Group Members: Chua Gerlynn Debra Eugenia

1.Introduction 2.Procedure 3.Data Table 4.Graph 5.Conclusion 6.Reflections 7.Credits

To find out the bacteria content in bottles under different conditions. Independent variables:  If soap was used to wash/rinse the bottles  How long the bottle has been used (no. of months) Dependent variables:  Number of bacteria colonies Our group thus wanted to find out how these variables will affect the cleanness of the water bottles, thus too the bacteria count.

Methods of extracting bacteria from bottles Method 1 1. Pour away all present water in the bottle. 2. Add distilled water to a bottle 3. Using a sterile cotton bud, swipe the inside of the bottle 4. Place the cotton bud into a Falcon Tube of water. Method 2 1. Pour all present water in the bottle. 2. Add distilled water to a bottle 3. Gently shake the bottle. 4. Pour the water into a Falcon Tube. Choice: Method 2  More convenient and easier  thus reducing the chances of making mistake in the experiment

Hypothesis (1) The longer the bottle was used for, the more bacteria will be present in the bottle. Hypothesis (2) The bacteria count in water bottles washed wish soap is lesser than the bacteria count in water bottles washed without soap. Constants - type of bottle: reusable - how often the bottle was washed - material of bottle: plastic

1.Pour all liquid in the bottles out 2.Measure 50ml of distilled water using the 50ml-Falcon Tube 3.Pour the 50ml of distilled water measured into each bottle 4.Shake the bottle up and down twice 5.Pour ‘contaminated’ water into 50ml-Falcon Tube and label 6.Pour 4 ml of distilled water into 6 Falcon Tubes for the 5x dilution 7.Pour 9ml of distilled water into 12 Falcon Tubes for the 10x and 100x dilution

8. Use the micro-pipetter to extract 1ml of ‘contaminated’ water from the 50ml-Falcon Tube and add into the 6 Falcon Tubes of 4ml of distilled water and shake. This gives us 5x-diluted water 9. Use the micro-pipetter to extract 1ml of ‘contaminated’ water from the 50ml-Falcon Tube and add into the 6 Falcon Tubes of 9ml containing distilled water and shake. This gives us 10x- diluted water 10. Use the micro-pipetter to extract 1ml of the 10x-diluted water and add into the remaining 6 Falcon Tubes containing 9ml of distilled water and shaker. This gives us 100x-diluted water 11. Label the nutrition agars with the person’s name, the date and the concentration of dilution

12. Use the micro-pipetter to extract 0.4ml (4 micro-litre) of the 5x-diluted water and add it to the correct nutrition agars plates 13. Dip the L-shaped glass rod in 95% alcohol 14. Place the rod over the fire for a few seconds then let it cool near the flame for another few seconds 15. Place the tip of the rod on the nutrition agar to check if it is cooled 16. Spread the water around the nutrition agar while using a finger to spin the plate of agar around

17. Use paraflim tape to seal the plates of nutrition agar 18. Repeat steps 11 to 17 for the 10x and 100x dilution 19. Leave the plates of nutrition agar into an incubator at 35  c overnight 20. Return the next day to count the number of colonies of bacteria visible 21. Record down observations

Range for the duration: A- Less than 1 month B- 1 to 3 months C- 4 to 6 months D- 7 to 9 months E- 10 to 12 months F- 13 to 15 months Range for bacteria colonies that were uncountable: A- wide spread, cloudy B- Too many to count, little space left on plate where bacteria have not grown on. C- Too many to count but some place still left on plate where bacteria have not grown on.

NameWash with soap? DurationBacteria Count JeanNoA46,875 Pan ChuenNoB12,890,625 Whai PengNoC1,409,375 MarielNoCA StellaNoE37,500 Timothy INoEA Timothy IINoEB EugeniaYesB2, Yi HuiYesB1,537,500 Jia YiYesD2,506,250 CleoYesEB Keat MunYesF453,125

Number of people against number of bacteria colonies (according to range)- comparing usage of soap ABCDEF Number of people Number of bacteria colonies With soap Without soap Range for number of bacteria colonies: A- A type B- B type C- below 10,000 D- 100,000 to 1,000,000 E- 1,000,001 to 3,000,000 F- More than 3,000,000

Number of people against number of bacteria colonies (according to range)- comparing duration of usage Range for number of bacteria colonies: A- A type B- B type C- below 10,000 D- 100,000 to 1,000,000 E- 1,000,001 to 3,000,000 F- 3,000,001 to 6,000,000 G- More than 6,000,000 4 to 6 months 1 to 3 months 7 to 9 months 10 to 12 months 13 to 15 months Less than one month ABCDEF Number of people Number of bacteria colonies

Reusable bottle Maojia: washes daily, without soap, metal bottle, contained green tea, used for 4 months  B (TNTC, little space left on plate where bacteria have not grown on ) Chen Yang: washes weekly, no soap, used for 6 months  bacteria colonies Non-reusable bottles Jaryl: does not wash bottle, Evian mineral bottle, used for less than a day  No bacteria colonies seen Junhan: washes daily, with soap, used for 1 month  bacteria colonies

From our data table  the longer the bottles were used (under the bottles not used with soap), the more bacteria was present in the bottles.  one result which does not match  Pan Chuen had approximately 13 million of bacteria  most likely experimental error For the experiment on the bottles with or without usage of soap  the results did not go along our hypothesis  results were irregular without any fixed pattern  results does not fit our second hypothesis (the bacteria count in water bottles washed wish soap is lesser than the bacteria count in water bottles washed without soap)

Yu Jia (Chua): The experiment was very interesting for we had fun trying out the experiments ourselves while learning new things. However, we might have made some experimental errors as they did not exactly match logically thinking. (E.g. more bacteria in bottles washed with soap compared to less bacteria washed without soap) Furthermore, halfway through all these discussions, we met up with a HUGE problem that we did not exactly solve… Gerlynn: Basically, I think that we really have learnt much from this project through all the hands-on experiments I have also learnt much about the importance of unity and planning in advance. The results collected were a little disappointing as they showed that we did make some experimental errors but I think that it’s the process that counts.

Debra: Well, this project has strengthened the bond between my friends and I as we spent a lot of time together, going through thick and thin. Although some of the results were quite disappointing, we manage to sort of tide over this crisis. Eugenia: I think that this project is very fun and interesting indeed as we got to use many new apparatus which we do not get to use as much during classes. We went through a lot of problems such as getting stuck at a point of time but I’m glad we overcame those problems. Furthermore, I enjoy mirobiology loads!

Ms Ivy Cheong, Teacher-In-Charge Mr Yap Jiawei, Biology Department Lab Officer Fung Ai Wei, Year 4 And everyone else who lent us your water bottles