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Host plant chemical composition critically shapes the performance of insect herbivores feeding on them. Some insects have become specialized on plant secondary metabolites, and even use them to their own advantage such as defense against predators. However, infection by plant pathogens can seriously alter the interaction between herbivores and their host plants. We tested whether the effects of the plant secondary metabolites, iridoid glycosides (IGs), on the performance and immune response of an insect herbivore are modulated by a plant pathogen. We used the IG-specialized Glanville fritillary butterfly Melitaea cinxia, its host plant Plantago lanceolata, and the naturally occurring plant pathogen, powdery mildew Podosphaera plantaginis, as model system. Pre-diapause larvae were fed on P. lanceolata host plants selected to contain either high or low IGs, in the presence or absence of powdery mildew. Larval performance was measured by growth rate, survival until diapause, and by investment in immunity. We assessed immunity after a bacterial challenge in terms of phenoloxidase (PO) activity and the expression of seven pre-selected insect immune genes (qPCR). We found that the beneficial effects of constitutive leaf IGs, that improved larval growth, were significantly reduced by mildew infection. Moreover, mildew presence downregulated one component of larval immune response (PO activity), suggesting a physiological cost of investment in immunity under suboptimal conditions. Yet, feeding on mildew-infected leaves caused an upregulation of two immune genes, lysozyme and prophenoloxidase. Our findings indicate that a plant pathogen can significantly modulate the effects of secondary metabolites on the growth of an insect herbivore. Furthermore, we show that a plant pathogen can induce contrasting effects on insect immune function. We suspect that the activation of the immune system toward a plant pathogen infection may be maladaptive, but the actual infectivity on the larvae should be tested.

Graphical abstract, Highlights • Baculovirus exposure reduces larval and pupal survival. • Development and mortality was affected in a concentration-dependent manner. • Sub-lethal infection has no effect on flight-related wing morphology. • Wing melanisation increases when individuals are exposed to baculovirus., Infections represent a constant threat for organisms and can lead to substantial fitness losses. Understanding how individuals, especially from natural populations, respond towards infections is thus of great importance. Little is known about immunity in the Glanville fritillary butterfly (Melitaea cinxia). As the larvae live gregariously in family groups, vertical and horizontal transmission of infections could have tremendous effects on individuals and consequently impact population dynamics in nature. We used the Alphabaculovirus type strain Autographa californica multiple nucleopolyhedrovirus (AcMNPV) and demonstrated that positive concentration-dependent baculovirus exposure leads to prolonged developmental time and decreased survival during larval and pupal development, with no sex specific differences. Viral exposure did not influence relative thorax mass or wing morphometric traits often related to flight ability, yet melanisation of the wings increased with viral exposure, potentially influencing disease resistance or flight capacity via thermal regulation. Further research is needed to explore effects under sub-optimal conditions, determine effects on fitness-related traits, and investigate a potential adaptive response of increased melanisation in the wings due to baculovirus exposure.

In butterflies, male reproductive success is highly related to the quality and the size of the spermatophore transferred to the female. The spermatophore is a capsule produced by the male during copulation, which in many species contains sperm in addition to a nuptial gift, and which is digested by the female after copulation. The nuptial gift may contribute to egg production and offspring quality, and in some cases also to female body maintenance. The production of the spermatophore, however, represents a cost for the male and, in polyandrous species, ejaculates are sometimes allocated adaptively across matings. Nonetheless, although the ecological factors affecting the reproductive success of female butterflies have been the topic of numerous studies, little information exists on the factors affecting males’ contribution to reproduction, and the indirect impacts on female fecundity and fitness. We used the Glanville fritillary butterfly, Melitaea cinxia (Linnaeus, 1758) (Nymphalidae), in order to assess variation in male allocation to matings. In this species, smaller males produce smaller spermatophores, but variation in spermatophore size is not correlated with female reproductive success. We show that spermatophore size increases with male age at first mating, decreases with mating frequency and adult food‐deprivation, and is not influenced by developmental food‐limitation. The length of copulation period does not influence the spermatophore size nor influences the polyandrous mating behavior in this species. Male contribution to his spermatophore size is clearly influenced by his condition and adult‐resource at the time of mating. Despite this variation, spermatophore size does not seem to have a direct impact on female reproductive output or mating behavior (Less)

The importance of trans-generational effects in shaping an individuals' phenotype and fitness, and consequently even impacting population dynamics is increasingly apparent. Most of the research on trans-generational effects still focuses on plants, mammals, and birds. In the past few years, however, increasing number of studies, especially on maternal effects, have highlighted their importance also in many insect systems. Lepidoptera, specifically butterflies, have been used as model systems for studying the role of phenotypic plasticity within generations. As ectotherms, they are highly sensitive to environmental variation, and indeed many butterflies show adaptive phenotypic plasticity in response to environmental conditions. Here, we synthesize what is known about trans-generational effects in Lepidoptera, compile evidence for different environmental cues that are important drivers of trans-generational effects, and point out which offspring traits are mainly impacted. Finally, we emphasize directions for future research that are needed for better understanding of the adaptive nature of trans-generational effects in Lepidoptera in particular, but potentially also in other organisms.

Down syndrome cell adhesion molecule 1 ( Dscam1 ) has wide-reaching and vital neuronal functions although the role it plays in insect and crustacean immunity is less well understood. In this study, we combine different approaches to understand the roles that Dscam1 plays in fitness-related contexts in two model insect species. Contrary to our expectations, we found no short-term modulation of Dscam1 gene expression after haemocoelic or oral bacterial exposure in Tribolium castaneum , or after haemocoelic bacterial exposure in Drosophila melanogaster. Furthermore, RNAi-mediated Dscam1 knockdown and subsequent bacterial exposure did not reduce T. castaneum survival. However, Dscam1 knockdown in larvae resulted in adult locomotion defects, as well as dramatically reduced fecundity in males and females. We suggest that Dscam1 does not always play a straightforward role in immunity, but strongly influences behaviour and fecundity. This study takes a step towards understanding more about the role of this intriguing gene from different phenotypic perspectives.

Flying insects have the highest known mass-specific demand for oxygen, which makes it likely that reduced availability of oxygen, as in hypoxia, may limit sustained flight instead of or in addition to the limitation due to metabolite resources. The Glanville fritillary butterfly (Melitaea cinxia) occurs as a large metapopulation in which adult butterflies frequently disperse between small local populations. Here, we examine how the interaction between oxygen availability and fuel use affects flight performance in the Glanville fritillary. Individuals were flown under either normoxic (21 kPa O2) or hypoxic (10 kPa O2) conditions and their flight metabolism was measured. To determine resource use, levels of circulating glucose, trehalose and whole-body triglyceride were recorded after flight. Flight performance was significantly reduced in hypoxic conditions. When flown under normoxic conditions, we observed a positive correlation among individuals between post-flight circulating trehalose levels and flight metabolic rate, suggesting that low levels of circulating trehalose constrains flight metabolism. To test this hypothesis experimentally, we measured the flight metabolic rate of individuals injected with a trehalase inhibitor. In support of the hypothesis, experimental butterflies showed significantly reduced flight metabolic rate, but not resting metabolic rate, in comparison with control individuals. On the other hand, under hypoxia there was no relationship between trehalose and flight metabolic rate. Additionally, in this case flight metabolic rate was reduced in spite of circulating trehalose levels that were high enough to support high flight metabolic rate under normoxic conditions. These results demonstrate a significant interaction between oxygen and energy availability in controlling flight performance.

With increasing global warming and habitat fragmentation, environmental conditions are becoming more variable, and the risk and prevalence of diseases may increase in nature. As a consequence, many species have to adapt or respond to rapid changes in their environment. Ectotherms, such as insects, are thought to be especially prone to such changes and thus are interesting and relevant species for assessing life-history responses towards variable conditions. In my thesis, I investigated how the Glanville fritillary butterfly responds to different environmental conditions, and how this translates to fitness-related traits and offspring performance. I was further interested in the potential positive and negative correlations between immunity and other life-history traits. I used experimental approaches conducted in the laboratory to generate changes in individual condition, resulting from variable environmental conditions that were induced via altered nutrition, bacterial and viral infection or via forced flight. Flight was included, as it represents a key life-history trait for this species and is a stressful and energy demanding physiological process. I compared female reproductive strategies, but also assessed how male contribution to female reproduction was affected by variable conditions. Moreover, I assessed potential trans-generational effects on the offspring, and whether the strength or direction of the effect depended on the type of poor condition the individual experienced. Overall the results of this thesis suggest that the Glanville fritillary butterfly is highly sensitive to its surrounding environment, and is able to rapidly change its reproductive strategy according to the conditions it experiences. While more optimal conditions lead to higher offspring quantity, poor or stressful conditions often result in the production of fewer but of higher quality offspring, indicated by e.g. increased hatching success. An increase in offspring quality further translates to increased offspring performance in response to nutritional deficit, thus implying adaptive trans-generational effects. Flight, a key trait in many organisms, has been previously shown to interact with immune response in the Glanville fritillary butterfly. Results from this thesis confirmed this relationship but suggest that the relationship is a general stress response rather than an adaptive response to cope with infections. Bacterial and viral infections drastically reduce lifespan, and hence have the potential to influence population dynamics. Responses to variable environmental conditions including risk of infection are complex in the wild, as several conditions are often acting simultaneously, making it difficult to generalize the observed results. However, the data of this thesis provides important insights in condition-dependent responses of the Glanville fritillary butterfly to stress, and how this translates to fitness-related traits and even offspring performance. Further studies on the adaptive nature and underlying mechanisms behind the observed responses will further contribute to our understanding of the drivers of maintenance of life-history variation and evolution in this species. Ilmaston lämpenemisen ja elinympäristöjen pirstoutumisen seurauksena elinolosuhteet luonnossa muuttuvat vaihtelevammiksi, ja riski erilaisten tautien yleistymiselle ja leviämiselle saattaa kasvaa. Lajien täytyy luonnossa pystyä sopeutumaan nopeasti muuttuvaan ympäristöönsä. Varsinkin vaihtolämpöisten eläinten, kuten hyönteisten, katsotaan olevan alttiita näille muutoksille, ja ne ovat siten erityisen mielenkiintoisia tutkimuskohteita, kun halutaan perehtyä lajien vasteisiin elinympäristön muuttuessa vaihtelevammaksi. Väitöskirjassani tutkin täpläverkkoperhosen vasteita erilaisiin ympäristöolosuhteisiin, ja miten ne siirtyvät sekä perhosen kelpoisuuteen liittyviin ominaisuuksiin että jälkikasvun menestykseen. Tutkimuksessani olin erityisesti kiinnostunut mahdollisista positiivisista ja negatiivisista korrelaatioista yksilöiden immuniteetin ja muiden elinkierron kannalta olennaisten ominaisuuksien välillä. Laboratoriokokeiden avulla muokkasin perhosten kuntoa jäljitellen tilannetta, jossa yksilöiden kunto olisi seurausta muuttuneista elinolosuhteista, mukaan lukien muutoksista ravinnon saannissa, bakteeri- ja virusinfektioissa tai perhosen lentämisestä koituvista kustannuksista. Perhosten lentäminen on elinkiertoon liittyvistä ominaisuuksista avainasemassa, sillä se on fysiologisena prosessina rasittava ja paljon energiaa vaativa. Tutkimuksissani vertailin naarasperhosten lisääntymisstrategioita, ja lisäksi myös missä määriin vaihteleva ympäristö vaikutti koiraiden panokseen naaraiden lisääntymiseen. Tutkin missä määrin vaihtelevien ympäristöolosuhteiden vaikutukset näkyivät jälkikasvussa sukupolvien yli ja missä määrin mahdolliset vaikutukset riippuivat elinolosuhteiden, ja siten perhosyksilöiden kunnon muutosten suunnista ja voimakkuuksista. Tutkimustulokseni viittaavat siihen, että täpläverkko-perhonen on erittäin herkkä elinympäristönsä suhteen. Se pystyy nopeasti muuttamaan lisääntymis-strategiaansa ympäristönsä mukaan — optimaalisissa olosuhteissa perhonen tuottaa enemmän jälkikasvua, kun taas heikkolaatuisessa ja muuten rasittavassa ympäristössä se tuottaa usein määrällisesti vähemmän mutta laadullisesti korkeatasoisempaa jälkikasvua, josta on merkkikä esim. kasvanut kuoriutumisen onnistuminen. Laadullisesta paremmuudesta johtuen jälkikasvu menestyy paremmin huonommissa ravinto-olosuhteissa, ja siten nämä muutokset perhosten lisääntymisstrategiassa välittyvät sukupolvien yli. Lentäminen on monille organismeille avainasemassa oleva osa elinkiertoa, ja aiemmat tutkimukset ovat osoittaneet yhteyden täpläverkkoperhosten lentämisen ja immuunivasteen välillä. Väitöskirjatutkimuksen tulosten perusteella kyseinen yhteys on olemassa, mutta yhteys liittyy enemmän perhosten yleiseen stressivasteeseen rasituksesta johtuen kuin niiden sopeutumiseen infektioihin. Bakteeri- ja virusinfektiot lyhentävät yksilöiden elinikää merkittävästi, ja siten ne voivat vaikuttaa merkittävästi lajien populaatiodynamiikkaan. Vasteet muuttuvaan elinympäristöön, mukaan lukien kasvavaan infektioriskiin, ovat monimutkaisia, ja useat eri olosuhteiden muutokset vaikuttavat näihin vasteisiin yhtäaikaisesti. Havaittujen ilmiöiden yleistäminen on siten haastavaa, mutta siitä huolimatta tämän tutkimuksen tulokset syventävät ymmärrystämme täpläverkkoperhosen vasteista ympäristön aiheuttamaan rasitukseen, ja miten ne näkyvät ominaisuuksissa sukupolvien yli sekä jälkikasvun suorituskyvyssä. Tutkimuksessa havaittujen prosessien taustalla olevien mekanismien ja niiden adaptiivisuuden arviointi vaatii lisätutkimuksia. Niiden avulla voitaisiin edelleen syventää ymmärrystämme täpläverkkoperhosen elinkierron vaihtelevuutta ja evoluutiota ajavista voimista.