Documente Academic
Documente Profesional
Documente Cultură
Research Project Proposal: The effect of pesticides and chemicals such as Acetamiprid,
Azoxystrobin, and Imidacloprid on bacteria found fruit and vegetables such as E.coli.
Hunter Galindo
Silva Topchyan
Biomedical Sciences
Period 8
West Career and Technical Academy
Purpose Statement
Methodology
Materials:
E.coli bacteria
Acetamiprid pesticide
Imidacloprid pesticide
Azoxystrobin pesticide
petri dishes
gloves
goggles
spray bottles
surgical/gas mask/fume hood
inoculation loop
bunsen burner
incubator
compound light microscope
Safranin
alcohol
Crystal Violet
slides
Iodine
bibulous paper
distilled water
ruler
notebook
calculator
writing utensil
Procedures :
Part I: Conducting experiment
1. Upon receiving the required materials, begin with recording and observing the basics of
the E.coli using standard classification methods. Before opening the bacteria, cleanse the
1. Begin observations with describing the colony and content of the petri dish. Begin Gram
staining to observe structure of E.coli .
Get sample for the Gram stain.
Transfer a drop of the sample to be examined on a slide with an inoculation loop. Add a
drop distilled water on the slide and aseptically transfer a small amount of a colony from
the petri dish. You only need a very small amount of sample. Add 1-2 drops of the
sample onto a glass slide. Spread it evenly on the slide to form a thin smear with an
inoculation loop, which can be done by sliding the edge of another glass slide across the
glass slide. Allow it to air dry.
Heat fix the smear, by quickly passing it two to three times through a bunsen burner.
Allow it to air dry.
Flood the smear with crystal violet, wait thirty seconds.
Gently rinse off the crystal violet with distilled water. Do not rinse too much, it will
remove the stain from the bacteria.
Flood the smear with iodine, Leave three seconds.
Gently rinse iodine with distilled water.
Decolorize by adding alcohol to the smear while holding the slide at an angle to drain the
coloring. Stop when runoff becomes clear, within seconds.
Gently rinse off excess color with distilled water.
Flood the smear with safranin counterstain.
Gently rinse off excess safranin with distilled water. Gently blot slide with bibulous paper
to remove distilled water and leave it to air dry. The Gram stain is done.
1. Once initial observations have been taken, separate the E.coli into four separate petri
dishes. Be sure to divide the bacteria into equal amounts.
2. Use petri dish A. Within the first dish, take the spray bottle containing acetamiprid
pesticide, and spray the bacteria three times to ensure that every inch of the bacteria is
fully washed by the acetamiprid pesticide. Immediately close the lid afterwards to
prevent contamination.
3. Use petri dish B. Within the second petri dish, take the spray bottle containing
imidacloprid pesticide, and spray the bacteria three times to ensure that every inch of the
bacteria is fully washed by the imidacloprid pesticide. Immediately close the lid
afterwards to prevent contamination.
4
1. After seven days take out the petri dish and begin collecting results.
2. Collect sample from each of the tree petri dishes and begin gram staining (refer to # 2).
*END OF DAY 14*
Safety:
Be sure to wear surgical gloves at all times when handling either E.coli bacteria or
pesticides as either one can cause harmful effects if brought into contact with the skin. If
so, wash your hands immediately and intensely, afterwards, get your instructors attention
for further instructions.
Wear goggles at all times as well when experimenting with one or both substances to
avoid getting either into your eyes. Contacts will need to be removed as potential mixing
with pesticide chemicals can cause detrimental effects. If pesticides make contact with
your eyes, flush them thoroughly with distilled water and seek medical attention
immediately
When working with the two types of pesticides, wear a mask that will protect you from
the strong fumes created from the application process to the bacteria - these pesticides are
hazardous and may cause severe harm if inhaled for an extended period of time.
Tie back long hair and avoid loose fitting clothing/jewelry when working with both the
bacteria and pesticides along with running the bunsen burner. Use all fire safety cautions
when using the bunsen burner, i.e never reach across an open flame, never leave a burner
unattended, make sure to turn the gas off when finished with all fire related proceedings,
etc.
Make sure to properly wash your hands upon entering and leaving the laboratory.
Washing your hands before getting started will protect samples from being exposed to
contamination, and washing your hands after all experimental proceedings will protect
you and other students who come into the vicinity from being exposed to potential
harmful agents within the pesticides and bacteria.
Reference:
10