@phdthesis{CastilloCajas2020,
  author    = {Castillo Cajas, Ruth},
  title     = {Evolution and diversity of cuticular hydrocarbon proﬁles of cuckoo wasps},
  doi       = {10.25972/OPUS-17341},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-173418},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2020},
  abstract  = {Cuticular hydrocarbons (CHC) abound on the surface of arthropods. In spite of their simple structure (molecules of carbon and hydrogen atoms), they provide pivotal functions in insects: their hydrophobic properties confer the insects a means to regulate water balance and avoid desiccation, whereas their diversity has enhanced their use as signals and cues in a wide range of communication and recognition processes. Although the study of CHC in insects over the past two decades has provided great insight into the wide range of functions they play, there is still a gap in understanding how they diversify and evolve. In this thesis, I have used members of the family Chrysididae to explore patterns of diversiﬁcation of CHC. Most of the species of cuckoo wasps in this study are specialized parasitoids or kleptoparasites of mainly solitary hymenopteran hosts. Other hosts of the family include butterﬂies or stick insects. Cuckoo wasps are a particular interesting model to study the evolution of cuticular hydrocarbons because of their chemical adaptations that allow them to remain unrecognized by their hosts. Chemical insigniﬁcance (the reduction of the total amount of CHC on the cuticle) and chemical mimicry (the de novo production of CHC proﬁles resembling those of their female host) have been described in some representatives of the family and unpublished evidence suggests chemical deception is widespread in Chrysididae (Chapter 2). Nonetheless, to trace the evolution of any trait of interest, a reliable phylogenetic reconstruction of the family is required. Therefore, the ﬁrst study of this thesis constitutes the largest and to-date most reliable phylogenetic reconstruction of the family Chrysididae, which includes representatives of 186 species of cuckoo wasps. While the results of this phylogenetic reconstruction are consistent with previous ideas on the relationships of subfamilies and tribes, it shows the existence of several non-monophyletic genera (Chapter 3). CHC are involved in intraspeciﬁc recognition, often acting as contact sex pheromones. Nevertheless, it is not yet understood to what extent CHC proﬁles diﬀer between the two sexes and whether some compound classes are more prevalent in one or the other sex. So far, no comparison of CHC proﬁles of males and females has been done for more than a dozen of related species. In Chapter 4, I describe and compare CHC proﬁles of females and males of 58 species of cuckoo wasps in order to evaluate whether and to what extent CHC proﬁles of these species diﬀer between the sexes. I demonstrated that CHC proﬁles of cuckoo wasps are frequently (more than 90\% of the species analyzed) and strongly dimorphic (both sexes of a given species tend to produce very diﬀerent CHC compounds). Methyl-branched compounds tend to be more prevalent in males (especially dimethyl-branched compounds) and unsaturated compounds prevail in females. Moreover, a sex-speciﬁc pattern in the distribution of the double bond position of alkenes was evident: internal double bond positions (> 11) occur predominantly in males, whereas alkenes with the doubl{\´e} bond at position 9 were more abundant and frequent in females (Chapter4). In Chapter5, I investigated how CHC proﬁles of cuckoo wasps diﬀer across species. Are CHC proﬁles of cuckoo wasps species-speciﬁc, enabling their use as cues for species recognition? How do CHC proﬁles resemble phylogenetic relatedness? In Chapter 5, I try to answer these questions by comparing CHC proﬁles of 59 species of cuckoo wasps. CHC proﬁles of cuckoo wasps are shown to be species (and sex-) speciﬁc. I show that CHC proﬁles are useful as a complementary tool to help delimiting taxonomically diﬃcult sibling species. Moreover, the evaluation of CHC proﬁles of ﬁve commonly occurring species within a genus, showed little or no geographical variation. However, CHC proﬁles of closely related species may diﬀer strongly among each other, not being useful to track the evolutionary history of species (Chapter 5). Sexual selection is generally credited for generating striking sexual dimorphism by causing changes in male traits. Most often, sexual selection has a stronger eﬀect on males, who compete for access to and may be selected by females, thus male traits may rapidly evolve. Nevertheless, in cuckoo wasps, it appears that it is the female sex the one evolving faster changes, with females of very closely related species showing extremely divergent proﬁles. One plausible reason for this disparity is that natural selection acting on female's CHC proﬁles may be stronger than sexual selection on males (Chapter 6). Since females of cuckoo wasps are most probably engaged in an evolutionary arms race with their female hosts, CHC proﬁles of female cuckoo wasps are likely rapidly evolving, thus explaining part of the strong observed sexual dimorphism of CHC (Chapter 6). In fact, Chapter 7 shows evidence of a possible ongoing evolutionary arms race between ﬁve cuckoo wasps of the genus Hedychrum and their hosts. Hedychrum species parasitize either Coleoptera-hunting or Hymenoptera-hunting digger wasps. Since the coleopteran prey of the former digger wasps is naturally better protected against fungus infestation, these wasps do not embalm their prey with alkene-enriched secretions as do the Hymenoptera-hunting digger wasps. Thus, Coleoptera-hunting digger wasps can apparently diversify their proﬁles to escape chemical mimicry. Interestingly, only female cuckoo wasps of these hosts have started producing the same compound classes and even the same CHC compounds as those of their hosts. Male cuckoo wasps, however retain an alkene-enriched CHC proﬁle that reﬂects the molecular phylogeny of the genus (Chapter 7). Whereas, a larger number of parasite-host comparisons may be needed to further conclude that an arms race between cuckoo wasps and their hosts is capable of generating sexual dimorphism of cuckoo wasps, this thesis constitutes the ﬁrst eﬀort towards this, providing a starting point for further studies. Finally, I provide some methodological tools that may help in speeding up the sometimes cumbersome process of analyzing and identifying CHC proﬁles. One of the most time-demanding steps in the processing of CHC data is the alignment of CHC chromatograms. This process is often done manually, because alignment programs are mostly designed for metabolomics or are just recently being developed. I analyzed CHC proﬁles using a combined approach with two freely available programs. I used AMDIS (Automated Mass Spectral Deconvolution and Identiﬁcation System, http://chemdata.nist.gov/mass-spc/amdis/) to deconvolute and automatically identify all CHC of interest present in a chromatogram. I then developed a series of R scripts to correct for potential, unavoidable errors while processing CHC chromatograms with AMDIS. Chapter 8 explains this procedure. In the next chapter, I developed a program that helps in the identiﬁcation of one commonly occurring class of hydrocarbons. The limited number of linear alkanes (only one per carbon atom) and their characteristic diagnostic ion allows a rapid and unambigous identiﬁcation of these substances. In opposition, unsaturated and methyl-branched compounds are more diﬃcult to identify, as a result of the much larger diversity of existing compounds. To identify unsaturated compounds a derivatization is necessary to determine the position of the double bond. Methyl-branched alkanes, however can be identiﬁed from the original chromatogram if their diagnostic ions are known. Nonetheless, polymethyl-branched alkanes (e.g., compounds with two or more methyl groups along the chain) are often diﬃcult to identify, because they may appear in mixes (e.g., 3,7 diMeC27 and 3,9 diMeC27), and tables containing the diagnostic ions are not easily available. Therefore, I developed a program that creates a table with all possiblemethyl-branched compounds containing up to 4 methyl groups, and that provides their diagnostic ions and a calculated retention index. This may allow a much faster identiﬁcation of the methyl-branched compound a researcher is dealing with, without having to lose time in the tedious calculations by hand. The program is able to correctly identify, or at least, greatly reduce the number of possible options for the identiﬁcation of an unknown methyl-branched compound. Thus, using this tool, most methyl-branched compounds can be readily identiﬁed (Chapter 9). This thesis ends with a general discussion (Chapter 10). Overall, this work provides a comprehensive overview of the diversity of cuticular hydrocarbons of cuckoo wasps. The analyses presented here shed light on the emergence and evolution of interspeciﬁc diversity and intraspeciﬁc sexual dimorphism of CHC proﬁles. In addition, two technical methods have been developed that could greatly facilitate the CHC analysis of insects.},
  language  = {en}
}