Lecture Set 1 : Part 2

Question 1

who was robert hooke?

Answer 1

-first to see and describe microbes
-used a compound microscope with 30x magnification (small but enough for discovery)
-observed cork cells, bread mold filaments (1st microbe), and the beginning of cell theory

Question 2

why could robert hooke not see bacteria?

Answer 2

-magnification of the microscope did not allow for it
-bacteria were still too small

Question 3

what is the basis of cell theory?

Answer 3

-all living things are composed of cells

Question 4

who was antoni van leeuwenhoek?

Answer 4

-lens maker
-built microscopes that maginfied specimen by 50-300x
-observed single celled microorganisms
-made the first discovery of bacteria (lens magnification was large enough)

Question 5

what did leeuwenhoek call the single celled microorganisms?

Answer 5

-animalcules

Question 6

what did leeuwenhoek do that stunted the evolution of microbiology?

Answer 6

-did not share the discovery of how to build his microscopes with high magnification

Question 7

what does it mean for a microscope to be a compound microscope?

Answer 7

-2 lenses to magnify the image

Question 8

who was louis pasteur?

Answer 8

-disproved spontaneous generation
-studied wine and beer production (fermentation)
-developed a method of gentle heating to kill unwanted bacteria (pasteurization)

Question 9

what is the process of fermentation that pasteur discovered?

Answer 9

-yeasts convert sugar to alcohol in the absence of oxygen
-la vie sans air (life without air)

Question 10

what was the idea of spontaneous generation? what countered this?

Answer 10

-living things can be created from nothing
-biogenesis countered this idea (ex: microbes can only arise from other microbes)

Question 11

what did pasteur discover about bacteria in wine?

Answer 11

-bacteria can sour wine by converting alcohols to acid
-shows that microbes can change their environment

Question 12

what was the experiment that pasteur did to disprove spontaneous generation?

Answer 12

-prepared meat infusions inside of long swan neck flasks
-put the infusion into a flask and boiled it to sterilize it
-used the heat to reshape the flask
-dust and microorganisms get trapped in the bend of the flask
-liquid remains sterile as its not exposed to the open air
-if the flask is tipped and the liquid contacts the dust and microorganisms, the liquid putrefies (gets DIRTY)

Question 13

what came from pasteur’s experiment?

Answer 13

-development of methods for controlling growth of microorganisms (aseptic technique)
-the steps for transferring microbes between growth media without contamination (air microbes, liquid droplets, or surfaces) to maintain pure culture

Question 14

who was robert koch?

Answer 14

-determined that a pathogen causes disease
-developed a system for determining this
-studied anthrax which helped him determine (epidemics in livestock caused by bacillus anthracis)

Question 15

how did robert koch study anthrax?

Answer 15

-he isolated bacteria from the diseased animal
-he then injected healthy animals with the bacterium
-those healthy animals then became ill with anthrax
-he then re-isolated the bacteria from the test subjects and showed that it was identical

Question 16

what did koch’s study allow him to do?

Answer 16

-establish a set of criteria for relating a specific microbe to a disease
-named them koch’s postulates (4)

Question 17

what is koch’s 1st postulate?

Answer 17

-the suspected pathogen must be present in all cases of the disease and absent from healthy animals
-technique used is microscopy and staining

Question 18

what is koch’s 2nd postulate?

Answer 18

-the suspected pathogen must be grown in pure culture
-technique used is laboratory cultures (streak plating)

Question 19

what is koch’s 3rd postulate?

Answer 19

-cells from a pure culture of the suspected pathogen must cause disease in a healthy animal
-technique used is experimental animals

Question 20

what is koch’s 4th postulate?

Answer 20

-the suspected pathogen must be re-isolated and shown to be the same as the original
-technique used is microscopy and laboratory cultures (streak plating)

Question 21

what are 4 exceptions to koch’s postulates?

Answer 21

-some pathogens cannot be put into pure culture
-HIV is a human specific disease, cannot use animals and humans would not want to be injected
-some pathogens can cause multiple diseases depending on how they enter the body
-a single disease can be caused by multiple pathogens

Question 22

what did koch realize about obtaining pure cultures?

Answer 22

-solid media was a simpler way to obtain them

Question 23

why is solid media better than liquid media?

Answer 23

-easier to see visually
-can immobilize cells to an extent
-cells will pile to form colonies

Question 24

how is liquid media solidified?

Answer 24

-adding agar

Question 25

what is agar? what are its characteristics that make it useful for creating solid media?

Answer 25

-polysaccharide derived from marine algae -melts at 97 degrees celcius (never incubating this high) -polymerizes (solidifies) at 43 degrees celcius (incubators are typically in a range from 25-40ish degrees celcius) -cannot be degraded by microorganisms (not a food source and no nutrients)

Question 26

what is the medium makeup of a typical petri plate (agar plate)?

Answer 26

-nutrient broth medium + 1.5% agar -broth medium is peptone, beef extract, and NaCl -the medium is brought up to 1 litre with distilled water

Question 27

what is the purpose of the different elements in a broth medium?

Answer 27

-peptone (sources of protein, carbon, nitrogen, and energy) -beef extract (sources of vitamins, growth factors, and proteins) -NaCl (for osmolarity)

Question 28

what is an isolated colony?

Answer 28

-one colony that is not touching other colonies

Question 29

what is the streak plate technique?

Answer 29

-primary method for isolating pure cultures -done with a solid medium (agar plate) -using an inoculate loop one edge of the plate is inoculated with a concentrated sample of bacteria -the sample is then diluted by streaking it across the surface of the plate (smearing peanut butter) typically in 3 sections -the plate is then incubated

Question 30

what is the purpose of diluting the sample as plating within a streak plate?

Answer 30

-deposits individual cells onto the plate that are separate from other cells -the individual cells will grow to form colonies

Question 31

what temperature are plates incubated at?

Answer 31

-25 - 42 degrees celcius -each temp has a different purpose

Question 32

what is a colony?

Answer 32

-a mass of cells that ideally arose from one single cell -can be used to create a pure culture

Question 33

what is the spread plate technique?

Answer 33

-sample is diluted before plating (serial dilution) -diluted sample is spread over the surface with a sterile glass spreader -separate cells grow into colonies on the SURFACE of the plate -use a 0.1mL volume (100 microlitres)

Question 34

what is the pour plate technique?

Answer 34

-sample is diluted before plating (serial dilution) -diluted sample is mixed with molten agar (liquid) -colonies will form embedded WITHIN the plate AND on the surface -use a 1mL volume (1000 microlitres)

Question 35

which techniques are used for counting and calculating the concentration of bacteria in a population?

Answer 35

-spread plate -pour plate

Question 36

which techniques are used for creating pure culture?

Answer 36

-streak plate

Question 37

what temperature is the molten agar used for pour plating?

Answer 37

-45 degrees celcius -we want to be able to touch without burning ourselves -we dont want to kill the bacteria

Question 38

what is a bacterial titre?

Answer 38

-concentration of bacteria in a population

Question 39

how do you calculate the titre for bacteria?

Answer 39

-titre = # colonies / volume x dilution factor -titre is expressed in cfu/mL or g

Question 40

what is cfu? what is the purpose of this measurement?

Answer 40

-cfu is colony forming units -used rather than individual viable cell #'s to account for clumps that contain more than one viable cell

Question 41

what is the purpose of a serial dilution?

Answer 41

-allows for us to obtain a countable number of colonies -10 fold dilution (# of colonies formed between dilutions should be a multiple of 10 apart, 7 > 70 > 700)

Question 42

what is the dilution liquid of a serial dilution?

Answer 42

-typically water -could also be a mineral salt solution or growth medium (can yield a higher recovery)

Question 43

what is the countable range?

Answer 43

-30-300 (greater than this will have crowding and growth issues) -less than 30 is not statistically significant -greater than 30 colonies will grow into each other (inaccurate counts)

Question 44

what do you do when you have more than one countable plate?

Answer 44

-calculate the titre of each and then take the average of them