deleterious effect
- advantageous effect
Bacterial transfer of genetic material
-2 things that can happen
- Genetic material can be transferred between bacteria - transferred gene can exist free in cytoplasm of the next bacteria (or recombination can occur)
- integration of new genetic info may occur OR
- replacement of areas of recipient DNA (reciprocal recombination)
Methods of Gene transfer (3)
- Transformation
- Transduction
- Conjugation
Gene transfer via transformation
-Artificial transformation
- Is the transfer of genetic info from one bacteria to another - part. where they are in close proximity
- competent strains can take up free DNA from enviro (req. energy - so is not random event)
-Artificial transformation: bacteria are treated to become competent so that they can take up free DNA
Gene Transfer via Transduction
-2 types of effects in cell
-A bacteriophage accidentally transducts bacterial genes into new bacterium
-many bacteriophages kill bacteria in process of reproducing, but some infected bacteria survive
2 types of bacteriophage effects on cell:
1. Lytic cycle: bacteriophage enters bacterium & takes over metabolism of cell - causes lysis of bacterium so many phages burst out & can affect other cells
2. Lysogenic cycle: temperate phages infect the cell & replicate slowly w/out causing lysis of bacteria
-DNA integrate as a prophage into recipient’s DNA
-when bacteria divides, progeny also contain the phage DNA
*both methods result in introduction of new sections of DNA
Transduction of bacterial DNA by phages (2 forms)
- Generalised: rare
- Phage inside bacterial cell accidentally picks up fragments of bacterial DNA at random - genes may be transferred packaged in phage head along w/ the phage genes
- Specialised:
- integration of temperate phage DNA into bacterial DNA
- when phage goes into lytic cycle and makes copies of itself, host DNA adjacent to phage may also get packaged up with phage
- phage still infective but can no longer replicate
Conjugation
- transfer of plasmid DNA through sex pilus
- only larger plasmids (w/ 2-3 copies per cell) are conjugative and can be transferred
- sex pillus = straight rod-like sturcture on surface of conjuagtive bacteria - locks onto another bacteria & pulls cells together
Conjugation; 2 types of plasmids
- Virulence plasmids: code for factors that make bacteria virulent - can be acquired by non-virulent strains
- R plasmids: Resistance
Genetic transfer by Moveable genetic elements
- small pieces of DNA that can move from one position in host DNA to another - provides organism with mechanism for DNA rearrangement
i) insertion sequence: Short DNA seq w/ inverted repeats - insert in multiple target sites in genome or plasmids
ii) Transposons: carry additional genetic material (may confer ability for antimicrobial resistance & toxin formation)
iii) Other moveable elements (e.g. conjugative transposons) - act as circular intermediates & transfer resistance genes directly following cell-cell contact
Bacterial Species
-strains
- Is a population of cells with similar characteristics -> each species can be made up of 1000s of strains
- strain = subset of a bacterial species differing from other bacteria of the same species by some identifiable difference
- strains can vary in their biological properties (e.g. virulence, antibiotic resistance)
- New strains continuously arising
Microbial evolution (5 factors that facilitate the evolution of microbes)
- High mutation rates of microbes
- Microbial interspecies genetic exchange
- Immunosupresion of host - avirulent strains survive
- host behaviour change - alter route/dose of infection
- change in ecology of host-microbe balance - more chance of infection
