Lecture 2- Observing Microbes

Question 1

Which scientist used rRNA gene sequencing to adapt Linnaeus’ five domains of life to just three?

Answer 1

Carl woese

Question 2

How are organisms grouped

Answer 2

Kingdom, phylum, class, order, family, genus, species

Question 3

What is binomial nomenclature

Answer 3

Genus and species e,g. Bacillus cereus.

Question 4

What denotes that the species is unknown?

Answer 4

Sp, after the genus e.g.

Clostridium sp.

Question 5

How was Carl woese’s method of creating the tree of the life into 3 domains of life different from the method of Linnaeus

Answer 5

He looked at things by a genotypic rather phenotypic way.

Question 6

What are the 3 domains of life

Answer 6

Bacteria (prokaryotes), eukaryotes (animals, fungi, plants) and archaea

Question 7

Who discovered archaea and and what makes them different?

Answer 7

Carl woese.

Live at extremes e,g, hot springs

Question 8

Humans are more similar to prokaryotes than archaea, true or false?

Answer 8

FALSE

Question 9

How many e.coli microorganisms are needed to kill a child?

Answer 9

ONLY TEN

Question 10

What are the ways to observe bacteria

Answer 10

Light microscopy, electron microscopy (transmission and scanning)

Question 11

What is the magnification and resolution of light microscopy?

Answer 11

Mag: 1000x
Res: 0.2um

Question 12

What is resolution.

Answer 12

The ability to be able to see two things seperately and not as one.

Question 13

What does the oil immersion in light microscopy do?

Answer 13

It immerses the objective and form a continuous barrel or tunnel for light to pass through.

Question 14

What is the difference between transmission and scanning electron microscopy?

Answer 14

Transmission allows us to see inside the organism studied, whereas scanning allows us to see the outside of the organism but in much higher detail than light microscopy

Question 15

What is the mag and resolution of electron microscopy

Answer 15

Mag - 100,000x

Resolution - 0.2nm

Question 16

Describe how electron microscopy works

Answer 16

Cathode ray to fire electrons down a tube. Wavelength much shorter.
Use magnets to guide and direct electron beam, get sample, in vaccum and observe electrons interacting with the sample and view on a screen.

Question 17

Describe the preparation process that must occur for transmission electron microscopy.

Answer 17

Take bacteria
Freeze it etc in matrix. Slice this matrix very thin (only 60nm) mounted in specialised frame to view them. Means we can see internal components,

Question 18

Hat two main stains are used to increase contrast in bright field microscopy

Answer 18

Methylene blue and crystal violet

Question 19

Describe the process of a gram stain

Answer 19

1- heat fixed smear, stick microbes to slide
2- take microscope slide and put on some sterile distilled water tapped on by a loop
3- take colonies off afar plate and emulsify on liquid or take liquid culture and place directly on. Only need 1 colony
4- expose to Bunsen burner for a second then press against hand. Should be warm to promote evaporation, if heated too much then can kill microbes.
5- white residual deposit is microbes ready for gram stain
6- add primary dye (crystal violet). Gram - peptidoglycan smaller and hidden from membrane so less accessible. Crystal violet impermeates through outer layer and peptidoglycan. LEAVE FOR A MINUTE THEN WASH OFF,
7- add second component called mordant - fix dyes into tissues. This is grams iodine (combines with crystal violet in cell walls and polymerises crystal violet into bugger crystals so embedded within tissue) all tissues gram + and - intense purple
8- alcohol wash (typically 90% ethanol sometimes acetone) and this de colourise gram -. Alcohol remove outer layer of gram - as it is lipid so lose the purple colour. Alcohol fixes peptidoglycan even further so dehydrating it so gram - de colourised. Gram + even more purple
9- counter stain - safranin or carbol fuschin(both very red) peptidoglycan in gram - pick it up but gram + already saturated with blue stain so stay that colour.

Question 20

Who in the 1700’s created the five domains of life, otherwise known as the tree of life?

Answer 20

Linnaeus