Categorisation Flashcards
Microbial culture
A collection of cells that have been grown in or on a nutrient medium
Medium
Plural = media
A liquid or solid nutrient mix that contains all of the nutrients required for a microorganism to grow
Growth
In microbiology the word growth is used to refer to the increase in cell number as a result of cell division - a single microorganism can grow into millions and billions of cells to form a visible colony
Microbes live in communities cooperatively and in conflict
- typically live in complex microbial communities and their activities are regulated by interactions with each other, with their environments and with other organisms
- Microorganisms were on the land and in the seas for billions of years before plants and animals appeared - they are very diverse
- microorganisms are a major part of the earth’s biomass and are essential in sustaining life
- for example, oxygen we breathe is the result of microbial activities
- evolution of plants and animals are heavily influenced by by microbial and symbioses and by pathogens
- there are many ways to study microorganisms
cylindrical
A cylindrically shaped cell is called a rod or a bacillus (plural, bacilli).
Spiral
A spiral-shaped cell is called a spirillum (plural, spirilla).
Curved
A cell that is slightly curved and comma-shaped is called a vibrio.
spirochete
A spirochete is a special kind of organism that has a spiral shape but which differs from spirilla because the cells of spirochetes are flexible, whereas cells of spirilla are rigid.
Irregular shape
Some bacteria are irregular in shape. Appendages, such as stalks and hyphae, are used by some cells for attachment or to increase surface area. In addition, asymmetrical cell division such as budding can result in irregular and asymmetrical cell shapes.
Cell division and morphology
Cell division impacts morphology because cells that remain attached together can form distinctive shapes, eg. Some cocci occur in pairs (diplodocus) and some form long chains (streptococci)
Filamentous bacteria
Filamentous bacteria are long thin rod shaped bacteria that divide terminally and then form long filaments composed of many cells attached end to end
Size
- the bigger it gets the more it is limited by the nutrients
- small cells have more surface area to volume ratio than larger cells
- as the cell size increases the S/V ratio decreases
- S/V ratio controls many processes in the cells, including its growth rate and shape
- cellular growth rate depends in part on the rate at which cells exchange nutrients and waste products with their environment
- as cell size decreases, the S/V ratio of the cell increases, and this means that small cells can exchange nutrients and wastes more rapidly (per unit cell volume)than can large cells
- free living cells that are smaller therefore tend to be more efficient than those larger and any given mass of nutrients will support the synthesis of more small cells than large cells.
- Cell morphology is often predicated on the effect of cell shape on S/V ratio.
- Eg. Cell shapes that increase the overall membrane are of the cell, such as those having thin appendages or invaginations, allow bacteria to increase their S/V ratio for any given mass of cytoplasm
Gram staining
- Flood the heat fixed smear with crystal violet for 1 min (all cells remain purple)
- Add iodine solution for 1 min (all cells remain purple)
- Decolorize with alcohol for 20 sec (gram positive cells are purple and gram negative are colourless)
- Counterstains with safranin in for 1-2 min (gram positive cells are purple and negative are pink to red
Example tests
morphology
physiology - what conditions are required for the growth of the microorganism?
biochemistry
serology
bacteriophage
pathogenicity
DNA analysis
Physiology:
- At either too hot or too cold a temperature microorganisms will not be able to grow and may even die.
- As temperature rises the rate of enzymatic reactions increases and growth becomes faster
- However, above a certain temp, proteins and other critical cell components may become denatured or otherwise irrevocably damaged.
Cardinal temperatures
- for every microorganism there is also a minimum temperature below which the organism cannot grow. Optimum temp - where growth is the most rapid, max temp - growth is not possible above this
- These three temperatures are called the cardinal temperatures
- they can differ largely between species
psychrophiles
low temperature optima
mesophiles
midrange temperature optima
thermophiles
high temperature optima
hyperthermophiles
very high temperature optima
Tolerance to chemical inhibitors
- respiratory inhibitors
- Gram -ve bacteria are sensitive to sodium azide (NaN3) as it inhibits cytochrome c oxidase. Whereas Enterococcus (e.g. E.faecalis) and other Gram +ve bacteria are resistant - chaotropic agents - mess up water which can lead to protein desaturation
- Pseudomonas resistant to phenol - Antibiotics
- Mycoplasma lack a cell wall therefore resistant to penicillin - we can characterise bacterial isolates by scoring the ‘minimal inhibitory concentration’ (MIC) for range of antibiotics
- alternatively, antibiotic impregnated discs
Biochemical tests can be used to distinguish specific enzymes or metabolic pathways
E.g sugar metabolism
Fermentation
anaerobic breakdown of the sugar into smaller molecules with the release of waste products (sometimes ethanol), sometimes organic acids, sometimes gases). Different waste products characteristic of different species
Oxidation: breakdown of sugar to CO2 and water. An aerobic process:
Check notes for result colours
Catalase test
Bubbles of oxygen = catalase + e.g. staphylococci
No bubbles = catalase - e.g streptococci
Urease test
Urea —-> NH3 (pH rises —-> colour change)
- these and numerous other biochemical tests can be miniaturised and automated
- test strips contain dehydrated substrates + indicators in microtubes
- just add the bacterial suspension and incubte
- different strains give characteristic colour patterns
Serology
- detection of bacteria antigens using specific antibodies
- Expose to specific antibodies (e.g from blood serum)
- Test in vitro for recognition of the cell- surface antigens (with ELISA, agglutination, precipitation, complement-fixation and fluorescent antibodies and chemiluminescence
- serology tests can be similarly miniturised e.g in 96 well microlitre plates (look at notes for diagram)
Bacteriophage type
Bacteriophages - bacterial viruses. Most have a specific host range
Check notes for diagram
Pathogenicity
Pathogens:
- Specific symptoms
- definite host range
sometimes symptoms alone are enough to indicate identity. for example -
1. Dipheria - formation of pseudomembrane in the throat - corynebacterium diptheriae
2. Leprosy - loss of fingers and toes - Mycobacterium leprae
3. Plague - buboes, skin darkening - Yersinia pestis
i. Genome sequencing
- Numerous bacterial species now completely sequenced
- sequences do not give the complete answer to how a bacterium functions but do provide a huge amount of comparative information on biochemistry, physiology, phylogenetic relationships and evolution
- note* - determining the sequence and gene content of a bacterial genome is still a major undertaking - not what you would do initially to characterise a new isolate. However complete genome sequences can be used as refernce points when characterising new isolates
ii. Polymerase chain reaction (PCR)
- simple in vitro technique to amplify short, specific DNA sequences. Now used regularly to diagnose bacterial/viral infections
- PCR is incredibly sensitive - can be used to detect single bacterial cells
- With DNA based tests, we can identify bacteria without culturing them first - makes clinical
diagnosis much faster - many of these test are used in a clinical context
Check notes for diagram
(Meta)genomics
- most of our knowledge of bacteria comes from pure strains in culture
- but >99% of species are thought to be uncultivatable
