Life History Implications of Symbiont Acquisition for the Leaffooted Bug Leptoglossus Zonatus

Abstract

Most insects are associated with symbiotic microbes acquired from their environment. These microbes are ingested from the host’s environment, are localized in the gut, and supplement the host’s diet with critical nutrients. While many insects can receive benefits from diverse microbial communities, reliance upon a single environmentally acquired symbiont is likely to present risks to the host, should the symbiont be absent or rare in the environment. The arboreal leaffooted bug, Leptoglossus zonatus (Hemiptera: Coreidae), forms a symbiosis with bacteria of the genus Caballeronia, which must be acquired by 2nd instar nymphs from their environment. Bugs that fail to acquire their symbiont rarely survive to adulthood, and those that do have reduced body size and do not reproduce. Due to the cost of failing to acquire their symbiotic partner, I asked 1) whether L. zonatus is able to readily acquire Caballeronia in the tree canopy, 2) If Caballeronia is rare, whether delaying symbiont acquisition influences host fitness, and 3) whether Caballeronia can be transmitted horizontally as an alternative to environmental acquisition. To investigate the first question, nymphs were caged on individual tree branches and allowed to forage and develop. Aposymbiotic (symbiont-free) nymphs were caged with potential environmental reservoirs (soil, water, phyllosphere) or horizontal sources (e.g. conspecific bugs) of the symbiont. I found that in this setting, acquisition of Caballeronia was uncommon, suggesting a cost to dependence on an environmentally-acquired partner. In addition to the risk that nymphs may not acquire the symbiont in the environment, should Caballeronia be rare or at low abundances, symbiont acquisition may be delayed, and delays may present consequences for host fitness. I then investigated the impact of delaying symbiont acquisition by providing nymphs with Caballeronia at different time points following development to the 2nd instar. To assess fitness costs, I recorded acquisition rates, survivorship, development time, adult weight, and development of the M4 symbiotic organ with fluorescence microscopy. I found that acquisition delay negatively influenced all measurements of performance, and the consequences of delay compounded as it was prolonged. The results suggest a finite window for Caballeronia acquisition, ~1 week following development to the 2nd instar, and these results highlight an additional potential cost for this type of symbiosis. To answer the third question, I investigated if L. zonatus released Caballeronia in its feces, and if feces could serve as a source of the symbiont. I assessed L. zonatus feces to determine the presence and abundance of Caballeronia, and assessed which life stages excreted bacteria. I found that Caballeronia was present in feces, and almost exclusively released by adult bugs. I then addressed whether nymphs provided with feces from Caballeronia-colonized adults were able to acquire the symbiont from feces, and whether Caballeronia acquired from feces is equivalent to cultured Caballeronia for bug performance. Bugs fed Caballeronia from fresh feces, like those fed cultured symbiont, were almost universally able to acquire the symbiont. However, feces-fed bugs showed significantly longer development times, reduced survivorship, and reduced adult mass. Together, this work indicates potential costs for insects that rely on free-living bacteria for critical nutrition, and informs future research into the ecological dynamics and evolutionary stability of environmentally transmitted, obligate symbioses.