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New Jersey Center for Teaching and Learning

Progressive Science Initiative

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www.njctl.org

BIOLOGY

Bacteria Growth Lab

Prokaryotes & Viruses

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Prokaryotes Unit Topics

· Bacterial Growth

Click on the topic to go to that section

· Analysis & Conclusion

· Antibacterial Agents

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Bacterial Growth

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Bacteria reproduce by binary fission. This proliferation by cell division results in rapid bacterial population growth.

Bacterial Growth

Bacterial Growth Animation

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As previously mentioned, bacteria can live almost anywhere and do live almost everywhere!

In the lab, bacteria are grown in a medium that contains the nutrients necessary for growth and survival.

Bacterial Growth

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Bacteria can also be grown on a gelatinous, polysaccharide substance called agar that is spread out on the surface of a petri dish. Bacteria are then "plated" on the agar.

Agar

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Agar contains:

Salts : provide essentia l e lements like magnesium, phosphorous, nitrogen, and sulfur needed for making prote ins and nucle ic acids.

Sugar: provides a source of carbon

Water: solvent

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Bacterial populations typically form colonies - groups of millions or billions of bacteria that live in a tightly packed area.

An individual bacterium can only be seen with a microscope, but a bacterial colony growing on a petri dish is visible to the naked eye.

Bacterial colonies growing on a plate of nutrient agar. Hans Knoll Institute. Jena, Germany

Bacterial Colonies

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Plating bacteria on agar allows one to visualize the different types of bacteria that may be present in a solution and to isolate a particular strain of bacteria for further use.

Plating Bacteria

Different types of bacteria can be distinguished by their distinct morphologies (shapes, structures, colors, or patterns).

Streak Plating Demonstration

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A culture is a solution that is optimal for the growth of a specific type of bacteria.

Bacterial cultures are used primarily to grow isolated strains of bacteria in the lab.

Cultures

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When bacteria are placed in an environment that provides all of their metabolic needs, bacterial populations grow rapidly until they have depleted all of the nutrients in their environment.

Bacterial Growth

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There are four phases to the bacterial growth cycle:*lag phase*log phase *stationary phase and *death phase.

http://www.ryancshaw.com/Files/micro/Animations/Bacteria lGrowth/PLAY_bacterial_growth.html

Bacterial Growth Phases

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During the lag phase, the bacterial cell grows and prepares for cell division

Lag Phase

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During the log phase the bacterial population grows rapidly - doubling in size with each generation.

http://www.ryancshaw.com/Files/micro/Animations/Bacteria lGrowth/PLAY_bacteria l_growth.ht

Log Phase

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watch the number of cells here

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watch the cell number take off!

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The stationary phase is reached when the bacteria start to run out of nutrients and the number of bacteria dying is equal to the number of bacteria reproducing.

Stationary Phase

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After nutrients have been used up and waste builds up, bacteria begin to die in large numbers and only very few reproduce.

Death Phase

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Calculating Bacterial Growth

It is possible to estimate the number of bacteria present in a culture after a period of time using the following formula:

(2n )(initial number of bacteria) = total number of bacteria

n = the number of generations.This can be determined by using the following formula:

total time generation time

(time it takes the bacteria to divide)

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At a fourth of July picnic, a bowl of potato salad was left out in the hot sun for 2 hours. If the salad originally contained 250 Salmonella enteritidis bacterial cells and Salmonella divides every 10 minutes, how many bacteria would be present at the end of 2 hours?

Givens:Initial number of bacteria = 250total time = 2hrsgeneration time = 10 minutesn = total time = 120 min. = 12 generation time 10 min.

Calculating Bacterial Growth

(2n )( initial number of bacteria) = final number of bacteria212 x 250 = 1,024,000 bacteria

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1 Bacillus cereus divides every 30 minutes. You inoculate a culture with exactly 100 bacterial cells. After 3 hours, how many bacteria are present?

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2 Staph. aureus, a disease causing bacteria, divides every 15 minutes at body temperature. A student gets a cut on her hand, is infected with 10 Staph. aureus bacterial cells, and does not clean the wound. After 1 hour, how many bacteria are present?

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Antibacterial Agents

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Antibacterial Agents

Antibacterial agents are chemical substances that either kill bacteria or inhibit growth. There are 3 main types:

antiseptics disinfectants

antibiotics

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Antibacterial Agentsantiseptics: chemicals used to kill or inhibit growth on living tissues (examples - hydrogen peroxide, soap, mouthwash)

disinfectants: used to kill or inhibit growth on nonliving objects (examples - bleach and boiling water)

antibiotics: chemicals produced by living organisms (fungi or bacteria) that inhibit growth of bacteria (examples - penicillin and tetracycline)

NOTE: some antiseptics are also disinfectants and vice versa

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How do we know they work?

An experiment was done to test the effectiveness of various disinfectants and antibiotics. The experiment had 5 steps.

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How do we know they work?Step 1

A sterile cotton swab was inserted into a culture of E. Coli bacteria.

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The tip of the cotton swab was then placed on the agar plate, streaking the sample of bacterial culture across the plate as show below to inoculate the plate with bacteria.

How do we know they work?Step 2

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3 filter paper disks were dipped in 3 different types of disinfectants and placed in the middle of each quadrant of the first petri dish. Distilled water was placed in the 4th quadrant as a control

Disinfectant 1: bleachDisinfectant 2: hydrogen peroxideDisinfectant 3: window cleaner

How do we know they work?Step 3

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3 disks coated in 3 different antibiotics were placed in the middle of each quadrant of the second petri dish. Distilled water was placed in the 4th quadrant as a control. Antibiotic 1: ampicillinAntibiotic 2: penicillinAntibiotic 3: tetracycline

How do we know they work?Step 4

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Both petri dishes were then taped shut, flipped upside down to prevent condensation from dripping onto the bacteria, and placed inside a 37 degree incubator for 48 hours.

How do we know they work?Step 5

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Predictions and ResultsMake your prediction about whether bacteria is killed...

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1

3

2

41

3

2

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Results Dish 2

The clearer areas surrounding the disks

are called zones of inhibition and

indicate that bacterial growth was

inhibited.

Predictions and Results

Results Dish 1

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Predictions and ResultsCHECK if a zone of inhibition is present on the plate

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Another mechanism of inhibiting bacterial growth is to infect the bacteria with a virus.

A drop of solution containing bacteriophage viruses was added to the bacterial culture below resulting in several clear zones.

These clear zones are called plaques. Viral plaques indicate that bacteriophages have infected and killed bacterial cells. Plaques may be counted and used to determine the number of viruses present in a bacterial culture.

Fighting Bacteria with Viruses

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Analysis & Conclusions

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1). Which disinfectant most effectively inhibited the growth of E. coli?

Analysis and Conclusions

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2). Which antibiotic was most effective in preventing the growth of E. coli?

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3). How do you know that any inhibition you have observed is due to the disinfectants and antibiotics on the disk?

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4). If the same experiment was repeated using a different strain of bacteria and a zone of inhibition was observed around the disk soaked in window cleaner, what could account for the different results?

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5). You wake up one morning with a severe sore throat and notice white dots lining your tonsils. You quickly visit the doctor, who diagnoses you with a Streptococcus infection (strep throat) and prescribes a 10-day antibiotic treatment.

After 3 days, you begin to feel better. Why should you continue taking the antibiotic for the full 10 days?

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6. What causes a zone of inhibition?