pH Indication of Respiration and Effects of Different Carbohydrates and Escherichia coli on Respiration Rates in Caenorhabditis elegans

Abstract

tions with a visible light spectrophotometer (Parrish amd Grammer, 2012). However, the shift in the absorption spectrum of the dye upon acidification, the pK of the shift, had to exactly match the pH range of the acidification, necessitating changes in dyes employed depending upon which pH decreases were observed. Moreover, the absorption properties of any colored additives to be investigated and the light scattering properties of any potential food sources, such as bacterial suspensions, made it difficult to interpret the spectral properties of the observed mixture due to the use of spectrophotometry. Thus, the use of pH probes was alternatively investigated. Due to the intracellular oxidation of glucose, phenol red and spectrophotometry can detect color change. It was assumed that exogenous glucose would produce significant acidification of the medium by the worms. As the sugar is oxidized through respiration, carbon dioxide is produced, which causes the medium to acidify. Previous studies have investigated the respiratory processes and demonstrated the dependence of absorption of pH indicators on time and weak dependence of respiration rate on glucose concentration (Parrish & Grammer, 2012). This study developed procedures to further detect respiration in C. elegans by using Vernier pH probes (Vernier Software and Technology, Beaverton, OR) and tested the effects of E. coli and different carbohydrates (glucose, fructose, and maltose) on respiration rates. It was hypothesized that Vernier pH probes could be utilized to detect respiration and that E. coli and glucose would show the highest respiration rates, even if all carbohydrates were metabolized to some degree. The addition of E. coli should reveal increased respiration rates because it serves as the main food source for C. elegans. Glucose was also expected to reveal high INTRODUCTION Caenorhabditis elegans are 1 mm-long, non-parasitic nematodes that can be found in the soil. They are utilized as model organisms because they are transparent, easy to maintain, inexpensive, have a sequenced genome, and have a rapid life cycle (Corsi et al., 2015). Their primary food source is Escherichia coli, a gramnegative bacterium. C. elegans provide chemotaxis indices via chemotaxis assays; chemotaxis is the movement towards or away from an attractant or a repellent, respectively (Bargmann, 2006). A mitochondrial inhibitor, sodium azide, is utilized to stop the worms once they move to the test or control spots during chemotaxis assays. Sodium azide blocks cytochrome c oxidase and adenosine triphosphate (ATP) synthase, which causes the worms to die due to inhibition of the respiration processes (Massie et al., 2003). C. elegans can also move by swimming. During the swimming period, the nematodes carry out cellular respiration, which acidifies the medium. The acidification can be detected by phenol red, a pH indicator (Parrish & Grammer, 2012). Phenol red changes colors from red to yellow as the pH level of the solution decreases; hence, the pH becomes acidic. Previously, the color change was analyzed by examining the absorbance of the experimental solupH Indication of Respiration and Effects of Different Carbohydrates and Escherichia coli on Respiration Rates in Caenorhabditis elegans