Week 3

Question 1

what does microbial growth rely on?

Answer 1

nutrition, growth cycle, how microbes are cultured

Question 2

what are the macronutrients that must be supplied from the environment

Answer 2

Macronutrients:
-Carbon, nitrogen, phosphorous, hydrogen, oxygen, sulfur to make up carbs, lipids, nucleic acids, proteins
-Magnesium, Iron, Potassium, Calcium to make up enzyme cofactors and regulatory molecules

Question 3

what are the components of metabolism

Answer 3

Metabolism=catabolism +anabolism

Anabolism=builds molecules, requires energy, endergonic reaction

Catabolism=breakdown of molecules, exergonic, releases energy, provides energy for anabolism

Question 4

describe how classification is based on metabolism

Answer 4

Phototrophs: energy from sunlight
-autotrophs
-Photoautotrophs
-Chemoautotrophs (chemolithotrophs)
- Heterotrophs (chemoautotrophs or chemolithotrophs)

Chemotrophs: energy from chemical compounds
-Autotrophs,
-Heterotrophs
-Photoheterotrophs
-Chemoheterotrophs (chemoorganotrophs)

Question 5

what are the micronutrients that must be supplied from the environment

Answer 5

Micronutrients:
-Cobalt, copper, manganese, molybdenum, nickel, zinc to make up components of cofactors or enzymes

Question 6

nitrogen cycling

Answer 6

• Nitrogen fixers (diazotrophs) have nitrogenase enzyme that concerts inorganic N2 to ammonium ions, only a few bacteria can do this
  ○ Rhizobium bacterial species are nitrogen fixing root endosymbionts of plants
  1. Nitrogenase fixes atmospheric N2 to ammonia
  2. Nitrifiers oxidise NH4+ to generate energy
  Denitrifies use oxidized forms as alternative e- accepters

Question 7

microbial growth cycle

Answer 7

• Some bacteria divide by binary fission, where a parent cell increases in size/cell volume and biomass before splitting into two equal daughter cells
  ○ DNA replication, formation of division septum ( FtZ ring forms), cell separation
  Population doubles at each division (generation)

Question 8

how does bacteria divide asymmetrically

Answer 8

○ Cell does not divide in the middle
Forms daughter cell on only one end that buds off mother cell

Question 9

what is generation time

Answer 9

time It takes for population to double
○ As cells grow in closed environment, might not have abundant nutrients produces products which are often toxic
○ In favourable environment, with unlimited resources, bacteria divide at a constant interval
○ For cells undergoing binary fission, N t = N 0 x 2n
where N t is the final cell number,
N 0 is the original cell number, and
n is the number of generations

Question 10

what are the 2 main forms of culture media

Answer 10

Liquid Media or broth:
Cells in suspension
Used to study growth characteristics of a pure culture to obtain large number of cells

Solid Media
Gelled/solidified with agar
Cells grow as colony forming units (CFU, pure culture) (a colony is a visible group of microbial cells that developed from the same mother cell
Used to try to separate bacteria in samples to isolate for pure culture and specific traits, amounts of cells, density

Question 11

explain the complex/rich, minimal defined, and enriched types of microbial culture media

Answer 11

Complex/rich: nutrient rich, don’t really know the exact composition

Minimal defined: contains only nutrients that are essential for growth, exact components, concentrations known

Enriched: complex media that have added specific factors, microbe cannot make it but needs it to grow

Question 12

explain the selective and differential types of microbial culture media

Answer 12

Selective: favours growth of one organism over another

Differential: exploits biochemical/physiological differences between 2 species that grow equally well

Question 13

how is the macconkey medium use ful

Answer 13

○ Selective: only gram-negative bacteria grow on lactose MacConkey
○ Differentials: only a species able to ferment lactose produces pink/red colonies, gram negatives appear white or uncoloured

Question 14

how is pure culture isolated, and what is the goal

Answer 14

2 techniques, dilution streaking and spread places

To obtain isolated single colonies to establish pure cultures or estimate total number of bacteria in a sample
Assumes one cell=one colony

Question 15

explain dilution streaking

Answer 15

○ Sterilized loop picks up small amount of sample
○ Dragged across surface of agar plate
○ Flame loop sterilized/cooled
○ Touch to end of last streak, pick up some bacteria, repeat
○ Flame loop and repeat
Dilution of the same of each streaking increases probability of single bacterium, producing a visible colony

Question 16

explain how to use spread plates

Answer 16

○ Set up 10-fold serial dilutions in liquid culture
○ A small amount of each dilution is plated on agar medium
○ Goal: obtain a dilution with separated colonies, calculate number of bacteria in original sample
○ Potential exam question: multiply number of countable colonies by 10m get number of cells in 1.0mL of the dilution (10 x10 to the negative 5)
By multiplying that number by the reciprocal of the dilution factor, we can calculate the number of
cells (colony-forming units, or CFUs) per milliliter in the original broth tube (107 ́ 10 1 ́ 10 5 =1.1 ́ 10 8 CFUs/ml)
TNTC = too numerous to count.10 th of a mL;10-fold dilution)

Question 17

why do we count microbes, and what are the techniques used?

Answer 17

• Food industry regulations, health applications
  
  • quality control (water, safety, contamination)
  • research
  • Methods:
    ○ Microbes/bacteria must be suspended in liquid to be counted
    ○ Expressed as number of bacteria/mL of liquid of number of bacteria/g of a solid
    ○ Counting procedures:
    § Plate counts (dilution plating)
    § Optical density with a spectrophotometer
    § Direct microscopic counts with hemocytometry
    □ Slide has grid with 9 large squares divided into 25 smaller boxes, consisting of 16 smaller boxes
    □ Has measurements to calculate volume, can get counts/volume, counts/mL

Question 18

what is the use of optical density

Answer 18

• Microbial cells in liquid suspension increases turbity of the liquid
  ○ Higher concentration of cells, the more turbid the suspension
  ○ Measured as light scattered by suspension at specific wavelength of the suspension, readings reflect density of the cell culture (higher reading=higher density)
  ○ Amount of refraction based on how big the cell is
  Density of the culture, not measure of number of cells