Simulating the Effect of Larval Dietary Restriction on C. elegans

One aspect of my model I tested in lab is whether the quantity of food available to a worm determines its transition to the next stage of development or if it is simply a matter of time passing. Studies show that life span is food-dependent, specifically adult worms have an increased life span if fed a restricted diet [e.g. Klass 1977, Lee et al. 2006]. However, it is unknown whether larval stages experience the same effect. To test this, I designed an experiment in lab with four treatment groups of L1 (first larval stage) worms, each fed a different concentration of E. coli (6 mg/mL, 4.8 mg/mL, 3.6 mg/mL, or 2.4 mg/mL). If the effect is the same on larvae as it is on adults, the expected outcome would be statistically different lengths of time spent in L1 before molting to L2/L2d (second larval stage). To determine when the worms are molting, I used a specific strain of C. elegans with a gfp (green fluorescent protein) marker on the molting gene that makes the worms glow under an epifluorescent microscope when they’re molting. The glow will last for about three hours, peaking in intensity when they molt [Monsalve et al. 2011]. One complication, however, is that if the worms are not completely synchronized in age (i.e. they don’t all start from time zero in their life stage), and depending on how far apart in age they are, the results may be confounded by this variation. I created a simulation to test whether unsynchronized worms can produce statistically significant results and found that they can, but not as well as worms that are synchronized in age. After performing this test in silico, I designed and conducted this experiment in the lab and I found that larval dietary restriction does lengthen the amount of time spent in that stage. This means that the amount of time it takes a worm to molt from one larval stage to the next is food-dependent and aligns with the assumption made in the model.

Files

• The file called "module.py" contains the functions needed to run this experiment in silico. The purpose was to determine whether unsynchronized experimental groups can produce statistically significant results.
• The file called "test_functions.py" contains a couple test functions used to ensure the functions in the model are working properly.
• The file called "requirements.py" lists the packages that need to be imported to run each of these files.
• The file called "Larval_DR_Sim.ipynb" explains the experiment in depth and provides a demonstration of how to use the model.
• The file called "data_analysis.py" contains functions to graph and determine statistical significance from data collected in the lab.

References

1. Klass, M. R. (1977). AGING IN THE NEMATODE CAENORHABDITIS ELEGANS: MAJOR BIOLOGICAL AND ENVIRONMENTAL FACTORS INFLUENCING LIFE SPAN. Mechanisms of Ageing and Development, 6, 413-429.
2. Lee, G. D., Wilson, M. A., Zhu, M., Wolkow, C. A., De Cabo, R., Ingram, D. K., & Zou, S. (2006). Dietary deprivation extends lifespan in Caenorhabditis elegans. Aging Cell, 5(6), 515-524.
3. Monsalve, G. C., Van Buskirk, C., & Frand, A. R. (2011). LIN-42/PERIOD controls cyclical and developmental progression of C. elegans molts. Current Biology, 21(24), 2033-2045.