Fat on plastic: Metabolic consequences of an LDPE diet in the fat body of the greater wax moth larvae (Galleria mellonella)

Highlights

• LDPE diet reduces fitness and growth of G. mellonella.

• Fat body is the only organ maintained in LDPE-fed G. mellonella.

• Oxidative ability is maintained in fat body mitochondria of LDPE-fed waxworms.

• LDPE fed caterpillars maintain lipid reserves at detriment of other metabolites.

Abstract

The caterpillar larvae of the greater wax moth (Galleria mellonella) are avid plastivores, as when provided a diet of low-density polyethylene (LDPE) they actively feed on it. Recent work has highlighted the importance of their microbiome in the putative biodegradation of this plastic polymer, though the impact of plastic metabolism on the insect host is less clear. In the present study, we undertook an integrative approach spanning all levels of biological organization to explore the effects of a plastic diet on the metabolic physiology of this animal model of plastic biodegradation. We demonstrate that an LDPE diet is not sufficient to maintain optimal larval growth and survival. In addition, we confirm that plastic fed waxworms retain their fat body lipid stores in a manner proportional to their individual polyethylene consumption suggesting a direct effect of LDPE biodegradation. At the functional level, the oxidative capacity of the fat body of LDPE-fed larvae is maintained reflecting unaltered metabolic function of the tissue. Finally, metabolomic analyses confirmed fat body lipid stores maintenance in LDPE-fed worms, but uncovered various other nutritional deficiencies. Overall, this work unveils novel insights in the complex interplay between LDPE biodegradation and the metabolic physiology of this model plastivore.

Introduction

The production and use of petroleum-based polymers have grown exponentially over the last decades. Owing to their characteristic resistance, versatility and inexpensive production cost, plastics have been integrated into almost every aspect of our everyday lives (Amaral-Zettler et al., 2020). Unfortunately, as a direct consequence of their widespread use and high resilience, global plastic production directly correlates with its appearance and accumulation in our environment (PlasticsEurope, 2019). Petro plastics in various forms have invaded most ecosystems, from the high artic to remote lakes and rivers, and accumulate at alarming rates (Lebreton et al., 2018, Rhodes, 2018, Borrelle et al., 2020). In particular, low density polyethylene (LDPE) is used in many single use consumer applications and thus is present in large quantities in our environment (Castaneda et al., 2014, Rodrigues et al., 2018, Wang et al., 2017, Imhof et al., 2013, Zbyszewski and Corcoran, 2011). As such, there is a great interest in the potential ecophysiological impact of plastic waste on aquatic, terrestrial and even aerial environments.

In the last decade, many studies have focused on the physiological and ecological impact of macro- and microplastics on a variety of taxa including birds, mammals, fish and various invertebrate species (Watts et al., 2014, Gregory, 2009, LeMoine et al., 2018). While plastics appear in many shapes and forms in our environment, the consensus is that exposure to plastics negatively impacts animal wellbeing at various levels of biological organization (e.g., Grigorakis et al., 2017, Redondo Hasselerharm et al., 2018, Puskic et al., 2020). For example, micro- and nanoplastics can affect cardiac function, behavior and induce broad changes in gene expression in larval zebrafish (Pitt et al., 2018, LeMoine et al., 2018). Of note, much of the previous work on plastic ecotoxicity involves animals that are passively exposed to plastics. With the exception of a few select “plastivores”, few animal species actively seek and voluntarily consume plastics.

Interestingly, a variety of insects, primarily lepidopterans and coleopterans, have been shown to voluntarily ingest various plastic polymers including polyethylene (Bombelli et al., 2017, Brandon et al., 2018, Yang et al., 2014, Yang et al., 2015). Not only do these plastivores ingest plastic, but there is evidence that plastic ingestion is accompanied by its biodegradation and bioassimilation (Kundungal et al., 2019a, Kundungal et al., 2019b; Lou et al., 2020; Kong et al., 2019). The precise biochemical pathways and respective contributions of the insect hosts and their respective microorganisms has not yet been elucidated; however, plastic biodegradation is accelerated by this symbiotic relationship when compared to in vitro processes (Bombelli et al., 2017). Considering the environmental cost of plastic pollution, the discovery of plastivores has fostered a very active field of research aimed at deciphering the various aspects of plastic biodegradation. Although there is a great deal of interest in the biodegradative capacity of these systems, as well as in harnessing the microbial species involved in these processes, relatively little attention has been given to the physiological impact of plastic ingestion in model plastic-eating animals. Indeed, since these insects actively seek and ingest plastics, it would suggest that this behavior could be associated with potential metabolic benefits to the organism (Lou et al., 2020). Furthermore, investigating the effects of plastic consumption on a representative plastivore species may reveal further insights into the biodegradation processes utilized by a rather unique group of animals.

Among the plastivores, the larvae of the greater waxworm (G. mellonella) are avid LDPE consumers. This charismatic lepidopteran resides as a pest in beehives around the globe, where voracious larvae continuously feed on wax, pollen and honey (Kwadha et al., 2017). As a holometabolous insect, G. mellonella exhibits rapid development during which the larvae grow and accumulate large energetic reserves within its fat body, a major metabolic hub (Ellis et al., 2013, Czaja-Topińska and Klekowski, 1970). Once reaching their final instar, the larvae pupate and metamorphose into non-feeding reproducing adults, placing a crucial importance on the accumulation of fat body reserves in order to ensure survival and reproductive success. The fat body serves multiple functions in lepidopterans, including immunity, endocrine, detoxification, and most importantly fuel storage (Arrese and Soulages, 2010). When food is plentiful, caterpillars capitalize on nutrient availability and accumulate reserves in the form of lipid droplets in their fat body. These reserves can then be mobilized when necessary, whether it is for intense activities (i.e. flight in adult moth) or periods of starvation (Walker, 1965, Wigglesworth, 1967).

The voracity of these larvae, along with the potential biochemical endowment enabling their use of large hydrocarbon chains (Kong et al., 2019) are likely strong drivers for their unique plastivorous ability. When fed polyethylene, G. mellonella caterpillars show unique molecular and biochemical signatures suggesting that polyethylene metabolism affects their physiology (Cassone et al., 2020, LeMoine et al., 2020). Shortly after being presented with LDPE, waxworm larvae start feeding voraciously on the plastic substrate and soon thereafter produce a liquid frass containing glycol, a putative LDPE metabolic by-product (Cassone et al., 2020). In addition, an LDPE diet affects the insects’ physiology, promoting an upregulation of a number of intestinal transcripts involved in lipid metabolism and an ability to maintain much higher lipid stores than their starved counterparts (LeMoine et al., 2020). Considering the importance of the fat body in developing insects and our limited understanding of how a plastic diet impacts the biology of a plastivore, we investigated the effects of a LDPE diet on the metabolic physiology of G. mellonella larvae.