Abstract
For most animals, feeding includes two behaviors: foraging to find a food patch and food intake once a patch is found. The nematode Caenorhabditis elegans is a useful model for studying the genetics of both behaviors. However, most methods of measuring feeding in worms quantify either foraging behavior or food intake, but not both. Imaging the depletion of fluorescently labeled bacteria provides information on both the distribution and amount of consumption, but even after patch exhaustion a prominent background signal remains, which complicates quantification. Here, we used a bioluminescent Escherichia coli strain to quantify C. elegans feeding. With light emission tightly coupled to active metabolism, only living bacteria are capable of bioluminescence, so the signal is lost upon ingestion. We quantified the loss of bioluminescence using N2 reference worms and eat-2 mutants, and found a nearly 100-fold increase in signal-to-background ratio and lower background compared to loss of fluorescence. We also quantified feeding using aggregating npr-1 mutant worms. We found that groups of npr-1 mutants first clear bacteria from within the cluster before foraging collectively for more food; similarly, during large population swarming, only worms at the migrating front are in contact with bacteria. These results demonstrate the usefulness of bioluminescent bacteria for quantifying feeding and generating insights into the spatial pattern of food consumption.
Highlights
Feeding behaviour plays an important role in fields ranging from ecology and evolution (Larsen 2003; MacArthur and Pianka 1966) to ageing and metabolism (Balasubramanian, Howell, and Anderson 2017; Trepanowski et al 2011) and health and disease (Djalalinia et al 2015; Mattson et al 2014)
C. elegans feeds by sucking bacteria into its mouth using rhythmic pumping of pharynx (Avery and You 2012), and pharyngeal pumping frequency is often used as a proxy for food intake
As fluorescent proteins form stable cooperatively folding structures, they are resistant to proteolytic cleavage (Nicholls and Hardy 2013; Bokman and Ward 1981). This results in high background fluorescence signal even after bacteria are digested by C. elegans, complicating both the quantification and the interpretation of feeding behaviour
Summary
Feeding behaviour plays an important role in fields ranging from ecology and evolution (Larsen 2003; MacArthur and Pianka 1966) to ageing and metabolism (Balasubramanian, Howell, and Anderson 2017; Trepanowski et al 2011) and health and disease (Djalalinia et al 2015; Mattson et al 2014). C. elegans feeds by sucking bacteria into its mouth using rhythmic pumping of pharynx (Avery and You 2012), and pharyngeal pumping frequency is often used as a proxy for food intake. Feeding can be measured using a non-food additive such as exogeneous luciferin (Rodríguez-Palero et al 2018), dye (You et al 2008), or fluorescent beads (Fang-Yen et al 2009; Kiyama et al 2012). Of the existing methods of quantifying feeding, imaging the consumption of fluorescently-labelled bacteria is of particular interest since it can provide information on both where and how much food has been consumed. This results in high background fluorescence signal even after bacteria are digested by C. elegans, complicating both the quantification and the interpretation of feeding behaviour. We use an E. coli strain with self-sustained bioluminescence to monitor both the rate of food intake and its spatial distribution in laboratory reference and mutant worms, worms treated with serotonin and naloxone, and in high-density worm swarms
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