The “Finding-The-Few-Spots-In-The World-Without-Phages-In-Them” Award Goes To…

Coming from a high school where getting A’s and quickly, efficiently working through labs was the norm for me, this class definitely was a little bit of a reality check. When assignments were dependent on intellect and consistency, hard work and studying was usually gifted with good grades. When labs required precise measurements, steady hands, and a calm demeanor, I was usually rewarded with accurate results. However, in great contrast, I’m infamous at my school for having the worst luck in everything else, what we call “RNG,” or random number generator. When I needed a 4, I would get a 67. I even recall my dad telling me why he’s not fond of investing in the stock market, because “his father’s genes don’t carry luck.”

This class requires some of that luck that I seem to lack, at least at the start. Come day one, I got my dirt sample, and was ready to do the direct isolation procedure, expecting to see plaques the next day in lab. When I did come in, I was already pretty sure that there was nothing there. However, I’m a very stubborn person, so I guess I called something that was not a plaque, a plaque, and tried to streak it. Not only did those plates get contaminated to add insult to injury, but I happened to basically incise the top agar when I was streaking. Like most, I also didn’t really believe that like a billion phages exist in each plaque, and that when you touched them with the streaking stick, half of them latch onto it, and that when you make a streak, you’re laying out almost all of those half billion phages onto the plate.

I happened to try four different dirt samples, looking for phages, before I finally adopted some of Sabrina’s plates. Some students had plaques on day one, and I felt so utterly left behind. However, that’s one of the strong points about the class. You get to work at your own pace, and don’t get graded down for things that aren’t really your fault. And technically, I wasn’t really behind at all. There are always going to be chunks of the class at the same place that you are, so you aren’t alone in that sense either. Phage hunting in my opinion, is one of the best classes that a freshman can take. It’s so welcoming in both procedure and people, and it’s a great way to learn about more advanced lab procedures that you haven’t learned in high school.
Now, that I have Sabrina’s adopted plates, I’ve been streaking for about four days now, and I’m just trying to perfect my streaking technique so that I can have three generations of a single type of phage in my plates. Coming in at the beginning of class and seeing that there are actually plaques on my plates never gets old. It’s like Christmas every Monday and Wednesday, and I think that by the end of the semester, I’ll be able to look back and say that I was really satisfied with what I was able to accomplish in this class.


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If At First You Don’t Succeed…

Try, try, try again.

A polluted river. What a perfect place to get a virus living in the dirt, one might think. The person who thought that was me. Because I had gone to Hampden previously and had seen the sign near the river warning people to not go into the river because it contains harmful runoff, I thought that this area would be crawling with phages. I collected my dirt, making sure not to step into the river for fear of contaminating myself with whatever lied within. I went back to my dorm, confident that there were millions of little phages ready to lyse some M. smeg the following Monday.

I was wrong.

My first attempt at direct isolation ended up in plates that looked just like the control with no plaques on it and I sure was bummed. Maybe I just got unlucky? Maybe the phages were just tired and didn’t feel like lysing the bacteria? I knew the latter was clearly not true, but maybe I had just been unlucky. So then I tried direct isolation with the same filtrate, thinking that this filtrate that I had inside my microcentrifuge tube must be home to many phages.

I was wrong. Again.

I held back my anger as I looked at my second round of plates and saw exactly what I had seen last time; nothing. Just more plates that looked like the control. I felt like I had wasted so much time, plating and filtering and shaking these tubes, and for what? For nothing! It was with a heavy heart that I took another sterile tube and went back behind the UTL’s, begrudgingly scooping up dirt, knowing that I had to start all over again. All I needed was one plaque.

I returned to class with this new dirt sample and was told to do a different procedure; enriched isolation. Direct isolation, the procedure I had done the two times previous, had a smaller phage yield. Enriched isolation would increase my chances of getting a phage by a large margin. For the first time in a while, I had a real feeling of hope as I mixed some enrichment broth and 7H9 with my dirt. I extracted and filtered the supernatant and did serial dilutions up to 10^-5, plated them, and left class with a smile on my face. I knew that next class I would have plaques on my plate ripe for the picking.

I was wrong. Again. For the third time.


This time, however, it wasn’t that I had no phages. The issue was the I had too many phages. All of my plates, even the 10^-5 plate, were basically all cleared, indicating a very high phage concentration. I scratched my head because I didn’t know how to feel. I guess this was a good thing? I mean, at least I had plaques. I decided that maybe streaking some of these plates would help dilute these phages. Maybe, next class, I would have some plaques!

I think you’re probably catching onto the theme here…

If this class gave credit for having plates that are as clear as they were before putting them into the incubator, I would have an A+. The streaked plates I had been so excited about were clear and one was contaminated. After yet another defeat, the amount I wanted to hang up my lab coat and leave forever was at an all time high, but not because I had not succeeded. It was because I had to do serial dilutions again. But this time, I had to do ten rounds. The thought of that made my stomach turn. There were so many ways I could mess up and contaminate my filtrate and have to restart, especially because I was still not very good at the procedure. Facing my fears and all of the odds favoring me messing up aseptic technique, I began the process of doing ten serial dilutions. After completing the serial dilutions, I plated the following plates: 10^0, 10^-1, and 10^-6 through 10^-10. As sweat dripped down my face from the stress and from the bunsen burner’s proximity, I placed my plates into the incubator.

Oh, what a feeling.

Plaques on 10^-6.jpg

Plaques. Plaques everywhere. On my 10^-6 plate, I had a beautiful array of clear dots, followed by fewer on my 10^-7 plate and only 2 on my 10^-8 plate. My dilutions had actually worked. All of my failure, all of my problems, were all worth it. These lessons that imposed themselves on me are definitely some that I can apply to my future in research, and, in life.

Sometimes, things don’t work. A hypothesis is refuted, a plaque doesn’t show up. The reason we experiment is to learn and to try new things and to fail. In my eyes, a failure in the scientific community may be just as valuable as success. Failure allows us to step back and view what we have done and see what could have gone wrong or what we can do to improve our procedures. Failure is a blessing in disguise, and it’s important to not fall victim of the nasty appearance that we perceive failure as having. Sometimes, following up on an unexpected finding or something that went wrong can be more beneficial than what one was looking for in the first place. I am happy to say that my failures have helped me learn more about laboratory procedures, and I have definitely learned that patience, in the scientific community, is definitely a virtue.

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Phun Stuph Phursure.

Getting more Phun Part 2…

My enriched isolation i mentioned in “Not so Phun Part 1”, yeah. That didn’t work out. But, thankfully, I think I found what was going on: The procedure indicated that we had to open the enriched isolation tube every 24 hours for air to get inside. however, we did not, so anaerobic bacteria started to grow, and most likely denatured my phages. The group of people doing the enriched isolation with me also did not yield any phages. My lab partner, Jake, found phages, BUT it was because he had used a flask with an open top, so that air could get in. I did not use a flask at first, because there weren’t any left.

Proof of Adoption:


After determining what went wrong, and why I could not yield a phage, I had the choice of going through the enriched isolation process again. However, I instead chose to “adopt” a phage from Jake, in hopes of saving time. I labeled 4 plaques I identified from his old plate (he had not streaked these plaques yet), and streaked them on agar plates. Hopefully, these streakings will confirm that these plaques are indeed phages when I check back in two days.

Well, two days after, my plates indeed had plaques… but it was horribly infected. Luckily, there were some noticeable colonies that did not seem to have touched the contaminants, so i streaked from those, and hoped for the best. I also streaked from an extra not-so-contaminated plaque and filtered it with a 0.22 micrometer filter. After plating all those new streaks, I came back five days later to completely contamination-free plaques. The method worked, and I can finally go onto streaking for my generation 2 phages.

Contaminated example:


No more contamination!!:

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Phage Phuns aren’t even that Phunny

My apartment is next to a hiking trail marked by two large boulders. This trail leads into Yang Ming Mountain National Park and connects to a complex network of trails that weaves throughout a dormant volcano. As I hike this trail, streams of sunlight penetrate the thick canopy and gently light up the thin layer of fog that permeated the entire forest. Growing up within the borders an of national park, I am no stranger to nature. In fact, my passion for science was instilled by the rainforest that surrounded me. Prior to the semester-long phage hunting class, I was enrolled in a decade long class that involved hunting for the various insects that lived in the trees and under the rocks of the forest. Digging in the soil, I would find myself returning home with the usual earthworm or, when I’m lucky, a grub that would eventually metamorphose in a stag beetle.

While I thought that my usual outdoor excursions would be cut-short by my move to Hopkins, I was proven wrong before I even started the first day of class. Opening up my email, I saw a message from Dr. Fisher detailing a sample collection process for potential phages in soil. We were to venture out in search of the most interesting soil samples we could find in hopes of uncovering new and exciting phages. The similarities to what I’ve been doing were uncanny.

To those unfamiliar to it, nature can seem quiet and lifeless. However, years of exploration have open up my eyes to a lively and exotic world. In almost every puddle, tree branch, or clump of soil, one can find some sort of organism living in its niche environment. What I didn’t realize before was that this under-appreciated world was larger than I could have ever imagined.

To identify or even “see” the myriad of biological entities that roam the micro-environments of the soil involves a little more than just careful observation. Phages are, if anything, extremely small and plentiful, making it hard for a single individual to be characterized. To start, the soil sample I collected was shaken up with an enrichment broth solution and filtered for a direct isolation solution. This solution was then plated with Mycobacterium Smegmatis. Ideally, a reaction between the phages and the Smegmatis would cause a plaque to form. This, however, did not occur in the first four weeks of class. For weeks, I would come back to class only to find a contaminated, disrupted, or perfectly Smegmatis covered plate.

After four weeks of futile attempt after futile attempt, I came to the conclusion that two variables had to be changed. First, I started using a new soil sample. The sample I had first collected did not yield any results, leading me to question that quality of the sample. I decided to collect new samples directly from the grounds right outside the undergraduate teach lab. Second, I also learned to be more efficient and effective with my plating procedure. I discovered that the time it takes for the agar to set can be longer than expected. Aseptic technique is also a very crucial step to execute correctly in order to yield non-contaminated plates.

Armed with the knowledge and experience, I set out round two with high hopes. With the new soil sample, I filtered and did a serial dilution. Then, I directly pipetted 30 microliters of filtrates onto marked spots of my different plates. After 48 hours, I observed that there were plaques forming on the marked spots! Finally, after four weeks of work I finally got to see my phages show themselves for the first time.

With the plaques, I can now finally start the streaking process to isolate a single phage. Unlike my bug collecting adventures as a child, the process of finding a viable plaque was a long but very rewarding one. The journey has only begun, however, as generations of streaking is required to isolate a single phage sample.

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Are We There Yet?

Not so Phun Part 1…

I jumped out of my chair when i received a notification from blackboard saying I got off the waitlist for Phage Hunting. By then I had already given up hope of getting in, yet here I was.

A week later, I received yet another notification; we received our first assignment: collect dirt. Eagerly, I found my friends Roy and Jake, and we decided to go to the polluted river near Hampden in hopes of getting many phages. We collected dirt into our conical 50mL tubes, and excitingly joked about how we would discover mutated versions of phages.


On the first day of lab, I quickly performed asceptic technique on our bench and religiously followed lab procedures to ensure that I do not mess up. I tried my best to avoid any contamination by staying close (too close) to the fire. Sadly, a couple of days later, I discovered that my first plaque assay yielded no plaques.


I immediately tried again, and yet no phage samples were yielded. So…. I began to think. Maybe it was because my environmental sample did not include phages? I proceeded to attempt to solve this problem by collecting a new environmental sample from the back of the UTL lab. I also tried the method of enriched isolation- yielded a more concentrated solution of phages. After, I tried serial dilutions and plated my solutions, and even made a spot test. Sadly, there were no plaques yielded.

Spot Test:


I began to wonder. “It probably wasn’t my samples. I tried my best to not contaminate anything. Could it be the materials I used?” I observed from my many plaque assays that the agar was broken up. I also observed that each assay was significantly different from each other, so I thought to myself “It must be from 1) the agar 2) my process in making the enriched isolation.”

So now, a month into the class, I’m back to doing enriched isolations. I admit, it does feel very disappointing not seeing any phages. However, at the same time, it’s also extremely fun to pursue and try to isolate what went wrong. I may not be there (discovering a phage) yet, but surely, I’ll get there- as many trials (smart trials) as it takes.

*advice for future hunters*

You should always take pictures of your plaque samples “UNDER” the light apparatus for it to show up better. Also, be thoughtful of what could have happened (errors) as you do the experiment.

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Third Time’s the Charm

I can’t take heat, period.
It was a hot Wednesday and I needed sample. Dr. Fisher had recommended that we go explore Baltimore to get something exotic, but I unfortunately did not have time over the weekend. Instead, my friend and I decided to take a walk to this pond that we saw on Google Maps. It looked as if it was only a 20 to 30-minute walk away from the Homewood campus, so we eagerly set off with high hopes of finding rich soil full of life.

It was 12 PM and the sun was shining relentlessly, rendering the white sidewalk so bright I could not open my eyes. My friend and I trudged on with sweaty backs until we saw this steep hill – was the pond up there? We ran up, both hopeful and doubtful, only to find a massive construction site. A dry, dead-looking, yellow, and hot construction site. The exact opposite of a pond.

Construction Site.JPG

The “pond” that my friend and I were looking for

Well, we certainly did not walk all the way out there for nothing, so I got some soil from a nearby area and walked into my first Phage Hunting class happily. The sight of our clean and well-equipped laboratory got me so excited because all I had throughout high school was a chemical-stained bench. Soon after Dr. Fisher showed us some techniques, it was our turn to be the scientists.

First class, and I accidentally set the tip of a pipette on fire. Plus, I could not light the Bunsen burner properly, and could not remove the plunger from the syringe barrel. I have to say, I was quite impressed at how differently my first class deviated from my own expectation. At the very least, though, I have an awesome lab partner. Plus, I did still successfully perform direct isolation, so I thought everything was still acceptable.

The next day went quite smoothly as well, and we left off at day one of plaque assay. I could not wait to pick the plaque – it did not occur to me that I would not be able to find one. And, of course, I did not find one the next day. Thankfully, I was not alone, and went out to the area outside UTL to get more soil with some other Phage Hunters. We re-did what we did in the previous days and bonded over how much we wanted to pick a plaque. An interesting method of socialization, but one that worked very effectively.

Somewhat shockingly, even after doing direct isolation and plaque assay a second time, I still could not find anything. For the third time, I had to hunt for soil sample. Was this phage hunting or soil hunting? This time, I performed enriched isolation and serial dilution rather than direct isolation. As much as I do enjoy research, it would be a lie if I said I kind of, just maybe, wanted to get this part over with.

Indeed, I suppose third time is the charm (for me)! For enriched isolation, I had 6 agar plates instead of 2, and my results from Monday showed that two of them have many interesting little circles. I put them under the magnifying glass, labelled them, and began streaking. Even though I’m not quite sure what I am looking for the next time I am in the lab, I know it is only uphill from here.

Plaque 10^-1.JPG

Agar dish showing plaques as seen under magnifying glass 

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Being a Scientist is Weird

I felt so embarrassed. I felt so weird; whimg_4516o sits and gathers dirt into a container in the middle of the day in a garden? This whole experience was not in the course summary, was not what I signed up for. Of course, I knew that around campus other Phage Hunters were similarly gathering soil samples to start our research with, but in this I was alone.

Walking into class that first day made me feel quite a bit better: many also had soil samples and dirt crusted under their fingernails, though a few had forgotten and now had to go outside and gather their own soil. This class was the first venture into lab work that I have ever had, and the feeling that I experienced when putting on my crisp white lab coat can only be described as satisfaction; satisfaction that I was in this class, that I could rightfully wear this coat with others who had perhaps performed more research than I, that it just felt so right.

I expected that first day to be slow, for the professor to discuss what we would do during the semester and what experiment we would follow the entire time. While we did spend about an hour doing just that, we were subsequently thrust into the lab and told to “go at it,” to immediately start work. Suffice to say, that first day did not end well: my desk mate and I set a pipette on fire, got a plunger stuck inside of a test tube, and we released gas while failing to light a Bunsen burner. But everything got better from then on out.

I was so terrified that I would be the one left behind in class, or that everyone else would know more than me. The class seemed something that would stress me out or have more work than all other classes, but sadly this is not the case as I would happily perform extra lab work for this course. As of now, I found plaques on my original plate of phage sample, and have successfully isolated and transferred phages (using a process called streaking) from a plaque on that original plate onto a new plate, and am now waiting – with bated breath – to see if the phage successfully transferred and isolated in a second round of streaking.

Even though we are only a few weeks into classes here, this is the class that I most look forward to every week and am most excited to talk about. I can’t wait to go further in the course and learn more amazing information and techniques, but above all I can’t wait to continue to nurture my little phage(s) and see my love of laboratory research grow.

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