Open-source flow setup for rapid and efficient [18 F]fluoride drying for automation of PET tracer syntheses.

One of the key strategies for radiochemical research facilities is the automation of synthesis processes. Unnecessary manual operations increase the radiation exposure of personnel, while simultaneously threatening the reliability of syntheses. We have previously reported an affordable open-source system comprising 3D-printed continuous flow reactors, a custom syringe pump, and a pressure regulator that can be used to perform radiofluorinations. In this paper, we address additional essential processes that are needed for radiotracer development and synthesis, with the aim of making laboratory work safer and research more efficient. We have designed and evaluated a fully automated system for rapidly and effectively processing and drying aqueous [18 F]fluoride that can be directly connected to the cyclotron. This process relies on triflyl fluoride gas generation and allows nucleophilic [18 F]fluoride to be prepared safely in a hotcell within 10 min and an activity recovery of 91.7 ± 1.6% (n = 5). Owing to the need for convenient radiofluorinated prosthetic ligands, we have adapted our continuous flow system to produce [18 F]fluoroethyl tosylate (FEOTs) and [18 F]fluoroethyl triflate (FEOTf), prosthetic groups that are widely used for late-stage fluoroethylation of PET tracers. The processes as well as the radiolabeling of different groups are compared and comprehensively discussed. Having a method providing [18 F]fluoroethyl tosylate (FEOTs) as well as [18 F]fluoroethyl triflate (FEOTf) quickly and highly efficiently is beneficial for radiochemical research.

Change of mechanical characteristics in spider silk capture threads after contact with prey.

Most spiders rely on specialized capture threads to subdue prey. Cribellate spiders use capture threads, whose adhesion is based on thousands of nanofibers instead of specialized glue. The nanofibers adhere due to van der Waals and hygroscopic forces, but the adhesion is strengthened by an interaction with the cuticular hydrocarbons (CHCs) covering almost all insects. The interaction between CHCs and cribellate threads becomes visible through migration of the CHCs into the thread even far beyond the point of contact. In this study, we were able to show that the migrated CHCs not only influence adhesion but also change the mechanical characteristics of the thread. While adhesion, extensibility and total energy decreased in threads treated with CHCs from different insects, we observed an increasing force required to break threads. Such mechanical changes could be beneficial for the spider: Upon the first impact of the insect in the web, it is important to absorb all the energy without breaking. Afterwards, a reduction in extensibility could cause the insect to stay closer to the web and thus become additionally entangled in neighboring threads. An increased tensile force would additionally ensure that for insects already in the web, it is even harder to free themselves. Taken together, all these changes make it unlikely that cribellate spiders reuse their capture threads, if not reacting rapidly and removing the prey insect before the CHCs can spread across the thread. STATEMENT OF SIGNIFICANCE: Cribellate spiders use capture threads that, unlike other spiders, consist of nanofibers and do not rely glue. Instead, prey adheres mainly because their surface compounds, so-called cuticular hydrocarbons (CHCs), interact with the thread, this way generating strong adhesion forces. Previous studies on biomechanics and adhesion of cribellate threads only dealt with artificial surfaces, neglecting any interaction with surface compounds. This study examines the dramatical mechanical changes of a cribellate thread after interaction with prey CHCs, showing modifications of the thread's extensibility, tensile force and total energy. Our results highlight the importance of studying mechanical properties of silk not only in an artificial context, but also in real life.

Why do ants differ in acclimatory ability? Biophysical mechanisms behind cuticular hydrocarbon acclimation across species.

Maintaining water balance is vital for terrestrial organisms. Insects protect themselves against desiccation via cuticular hydrocarbons (CHCs). CHC layers are complex mixtures of solid and liquid hydrocarbons, with a surprisingly diverse composition across species. This variation may translate into differential phase behaviour, and hence varying waterproofing capacity. This is especially relevant when temperatures change, which requires acclimatory CHC changes to maintain waterproofing. Nevertheless, the physical consequences of CHC variation are still little understood. We studied acclimatory responses and their consequences for CHC composition, phase behaviour and drought survival in three congeneric ant species. Colony sub-groups were kept under cool, warm and fluctuating temperature regimes. Lasius niger and Lasius platythorax, both of which are rich in methyl-branched alkanes, showed largely predictable acclimatory changes of the CHC profile. In both species, warm acclimation increased drought resistance. Warm acclimation increased the proportion of solid compounds in L. niger but not in L. platythorax. In both species, the CHC layer formed a liquid matrix of constantly low viscosity, which contained highly viscous and solid parts. This phase heterogeneity may be adaptive, increasing robustness to temperature fluctuations. In Lasius brunneus, which is rich in unsaturated hydrocarbons, acclimatory CHC changes were less predictable, and warm acclimation did not enhance drought survival. The CHC layer was more homogeneous, but matrix viscosity changed with acclimation. We showed that ant species use different physical mechanisms to enhance waterproofing during acclimation. Hence, the ability to acclimate, and thus climatic niche breadth, may strongly depend on species-specific CHC profile.

Insect cuticular hydrocarbon composition influences their interaction with spider capture threads.

Insects represent the main prey of spiders, and spiders and insects co-diversified in evolutionary history. One of the main features characterizing spiders is their web as a trap to capture prey. Phylogenetically, the cribellate thread is one of the earliest thread types that was specialized to capture prey. In contrast to other capture threads, it lacks adhesive glue and consists of nanofibres, which do not only adhere to insects via van der Waals forces but also interact with the insects' cuticular hydrocarbon (CHC) layer, thus enhancing adhesion. The CHC layer consists of multiple hydrocarbon types and is highly diverse between species. In this study, we show that CHC interaction with cribellate capture threads is affected by CHC composition of the insect. We studied the interaction in detail for four insect species with different CHC profiles and observed a differential migration of CHCs into the thread. The migration depends on the molecular structure of the hydrocarbon types as well as their viscosity, influenced by the ambient temperature during the interaction. As a consequence, adhesion forces to CHC layers differ depending on their chemical composition. Our results match predictions based on biophysical properties of hydrocarbons, and show that cribellate spiders can exert selection pressure on the CHC composition of their insect prey.

Cuticular hydrocarbon profiles differ between ant body parts: implications for communication and our understanding of CHC diffusion.

Insect cuticular hydrocarbons (CHCs) serve as communication signals and protect against desiccation. They form complex blends of up to 150 different compounds. Due to differences in molecular packing, CHC classes differ in melting point. Communication is especially important in social insects like ants, which use CHCs to communicate within the colony and to recognize nestmates. Nestmate recognition models often assume a homogenous colony odor, where CHCs are collected, mixed, and redistributed in the postpharyngeal gland (PPG). Via diffusion, recognition cues should evenly spread over the body surface. Hence, CHC composition should be similar across body parts and in the PPG. To test this, we compared CHC composition among whole-body extracts, PPG, legs, thorax, and gaster, across 17 ant species from 3 genera. Quantitative CHC composition differed between body parts, with consistent patterns across species and CHC classes. Early-melting CHC classes were most abundant in the PPG. In contrast, whole body, gaster, thorax, and legs had increasing proportions of CHC classes with higher melting points. Intraindividual CHC variation was highest for rather solid, late-melting CHC classes, suggesting that CHCs differ in their diffusion rates across the body surface. Our results show that body parts strongly differ in CHC composition, either being rich in rather solid, late-melting, or rather liquid, early-melting CHCs. This implies that recognition cues are not homogenously present across the insect body. However, the unequal diffusion of different CHCs represents a biophysical mechanism that enables caste differences despite continuous CHC exchange among colony members.

Extreme lifespan extension in tapeworm-infected ant workers.

Social insects are hosts of diverse parasites, but the influence of these parasites on phenotypic host traits is not yet well understood. Here, we tracked the survival of tapeworm-infected ant workers, their uninfected nest-mates and of ants from unparasitized colonies. Our multi-year study on the ant Temnothorax nylanderi, the intermediate host of the tapeworm Anomotaenia brevis, revealed a prolonged lifespan of infected workers compared with their uninfected peers. Intriguingly, their survival over 3 years did not differ from those of (uninfected) queens, whose lifespan can reach two decades. By contrast, uninfected workers from parasitized colonies suffered from increased mortality compared with uninfected workers from unparasitized colonies. Infected workers exhibited a metabolic rate and lipid content similar to young workers in this species, and they received more social care than uninfected workers and queens in their colonies. This increased attention could be mediated by their deviant chemical profile, which we determined to elicit more interest from uninfected nest-mates in a separate experiment. In conclusion, our study demonstrates an extreme lifespan extension in a social host following tapeworm infection, which appears to enable host workers to retain traits typical for young workers.

Candidate genes involved in cuticular hydrocarbon differentiation between cryptic, parabiotic ant species.

Insect cuticular hydrocarbons (CHCs) are highly diverse and have multiple functions, including communication and waterproofing. CHC profiles form species-specific, complex blends of up to 150 compounds. Especially in ants, even closely related species can have largely different profiles, raising the question how CHC differences are mirrored in the regulation of biosynthetic pathways. The neotropical ants Crematogaster levior and Camponotus femoratus both consist of two cryptic species each that are morphologically similar, but express strongly different CHC profiles. This is ideal to study the molecular basis of CHC differences. We thus investigated gene expression differences in fat-body transcriptomes of these ants. Despite common garden conditions, we found several thousand differentially expressed transcripts within each cryptic species pair. Many of these were related to metabolic processes, probably accounting for physiological differences. Moreover, we identified candidate genes from five gene families involved in CHC biosynthesis. By assigning candidate transcripts to orthologs in Drosophila, we inferred which CHCs might be influenced by differential gene expression. Expression of these candidate genes was often mirrored in the CHC profiles. For example, Cr. levior A, which has longer CHCs than its cryptic sister species, had a higher expression of elongases and a lower expression of fatty acyl- CoA reductases. This study is one of the first to identify CHC candidate genes in ants and will provide a basis for further research on the genetic basis of CHC biosynthesis.

How does climate change affect social insects?

Climate change poses a major threat to global biodiversity, already causing sharp declines of populations and species. In some social insect species we already see advanced phenologies, changes in distribution ranges, and changes in abundance Rafferty (2017) and Diamond et al. (2017). Physiologically, social insects are no different from solitary insects, but they possess a number of characteristics that distinguish their response to climate change. Here, we examine these traits, which might enable them to cope better with climate change than solitary insects, but only in the short term. In addition, we discuss how climate change will alter biotic interactions and ecosystem functions, and how it will affect invasive social insects.

Little parallelism in genomic signatures of local adaptation in two sympatric, cryptic sister species.

Species living in sympatry and sharing a similar niche often express parallel phenotypes as a response to similar selection pressures. The degree of parallelism within underlying genomic levels is often unexplored, but can give insight into the mechanisms of natural selection and adaptation. Here, we use multi-dimensional genomic associations to assess the basis of local and climate adaptation in two sympatric, cryptic Crematogaster levior ant species along a climate gradient. Additionally, we investigate the genomic basis of chemical communication in both species. Communication in insects is mainly mediated by cuticular hydrocarbons (CHCs), which also protect against water loss and, hence, are subject to changes via environmental acclimation or adaptation. The combination of environmental and chemical association analyses based on genome-wide Pool-Seq data allowed us to identify single nucleotide polymorphisms (SNPs) associated with climate and with chemical differences. Within species, CHC changes as a response to climate seem to be driven by phenotypic plasticity, since there is no overlap between climate- and CHC-associated SNPs. The only exception is the odorant receptor OR22c, which may be a candidate for population-specific CHC recognition in one of the species. Within both species, climate is significantly correlated with CHC differences, as well as to allele frequency differences. However, associated candidate SNPs, genes and functions are largely species-specific and we find evidence for minimal parallel evolution only on the level of genomic regions (J = 0.04). This highlights that even closely related species may follow divergent evolutionary trajectories when expressing similar adaptive phenotypes.

Correction to: Ant-mediated ecosystem processes are driven by trophic community structure but mainly by the environment.

Unfortunately, the given and family names of author "Mickal Houadria" was incorrectly published in the original.

Non-consumptive effects between predators depend on the foraging mode of intraguild prey.

Predator non-consumptive effects (NCE) are a suite of phenotypic responses of prey to avoid predation that cascade down food webs and can have a stronger impact than predation itself. However, the role of NCE in intraguild interactions remains understudied. Thus, multi-species experiments based on species traits are needed to improve our understanding of the impact of NCE on community structure. We investigated NCE of ants on 17 spider species as intraguild prey (8 cursorial and 9 sedentary) to test the influence of spider hunting mode on the strength of NCE. Sedentary spiders select a habitat patch and wait for prey, whereas cursorial spiders roam around searching for prey. Because actively foraging organisms have higher encounter rates with predators, cursorial spiders should show stronger reactions to ant presence. We collected cues (footprint hydrocarbons) of the black garden ant Lasius niger on filter papers. Then, we placed spiders individually on the papers and filmed their behaviour once on a paper with cues and once on a cue-free one. We used the increase in walking activity in the presence of cues as an indicator of avoidance behaviour towards ants. Cursorial spiders spent a longer time moving and being mobile, and had a higher overall mean mobility when exposed to ant cues compared with blank filter papers, whereas sedentary spider species did not react to ant cues at all. The avoidance of ant cues by cursorial spiders corresponds to their higher risk of encountering ants compared with sedentary species, in accordance with the threat-sensitivity hypothesis. Our multi-species study shows the relevance of NCE for intraguild interactions and highlights the importance of experimental trait-based approaches to improve predictability of species interactions and the role of NCE in ecological communities.

Communication versus waterproofing: the physics of insect cuticular hydrocarbons.

Understanding the evolution of complex traits is among the major challenges in biology. One such trait is the cuticular hydrocarbon (CHC) layer in insects. It protects against desiccation and provides communication signals, especially in social insects. CHC composition is highly diverse within and across species. To understand the adaptive value of this chemical diversity, we must understand how it affects biological functionality. So far, CHCs have received ample research attention, but their physical properties were little studied. We argue that these properties determine their biological functionality, and are vital to understanding how CHC composition affects their adaptive value. We investigated melting behaviour and viscosity of CHCs from 11 ant species using differential scanning calorimetry and a novel microrheological technique. CHCs began melting below -45°C, and often were entirely liquid only above 30°C. Thus, they formed a solid-liquid mixture under ambient conditions, which contrasts to previous assumptions of entirely solid layers in many species. This may be adaptive as only biphasic CHC layers ensure uniform coating of the insect body, which is necessary for waterproofing. CHC viscosity was mostly between 0.1 and 0.2 Pa s-1, thus similar to motor oils. Surprisingly, chemically different CHC profiles had similar viscosities, suggesting that a certain viscosity level is adaptive and ensures that communication signals can be perceived. With this study, we draw attention to the importance of studying the physics of CHC layers. Only by understanding how chemical and physical mechanisms enable CHC functionality can we understand the causes and consequences of CHC diversification.

Cuticular hydrocarbons as potential mediators of cryptic species divergence in a mutualistic ant association.

Upon advances in sequencing techniques, more and more morphologically identical organisms are identified as cryptic species. Often, mutualistic interactions are proposed as drivers of diversification. Species of the neotropical parabiotic ant association between Crematogaster levior and Camponotus femoratus are known for highly diverse cuticular hydrocarbon (CHC) profiles, which in insects serve as desiccation barrier but also as communication cues. In the present study, we investigated the association of the ants' CHC profiles with genotypes and morphological traits, and discovered cryptic species pairs in both genera. To assess putative niche differentiation between the cryptic species, we conducted an environmental association study that included various climate variables, canopy cover, and mutualistic plant species. Although mostly sympatric, the two Camponotus species seem to prefer different climate niches. However in the two Crematogaster species, we could not detect any differences in niche preference. The strong differentiation in the CHC profiles may thus suggest a possible role during speciation itself either by inducing assortative mating or by reinforcing sexual selection after the speciation event. We did not detect any further niche differences in the environmental parameters tested. Thus, it remains open how the cryptic species avoid competitive exclusion, with scope for further investigations.

Niche differentiation in rainforest ant communities across three continents.

A central prediction of niche theory is that biotic communities are structured by niche differentiation arising from competition. To date, there have been numerous studies of niche differentiation in local ant communities, but little attention has been given to the macroecology of niche differentiation, including the extent to which particular biomes show distinctive patterns of niche structure across their global ranges. We investigated patterns of niche differentiation and competition in ant communities in tropical rainforests, using different baits reflecting the natural food spectrum. We examined the extent of temporal and dietary niche differentiation and spatial segregation of ant communities at five rainforest sites in the neotropics, paleotropics, and tropical Australia. Despite high niche overlap, we found significant dietary and temporal niche differentiation in every site. However, there was no spatial segregation among foraging ants at the community level, despite strong competition for preferred food resources. Although sucrose, melezitose, and dead insects attracted most ants, some species preferentially foraged on seeds, living insects, or bird feces. Moreover, most sites harbored more diurnal than nocturnal species. Overall niche differentiation was strongest in the least diverse site, possibly due to its lower number of rare species. Both temporal and dietary differentiation thus had strong effects on the ant assemblages, but their relative importance varied markedly among sites. Our analyses show that patterns of niche differentiation in ant communities are highly idiosyncratic even within a biome, such that a mechanistic understanding of the drivers of niche structure in ant communities remains elusive.

Influence of Mutualistic Lifestyle, Mutualistic Partner, and Climate on Cuticular Hydrocarbon Profiles in Parabiotic Ants.

A vital trait in insects is their cuticular hydrocarbon (CHC) profile, which protects the insect against desiccation and serves in chemical communication. Due to these functions, CHC profiles are shaped by both climatic conditions and biotic interactions. Here, we investigated CHC differentiation in the neotropical parabiotic ant species Crematogaster levior and Camponotus femoratus, which mutualistically share a nest. Both consist of two cryptic species each (Cr. levior A and B and Ca. femoratus PAT and PS) that differ genetically and possess strongly different CHC profiles. We characterized and compared CHC profiles of the four cryptic species in detail. Our results suggest that Cr. levior A, Ca. femoratus PAT and Ca. femoratus PS adapted their CHC profiles to the parabiotic lifestyle by producing longer-chain CHCs. At the same time, they changed their major CHC classes, and produce more alkadienes and methyl-branched alkenes compared to Cr. levior B or non-parabiotic species. The CHC profiles of Cr. levior B were more similar to related, non-parabiotic species of the Orthocrema clade than Cr. levior A, and the chain lengths of B were similar to the reconstructed ancestral state. Signals of both the parabiotic partner (biotic conditions) and climate (abiotic conditions) were found in the CHC profiles of all four cryptic species. Our data suggest that mutualisms shaped the CHC profiles of the studied species, in particular chain length and CHC class composition. Beside this, signals of the parabiotic partners indicate potential impacts of biotic interactions, via chemical mimicry or chemical camouflage.

Hybrid Genome Assembly of a Neotropical Mutualistic Ant.

The success of social insects is largely intertwined with their highly advanced chemical communication system that facilitates recognition and discrimination of species and nest-mates, recruitment, and division of labor. Hydrocarbons, which cover the cuticle of insects, not only serve as waterproofing agents but also constitute a major component of this communication system. Two cryptic Crematogaster species, which share their nest with Camponotus ants, show striking diversity in their cuticular hydrocarbon (CHC) profile. This mutualistic system therefore offers a great opportunity to study the genetic basis of CHC divergence between sister species. As a basis for further genome-wide studies high-quality genomes are needed. Here, we present the annotated draft genome for Crematogaster levior A. By combining the three most commonly used sequencing techniques-Illumina, PacBio, and Oxford Nanopore-we constructed a high-quality de novo ant genome. We show that even low coverage of long reads can add significantly to overall genome contiguity. Annotation of desaturase and elongase genes, which play a role in CHC biosynthesis revealed one of the largest repertoires in ants and a higher number of desaturases in general than in other Hymenoptera. This may provide a mechanistic explanation for the high diversity observed in C. levior CHC profiles.

Ant behaviour and brain gene expression of defending hosts depend on the ecological success of the intruding social parasite.

The geographical mosaic theory of coevolution predicts that species interactions vary between locales. Depending on who leads the coevolutionary arms race, the effectivity of parasite attack or host defence strategies will explain parasite prevalence. Here, we compare behaviour and brain transcriptomes of Temnothorax longispinosus ant workers when defending their nest against an invading social parasite, the slavemaking ant Temnothorax americanus. A full-factorial design allowed us to test whether behaviour and gene expression are linked to parasite pressure on host populations or to the ecological success of parasite populations. Albeit host defences had been shown before to covary with local parasite pressure, we found parasite success to be much more important. Our chemical and behavioural analyses revealed that parasites from high prevalence sites carry lower concentrations of recognition cues and are less often attacked by hosts. This link was further supported by gene expression analysis. Our study reveals that host-parasite interactions are strongly influenced by social parasite strategies, so that variation in parasite prevalence is determined by parasite traits rather than the efficacy of host defence. Gene functions associated with parasite success indicated strong neuronal responses in hosts, including long-term changes in gene regulation, indicating an enduring impact of parasites on host behaviour. This article is part of the theme issue 'The coevolutionary biology of brood parasitism: from mechanism to pattern'.

Unveiling community patterns and trophic niches of tropical and temperate ants using an integrative framework of field data, stable isotopes and fatty acids.

BACKGROUND: The use and partitioning of trophic resources is a central aspect of community function. On the ground of tropical forests, dozens of ant species may be found together and ecological mechanisms should act to allow such coexistence. One hypothesis states that niche specialization is higher in the tropics, compared to temperate regions. However, trophic niches of most species are virtually unknown. Several techniques might be combined to study trophic niche, such as field observations, fatty acid analysis (FAA) and stable isotope analysis (SIA). In this work, we combine these three techniques to unveil partitioning of trophic resources in a tropical and a temperate community. We describe patterns of resource use, compare them between communities, and test correlation and complementarity of methods to unveil both community patterns and species' niches. METHODS: Resource use was assessed with seven kinds of bait representing natural resources available to ants. Neutral lipid fatty acid (NLFA) profiles, and δ15N and δ13C isotope signatures of the species were also obtained. Community patterns and comparisons were analyzed with clustering, correlations, multivariate analyses and interaction networks. RESULTS: Resource use structure was similar in both communities. Niche breadths (H') and network metrics (Q and H2') indicated similar levels of generalization between communities. A few species presented more specialized niches, such as Wasmannia auropunctata and Lasius fuliginosus. Stable isotope signatures and NLFA profiles also indicated high generalization, although the latter differed between communities, with temperate species having higher amounts of fat and proportions of C18:1n9. Bait use and NLFA profile similarities were correlated, as well as species' specialization indices (d') for the two methods. Similarities in δ15N and bait use, and in δ13C and NLFA profiles, were also correlated. DISCUSSION: Our results agree with the recent view that specialization levels do not change with latitude or species richness. Partition of trophic resources alone does not explain species coexistence in these communities, and might act together with behavioral and environmental mechanisms. Temperate species presented NLFA patterns distinct from tropical ones, which may be related to environmental factors. All methods corresponded in their characterization of species' niches to some extent, and were robust enough to detect differences even in highly generalized communities. However, their combination provides a more comprehensive picture of resource use, and it is particularly important to understand individual niches of species. FAA was applied here for the first time in ant ecology, and proved to be a valuable tool due to its combination of specificity and temporal representativeness. We propose that a framework combining field observations with chemical analysis is valuable to understand resource use in ant communities.

Coping with the climate: cuticular hydrocarbon acclimation of ants under constant and fluctuating conditions.

Terrestrial arthropods achieve waterproofing by a layer of cuticular hydrocarbons (CHCs). At the same time, CHCs also serve as communication signals. To maintain waterproofing under different climate conditions, insects adjust the chemical composition of their CHC layer, but this may affect the communication via CHCs. The detailed acclimatory changes of CHCs and how these influence their physical properties are still unknown. Here, we studied acclimation in two closely related ant species with distinct CHC profiles, Myrmica rubra and Myrmica ruginodis, in response to constant or fluctuating temperature and humidity regimes. We measured how acclimation affected CHC composition and viscosity, and the ants' drought survival. In both species, CHC composition showed strong, predictable responses to temperature regimes. Warm-acclimated individuals had higher proportions of linear alkanes, and less methyl-branched or unsaturated CHCs. These changes coincided with higher solid content and viscosity of CHCs in warm-acclimated ants. Temperature fluctuation caused effects similar to those observed under constant-cool conditions in Mrubra, but led to entirely different profiles in Mruginodis, suggesting that fluctuating and constant conditions pose very different challenges. Acclimation to dry conditions led to higher absolute amounts of CHCs, which increased the ants' drought survival, whereas temperature acclimation did not. Hence, the temperature-induced CHC changes cannot be explained by the need for waterproofing alone. Although these changes could be non-adaptive, we propose that they serve to maintain a constant CHC viscosity, which may be essential for communication and other functions.