One Pathway Is Not Enough: The Cabbage Stem Flea Beetle Psylliodes chrysocephala Uses Multiple Strategies to Overcome the Glucosinolate-Myrosinase Defense in Its Host Plants

Abstract

The cabbage stem flea beetle (Psylliodes chrysocephala) is a key pest of oilseed rape in Europe, and is specialized to feed on Brassicaceae plants armed with the glucosinolate-myrosinase defense system. Upon tissue damage, the β-thioglucosidase enzyme myrosinase hydrolyzes glucosinolates (GLS) to form toxic isothiocyanates (ITCs) which deter non-adapted herbivores. Here, we show that P. chrysocephala selectively sequester GLS from their host plants and store these throughout their life cycle. In addition, P. chrysocephala metabolize GLS to desulfo-GLS, which implies the evolution of GLS sulfatase activity in this specialist. To assess whether P. chrysocephala can largely prevent GLS hydrolysis in ingested plant tissue by sequestration and desulfation, we analyzed the metabolic fate of 4-methylsulfinylbutyl (4MSOB) GLS in adults. Surprisingly, intact and desulfo-GLS together accounted for the metabolic fate of only 26% of the total ingested GLS in P. chrysocephala, indicating that most ingested GLS are nevertheless activated by the plant myrosinase. The presence of 4MSOB-ITC and the corresponding nitrile in feces extracts confirmed the activation of ingested GLS, but the detected amounts of unmetabolized ITCs were low. P. chrysocephala partially detoxifies ITCs by conjugation with glutathione via the conserved mercapturic acid pathway. In addition to known products of the mercapturic acid pathway, we identified two previously unknown cyclic metabolites derived from the cysteine-conjugate of 4MSOB-ITC. In summary, the cabbage stem flea beetle avoids ITC formation by specialized strategies, but also relies on and extends the conserved mercapturic acid pathway to prevent toxicity of formed ITCs.

Ontogenetic differences in the chemical defence of flea beetles influence their predation risk

“…Adapted herbivores and pathogens use different strategies such as sequestration, metabolic detoxification and excretion to overcome this plant defence (Jeschke et al, 2016;van den Bosch, Niemi, & Welte, 2019;Vela-Corcía et al, 2019). The ability to sequester intact GLS evolved independently in specialized insects belonging to the orders Hemiptera, Hymenoptera and Coleoptera, but not all GLS-sequestering species evolved endogenous myrosinase activity (Beran et al, 2018;Müller & Wittstock, 2005;Opitz & Müller, 2009). One of these is the turnip sawfly Athalia rosae where rapid GLS sequestration has been suggested to function as a detoxification mechanism by preventing hydrolysis by plant myrosinases (Abdalsamee, Giampà, Niehaus, & Müller, 2014;van Geem, Harvey, & Gols, 2014).…”

“…GLS were extracted, converted to desulfo-GLS and analysed using high performance liquid chromatography coupled to diode array detection (HPLC-DAD) as described in Beran et al (2014). Myrosinase activity was determined in crude protein extracts prepared from different P. armoraciae life stages using 0.5 mM allyl GLS (Carl Roth) as a substrate as described in Beran et al (2018).…”

The phytopathogenic fungus Sclerotinia sclerotiorum detoxifies plant glucosinolate hydrolysis products via an isothiocyanate hydrolase

“…Subsequently, SaxA was found in some gutassociated bacteria from herbivores feeding on Brassicales plants, where it was proposed to facilitate tolerance of host larvae toward dietary ITCs 20,21 . Recently, an amine and its corresponding acetylated derivative (acetamide) were detected as minor products of ITC metabolism in the flea beetle Psylliodes chrysocephala 22 , but it is not known whether this conversion was performed by the insect itself or by its associated microbiota. Despite our knowledge on GL metabolism in insects and bacteria, we know very little about fungal metabolism of GLs and how pathogenic fungi colonizing Brassicaceae plants cope with GLs and their toxic hydrolysis products.…”

Performance of cabbage stem flea beetle larvae (Psylliodes chrysocephala) in brassicaceous plants and the effect of glucosinolate profiles

“…The effects of insect feeding on GSL composition in plants have been repeatedly reported in the literature (Koritsas et al, 1989, 1991; Birch et al, 1992; Bartlet et al, 1999; Textor & Gershenzon, 2009; Beran et al, 2018), but the results were not consistent. In our experiments, infestation by CSFB larvae had only a minor effect on indole GSL whereas aliphatic GSL increased in most of the genotypes tested.…”

“…However, as the effect of a single GSL on various insects is very variable (Hopkins et al, 2009), further research is needed on this topic. Recent studies showed that adult CSFBs have developed different pathways to overcome the GSL myrosinase defence of their host plants (Beran et al, 2018). It can be assumed that this is also true for CSFB larvae and that different GSL profiles or contents in the brassicaceous plants tested may exert different effects and toxicities on CSFB larvae, thereby impacting larval development.…”