Microbiology MOOC

Question 1

gram-positive versus gram-negative bacteria

Answer 1

utilize different forms of quorum sensing systems.

Question 2

bacterium can turn pathogenic when different combinations of the quorum sensing genes are activated

Answer 2

e.g. the virulence of S. aureus increases significantly.

Question 3

Commensal bacteria can produce a quorum sensing effect on the Vibrio cholerae

Answer 3

that changes its gene expression and restricts its ability to become established in the gut and cause disease

Question 4

fimbriae

Answer 4

which enable them to stick to surfaces.

Question 5

containment measures

Answer 5

prevent transmission of pathogens

Question 6

autoclave, incineration or chemical disinfectants

Answer 6

decontamination

Question 7

biological safety cabinet

Answer 7

BSC

Question 8

HEPA filters in the cabinet

Answer 8

remove bacteria and viruses from the air as it exits the ventilated workspace inside, preventing pathogens from being released into the environment.

Question 9

biosafety levels

Answer 9

depend on:

• infectivity
• potential severity
• transmissibility
• prevention/treatments

Question 10

when using water

Answer 10

it should be sterile

Question 11

aseptic techniques:

Answer 11

• close to a Bunsen burner; sterilise air

- flame the neck of the vial

Question 12

pipettes

Answer 12

do not touch glass vial

Question 13

use sterile spreader

Answer 13

to cover agar with suspension

Question 14

colony morphology

Answer 14

unique

Question 15

Streak plate

Answer 15

• culture of one species
• sterilise innoclation loop in bunsen
• cool the loop to touch the colony
• transfer to agar plate
• repeat then streak

Question 16

Tryptone Soya Agar

Answer 16

TSA

Question 17

vortexer

Answer 17

to release microbes

Question 18

Microscope slide preparation

Answer 18

• pure culture
• sterilise innoculation loop to place sterile water onto slide
• take a single sample from colonies
• spread into water to create a smear
• pass slide through the bunsen burner to heat-fix it

Question 19

heat-fixing

Answer 19

firmly attaches bacterial cells to the slide so that the stain doesn’t wash them off

Question 20

Gram staining

Answer 20

• flood the smear with crystal violet for on minute
• tip off excess
• flood with gram iodine and leave for a minute
• tip of excess and keep it tilted, decolurising with ethanol until it runs almost clear
• rinse with tap water
• flood with saphronin to counterstain the smear for 45s
• rinse with water
• blot dry with absorbant paper

Question 21

Identification of gram positive bacteria

Answer 21

• immersion oil on stain smear
• x100 objective lens
• cells are stained purple and rectangular - rod

Question 22

Gram positive bacteria appearance explanation

Answer 22

have a wall that has a thick layer of peptidoglycan, which retains the crystal violet-iodine dye after washing with ethanol; appear purple

Question 23

Gram negative bacteria appearance explanation

Answer 23

• have a much thinner layer of peptidoglycan and an outer membrane that contains porin channels
• when the cells are washed with ethanol the crystal violet-iodine dye is washed out and the cells decolourize.

Question 24

Further gram negative identification

Answer 24

• A pink dye (safranin or fuchsine) is used to counterstain, so that they are visible under the light microscope.
• Gram positive cells also take up the counterstain but it is not noticeable because the crystal violet stain is much darker

Question 25

Gram negative bacteria

Answer 25

- outer membrane phospholipid bilayer - porins - LPS in outer membrane - thin peptidoglycan layer between inner and outer membranes - high abundance of Braun's lipoprotein - periplasmic space - inner cell membrane (cytoplasmic membrane) phospholipid bilayer

Question 26

lipopolysaccharide (endotoxin)

Answer 26

- composed of a chain of polysaccharides that are anchored to the outer membrane via lipid A. - chemical composition differs between bacterial strains. - can induce a strong immune response and cause toxic shock.

Question 27

peptidoglycan

Answer 27

- composed of amino sugars that are cross-linked to form a crystal lattice - provides structural support to the cell and helps to withstand osmotic pressure

Question 28

Braun’s lipoprotein

Answer 28

links the outer membrane to the peptidoglycan layer and helps provide structural support to the cell

Question 29

Periplasmic space

Answer 29

contains periplasm

Question 30

periplasm

Answer 30

- gel-like matrix that fills the space between the outer membrane and the inner cell membrane - contains a variety of different proteins that play roles in important cellular processes including nutrient uptake, protein folding and the elimination of waste and toxic compounds.

Question 31

colony forming units per ml =

Answer 31

number of colonies / (dilution x volume plated)

Question 32

underestimate inaccuracies:

Answer 32

- did not count all colonies (repeat!) | - laboratory culture conditions

Question 33

effect of laboratory culture conditions

Answer 33

- e.g. oxygen conditions might exclude anaerobes or microaerophiles - e.g. temperature may not be at optimum

Question 34

how many microbes are there in soil?

Answer 34

between 1 and 100 billion (1x10^9 - 1x10^11)

Question 35

BLAST

Answer 35

basic local alignment search tool; searches a large database to find nucleotide sequences related to or similar to a sequence you input

Question 36

want a low e value

Answer 36

0.0001 - homologous | expected value - how similar the input to the output

Question 37

Sequencing

Answer 37

PCR

Question 38

Antibiotics act by

Answer 38

disrupting a specific cellular component (eg cell wall, cell membrane) or biosynthetic pathway (protein synthesis, nucleic acid synthesis, folate synthesis) within a bacterial cell.

Question 39

Penicillin

Answer 39

- a Beta-lactam antibiotic - inhibits penicillin binding protein enzyme that cross-links peptidoglycan layers in the bacterial cell wall. - specific to bacteria because only they have this polymer in their cell wall - more effective against Gram positive bacteria because they have a much thicker layer of peptidoglycan in their cell wall - when a bacterial cell replicates, it makes small cuts in the peptidoglycan layer in order to divide and to incorporate more peptidoglycan into the cell wall as it grows. - when a bacterial cell replicates in the presence of penicillin the peptidoglycan layer becomes weaker and weaker, and the cell ultimately bursts when it can no longer withstand the internal osmotic pressure.

Question 40

defence mechanisms against antibiotics

Answer 40

- produce a slightly modified cellular component that is no longer inactivated by the antibiotic (target modification) - produce an inactivating enzyme that modifies or destroys the antibiotic - produce an entirely new enzyme that is not sensitive to the antibiotic to bypass the inactivated component - reducing the ability of the antibiotic to reach the target by either decreasing uptake of the antibiotic or actively removing it from the cell using an efflux pump

Question 41

Penicillin resistance

Answer 41

- in Gram positive bacteria is predominantly via target modifications that prevent penicillin from binding to the PBP - in Gram negative bacteria is more commonly achieved by production of inactivating enzymes (𝛽-lactamases)

Question 42

Campylobacter jejuni is...

Answer 42

... a Gram negative corkscrew-shaped bacterium that lives as a commensal in the intestines of healthy birds, causing no obvious harm or benefit to the host.