Drs. Benoit and Wilson have long had a love affair with Bacillus thuringiensis (Bt) spores and their study. Their many papers on the subject have often centered on a germination assay that follows the conversion of dormant and resistant spores back to vegetative growth. The assay is based on watching a change in the optical density (absorbance) of spores using a spectrophotometer. It is fast and easy and dependable. At least it was until about 12 years ago when that assay, for some reason, stopped working.
The first indication something was wrong came in the form of an email from a researcher in New Zealand in the fall of 2001. The graduate student was studying Bt germination and ran into difficulty replicating the results Wilson and Benoit had reported. The email simply asked Dr. Wilson whether there were any special tricks involved in the assay. Since neither Wilson or Benoit was involved in research due to administrative responsibilities at the time and had not experienced similar problems, they had no advice to offer.
Fast forward a few years to the creation of the BIMS program and a new graduation requirement that every student must participate in capstone research, an Honors project, or an internship. The department needed research ideas, and Benoit and Wilson resurrected the germination assay as a means to engage students in studying new aspects of spores physiology. The number of unique projects this system would provide for future students was enormous. But the assay failed to work. Even with new facilities and new resources to support research, several student projects failed to recreate results from earlier papers. Nothing in the literature and no one in the field had an answer.
This summer, Honors student Heather Rawls became the most recent student to attempt the assay. Through the summer she tried differences in media, growth temperature, ways of collecting and processing spores, water quality being used, activation techniques, a variety of germinants, and at least five different spectrophotometers with no predictability or consistency in the results. If anything, fewer and fewer spores were germinating with each attempt.
In August, Heather and Dr. Wilson had a research pow wow and developed an alternate project for her Honors thesis. Time was running out to complete her research before starting the writing, and moving to something with a higher probability for success was needed. But Heather wasn’t ready to give up that easily. At an impromptu meeting a couple of weeks ago, a new thought emerged when it seemed every variable possible already had been tested. Glassware! In their graduate programs and during their prior research using the assay, Benoit and Wilson always collected the spores in glass containers and the germinants and all other chemicals used had been stored in glass containers. In our growing emphasis on research, McMurry had improved the funding of science programs so much that the use of disposable plastics was now the norm. Maybe the plastic centrifuge tubes used to collect spores and store germinants were coated with something inhibiting spore germination? Maybe some chemical was leaching out of the plastic?
Over the last two weeks, Heather switched to glassware for collecting spores and making reagents. Her results were dramatically different. We now are certain plasticware has an inhibitory effect on germination. A decade of frustration was caused by our affluence and the use of disposable labware rather than old-school glass.
With a working assay, Heather will turn back to the project she intended to complete this summer looking at the germination of a variety of genetically-engineered and wildtype strains of Bacilli. Another Honors student just beginning her work will investigate what the mystery chemical from the plastics might be and how it inhibits germination, or the range of spore-forming species affected. One mystery is solved, more are uncovered…
BIMS Honors students prove to us on a daily basis that they are among the best thinkers and hardest workers on campus. This commitment to uncovering the truth is what will drive them to become leaders in biomedical science and healthcare provision in the future.