RNA allows identifying the consumption of carrion prey

Abstract

Facultative scavenging by predatory carnivores is a prevalent but frequently underestimated feeding strategy. DNA-based methods for diet analysis, however, do not allow to distinguish between scavenging and predation, thus, the significance of scavenging on population dynamics and resource partitioning is widely unknown. Here, we present a methodological innovation to differentiate between scavenging and fresh prey consumption using prey RNA as a target molecule. We hypothesized that the rapid post-mortem breakdown of RNA in prey tissue should lead to a significantly lower detection probability of prey RNA than DNA when carrion rather than fresh prey is consumed. To test this hypothesis, ground beetles (Pseudoophonus rufipes [De Geer]) were offered either fresh or 1-day-old dead Drosophila melanogaster fruit flies (carrion). The detectability of prey RNA and DNA in the beetles' regurgitates was assessed with diagnostic Drosophila-specific RT-PCR and PCR assays at 0, 6, 12, 24 and 48 h post-feeding. After fresh fly consumption, prey RNA and DNA were detectable equally well at all times. When carrion prey was consumed, the detection strength of prey RNA immediately after feeding was significantly lower than that of prey DNA and reached zero in most samples within 6 h of digestion. Our findings provide evidence that prey RNA allows distinguishing between the consumption of fresh and scavenged prey, thereby overcoming a long-known weakness of molecular diet analysis. The assessment of prey RNA offers a generally applicable approach for examining the importance of scavenging in food webs to unravel its functional consequences for populations, communities, and ecosystems.

Keywords: food webs; molecular diet analysis; scavenging; top-down control; trophic cascades; trophic linking.

FIGURE 1

Detection rate and probability of Drosophila melanogaster DNA (left panels) and RNA (right panels) in regurgitates of Pseudoophonus rufipes, 0, 6, 12, 24, and 48 h after feeding on one fresh (top) or one 24‐h‐dead fly (bottom). The proportion of samples positive for the molecular target (●) is plotted along the detection probability (solid lines) with 95% CIs (dotted lines) calculated with generalized linear models (GLM)

FIGURE 2

Comparison of prey DNA and prey RNA signal strengths, measured as relative fluorescent units (RFU), in regurgitates of Pseudoophonus rufipes within the same prey treatment, fresh (left panel) or carrion prey (right panel) after 0, 6, 12, 24 and 48 h of digestion. Significant differences revealed by Wilcoxon signed‐rank test are indicated with (***) for p < .001 and (**) for p < .01

FIGURE 3

Boxplots of the ratios RFU_RNA:RFU_DNA, measured for capillary electrophoresis PCR (celPCR) products of regurgitates of Pseudoophonus rufipes 0, 6, 12, 24 and 48 h after consumption of one fresh (red) or one carrion fruit fly (blue). Significant differences between ratios of different prey types as revealed by Wilcoxon rank sum test are indicated with (**) for p < .01. All samples with carrion prey were negative for the target prey at 24 h after feeding