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Cameron Galt

2013-2014 Period 4 Biology

Science Fair Project

 

Title:  Can Sunlight Effectively Disinfect Water?

 

Abstract:

Over 5,000 people die each day in developing countries from diseases caused by bacteria contaminated drinking water.  A simple and cheap method of effectively purifying water could save lives and improve health.  SODIS (Solar DISinfection) is simple, cheap and effective.  SODIS uses disposable PET plastic water bottles filled with contaminated water and placed in the sun for hours.  The ultraviolet (UVA) radiation in sunlight kills the bacteria over time and makes the water safe to drink.  This experiment investigated whether SODIS treatment for 12 hours or more is as effective at killing bacteria as boiling water for 5 minutes.  Small plastic water bottles were filled with bacteria contaminated water and placed under a UVA light bulb to simulate sunlight.  UVA exposures of 12-48 hours were tested.  Samples of SODIS treated water were cultured in petri dishes.  Samples of contaminated water were boiled for 5 minutes, cooled, then cultured.  Samples of distilled water were also cultured.  An incubator was constructed to provide an optimal environment for the cultures to grow.  Bacteria counts were made daily.  The experiment showed that zero bacteria growth occurred in the samples treated by SODIS for 12+ hours, boiling for 5 minutes, or distilling, but untreated samples resulted in an average count of 3.8 bacteria colonies.  This experiment demonstrated that SODIS works.  Exposing bacteria contaminated water to 12 hours of sunlight kills bacteria, making the water safe to drink.  Using SODIS to treat contaminated water in developing countries could save lives and improve health.

 

Objective/Investigative Question:

Can UVA radiation in sunlight effectively disinfect water contaminated with bacteria?

 

Variables:

Phase 1:

Independent variable - Water treatment process

Dependent variable -  Bacteria count

In the first phase of the experiment, the water treatment process used will be varied and the bacteria resulting will be counted.

Phase 2:

Independent variable -  UVA exposure time for SODIS treatment

Dependent variable - Bacteria count

In the second phase of the experiment, the time the contaminated water is exposed to UVA light will be varied and the bacteria resulting will be counted.

Controls:

Distilled water was cultured to detect accidental bacterial contamination of materials.

Sterile procedures were used to avoid accidental bacterial contamination.

Incubator temperature was maintained at optimal growth temperature for all samples.

 

Hypothesis:

If bacteria contaminated water is exposed to sunlight (UVA radiation) for 12 hours then it will be disinfected as effectively as by boiling.

 

Materials:

 

  • Harmless bacteria sample (Ward’s Science #85 W 1664 Escherichia coli K12)

  • Aquarium water sample

  • Tryptic soy agar growth medium (Ward’s science Agar Powder 28 grams)

  • Petri dishes, 24 count, sterile (Amazon.com)

  • Disposable gloves (Target)

  • Ziploc bags, quart size (Target)

  • Refrigerator

  • Glass medicine droppers, 4 (Target Pharmacy)

  • Glass stirring rods, 4 (Amazon.com)

  • Computer

  • Distilled water, two 1 gallon bottles

  • Clear disposable drinking water bottles, 8 oz. Arrowhead

  • Daylight Blue Reptile Bulb UVA source, 150 watts, Zoo Med item #DB-150 (Petco)

  • Heat lamp socket, 10.5 inch hood heavy duty (Home Depot)

  • Lamp stand

  • Digital thermometer, 0.1 degree F accuracy, Taylor 9831-21 (Target)

  • Heating pad, Sunbeam Xpress Heat XL Model 2013-912, 180 watts, with stay-on feature (Walgreens)

  • Tape measure

  • Metal straight edge

  • Large cardboard box, approximately 24” x 22” x 24”

  • Styrofoam sheet, 2” thick

  • Long razor knife with 7” retractable blade, Dewalt (Home Depot)

  • Wire lath or hardware cloth (Home Depot)

  • Rebar tying wire (Home Depot)

  • Large nails, 4

  • Needle nose pliers (Home Depot)

  • Tin snips (Home Depot)

  • Duct tape (Home Depot)

  • Blankets

  • Aluminum foil

  • Dark metal cookie sheet

  • Cooking pot

  • Range top and oven

  • Anti-bacterial cleaner, Formula 409 (Target)

  • Metal tongs

  • Lab notebook

  • Clock

  • Digital scale accurate to 1 gram (Amazon.com)

  • Permanent marker

 

Procedure:

1. Order Harmless Bacteria

2. Order tryptic soy agar growth medium

3.   Order petri dishes

4. Collect the remaining items in the materials list.

5. Wear disposable gloves whenever contaminated or sterile items are handled.

6. Sterilize all of the tools to prevent them from adding bacteria to the sample:

a) Preheat an oven to 225 degrees F.

b) Cut a piece of aluminum foil and bend it into a shallow sheet pan.

c) Put the utensils, including medicine droppers, glass rods, glass jar and its lid in a pot and fill it with tap water.

d) Place the pot on a burner, heat it to boiling, boil for 5 minutes, and turn it off.

e) Use metal tongs to move the utensils from the pot to the foil sheet.

f)   Place the foil sheet in the oven for 15 minutes to dry the utensils.

g) Remove the foil sheet from the oven.

h) Wrap the utensils completely in the foil to keep them clean.

7. Mix the agar and assemble the petri dishes by following the instructions included with the agar medium:

a) Weigh 20 grams of Agar

b) Measure 500 ml distilled water

c) Combine the Agar and distilled water in a sterile cooking pot and mix thoroughly using a sterile glass rod.

d) Heat the mixture on a cooktop while stirring continuously.

e) Boil for one minute.

f) Remove from heat, cover and allow to cool for 10 minutes.

g) Carefully pour melted agar mixture into each petri dish to form a ⅛ inch deep layer.  Cover each petri dish with its lid immediately after filling.

h) Allow the petri dishes to cool to room temperature on a countertop and the agar to set to a stiff gelatin texture.

i) Stack the petri dishes in Ziploc bags, seal the bags, turn them upside down and place them in a refrigerator until needed.

8. Construct the incubator:

a) Measure cardboard box

b) Lay out the cuts on the styrofoam sheet  in order to cover the outside of the cardboard box.  Styrofoam should overlap at the corners.  This will require cutting the top and bottom sheet to the same size as the box.  The sides will be cut 4 inches taller than the box dimension and 2 inches wider than the box dimension.  Use the razor knife and straightedge to cut the styrofoam.

c) Construct the insulating box cover using the cut pieces of styrofoam sheet and the duct tape.  Leave the top styrofoam piece unattached and construct a handle using duct tape so that the top can be easily removed to open the box.

d) Cut the metal lath or hardware cloth 4 inches larger than the dimensions of the top of the box using tin snips.

e) Using the straight edge, bend the metal lath up 90 degrees 2 inches  from each edge in order to form a rack which will fit into the box.  Cut out the corners of the lath as necessary.

f) On the front of the cardboard box, measure down 4 inches and in from the side 4 inches. Mark the spot. Do the same on the opposite edge of the front.  Repeat on the back of the box.

g) Using a nail, poke holes at the marked locations on the front and back of the box.

h) Two wires will support the metal lath rack in the box.  String a wire through a hole in the front of the box to the corresponding hole in the back of the box.  Cut the wire 12 inches longer.  Do the same for the second wire.

i) Using a nail and the needle nose pliers, twist an end of a wire around a nail.  Stretch the wire by pulling from its other end and twist it around a nail to hold it in place.  Do the same to the other wire.

j) Place the heating pad in the bottom of the box.

k) Using the razor knife, cut a small slit in the front of the box and thread the heating pad’s power cord through the slit.

l) Place the wire lath rack in the box on top of the wire supports.

m) Connect the heating pad’s power cord to a power source.

n) Set the heating pad to its highest temperature setting (6) and select “Stay On” on the control pad.

o) On the front of the cardboard box, measure across to its horizontal center and measure down 2 inches from the top of the cardboard box and poke the thermometer through the styrofoam and cardboard into the box.

p) Monitor the thermometer until it registers 98.6 degrees F.

q) Reduce the heater setting to medium low (2).

r) Continue monitoring the temperature for the next few hours until a stable 98.6 degrees F can be maintained.  It may be necessary to wrap the box in blankets in order to maintain a correct and stable temperature.

s) When a stable temperature of 98.6 degrees F is established, the incubator is ready to use.

9. Install the Daylight Blue Reptile Bulb in the clip-on light socket, clip it to the stand with the bulb facing down 8 inches above the work surface, and place it in a location on the work bench near an electrical outlet and where it will not be disturbed.  The Daylight Blue Reptile Bulb will provide a consistent UVA radiation source, to simulate sun exposure.

10. Prepare the SODIS Samples:

a) Prepare the bacteria sample as directed on its packaging.

b) Using a new, unopened bottle of distilled water, open the bottle and discard approximately ½ gallon of water.

c) Label this bottle “Contaminated e coli.”

d) Add the bacteria sample to this bottle.  Replace the cover and shake thoroughly.

e) Remove the labels from 3 new 8 oz. clear plastic water bottles.  Open and empty these bottles.  Rinse the interior of the bottles using distilled water.

f) Fill the 3 clean 8oz. clear plastic water bottles full with water from the 1 gallon jug labeled “Contaminated e coli” and screw on the caps.

g) Keep the “Contaminated e coli” jug in a cool, dark place to prevent UVA exposure and excessive bacteria growth.

11. Treat the SODIS Samples:

a) Turn on the lamp.

b) Adjust the lamp position as necessary to 8 inches from the work surface.

c) Put the dark metal sheet under the light.

d) Put the water bottles containing the contaminated samples under the lamp on top of the dark metal sheet.  Make sure the light is aimed directly at the samples and that each sample receives the same light exposure.

e) Record the start time in the lab notebook.

f) Leave the samples sitting under the light undisturbed for the lowest SODIS exposure time being tested.

g) The next section can be completed while the SODIS samples are being treated.

12. Prepare and Test the Contaminated, boiled and distilled Water Samples:

a) Measure 1 cup of water from the “Contaminated e coli” gallon bottle into the pot and put it on the burner.

b) Set the burner to high and bring the water to a boil.

c) Continue boiling for 5 minutes.

d) Remove the pot from the burner, cover the pot and let the water cool to room temperature.

e) Put on fresh disposable gloves.

f) Get 3 tryptic soy agar plates.

g) Get a clean glass stirring rod and a clean medicine dropper from the foil packet.  Fill the dropper with the room temperature boiled water.

h) Remove the cover from one of the plates and place 3 well-separated drops of the boiled water on the soy agar.

i) Use the glass stirring rod to smear the water drops in a zigzag pattern on the surface of the soy agar.

j) Replace the plate cover.

k) Repeat steps h) through j) with two more plates and the same medicine dropper.

l) Use a permanent marker to write on the top of each plate the sample number, start time, date and treatment process (Distilled, Boiled, Untreated, or SODIS.)

m) Place the plates in the incubator where they can sit undisturbed.

n) Repeat steps e) through m) using distilled and contaminated water.  At this point, there will be a total of 9 plates in use.

o) Allow the 9 plates to sit for 24 hours in the incubator.

p) Remove the plates from the incubator.

q) Look at the plates and count the number of bacterial colonies.

r) Record in the lab notebook: Sample Number, Date, Time, Number of Bacterial Colonies, Observations.

s) Repeat steps p) and q) at 24 hour intervals (48, 72, 96 hours.)

13. Test the SODIS Samples:

a) After exposure to the UVA light for the SODIS time being tested, remove one of the sample bottles from under the light.

b) Using this sample, repeat steps 6 e) through 6 q).

c) Repeat 13 a) and b) at the selected SODIS test time increments.

14. Analyze the Data:

a) Use a scatter plot to display the data.  Plot on the x axis “Treatment Process” and on the y axis “Bacterial Count at 96 Hours.”

b) Use another scatter plot to display the data. Plot on the x axis “Time” and on the y axis “Bacterial Count.”

c) Note how the bacterial count changes for the different treatment types.

d) Use the data to determine whether SODIS is an effective treatment for reducing bacteria count and the minimum effective UVA exposure time.

e) Use a line graph to display the incubator temperature data.  Plot on the x axis “Time” and on the y axis “Incubator Temperature.”

f) Use the data to determine the temperature consistency of the constructed incubator.

15. Cleanup:

a) Soak all materials used, including plates and tools, in a 10% chlorine bleach solution for 2 hours.

b) Dispose of Petri dishes in the trash.

c) Thoroughly clean the work area with disinfectant such as 10% bleach solution, 70% ethyl alcohol, or commercial antibacterial cleaner (like Formula 409).


 

Results/Data Discussion/Calculations:

 

The graph “Bacteria Count by Treatment Method” shows that bacteria growth was observed only in the untreated samples.  All treatment methods investigated resulted in zero bacteria counts.  This suggests that SODIS treated water (12-48 hours of UVA radiation exposure), boiled water and distilled water are all free of bacteria and that the untreated water contains bacteria.  Culturing samples of distilled water was used as a  control to detect accidental contamination of utensils and materials.

 

The graph “Incubator Temperature vs Elapsed Time” shows the changes to the internal temperature of the constructed incubator over time.  Frequent changes in incubator temperature could result in error due to slowing of bacterial culture growth.  The target temperature was 98.6 degrees F, which is the optimal growth temperature for the e coli bacteria contaminating the water sample.  The temperature was consistent to within 1 degree F when the incubator remained unopened.  Temperature drops of up to 11 degrees F were observed when the incubator top was opened to put in or remove petri dishes.  This was necessary when starting new cultures or removing petri dishes for examination and counting bacteria.  The incubator temperature typically regained the target temperature of 98.6 degrees F within 1 hour of opening its top after temporarily increasing the heater setting to high.  To maintain the most constant incubator temperature, it was determined that all of the cultures should be examined at the same time and only once daily to avoid opening the incubator top more frequently.

 

This experiment demonstrated that the SODIS process with 12 hours of simulated sunlight is capable of killing bacteria in water.  People in areas with contaminated water could greatly benefit from using the SODIS process.  According to the World Health Organization, over one billion people have unsafe drinking water.  Over 5000 deaths a day are a direct result of diseases caused by contaminated drinking water.  The SODIS process is a cheap and simple method of providing safe drinking water and would lead to improvements in the health of people in developing countries.

 

The minimum SODIS treatment time investigated was 12 hours.  This resulted in a zero bacteria count in the cultures.  Further research suggests that a minimum SODIS treatment time of 6 hours may be effective in disinfecting bacteria contaminated water.  Additional testing at SODIS treatment times ranging from 2 to 8 hours would be needed to confirm or disprove that hypothesis.

 

The only calculations performed were to determine the mean of bacteria counts for similar bacteria cultures.  To determine the mean, the data values were added and the total was divided by the number of data values. For example:

(C1 + C2 + C3)/3 = C mean

 

 

Conclusion:

 

The experimental results show that SODIS treated water (for 12 or more hours), boiled water and distilled water are all equally free of bacteria.  This would suggest that each of these treatment processes results in water which is safe to drink.  The SODIS process of water treatment was successful in killing all of the bacteria present in the contaminated sample water when the sample was exposed to 12 hours to 48 hours of UVA radiation, which is present in sunlight.  For experimental consistency, a UVA light bulb was used to represent sunlight.  It is reasonable to conclude that sunlight would produce a similar result.

 

The constructed incubator proved to be able to maintain a constant temperature near the target temperature.  This provided a suitable environment for bacteria culture growth.  It was determined that opening the incubator top only once daily resulted in a more constant temperature environment for the cultures.   

 

The experimental results agree well with the hypothesis.  Exposing bacteria contaminated water to UVA radiation with the SODIS process for 12 hours resulted in disinfection.  Boiling bacteria contaminated water for 5 minutes also resulted in disinfection.  The hypothesis, “If bacteria contaminated water is exposed to sunlight (UVA radiation) for 12 hours then it will be disinfected as effectively as by boiling” was confirmed.

Research:

 

Over one billion people do not have safe drinking water, according to the World Health Organization.  Bacterial contamination of drinking water is a widespread problem in developing countries.  About 80 percent of disease is a result of bad water and poor sanitation.  These can lead to diseases such as cholera, dysentery and typhoid.  These diseases cause over 5000 deaths each day.  Bacteria contaminated water causes diarrhea.  In these areas, diarrhea is the most frequent cause of death in infants.  It also leads to serious illnesses in children and adults and often death.

 

In order for water treatment to be practical in developing countries, it must be inexpensive and simple to do.  People in developing countries generally cannot afford to pay for expensive water treatment processes.  Even if a highly technical water process is setup by an outside group, maintenance of the treatment plant will usually fail in the long run due to a lack of trained local workers and money.

 

Research by EAWAG, the Swiss Federal Institute of Aquatic Sciences and Technology, suggests that a minimum of 6 hours of sunlight exposure is needed to kill bacteria in water.  The process they developed for treating water with sunlight is called SODIS, which stands for SOlar water DISinfection.

 

SODIS is simple and cheap to use.  Disposable PET plastic water bottles can be filled with water from bacteria contaminated local wells, rivers or lakes and put in the sun.  The UVA radiation in the sunlight will kill the bacteria within about six hours.  Local people can easily use SODIS with very little training and expense to treat their own water.

 

SODIS kills bacteria, viruses and parasites like giardia and cryptosporidia.  One researcher said “We have yet to find a waterborne disease that’s not significantly affected by solar disinfection.”  The method also works with low water and air temperatures.

 

SODIS works to kill disease causing organisms because UVA radiation damages the pathogens’ DNA and cell walls.  The sun also heats the water in the bottle making the pathogens’ cell repair mechanisms less effective.  Hydroxyl radicals are also created and these break the long chain molecules which make up the pathogens’ cells.

 

Studies have shown that leaving plastic water bottles in the sun does not release carcinogens into the water.  This disproves one of the most common objections to SODIS.

 

SODIS only works when there is sunlight.  Because of this limitation, chlorination is a better, long term solution if it is affordable for the local people.

 

Where people are trained to use SODIS the health of the population generally improves.  According to the World Health Organization “Solar disinfection is an example of another measure with proven health impact that requires little capital investment on the part of end users, and is thus appropriate for the very poor.”

 

SODIS is now in daily use in 33 countries by about 2.5 million people, but it could be useful in many more places.  It has been difficult to convince people that SODIS is effective, even though research supports this.  People often do not believe that putting water in an old plastic bottle in the sun will do anything to make the water safer.  This has kept SODIS from being used in more of the places where it could be helpful.  Many lives could be saved if the use of SODIS could be expanded to more areas.

 

Bibliography/Works Cited:

 

Bachus, Sue E. "Tryptic Soy Agar Powder - Preparation & Equipment Use." Agar Powder Preparation Recipe. Science Stuff Inc., 1 Jan. 2013. Web. 11 Oct. 2013. <http://www.sciencestuff.com/playground/ts_agar_powder.shtml>.

 

Editor in Chief. "SODIS - A Place in the Sun | Latest Features | Physics.org." SODIS - A Place in the Sun | Latest Features | Physics.org. Institute of Physics, 7 Oct. 2009. Web. 11 Oct. 2013. <http://www.physics.org/featuredetail.asp?id=39>.

 

"Simple Way to Remove Mud from Drinking Water." ScienceDaily. Michigan Technological University, 01 May 2012. Web. 04 Jan. 2014. <http://www.sciencedaily.com/releases/2012/05/120501134315.htm>.

 

"SODIS METHOD." SODIS:. Swiss Federal Institute of Aquatic Science and Technology, 24 May 2011. Web. 11 Oct. 2014. <http://www.sodis.ch/methode/index_EN>.

 

McGuigan, Kevin G. "Solar Water Disinfection (SODIS): A Review from Bench-top to Roof-top." Solar Water Disinfection (SODIS): A Review from Bench-top to Roof-top. Journal of Hazardous Materials, 15 Oct. 2012. Web. 04 Sept. 2013. <http://www.sciencedirect.com/science/article/pii/S0304389412007960>.

 

Zeitung, Schwabischen. "Eawag: Welcome." Eawag: Welcome. Eawag Aquatic Research, 11 Dec. 2013. Web. 04 Oct. 2013. <http://www.eawag.ch/index_EN>.

 

Fisher, Michael B. "Solar Water Disinfection (SODIS) of Escherichia Coli, Enterococcus Spp., and MS2 Coliphage: Effects of Additives and Alternative Container Materials." Solar Water Disinfection (SODIS) of Escherichia Coli, Enterococcus Spp., and MS2 Coliphage: Effects of Additives and Alternative Container Materials. Water Research, 15 Apr. 2012. Web. 04 Oct. 2014. <http://www.sciencedirect.com/science/article/pii/S0043135411008426>.

 

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