Rustling ants: Vibrational communication performed by two Camponotus species in Borneo.

Interactions between ants and plants are classic examples of cooperation between individuals of different species. Usually, plants provide shelter or food for ants and in turn are defended against herbivores by their insect allies. To coordinate attacks, ants use multi-modal alarm signals consisting of vibrational and chemical components. This can also be observed in Borneo, where two Camponotus species inhabit the ocreas (diverging, tubular leaf sheaths) of the rattan palm Korthalsia robusta. When ants are disturbed, they beat or scratch mandibles and abdomens on the plant surface resulting in loud rustling sounds. To describe the characteristics of these signals, we recorded them with a Laser-Doppler-vibrometer in the field. Analyses of temporal patterns and dominant frequency revealed that the signals of the two species differ fundamentally. To assess transmission characteristics of the rattan palm, we conducted experiments under controlled lab-conditions. We show that the ocrea is an adequate structure for converting airborne sound into substrate vibrations, acting as a mediator between these two modalities. We hypothesize that the ants' vibratory signal has multiple functions, with the substrate-borne component used as an alarm signal for conspecifics, and the airborne component acting as vibro-acoustic aposematism against predators or herbivores to protect the host plant.

Limited dispersal and local adaptation promote allopatric speciation in a biodiversity hotspot.

Recently diverged or diverging populations can offer unobstructed insights into early barriers to gene flow during the initial stages of speciation. The current study utilised a novel insect system (order Mantophasmatodea) to shed light on the early drivers of speciation. The members of this group have limited dispersal abilities, small allopatric distributions and strong habitat associations in the Cape Floristic Region biodiversity hotspot in South Africa. Sister taxa from the diverse family Austrophasmatidae were chosen as focal species (Karoophasma biedouwense, K. botterkloofense). Population genetics and Generalized Dissimilarity Modelling (GDM) were used to characterise spatial patterns of genetic variation and evaluate the contribution of environmental factors to population divergence and speciation. Extensive sampling confirmed the suspected allopatry of these taxa. However, hybrids were identified in a narrow region occurring between the species' distributions. Strong population structure was found over short geographic distances; particularly in K. biedouwense in which geographic distance accounted for 32% of genetic variation over a scale of 50 km (r = .56, p < .001). GDM explained 42%-78% of the deviance in observed genetic dissimilarities. Geographic distance was consistently indicated to be important for between species and within population differentiation, suggesting that limited dispersal ability may be an important neutral driver of divergence. Temperature, altitude, precipitation and vegetation were also indicated as important factors, suggesting the possible role of adaptation to local environmental conditions for species divergence. The discovery of the hybrid-zone, and the multiple allopatric species pairs in Austrophasmatidae support the idea that this could be a promising group to further our understanding of speciation modes.

Asymmetry of the male internal reproductive organs in Mantophasmatodea.

BACKGROUND: Asymmetries are a widespread phenomenon in otherwise bilaterally symmetric organisms, and investigation of asymmetric structures can help us gather insights into fundamental evolutionary processes such as the selection for morphological novelties caused by behavioural changes. In insects, asymmetric genitalia have evolved in almost every order, and usually it's the sclerotized parts and most conspicuous male phallic organs that are known to exhibit asymmetries. While external copulatory organs in insects have often been subject to investigations concerning asymmetries and the evolution thereof, internal reproductive structures have received far less attention. Here we describe the internal and external male genitalia in three species of Austrophasmatidae, Mantophasmatodea, using µ-CT imaging and light microscopy. Mantophasmatodea is the most recently discovered insect order, and with 21 species described to date, it is among the smallest insect orders currently known. RESULTS: We confirm that male heelwalkers exhibit asymmetries in the external genitalia and associated structures, represented by asymmetric phallic lobes and cerci. Moreover, we found an extreme asymmetry within the internal male genitalia: in all adult males investigated (N = 5), the seminal vesicle, a dilatation of the vas deferens, was only developed on the right side of the male while missing on the left side. CONCLUSION: The false-male-above mating position exhibited by Mantophasmatodea and especially the long copulation duration of ca. 3 days might select for this unusual absence asymmetry of the left seminal vesicle. If this holds true for all heelwalker species, this absence asymmetry constitutes another autapomorphy for Austrophasmatidae or even the insect order Mantophasmatodea.

Comparative transcriptomics of ice-crawlers demonstrates cold specialization constrains niche evolution in a relict lineage.

Key changes in ecological niche space are often critical to understanding how lineages diversify during adaptive radiations. However, the converse, or understanding why some lineages are depauperate and relictual, is more challenging, as many factors may constrain niche evolution. In the case of the insect order Grylloblattodea, highly conserved thermal breadth is assumed to be closely tied to their relictual status, but has not been formerly tested. Here, we investigate whether evolutionary constraints in the physiological tolerance of temperature can help explain relictualism in this lineage. Using a comparative transcriptomics approach, we investigate gene expression following acute heat and cold stress across members of Grylloblattodea and their sister group, Mantophasmatodea. We additionally examine patterns of protein evolution, to identify candidate genes of positive selection. We demonstrate that cold specialization in Grylloblattodea has been accompanied by the loss of the inducible heat shock response under both acute heat and cold stress. Additionally, there is widespread evidence of selection on protein-coding genes consistent with evolutionary constraints due to cold specialization. This includes positive selection on genes involved in trehalose transport, metabolic function, mitochondrial function, oxygen reduction, oxidative stress, and protein synthesis. These patterns of molecular adaptation suggest that Grylloblattodea have undergone evolutionary trade-offs to survive in cold habitats and should be considered highly vulnerable to climate change. Finally, our transcriptomic data provide a robust backbone phylogeny for generic relationships within Grylloblattodea and Mantophasmatodea. Major phylogenetic splits in each group relate to arid conditions driving biogeographical patterns, with support for a sister-group relationship between North American Grylloblatta and Altai-Sayan Grylloblattella, and a range disjunction in Namibia splitting major clades within Mantophasmatodea.

Causes of variability in male vibratory signals and the role of female choice in Mantophasmatodea.

Communication systems that involve substrate vibrations are increasingly a focus of research since this communication mode - recently termed biotremology - has been found to be remarkably widespread in the animal kingdom. Vibrational signals are often used during courtship and therefore underlie both natural and sexual selection. Mantophasmatodea use species- and sex-specific substrate vibrational signals during courtship. We explored whether male vibrational signals of the South African heelwalker Karoophasma biedouwense vary with temperature, body condition and age, and tested female preference towards various signal pattern combinations. We recorded male signals under varying temperatures and over 3.5 weeks after onset of signaling. Our results show that the temporal structure of male signals is modified by changes in temperature, and changes with male age. Other characteristics, especially duty cycles, are less affected, but correlate with body condition. Females responded along a broad spectrum of signaling patterns, indicating that they do not favor signals of males of a certain age or condition. They were selective towards the fine structure of vibratory signals, suggesting that pulse repetition times carry species-specific information. Mantophasmatodea thus use vibrational signals to identify and localize a mating partner, but presumably not for precopulatory mate selection.

Variable Molecular Markers for the Order Mantophasmatodea (Insecta).

The recently discovered insect order Mantophasmatodea currently comprises 19 Southern African species. These mainly occur in allopatry, have high levels of color polymorphism and communicate via species- and gender-specific vibratory signals. High levels of interspecific morphological conservatism mean that cryptic species are likely to be uncovered. These aspects of Mantophasmatodean biology make them an ideal group in which to investigate population divergence due to habitat-specific adaptation, sexual selection, and potentially sensory speciation. Lack of appropriate genetic markers has thus far rendered such studies unfeasible. To address this need, the first microsatellite loci for this order were developed. Fifty polymorphic loci were designed specifically for Karoophasma biedouwense (Austrophasmatidae), out of which 23 were labeled and tested for amplification across the order using 2-3 individuals from 10 species, representing all 4 currently known families. A Bayesian mitochondrially encoded cytochrome c oxidase I (COI) topology was reconstructed and divergence dates within the order were estimated for the first time. Amplification success and levels of polymorphism were compared with genetic divergence and time since divergence. In agreement with studies on vertebrate taxa, both amplification and variability were negatively correlated with distance (temporal and genetic). The high number of informative loci will offer sufficient resolution for both broad level population genetic analysis and individual based pedigree or parentage analyses for most species in Austrophasmatidae, with at least some loci available for the other families. This resource will facilitate research into the evolutionary biology of this understudied but fascinating group.

A temperature rise reduces trial-to-trial variability of locust auditory neuron responses.

The neurophysiology of ectothermic animals, such as insects, is affected by environmental temperature, as their body temperature fluctuates with ambient conditions. Changes in temperature alter properties of neurons and, consequently, have an impact on the processing of information. Nevertheless, nervous system function is often maintained over a broad temperature range, exhibiting a surprising robustness to variations in temperature. A special problem arises for acoustically communicating insects, as in these animals mate recognition and mate localization typically rely on the decoding of fast amplitude modulations in calling and courtship songs. In the auditory periphery, however, temporal resolution is constrained by intrinsic neuronal noise. Such noise predominantly arises from the stochasticity of ion channel gating and potentially impairs the processing of sensory signals. On the basis of intracellular recordings of locust auditory neurons, we show that intrinsic neuronal variability on the level of spikes is reduced with increasing temperature. We use a detailed mathematical model including stochastic ion channel gating to shed light on the underlying biophysical mechanisms in auditory receptor neurons: because of a redistribution of channel-induced current noise toward higher frequencies and specifics of the temperature dependence of the membrane impedance, membrane potential noise is indeed reduced at higher temperatures. This finding holds under generic conditions and physiologically plausible assumptions on the temperature dependence of the channels' kinetics and peak conductances. We demonstrate that the identified mechanism also can explain the experimentally observed reduction of spike timing variability at higher temperatures.

Temperature effects on the tympanal membrane and auditory receptor neurons in the locust.

Poikilothermic animals are affected by variations in environmental temperature, as the basic properties of nerve cells and muscles are altered. Nevertheless, insect sensory systems, such as the auditory system, need to function effectively over a wide range of temperatures, as sudden changes of up to 10 °C or more are common. We investigated the performance of auditory receptor neurons and properties of the tympanal membrane of Locusta migratoria in response to temperature changes. Intracellular recordings of receptors at two temperatures (21 and 28 °C) revealed a moderate increase in spike rate with a mean Q10 of 1.4. With rising temperature, the spike rate-intensity-functions exhibited small decreases in thresholds and expansions of the dynamic range, while spike durations decreased. Tympanal membrane displacement, investigated using microscanning laser vibrometry, exhibited a small temperature effect, with a Q10 of 1.2. These findings suggest that locusts are affected by shifts in temperature at the periphery of the auditory pathway, but the effects on spike rate, sensitivity, and tympanal membrane displacement are small. Robust encoding of acoustic signals by only slightly temperature-dependent receptor neurons and almost temperature-independent tympanal membrane properties might enable locusts and grasshoppers to reliably identify sounds in spite of changes of their body temperature.

Structure and function of the arolium of Mantophasmatodea (Insecta).

All species of the insect order Mantophasmatodea characteristically keep the 5th tarsomere and pretarsus (arolium plus two claws) turned upwards and off the substrate. The unusually large arolium was studied in two species of Mantophasmatodea using bright field light microscopy, reflection microscopy, fluorescence microscopy, TEM, SEM, and Cryo-SEM. It contains an epithelial gland, numerous tracheoles, and nerves. The gland consists of enlarged epithelial cells with large nuclei, mitochondria, RER, golgi complexes, microtubules, and numerous secretion vesicles. Evidence for exocytosis of the vesicles into the gland reservoir between the epithelial gland and the thick cuticle could be observed. Cryo-SEM revealed that the ventral side of the arolium and distal part of its dorsal side are covered with a liquid film. Fluid footprints of arolia of individuals walking on a glass plate also indicate the presence of secretory fluid on the arolium surface. Behavioral experiments using animals with ablated arolia showed that representatives of Mantophasmatodea do not need their arolia to detect and respond to vibratory communication signals nor to catch small to medium-sized prey. Individuals with ablated arolia were not able to move upside down on a smooth glass plate. We conclude that Mantophasmatodea use their arolia for attachment when additional adhesion force is required (e.g. windy conditions, handling large prey, mating). They can bring their arolia in contact with the surface in a very fast reflex (18.0 +/- 9.9 ms). The secretory fluid found on the surface is produced by the gland and transported to the outside, presumably through small pore channels, to enhance adhesion to the substrate.

Mechanical damage to pollen aids nutrient acquisition in Heliconius butterflies (Nymphalidae).

Neotropical Heliconius and Laparus butterflies actively collect pollen onto the proboscis and extract nutrients from it. This study investigates the impact of the processing behaviour on the condition of the pollen grains. Pollen samples (n = 72) were collected from proboscides of various Heliconius species and Laparus doris in surrounding habitats of the Tropical Research Station La Gamba (Costa Rica). Examination using a light microscope revealed that pollen loads contained 74.88 ± 53.67% of damaged Psychotria pollen, 72.04 ± 23.4% of damaged Psiguria/Gurania pollen, and 21.35 ± 14.5% of damaged Lantana pollen (numbers represent median ± first quartile). Damaged pollen grains showed deformed contours, inhomogeneous and/or leaking contents, or they were empty. Experiments with Heliconius and Laparus doris from a natural population in Costa Rica demonstrated that 200 min of pollen processing behaviour significantly increased the percentage of damaged pollen of Psychotria compared to pollen from anthers (P = 0.015, Z = -2.44, Mann-Whitney U-test). Examination of pollen loads from green house reared Heliconius butterflies resulted in significantly greater amounts of damaged Psiguria pollen after 200 min of processing behaviour compared to pollen from flowers (P < 0.001, Z = -4.583, Mann-Whitney U-test). These results indicate that pollen processing functions as extra oral digestion whereby pollen grains are ruptured to make the content available for ingestion.