Max A.

Sutters

The Dirt Report: Microbiomes of Quest University

Purpose:
In this lab we investigated Quest microbiomes by growing and identifying bacteria on often
infrequently washed surfaces from several locations on campus. Participants also drilled general
Hmm from students bodies! We didn't collect any
campus samples!

laboratory protocol and practiced a staining technique for telling apart Gram positive and negative
bacteria.

Methods and Observations:

Plate Preparation:
Petri dishes filled with agar growth medium were prepared and marked into four quadrants prior to the
experiment. I chose three surfaces to swab: the belly button, underside of my left hand, and the surface
of my laptop keyboard. I prepared two sterile, double-ended cotton swabs: three ends for the bacteriacarrying quadrants, and a fourth for a negative control (NC). After swabbing, I gently wiped each end
of the Q-tips in a cross-hatch pattern to cover most of their respective quadrant. For the order of
samples swabbed, refer to Figure 2. After bacteria collection and labeling the petri dishes were stored at
What happened to figure 1?

37 C to incubate for roughly 24-48 hours.

We incubated them for 48 hours!

Colony Analysis:

Reverse order

The bacteria present were analyzed based on the appearance, form, and structure of the colonies. We
were given a list of terms to describe qualitative data. Refer to Figure 1 below for raw data gathered. I
estimated the number of colonies for the most populated quadrant by counting colonies in a small area
and assuming colony density remained constant for the rest of the quadrant. I measured colony
diameter from the largest colony present in the quadrant. Staining and viewing under a microscope to
What is the limitation with this?

determine shape and gram positivity of bacteria are explained in the next section. Cells in the table
marked with an X represent unclear data due to experimental error.

Max A. Sutters

Sample

Belly Button

Hand (underside) Laptop Keyboard NC

Number of colonies

500

18

13

Diameter (mm)

0.5

1.6

Form

Punctiform,
circular,
irregular

circular

circular

circular

Elevation:

flat

flat, convex, raised flat, convex

convex

Margin:

entire,
undulate,
lobate

entire

entire

entire

Texture

moist

moist

moist

moist

Colour

dull, opaque

shiny, opaque

dull, opaque

yellow, opaque

Shape

Gram

Positive

Figure 1: Raw quantitative and qualitative data gathered for bacteria samples and control. This

is a table not figure

As seen in the petri dish photograph in Figure 2, the hundreds of cultures incubated from the belly
button sample were largely punctiform, with few circular and irregular colonies. The margins of the
punctiform colonies were entire. I observed undulate and lobate margins on the circular and irregular
colonies. The colonies were flat, dull, and opaque. The 18 colonies incubated from the underside of my
left hand were all circular in form. Most were flat, but convex and raised colonies were observed with
some difficulty. Bacteria gathered from the laptop keyboard formed 13 circular colonies. They were flat
and convex in elevation, and dully opaque in color. Colonies in all quadrants were moist in texture, and
all colonies in quadrants 2, 3, and NC had entire margins. The presence of a bacteria colony in the
negative control quadrant could be attributed to accidental contamination of the cotton swab,
condensation dripping onto the quadrant when flipped, or simply that the Q-tip was not as sterile as the
packaging promised.

Max A. Sutters

Can you onto the quadrants with arrows?

Figure 2: Bacteria cultures after incubation,

swabbing, and further non-experimental
incubation.

Gram Staining:
Gram staining is a useful tool for determining the complexity of a bacteria's cell wall. Bacteria that
appear purple after a Gram test have been stained with crystal violet and have not been decolorized by
alcohol. Red-stained bacteria in the test have been stained with Safranin after decolorization. The cell
walls of Gram positive bacteria are thicker, simpler, have more peptidoglycan, and lack a
So would gram positive or negative be blue?

lipopolysaccharide layer compared to Gram negative bacteria.

Max A. Sutters

Put images after you refer to them in text.

Great idea to take pictures, but visibility
would be enhanced with different colour
background

Figure 3: Gram stained bacteria smears. From left to right: belly button, hand, laptop keyboard, NC (in corner).

I placed four drops of water on a clean slide with a sterile loop. Re-sterilizing the loop in a flame after
each sample, I gathered bacteria from a variety of colonies within each quadrant and mixed it into the
corresponding droplet of water. Once the water appeared cloudy in each droplet, I distributed it thinly
and evenly in a rectangular, upright smear (Fig. 3). In hindsight the ratio of culture to water was too
high, a mistake that became obvious later when bacteria cells were very tightly packed and mostly
unstained. The fourth smear on the same slide in particular was prepared with less water and the same
amount of bacteria compared to the previous smears. Swiping the slide through a flame too many times
while heat fixing destroyed cells and further explained the dearth of stained bacteria.

Analysis and Results:

To streamline the data I standardized qualitative descriptions (e.g. smooth and entire, coccus/cocci,
etc.) and selected the primary description in qualitative cells with multiple values (e.g.
convex(2),flat(6) flat). Cleaning data that way is very is extremely quick and dirty, but it would be
inconsistent and unworkable otherwise without separate categories for each description (with a yes or
no for each).

Not sure what you did here

Max A. Sutters

Conclusions:

R T test: one continuous variable, one categorical variable, only two categories
P: confidence interval

Max:

The start of this lab is well written. See
comments throughout to help with clarity and
formatting. Unfortunately, you don't have any of
the analyses which was a big component of this
lab or the conclusions/reflections. Please let
me know if you don't understand them, happy to
go over it with you again.

I know you took your 24 hour extension, but I
didn't receive the lab until Monday. This time I
won't deduct points for lateness, but in the
future please make sure your emails send!

C