The most notable aspect of our results is the apparently sexually dimorphic occurrence of adalinine in A. decempunctata. Adalinine is a minor alkaloid of A. bipunctata (Lognay et al. 1996; Oudendijk & Sloggett 2022), apparently in all life history stages (Lognay et al. 1996; J.J. Sloggett unpub. data). Adalinine is also already known from adult A. decempunctata (Lognay et al. 1996) but the abundance described here in males makes it the dominant alkaloid and much more abundant than in females or eggs. Such quantitative dimorphism in the relative amounts of different alkaloids has not been described in ladybirds before, although in general females tend to have higher concentrations of major alkaloids than males including in A. bipunctata (Holloway et al. 1991, 1993; de Jong et al. 1991, but see Oudendijk & Sloggett 2022).
This might in part be related to female egg production: in A. bipunctata, after reflex bleeding, females accumulate alkaloid more slowly than males, probably because they allocate alkaloids to eggs (Oudendijk & Sloggett 2022). If, as appears to be the case here, females allocate proportionately more adalinine to eggs than they have themselves, but both sexes synthesize the two alkaloids at the same rate, then ultimately males could end up with a higher proportion of adalinine in their bodies. However, no such marked phenomenon has been observed in A. bipunctata (Oudendijk & Sloggett 2022) and it is hard to envisage such a large difference developing by solely by these means, suggesting that the dimorphism more likely occurs predominantly through differences in synthesis.
It appears that adalinine is synthesized from adaline in Adalia spp. (Laurent et al. 2001): if this is the case, then adalinine must be more energetically costly to produce due to the additional synthetic steps. Possibly males invest more in this defensive molecule than females due to the latter’s high reproductive costs. Males could even transfer adalinine to females as a nuptial gift, given its higher occurrence in eggs: alkaloid transfer from male to female is known in the ladybird Epilachna paenulata (Camarano et al. 2009). Alternatively, the costs of storing adalinine could be lower if, for example, adaline presents a serious risk of self-poisoning: this synthesis of a less toxic storage compound is known from other chemically defended organisms (e.g. Hartmann 2004; Brückner et al. 2017).
The sexual difference in the abundance of the two alkaloids in A. decempunctata poses a potential issue in understanding the role of colour pattern in signalling defensive capability. A weak, borderline significant relationship (P = 0.06) exists for adaline and total alkaloid. Bearing in mind that this is non-significant, this could be a consequence of our relatively small sample sizes, exacerbated by the fact that all morphs of field-collected adults will exhibit alkaloid variation in response to a diversity of factors. These include diet, reflex bleeding, age, temperature and parasite infection (e.g. de Jong et al. 1991; Steele et al. 2020, 2023; Oudendijk & Sloggett 2022; Sakaki & Nedvěd 2023). A laboratory study could eliminate this variation. However, in the absence of an understanding of the roles of the two alkaloids, it is hard from the perspective of a predator to interpret whether the different colour pattern morphs really convey any information about relative distastefulness and toxicity. It should also be borne in mind that many other factors play a role in colour pattern polymorphism apart from chemical defence (Sloggett & Honěk 2012; Briolat et al. 2019)
There was no clear relationship between maternal alkaloid and egg alkaloid in this study, except possibly for adalinine, where the relationship was positive. Paul et al. (2015, 2018) reported contradictory results for adaline: in A. bipunctata in their experiments: in one study, they found no correlation, whereas in another there was a positive relationship. In this study of A. decempunctata, there was a tendency for alkaloids to decline with female fecundity, though not significantly. Data on adaline in A. bipunctata is also contradictory on this point (Paul et al. 2015, 2018) but in one case, they also observed a negative relationship (Paul et al. 2018). In both studies of A. bipunctata, adalinine was not quantified, making a direct comparison with our study difficult, but if a relationship, albeit weak, with female parameters exists, for adaline in A. bipunctata and adalinine (but not adaline) in A. decempunctata, then this might reflect the relative importance of the alkaloids in the two species.
Ladybirds display a high taxon-related alkaloid diversity (Daloze et al. 1994; King & Meinwald 1996; Laurent et al. 2005) and it is not yet clear why such diversity exists. Presumably, new major alkaloids evolve initially as minor components. The alkaloids of only two Adalia species have been identified, with other “species” (e.g. Tursch et al. 1973) merely representing morphs of A. bipunctata and A. decempunctata. However, cautiously assuming that adaline as the major and adalinine as the minor alkaloid is the ancestral condition, the altered situation in A. decempunctata could throw light on how changes in chemical defences evolve in taxa such as ladybirds, where autogenous production predominates.