1.Introduction
The interplay between plants and insects represents one of nature’s most
common species interactions (Bernays and Chapman 1994; Stork 2018; Joneset al. 2022). Insects rely upon their host plants, as they use
them as a food source, for reproduction or as a shelter (Futuyma and
Moreno 1988; Kergoat et al. 2017). When feeding on a particular
plant, herbivorous insects are exposed to a variety of different
structural and molecular components that can affect their cellular
homeostasis and in further consequence their fitness (for a summary of
different plant defense mechanisms see e.g. Fürstenberg-Hägg et
al. 2013). The efficient utilization of a host, thus, requires genetic
and physiological “adjustments” to the different properties of the
respective plant(s) (Ehrlich and Raven 1964; Després, David and Gallet
2007; Heidel-Fischer and Vogel 2015; Simon et al. 2015; Xi, Guo
and Hu 2024). Within the order Lepidoptera, the majority of butterflies
(superfamily Papilionoidea) are purely herbivorous. The associations
with a host can be very exclusive, with a specialist insect species
using only one or a few host plant species (Bernays and Graham 1988; Via
1990; Bernays 1991; Forister et al. 2015; Hardy et al.2020). In contrast, phytophagous generalists are characterized by a
broader host repertoire that can cover multiple plant families and
sometimes even different orders (polyphagy; Bernays and Graham 1988; Via
1990; Forister et al. 2015; Hardy et al. 2020). While
specialists should be able to develop strong adaptations to their hosts,
generalists encounter a much broader diversity of physically and
chemically different hosts and should require a more flexible set of
adaptations.
As suggested by Ehrlich and Raven (1964), the broad host repertoire of
polyphagous butterflies can be achieved by including plants that are
chemically similar. Chemical similarity in plants can either result from
shared ancestry or convergent evolution (Janz and Nylin 1998; Janz,
Nyblom and Nylin 2001; Kergoat et al. 2005; Heidel-Fischeret al. 2009). Adaptations to one specific host could, thus, also
serve as preadaptations for another plant with a similar chemical
profile and allow an expansion of the host repertoire (Janz and Nylin
2008; Agosta and Klemens 2008; Agosta et al. 2010). This idea has
received some support from simulation studies (Araujo et al.2015) and at the level of gene expression (Heidel-Fischer et al.2009; Celorio-Mancera et al. 2023). Using the polyphagous comma
butterfly (Polygonia c-album ), Heidel-Fisher et al. (2009)
showed that similarities in the gene expression profiles on different
hosts could be explained by a close phylogenetic relatedness or the same
growth form of the plants. Despite these similarities in the gene
expression profiles, the overall patterns indicated a very host-specific
transcriptional response in the larval gut (Heidel-Fisher et al.2009). Such host-specific gene expression profiles in P. c-albumwere further supported by Celorio-Mancera et al. (2013). Based on
this, it has been suggested that adaptations to host plants can
profitably be viewed as “modules of co-expressed genes and their
resulting phenotypic appearance ” (cf. Celorio-Mancera et al.2023). The latter study, which examined the overall gene expression
patterns on different plants in P. c-album and related species,
furthermore, showed that these specific modules can be conserved over
evolutionary time and can show quite distinct gene expression profiles
that differ in the degree of overlap among plants (Celorio-Manceraet al. 2023). These similarities or differences could potentially
be used as an indicator for overlapping mechanisms to cope with a
plant’s properties.
Another important factor when describing environment-specific expression
patterns is how quickly insects can adapt their transcriptional response
to a new host. A high degree of plasticity and rapid responses to a host
switch can ensure that juvenile stages are able to react quickly to a
changing environment (cf. Xiao et al. 2019). In many phytophagous
insects in which the host is consumed during their larval stages, the
actual host plant is determined by the mother (König et al. 2016;
see also Nylin and Janz 1993; Gripenberg et al. 2010; Refsnider
and Janzen 2010). It is still largely unknown if and to which extent
larvae of such species can leave their host plant and look for
alternatives, in case of deteriorating plant quality or oviposition
mistakes (but see Nylin et al. 2000; Schäpers et al.2016). Although such scenarios are not well documented, larval movement
within plants should often be necessary for similar reasons. Plastic
adaptations to different plants and plant parts without a long temporal
lag could ensure that larvae survive and complete their development
without severe fitness costs. Thus, an experimental shift of host plant
during larval development should give some indication of how difficult
an evolutionary host plant shift would be for caterpillars. Furthermore,
previous studies have mainly focused on gene expression in larvae that
were reared on the same host plant during their entire development (for
a summary see Birnbaum and Abbot 2020). The transcriptional profiles
could, thus, not exclusively be associated with the direct responses to
a certain host but also were the product of different developmental
trajectories linked to a particular plant (cf. Celorio-Mancera et al.
2023). Experimental shifts could now help to further identify genes and
transcriptional adjustments that are crucial for the use of a specific
host plant.
Polygonia c-album (Linnaeus, 1758), the comma butterfly, is a
polyphagous butterfly species in the family Nymphalidae that feeds on
trees, bushes and herbs from four different orders: Rosales, Fagales,
Malphigiales and Saxifragales (Nylin 1988). Within these orders the
repertoire of the comma comprises Urtica , Humulus andUlmus (Urticaceae and other “urticalean rosids”), Betulaand Corylus (Betulaceae) as well as Salix (Salicaceae) andRibes (Grossulariaceae) (Nylin 1988). For some of these plants,
quite characteristic chemical profiles have been described. Urticaceae,
for instance, are rich in alkaloids and phenols, leaves of the Salicaea
contain flavonoids and phenolic glycosides, while the presence of
cyanogenic compounds has been reported for the genus Ribes (Hegnauer,
1973). Ribes is particularly unusual as a host, as its use is
rather rare among butterflies. Within the family of Nymphalidae
(~6,100 species; Van Nieukerken et al. 2011), for
instance, Ribes is only consumed by a few species of the genusPolygonia (Gamberale-Stille et al. 2019), which together
with the distinct gene expression patterns (Celorio-Mancera et
al. 2013, 2023) indicates very specific and exclusive adaptations to a
likely chemical divergent host. Such plant-specific profiles now
represent a good prerequisite for investigating the plasticity of
adaptations.
In this study, the larvae’s ability to adjust to a new host within one
generation was investigated. Special attention was paid to how, and how
fast, polyphagous butterfly larvae can react to a new plant in terms of
gene expression. Earlier studies already showed that larvae can be moved
to another host plant during their development without major effects on
their performance (Söderlind 2012). It is now assumed that also the
underlying transcriptional patterns can be adjusted very rapidly in
response to the new environment. Based on the patterns found in previous
studies, it was expected that diet shifts between hosts that strongly
overlap in their gene expression profiles should be easier (i.e. faster
and with fewer responding genes). In contrast, a slower and more drastic
adjustment is assumed for switches between plants with very opposing
patterns (e.g. Urtica vs. Ribes ; see Celorio-Manceraet al. 2023). To test this, two host switch experiments were
combined with a performance screening. It was predicted that:
(1) There is a temporal course of transcriptional adjustments. Shortly
after the host switch, the gene expression profiles will still
correspond to the first host, but can change quickly to regain cellular
homeostasis on the new host plant.
(2) There is a lower number of transcriptional differences between hosts
with similar chemical properties. It is assumed that overlapping
processes are involved when feeding on host plants that show
similarities in their chemical composition, either due to a close
phylogenetic relatedness or a shared growth form (e.g. Switch A). These
overlaps will result in a lower number of differentially expressed
genes.
(3) Switching to chemically more different hosts is more challenging.
(in contrast to (2)) The use of hosts with very distinct structural and
chemical properties (e.g. Switch B), often requires more transcriptional
and physiological changes to regain cellular homeostasis that will
manifest in a higher number of differentially expressed genes.
(4) Differences in gene expression profiles will have a physiological
correspondence. Depending on the extent of required adjustments, the
switch to certain hosts can be associated with higher costs, which can
result in a reduced larval performance.