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ORAL MICROBIOLOGY > Lecture 1 - Introduction to microorganisms > Flashcards

Lecture 1 - Introduction to microorganisms Flashcards

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1
Q

What are relative sizes of the following:
Bacteria
Fungi
Viruses

A

Bacteria - 0.5-5 um
Fungi - 5-100 um
VIruses - 0.002 - 0.05 um

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2
Q

What is the relation of micron (uL) to mL

A

um = micron = micrometer

A MICRON IS 1/1000 of a MM

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3
Q

How much bacteria is on earth?

A

10^30

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4
Q

How much bacteria is in the oral cavity?

A

10^10

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5
Q

How much bacteria is in 1 mL of saliva

A

10^8 cells/mL of saliva

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6
Q

What is the approximate amount of bacteria in body weight?

A

1-2% of body weight consists of bacteria

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7
Q

What is a thermophile?

A

High in temperature bacteria
Ex: live in hot springs, volcanoes
*this will be important in PCR

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8
Q

What is a psychrophile?

A

Low temperature bacteria

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9
Q

What is an acidophile?

A

Low pH bacteria (pH 1-5)
*this will be important when we talk about dental caries - those bacteria involved are acidophiles.
Acidogenic bacteria like Streptococcus mutans utilize sucrose in food to convert it to acid

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10
Q

High temperature bacteria

A

THERMOPHILE

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11
Q

Low temperature bacteria

A

PSYCHROPHILE

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12
Q

Low pH bacteria

A

ACIDOPHILE

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13
Q

What is a halophile?

A

High salt bactera

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14
Q

High salt bactera

A

HALOPHILE

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15
Q

High pressure bacteria

A

BAROPHILE

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16
Q

What is a barophile?

A

High pressure bacteria

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17
Q

We usually think of pathogenic bacteria = bad bacteria = cause disease, however, most bacteria in nature are NOT bad. THey serve vital functions like:

A
  1. ) Producing oxygen (cyanobacteia believed to be first oxygen producing organisms on the planet)
  2. ) Recycling Nutrients
  3. ) Decomposition of dead organic matter
  4. ) Digest food by producing enzymes that break down food
  5. ) Provide vitamins (Ex: vitamin B-12 produced by E. coli)
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18
Q

Not all bacteria in our body are bad, including in the mouth - they are required to prevent

A

Pathogens from colonizing

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19
Q

Skin, GI tract, mouth bacteria are all there for a reason, they have established their presence for a long time. If you wipe them out (ie. taking antibiotics) what happens?

A

You become prone to fungal infections such as oral thrush (candidiaasis) because you wipe you bacteria that normally prevents them from colonizing

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20
Q

If you wipe out flora with antibiotics and then go off antibiotics, what happens?

A

They come back in the same composition

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21
Q

Describe proper bacterial nomenclature using example (Staphylococcus aureus)
Describe what is identified where a

A

Genus species
Genus: Staphylococcus (First letter capitalized)
Species: aureus (lower case)
whole thing italicized

staphy = cluster of grapes
coccus = sphere
aureus = gold
The actual colonies on a plate look golden and in clusters of spheres

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22
Q

How are bacteria named?

A

Sometimes nomenclature describes shape/color but sometimes discovers the person who discovered the organism Ex: Yersinia pestis (Alexandre Yersin discovered it) causes Bubonic plague

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23
Q

When did the study of bacteria begin?

A

Late 1600’s

24
Q

Who was Anthony von Leeuwenhoek

A

He was a glass lens maker. In Netherlands, 1684.
Considered the father of microscopes
He would goto friends and ask for swab of plaque from their teeth , place it on his microscope:
brass panel w/ glass lens in middle. He would put sample on rod in front of glass and screw rod close until it came into focus.
His primitive microscope produced images that we can still recognize today.

25
Q

Is identifying bacteria a definite way of identification?

What are the shapes of bacteria?

A 
There are only so many shapes bacteria can be so that is not a definite way to identify bacteria. 
They can be:
Coccoid (round)
Bacillus (rod) 
Spirochete (spiral)

Shapes are not definitive way to identify, THEREFORE, you must ISOLATE THEM!

26
Q

Who was Robert Koch?

A

In 1880, Robert Koch developed the PURE CULTURE TECHNIQUE.
He put bacteria sample (from infected skin of a person) into beef broth. You get a turbid suspension. He found out that surfaces of potato wedges also grew bacteria well. One bacterium divides millions of times forming colonies of bacteria, visible as a small patches on the surface of the potato wedge. Different bacteria form different looking colonies (Staph aureus colonies look different from Strep mutans colonies) This is not enough to identify the bacteria but a good way to estimate how many different types are present in the sample.
Koch then began using gelatin and added it to beef broth to form cultures. Problematic w/ regard to gelatin withstanding temperature that was beset for bacterial growth (37 C).

27
Q

Who developed the PURE CULTURE TECHNIQUE?

A

Robert Koch

28
Q

Why was beef broth often used in culturing bacteria?

A

because it is very rich in nutrients and will grow lots of bacteria

29
Q

What is a turbid suspension?

A

A cloudy suspension of bacteria

30
Q

Why were potato wedges initially considered a good place to grow bacteria and why were they later ruled out?

A

Robert Koch found out that potato wedges could grow bacteria well because of the starch in them - it served as food for bacteria.

Problem with potato wedges is that they aren’t sterile - susceptible to fungi, etc…

31
Q

One bacteria divides millions of times and forms

A

A colony

32
Q

What is the optimal temperature for most bacteria?

A

37 degrees celcius

33
Q

What was the problem with using gelatin for culturing the bacteria?

A

37 degrees Celsius is optimal temp for bacterial growth - problem with heating gelatin to that temperature is that it melts.

34
Q

Who was Walter Hesse?

A

An associate of Robert Koch, when problems with gelatin culture melting, spoke w/ wife about cooking w/ agar - His wife, Fannie Hesse suggested using agar in the lab which remains semi-solid up to 55 degrees Celsius therefore bacteria can be incubated on agar medium.

35
Q

What is agar and benefit of it as a medium.

A

Agar comes from red seaweed - it is an excellent media because it can be melted but remains semi-solid up to temperatures of 55 degrees Celsius.
Also, bacteria don’t utilize agar as food so you can add different types of nutrients to the sample and no inference form original culture will occur.
Bacteria need glucose,, sucrose, galactose as carbon source and certain amino acids to grow. The agar is NOT a nutrient - it is just used as a semi-solid surface.

36
Q

Who was William Petri

A

William Petri invented the Petri dish to culture bacteria.
Petri dishes provided a solid surface to culture and study bacteria on. Agar - semisolid is often placed inside of petri dishes for use as a growth medium. The bacterial will grow on the agar as long as there are other nutrients on it - required by bacteria (Ex: glucose/ sucrose)

37
Q

What is a pure culture method?

A

Method of culturing to produce single colonies of bacteria - with the use of agar and petri dish generally.

38
Q

What must be done in order to isolate bacteria?

A

When you have a sample, in order to isolate individual colonies, you:

  1. Use a sterile toothpick/cotton swab/inoculating loop
  2. Use loop to to touch soil/patient’s mouth and streak a section in the plate
  3. Sterlizie the loop (or get new one depending on type you used)
  4. Turn plate 90 degrees and pull streaks out from the section of bacteria.
  5. Repeat this and thus you are DILUTING the sample of bacteria until you get isolated colonies of bacteria.
39
Q

What are Koch’s Postulates?

A
  1. Bacterium must be present in every disease case
    (Ex: if a group of patients all died from cholera, you must be able to isolate that bacteria from each person)
  2. Bacteria must be isolated and grown in pure culture.
    (About 50% of bacteria out there, we don’t know what they require for growth)
  3. You should be able to reintroduce the bacteria into a susceptible host and cause the disease to occur.
    (Ethically, we cannot do this in humans most of the time, but can use animal models,. the downside is that certain bacteria do not cause the same disease in humans as in animals.)
  4. You must be able to recover bacteria from the susceptible host.
    (In other words, you must be able to culture bacteria from the disease host that you infected)
40
Q

Some ______ bacteria cannot be grown outside of the host so animal tissue is used as a host rather than agar medium.

A

SPIRILLA

41
Q

What are the ways to enumerate bacteria?

A

Direct counting method (microscopy count)

Viable cell count

42
Q

What is the direct counting method?

A

This method uses a microscope and a counting chamber. The counting chamber is a glass slide with a grid printed on it.
1. Pipette a volume of your sample onto the grid (generally ~ 5 uL) and look at it under a microscope. Bacterial cells will be visible in quadrants of the grid.
2. Count the number of bacteria per grid - average it if number varies in grids
3. To get to total number of bacteria and calculate bacteria concentration do the following:
i. If for ex: 3 bacterial cells / quad
ii. If there are 100 quadrants on the grid, then there were about 300 bacteria on the slide (3 bacterial cells/quad X 100 quadrants on grid = 300 bacteria per sample.)
iii. Out sample was 5 uL so the concentration is 300 bacterial cells / 5 uL - we must REPORT BACTERIAL CONCENTRATION IN ML!!!!!!!!!!!!
To convert 5 uL to mL,, you have to multiply by the number that will give you 1000 uL (1 mL). Multiply 300 by 200 and you get 60,000 bacteria. The final concentration is 60,000 bacteria/mL or 6.0x10^4 bacteria/mL

43
Q

What is the serial dilution aka Viable Cell Count Method?

A

This method tells you how many are alive (viable) cells. we plate out 0.1 mL (100 uL) ALWAYS.
If you get a lawn (covered in bacteria and no distinguishable colonies) that means you have too many bacteria 20-200 is the range you want.

  1. From your source of bacteria, (stock solution) plate out 100 uL (0.1 mL) onto a solid medium in a petri dish. This is the typical size of a sample, since it is enough to get a statistically significant number of bacteria without overflowing the plate.
    This will produce a lawn of bacteria - not individual colonies, meaning you have far too many bacteria to be able to determine how many there are. TO determine this, serial dilution is done.

Serial dilutions are performed as follows:

i. stock solution has 5 mL, take 1 mL from the stock and put it in a new tube (lets call it TUBE 1) that contains 9 mL of sterile medium. The volume of this tube 1 now contains 10 mL. The dilution is 10x more diluted than the stock (a 1/10 dilution or 10^-1)
ii. Take a 0.1 mL sample of tube 1 and plate it. most likely, it is still too concentrated to produce isolated colonies, so you have to make more serial dilutions until you get a solution that is dilute enough to grow single colonies.
iii. Take a 1 mL sample from Tube 1 and put it into 9 mL of sterile medium in a new tube (tube 2) and place into 9 mL of medium in tube 3 - Dilution for tube 3 = 10^-3

Calculating concentration
iv. Lets say that the plate from tube 3 gives you 150 individual colonies. The bacteria are dilute enough WHEN YOU HAVE 20 - 200 INDIVIDUAL COLONIES!
After 200 - become too difficult to distinguish
Below 20 - too few to be significant
v. if you had 150 colonies on your third plate, then on the second plate you had 1500 (since 3rd plate is 1/10 dilution of the second)
vi. On the first plate, you had 15,000 colonies / 0.1 mL this MUST BE EXPRESSED AS COLONIES / ML THIS WAS CALCULATED PER 0.1 ML – so multiply by 10 to get 150,000/ML = 1.5X10^5
TO GET CONCENTRATION FO THE STOCK YOU MUST MULTIPLE BY 10 SINCE THE FIRST PLATE WAS A 1/10 DILUTION OF THE STOCK.
THIS GIVES YOU A STOCK CONCENTRATION OF 1.5 X 10^6 COLONIES/ML.

44
Q

Why is the Serial Dilution method preferred over Direct Cell Count?

A

The serial dilution method allows you to get viable cell counts - which is not possible with the direct counting method, which doesn’t tell you if the bacteria you are counting are dead or alive. A viable cell count will tell you the concentration of live bacteria in a sample.

45
Q

How much do we plate out in Serial Dilutions and why?

A

We ALWAYS plate out 0.1 mL = 100 uL of solution, because it is enough to get statistically significant numbers wihtout overflowing the Petri Dish

46
Q

What is the range of colonies that you want on a plate and why?

A

It turns out that the number of colonies are countable on a plate are between 20 and 200

< 20 – becomes insignificant (# is too low)
> 20 – becomes too difficult to count because they’re so close together and so many that you cannot distinguish them

YOU ARE SHOOTING FOR THE 20 – 200 RANGE!

47
Q

Why study microbiology?

A

All dental diseases are caused by microorganisms. Ex: caries, periodontal disease, gingivitis, root canal infections - all caused by bacterial infections.
Usually these infections are treated when it becomes a problem, here we learn how they case the disease and what the etiology of disease is.

48
Q

Bacteria that cause caries are _____ and ______

A

acidophilic and aciogenic

49
Q

What is acidogenic?

A

means acid producing, (in bacterial of the mouth this generally is bacteria that degrades enamel)

50
Q

What are the important bacterial functions?

A
  1. oxygen production
  2. recycling of nutrients
  3. Decomposition of organic matter
  4. Prevent pathogens from colonizing
  5. Digestion
  6. Provide Vitamins
51
Q

Oxygen production in bacteria

A

Because of biomass of bacteria, the amount of oxygen they produce is significant!!!
Arguably the most important function of bacteria.
Cyanobacteria are believed to be on the of the first oxygen producing organisms on the planet. Started to produce molecular oxygen about 1-2 billion years after the early was formed.

52
Q

Recycling of nutrients in bacteria

A

The carbon cycle, nitrogen cycle, phosphorus cycles are processes mainly accomplished by bacteria

53
Q

Prevention of pathogens from colonizing by bacteria

A
  1. Most of our mouth is colonized by “good” bacteria required to prevent pathogens from gaining a foothold or colonizing.
  2. The bacteria on our skin and in our GI tract and mouth are there for a reason and have established their presence over a long period of time. If you wipe them out, other organisms can colonize and cause disease.
    Ex: oral thrush in people on antibiotics
  3. The percentage and types of bacteria that you have in your mouth / GI tract is different than other people - it is difficult to make new bacteria colonize.
54
Q

Why does oral thrush (candidiasis) form in oral cavities of those who are taking antibiotics?

A

People on antibiotics over a long period become prone to fungal infections like thrush because the bacteria that normally prevent them from colonizing have been wiped out. if you get off of the antibiotics the flora comes back with the same composition as before.

55
Q

Bacteria help with digestion

A

some bacteria produce enzymes that help to digest food and obtain nutrients

56
Q

Bacteria provide vitamins example

A

E coli produces vitamin B12

ORAL MICROBIOLOGY (5 decks)

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