Who Lives in Your Water Bottle: A Lab Exercise With Student Relevance!

I often tell students, when teaching sterile procedure, to imagine that it is raining microbes all around them, all the time.  Tragically, when I think of this, I seem to hear "It's Raining Microbes" to the tune of The Weather Girls' "It's Raining Men" (written inexplicably by David Letterman's bandleader Paul Schaffer)---which would make a fine humorous microbiological-themed video!  

1980s humor aside, it is one thing to hear that microbes are in a cloud all around us, and another thing entirely to observe the reality firsthand.

Some investigators have approached this recently with the "Belly Button Microbiota" project (notice that they have now branched out into armpit microbiota).  That's well and good, and they have received a lot of positive press.  For some years, I have been assigning my own students the task of learning what is living in their water bottles. You know the reusable ones that students seem to carry everywhere?  "It's just water" suddenly becomes a fascinating oversimplification in my lab sections!

So this year, my students are thinking about their water bottles in earnest...and what microbes reside in them.  Heck, Kaitlin Reiss even made us a fine logo for our efforts!

In my first lab session, I have students bring in their water bottles, and we talk about how they treat those bottles (some never wash them, others are metaphorically quite religious on the topic).  I ask where microbes might come from in their water bottles.  I even discuss biofilms in relation to that topic---that is, that biofilm-forming microbes will have a great advantage under these conditions, especially for folks who simply rinse out their water bottles.  Then, I have the students do a simple swab of the inside surface (and sometimes the impossible-to-clean mouthpieces) of their water bottles onto various plates, parafilm them (I don't expect danger, but caution is always a good thing), and incubate at room temperature.

The students swab onto LB (rich in nutrients), PPYE (very low in nutrients), R2a (commonly used in the study of water quality), as well as MacConkey and EMB (to check for enterics and fecal coliforms).  I tell the students that some organisms actually prefer low nutrient conditions (nutrient broth and LB are not universal media!).  The results are interesting, if qualitative!

As you can see, student reactions run the gamut from unconcerned, to deeply worried, to mildly aggressive, to the level of microbial glee I personally feel!  Not to mention the frequent student who seems concerned with the odors of the plates.  Here are some examples of proud students in my microbiology course:

And the plates? I am no "fancy plate photographer," but here are some attempts I made with my own camera.  Everyone was a "winner" this year (in terms of finding microbes in their water bottles), from the person who bought their water bottle a few days before lab to the person who swears he or she obtained it from the Lost and Found in the gymnasium.  Some plates were covered with, um, a rich microbiota, while others were less populated.  Diverse colony morphologies were seen, as well as pigmented colonies and what appeared to be gliding microbes.

Some coliforms appeared here!

Here we have some collapsing colonies (Bacillus?)

Lots of diversity here, and wonderful colonial colors!

Another interesting group of colonies.

Some more yellow colonies, plus a gliding bacterium?

Lots of diversity here, again!

Mostly pink colonies, plus a fascinating colonial type in yellow!

And here is a student trying something extra!  Half the plate was swabbed from the interior surface of the water bottle, and half was swabbed from the mouthpiece---very different populations, it would appear!

Thus, it was a very direct demonstration to students of the prevalence of microbes in their own lives.  It was fascinating to hear their reactions, discuss the "histories" of their water bottles, and so forth.

But "who goes there?"  In the past, I have made 16s rRNA libraries from a water bottle eDNA sample, and had students look at one or two candidate clones (using the RDP "Classifier" program, as well as simple BLAST).  I have also had students do colony PCR of a given "pet microbe" with universal 16s rRNA primers, clean up the reaction, obtain sequence data, and classify.  But we couldn't afford to look at the populations that exist within water bottles.

This year, we hope to really expand this process, and teach my students (and me!) how to DEEPLY analyze what microbes are present in every water bottle.  I have a collaborator who is in the process of helping us with this...and I cannot wait to write about the process and what we learn!

For now, it gratifies me to know that my students can look at their water bottles and realize that they have microbial friends where they least expect them!

And...when is the last time you washed your water bottle, and how well?