Search

Dyadic contests for possession of resources occur across a wide range of animal taxa, with contest outcome often being heavily influenced by the energetic reserves of the competitors. The majority of studied parasitoid wasps lack de novo lipogenesis, with adult lipid reserves being limited to those acquired throughout larval development. Carbohydrate- and lipid-rich adult diets can increase adult parasitoid lifespan and fecundity by potentially acting as a maintenance energy store. However, the effects of such diets on fat reserve compositions and some aspects of parasitoid behaviour contest outcome dynamics have not been examined. This study assesses the effect of a carbohydrate rich diet (honey) on the longevity, metabolic state and contest performance of Goniozus legneri, a bethylid wasp. The longevity of honey-fed diet adults was typically more than twice that of starved wasps. Compared to similarly-aged starved wasps, honey feeding increased concentrations of common haemolymph sugars and amino acids in a 3-day old and 7-day old wasps, and higher concentrations of stored lipid in7-day old wasps. However, nutritional status did not affect the outcome of dyadic contests over host possession when both contestants were either 3 days or 7 days old. While contest dynamics outcome may be unaffected by diet, it remains possible that an enhancing effect of feeding on contest ability is matched by an effect of higher value being placed on winning by starved wasps.

Metabolomic analyses can reveal associations between an organism's metabolome and further aspects of its phenotypic state, an attractive prospect for many life-sciences researchers. The metabolomic approach has been employed in some, but not many, insect study systems, starting in 1990 with the evaluation of the metabolic effects of parasitism on moth larvae. Metabolomics has now been applied to a variety of aspects of insect biology, including behaviour, infection, temperature stress responses, CO2 sedation, and bacteria–insect symbiosis. From a technical and reporting standpoint, these studies have adopted a range of approaches utilising established experimental methodologies. Here, we review current literature and evaluate the metabolomic approaches typically utilised by entomologists. We suggest that improvements can be made in several areas, including sampling procedures, the reduction in sampling and equipment variation, the use of sample extracts, statistical analyses, confirmation, and metabolite identification. Overall, it is clear that metabolomics can identify correlations between phenotypic states and underlying cellular metabolism that previous, more targeted, approaches are incapable of measuring. The unique combination of untargeted global analyses with high-resolution quantitative analyses results in a tool with great potential for future entomological investigations.

Many animals avoid attack from predators through toxicity or the emission of repellent chemicals. Defensive mimicry has evolved in many species to deceive shared predators, for instance through colouration and other morphological adaptations, but mimicry hardly ever seems to involve multi-trait similarities. Here we report on a wingless parasitoid wasp that exhibits a full spectrum of traits mimicing ants and affording protection against ground-dwelling predators (wolf spiders). In body size, morphology and movement Gelis agilis (Ichneumonidae) is highly similar to the black garden ant (Lasius niger) that shares the same habitat. When threatened, G. agilis also emits a volatile chemical that is similar to an ant-produced chemical that repels spiders. In bioassays with L. niger, G. agilis, G. areator, Cotesia glomerata and Drosophila melanogaster, ants and G. agilis were virtually immune to spider attack, in contrast the other species were not. Volatile characterisation with gas chromatography-mass spectrometry identified G. agilis emissions as 6-methyl-5-hepten-2-one, a known insect defence semiochemical that acts as an alarm pheromone in ants. We argue that multi-trait mimicry, as observed in G. agilis, might be much more common among animals than currently realized.

Metabolomics studies of low-biomass organisms, such as small insects, have previously relied on the pooling of biological samples to overcome detection limits, particularly using NMR. We show that the differentiation of metabolite profiles of individual 1 mg parasitoid wasps of different ages is possible when using a modified sample preparation and a combination of untargeted NMR and LC-MS based metabolomics. Changes were observed between newly emerged and older wasps in glycerolipids, amino acids and circulatory sugars. This advance in chemical profiling has important implications for the study of the behaviour and ecology of parasitoids and many other species of small organisms because predictions and observations are typically made at the level of the individual. Thus, the metabolomic state of low-biomass individuals can now be related to their behaviour and ecological performance. We discuss specifically the utility of age-related metabolomic profiling but our new approach can be applied to a wide range of biological research.

The current decline of the European Honey Bee (Apis Mellifera) has been linked to the increasing use of neonicotinoid pesticides within agriculture. Whilst the toxicity of these pesticides to Apis has long been established, the possibility of low dosages inducing molecular stress has not yet been fully explored. Of particular interest is the action of these nicotine derivatives on the nicotinic acetylcholine receptor, and its association with the DNA methyltransferase family (Dnmts). An experimental group of three hives were exposed to sugar water contaminated with a low concentration of imidacloprid (2µg/l). From these hives, 12 third instar larvae were selected. A corresponding number of larvae were also selected from three control hives, for a total of 12 samples. Using quantitative reverse transcription-PCR, known Dnmt transcripts were detected and amplified from these larval samples. Specially designed oligonucleotide primers were used containing gene specific sequences that linked to universal DNA sequences, ensuring that PCR amplification products were of predetermined sizes. Products of this amplification were resolved by capillary electrophoresis and detected by fluorescence spectrophotometry. Simultaneously, the transcriptomes of 3 larval samples each from the control and experimental groups were generated using SOLiD platform sequencing. The statistical package EdgeR was then utilised to identify differential candidates of known honey bee microRNA’s. Statistical analysis utilising a one-way Analysis of Variance (ANOVA) found no significant differences in the expression levels of known Dnmt transcripts between control and experimental groups. However, comparisons of sequenced control and experimental transcriptomes identified a number of differential microRNA candidates, most notably miR-9/14.

The survival of a species depends on its capacity to adjust to changing environmental conditions, and new stressors. Such new, anthropogenic stressors include the neonicotinoid class of crop-protecting agents, which have been implicated in the population declines of pollinating insects, including honeybees (Apis mellifera). The low-dose effects of these compounds on larval development and physiological responses have remained largely unknown. Over a period of 15 days, we provided syrup tainted with low levels (2 µg/L−1) of the neonicotinoid insecticide imidacloprid to beehives located in the field. We measured transcript levels by RNA sequencing and established lipid profiles using liquid chromatography coupled with mass spectrometry from worker-bee larvae of imidacloprid-exposed (IE) and unexposed, control (C) hives. Within a catalogue of 300 differentially expressed transcripts in larvae from IE hives, we detect significant enrichment of genes functioning in lipid-carbohydrate-mitochondrial metabolic networks. Myc-involved transcriptional response to exposure of this neonicotinoid is indicated by overrepresentation of E-box elements in the promoter regions of genes with altered expression. RNA levels for a cluster of genes encoding detoxifying P450 enzymes are elevated, with coordinated downregulation of genes in glycolytic and sugar-metabolising pathways. Expression of the environmentally responsive Hsp90 gene is also reduced, suggesting diminished buffering and stability of the developmental program. The multifaceted, physiological response described here may be of importance to our general understanding of pollinator health. Muscles, for instance, work at high glycolytic rates and flight performance could be impacted should low levels of this evolutionarily novel stressor likewise induce downregulation of energy metabolising genes in adult pollinators.

This thesis examines the relationship between insect life history and behavioural decisions and underlying cellular biochemistry, with particular focus on bethylid parasitoid wasps in the genus Goniozus. This comprises the first major body of work attempting to draw links between the underlying metabolome of an organism and its behaviour. This thesis further optimised the first known example of a combined LC-MS and NMR metabolomic approach capable of analysing extremely low biomass samples (<1 mg), a vital requirement when studying the behaviour of individual organisms. Part 1 of this thesis details the optimisation and validation of this metabolomic approach, whilst also examining the effects of aging on the metabolome of adult Goniozus wasps. Part 2 applies this approach to examine the effects of diet, host species and host aging on Goniozus wasp behaviour and biochemistry. Comparisons of the metabolomes of starved and honey fed wasps indicate that G. legneri is capable of utilising a carbohydrate rich diet as an energy source. Aged honey fed wasps possessed higher levels of large storage lipids, such as tri- and diacylglycerides, than starved wasps of the same age. Metabolomic analysis also detected a legacy effect on the metabolome of G. legneri associated with differences in the species of host each wasp was reared on. A similar legacy effect was confirmed when examining the metabolomes of wasps reared on artificially aged hosts. Whilst Goniozus wasp oviposition behaviour was altered by the species of host presented, no links between changes in a wasp’s metabolome and its resulting contest behaviour were found. Part 3 of this thesis examines the morphological, behavioural and chemical mimicry of another wasp, the hyperparasitoid Gelis agilis. G. agilis demonstrated an enhanced predation avoidance rate compared with control species, similar to that of the black garden ant Lasius niger. Agitation of G. agilis also resulted in the chemical emission of a known ant

The current decline of the European Honey Bee (Apis Mellifera) has been linked to the increasing use of neonicotinoid pesticides within agriculture. Whilst the toxicity of these pesticides to Apis has long been established, the possibility of low dosages inducing molecular stress has not yet been fully explored. Of particular interest is the action of these nicotine derivatives on the nicotinic acetylcholine receptor, and its association with the DNA methyltransferase family (Dnmts). An experimental group of three hives were exposed to sugar water contaminated with a low concentration of imidacloprid (2µg/l). From these hives, 12 third instar larvae were selected. A corresponding number of larvae were also selected from three control hives, for a total of 12 samples. Using quantitative reverse transcription-PCR, known Dnmt transcripts were detected and amplified from these larval samples. Specially designed oligonucleotide primers were used containing gene specific sequences that linked to universal DNA sequences, ensuring that PCR amplification products were of predetermined sizes. Products of this amplification were resolved by capillary electrophoresis and detected by fluorescence spectrophotometry. Simultaneously, the transcriptomes of 3 larval samples each from the control and experimental groups were generated using SOLiD platform sequencing. The statistical package EdgeR was then utilised to identify differential candidates of known honey bee microRNA’s. Statistical analysis utilising a one-way Analysis of Variance (ANOVA) found no significant differences in the expression levels of known Dnmt transcripts between control and experimental groups. However, comparisons of sequenced control and experimental transcriptomes identified a number of differential microRNA candidates, most notably miR-9/14.

The current decline of the European Honey Bee (Apis Mellifera) has been linked to the increasing use of neonicotinoid pesticides within agriculture. Whilst the toxicity of these pesticides to Apis has long been established, the possibility of low dosages inducing molecular stress has not yet been fully explored. Of particular interest is the action of these nicotine derivatives on the nicotinic acetylcholine receptor, and its association with the DNA methyltransferase family (Dnmts). An experimental group of three hives were exposed to sugar water contaminated with a low concentration of imidacloprid (2µg/l). From these hives, 12 third instar larvae were selected. A corresponding number of larvae were also selected from three control hives, for a total of 12 samples. Using quantitative reverse transcription-PCR, known Dnmt transcripts were detected and amplified from these larval samples. Specially designed oligonucleotide primers were used containing gene specific sequences that linked to universal DNA sequences, ensuring that PCR amplification products were of predetermined sizes. Products of this amplification were resolved by capillary electrophoresis and detected by fluorescence spectrophotometry. Simultaneously, the transcriptomes of 3 larval samples each from the control and experimental groups were generated using SOLiD platform sequencing. The statistical package EdgeR was then utilised to identify differential candidates of known honey bee microRNA’s. Statistical analysis utilising a one-way Analysis of Variance (ANOVA) found no significant differences in the expression levels of known Dnmt transcripts between control and experimental groups. However, comparisons of sequenced control and experimental transcriptomes identified a number of differential microRNA candidates, most notably miR-9/14.

The current decline of the European Honey Bee (Apis Mellifera) has been linked to the increasing use of neonicotinoid pesticides within agriculture. Whilst the toxicity of these pesticides to Apis has long been established, the possibility of low dosages inducing molecular stress has not yet been fully explored. Of particular interest is the action of these nicotine derivatives on the nicotinic acetylcholine receptor, and its association with the DNA methyltransferase family (Dnmts). An experimental group of three hives were exposed to sugar water contaminated with a low concentration of imidacloprid (2µg/l). From these hives, 12 third instar larvae were selected. A corresponding number of larvae were also selected from three control hives, for a total of 12 samples. Using quantitative reverse transcription-PCR, known Dnmt transcripts were detected and amplified from these larval samples. Specially designed oligonucleotide primers were used containing gene specific sequences that linked to universal DNA sequences, ensuring that PCR amplification products were of predetermined sizes. Products of this amplification were resolved by capillary electrophoresis and detected by fluorescence spectrophotometry. Simultaneously, the transcriptomes of 3 larval samples each from the control and experimental groups were generated using SOLiD platform sequencing. The statistical package EdgeR was then utilised to identify differential candidates of known honey bee microRNA’s. Statistical analysis utilising a one-way Analysis of Variance (ANOVA) found no significant differences in the expression levels of known Dnmt transcripts between control and experimental groups. However, comparisons of sequenced control and experimental transcriptomes identified a number of differential microRNA candidates, most notably miR-9/14.

The current decline of the European Honey Bee (Apis Mellifera) has been linked to the increasing use of neonicotinoid pesticides within agriculture. Whilst the toxicity of these pesticides to Apis has long been established, the possibility of low dosages inducing molecular stress has not yet been fully explored. Of particular interest is the action of these nicotine derivatives on the nicotinic acetylcholine receptor, and its association with the DNA methyltransferase family (Dnmts). An experimental group of three hives were exposed to sugar water contaminated with a low concentration of imidacloprid (2µg/l). From these hives, 12 third instar larvae were selected. A corresponding number of larvae were also selected from three control hives, for a total of 12 samples. Using quantitative reverse transcription-PCR, known Dnmt transcripts were detected and amplified from these larval samples. Specially designed oligonucleotide primers were used containing gene specific sequences that linked to universal DNA sequences, ensuring that PCR amplification products were of predetermined sizes. Products of this amplification were resolved by capillary electrophoresis and detected by fluorescence spectrophotometry. Simultaneously, the transcriptomes of 3 larval samples each from the control and experimental groups were generated using SOLiD platform sequencing. The statistical package EdgeR was then utilised to identify differential candidates of known honey bee microRNA’s. Statistical analysis utilising a one-way Analysis of Variance (ANOVA) found no significant differences in the expression levels of known Dnmt transcripts between control and experimental groups. However, comparisons of sequenced control and experimental transcriptomes identified a number of differential microRNA candidates, most notably miR-9/14.

This thesis examines the relationship between insect life history and behavioural decisions and underlying cellular biochemistry, with particular focus on bethylid parasitoid wasps in the genus Goniozus. This comprises the first major body of work attempting to draw links between the underlying metabolome of an organism and its behaviour. This thesis further optimised the first known example of a combined LC-MS and NMR metabolomic approach capable of analysing extremely low biomass samples (<1 mg), a vital requirement when studying the behaviour of individual organisms. Part 1 of this thesis details the optimisation and validation of this metabolomic approach, whilst also examining the effects of aging on the metabolome of adult Goniozus wasps. Part 2 applies this approach to examine the effects of diet, host species and host aging on Goniozus wasp behaviour and biochemistry. Comparisons of the metabolomes of starved and honey fed wasps indicate that G. legneri is capable of utilising a carbohydrate rich diet as an energy source. Aged honey fed wasps possessed higher levels of large storage lipids, such as tri- and diacylglycerides, than starved wasps of the same age. Metabolomic analysis also detected a legacy effect on the metabolome of G. legneri associated with differences in the species of host each wasp was reared on. A similar legacy effect was confirmed when examining the metabolomes of wasps reared on artificially aged hosts. Whilst Goniozus wasp oviposition behaviour was altered by the species of host presented, no links between changes in a wasp’s metabolome and its resulting contest behaviour were found. Part 3 of this thesis examines the morphological, behavioural and chemical mimicry of another wasp, the hyperparasitoid Gelis agilis. G. agilis demonstrated an enhanced predation avoidance rate compared with control species, similar to that of the black garden ant Lasius niger. Agitation of G. agilis also resulted in the chemical emission of a known ant