Stick Insect Research Articles

Genetic variation within a stick insect species associated with community-level traits.

Phenotypic variation within species can affect the ecological dynamics of populations and communities. Characterizing the genetic variation underlying such effects can help parse the roles of genetic evolution and plasticity in 'eco-evolutionary dynamics' and inform how genetic variation may shape patterns of evolution. Here we employ genome-wide association (GWA) methods in Timema cristinae stick insects and their co-occurring arthropod communities to identify genetic variation associated with community-level traits. Previous studies have shown that maladaptation (i.e., imperfect crypsis) of T. cristinae can reduce the abundance and species richness of other arthropods due to an increase in bird predation. Whether genetic variation that is independent from crypsis has similar effects is unknown and was tested here using genome-wide genotyping-by-sequencing data of stick insects, arthropod community information, and GWA mapping with Bayesian sparse linear mixed models. We find associations between genetic variation in stick insects and arthropod community traits. However, these associations disappeared when host-plant traits are accounted for. We thus use path analysis to disentangle interrelationships among stick-insect genetic variation, host-plant traits and community traits. This revealed that host-plant size has large effects on arthropod communities, while genetic variation in stick insects has a smaller, but still significant effect. Our findings demonstrate that: (1) genetic variation in a species can be associated with community-level traits, but that (2) interrelationships among multiple factors may need to be analyzed to disentangle whether such associations represent causal relationships. This work helps to build a framework for genomic studies of eco-evolutionary dynamics.

Evolution of alternative reproductive systems in Bacillus stick insects.

Reproduction is a key feature of all organisms, yet the way in which it is achieved varies greatly across the tree of life. One striking example of this variation is the stick insect genus Bacillus, in which five different reproductive modes have been described: sex, facultative and obligate parthenogenesis, and two highly unusual reproductive modes: hybridogenesis and androgenesis. Under hybridogenesis, the entire genome from the paternal species is eliminated and replaced each generation by mating with the corresponding species. Under androgenesis, an egg is fertilized, but the developing diploid offspring bear two paternal genomes and no maternal genome, as a consequence of unknown mechanisms. Here, we reevaluate the previous descriptions of Bacillus lineages and the proposed F1 hybrid ancestries of the hybridogenetic and obligately parthenogenetic lineages (based on allozymes and karyotypes) from Sicily, where all these reproductive modes are found. We generate a chromosome-level genome assembly for a facultative parthenogenetic species (B. rossius) and combine extensive field sampling with RADseq and mtDNA data. We identify and genetically corroborate all previously described species and confirm the ancestry of hybrid lineages. All hybrid lineages have fully retained their F1 hybrid constitution throughout the genome, indicating that the elimination of the paternal genome in hybridogens is always complete and that obligate parthenogenesis in Bacillus hybrid species is not associated with an erosion of heterozygosity as known in other hybrid asexuals. Our results provide a stepping stone toward understanding the transitions between reproductive modes and the proximate mechanisms of genome elimination.

Divergent dynamics of sexual and habitat isolation at the transition between stick insect populations and species

Speciation is often viewed as a continuum along which populations diverge until they become reproductively-isolated species. However, such divergence may be heterogeneous, proceeding in fits and bursts, rather than being uniform and gradual. We show in Timema stick insects that one component of reproductive isolation evolves non-uniformly across this continuum, whereas another does not. Specifically, we use thousands of host-preference and mating trials to study habitat and sexual isolation among 42 pairs of taxa spanning a range of genomic differentiation and divergence time. We find that habitat isolation is uncoupled from genomic differentiation within species, but accumulates linearly with it between species. In contrast, sexual isolation accumulates linearly across the speciation continuum, and thus exhibits similar dynamics to morphological traits not implicated in reproductive isolation. The results show different evolutionary dynamics for different components of reproductive isolation and highlight a special relevance for species status in the process of speciation.

The only species of Calvisia (Phasmatodea, Necrosciinae) from China, with a new synonymy and additional descriptions.

Calvisia is a colorful winged stick insect genus consisting of 6 subgenera and 44 species widely distributed in temperate and tropical Asia. C. medogensis syn. nov. was discovered in Mdog, Xizang (Tibet), China and is so far the only species recorded from China. We here propose that C. medogensis syn. nov. is a synonym of C. fuscoalata after checking type specimens of both species. New materials studied are deposited in Yunnan Agricultural University, China (YNAU).

The synaptic drive of central pattern-generating networks to leg motor neurons of a walking insect is motor neuron pool specific.

Rhythmic locomotor activity, such as flying, swimming, or walking, results from an interplay between higher-order centers in the central nervous system, which initiate, maintain, and modify task-specific motor activity, downstream central pattern-generating neural circuits (CPGs) that can generate a default rhythmic motor output, and, finally, feedback from sense organs that modify basic motor activity toward functionality.1,2,3 In this context, CPGs provide phasic synaptic drive to motor neurons (MNs) and thereby support the generation of rhythmic activity for locomotion. We analyzed the synaptic drive that the leg MNs supplying the three main leg joints receive from CPGs in pharmacologically activated and deafferented preparations of the stick insect (Carausius morosus). We show that premotor CPGs pattern the tonic activity of five of the six leg MN pools by phasic inhibitory synaptic drive. These are the antagonistic MN pools supplying the thoraco-coxal joint and the femur-tibial joint4,5 and the levator MN pool supplying the coxa-trochanteral (CTr) joint. In contrast, rhythmic activity of the depressor MN pool supplying the CTr joint was found to be primarily based on a phasic excitatory drive. This difference is likely related to the pivotal role of the depressor muscle in generating leg stance during any walking situation. Thus, our results provide evidence for qualitatively differing mechanisms to generate rhythmic activity between MN pools in the same locomotor system.

Mindanao broken stick insect: a new species and first country record of the genus Baculofractum Zompro, 1995 (Phasmatodea: Lonchodidae: Lonchodinae: Lonchodini) from the Philippines

Baculofractum Zompro 1995 is a genus of stick insects in the family Lonchodidae, subfamily Lonchodinae represented by three species: B. insigne, B. shelfordi, and B. williemsei, distributed in Indonesia and Malaysia. This paper reports the discovery of a new species of Baculofractum from Mt. Malimumu, Pantaron Range, Bukidnon which also represents the first record of the genus in the Philippines. The new species can be distinguished by the absence of four tubercles at the back of the head’s median ridges and its distinctly elongated operculum exceeding the end of the abdomen.

A new species of Jeremia Redtenbacher (Phasmatodea: Phasmatidae: Cladomorphinae), a stick insect from the northern Brazilian Amazon Basin.

A new species of Jeremia Redtenbacher (Phasmatodea: Phasmatidae: Cladomorphinae) is described based on both male and female specimens from the northern Brazilian Amazon Basin and named Jeremia megaplax sp. nov. The new species can be differentiated from other species of this genus by the length of the female subgenital plate, which is two times longer than the length of tergites 810. Photos and description from subadult male and not fully developed eggs are also provided. The studied material is deposited at the Invertebrate Collection of the Instituto Nacional de Pesquisas da Amaznia (INPA).

Mechanosensory encoding of forces in walking uphill and downhill: force feedback can stabilize leg movements in stick insects.

Force feedback could be valuable in adapting walking to diverse terrains, but the effects of changes in substrate inclination on discharges of sensory receptors that encode forces have rarely been examined. In insects, force feedback is provided by campaniform sensilla, mechanoreceptors that monitor forces as cuticular strains. We neurographically recorded responses of stick insect tibial campaniform sensilla to "naturalistic" forces (joint torques) that occur at the hind leg femur-tibia (FT) joint in uphill, downhill, and level walking. The FT joint torques, obtained in a previous study that used inverse dynamics to analyze data from freely moving stick insects, are quite variable during level walking (including changes in sign) but are larger in magnitude and more consistent when traversing sloped surfaces. Similar to vertebrates, insects used predominantly extension torque in propulsion on uphill slopes and flexion torques to brake forward motion when going downhill. Sensory discharges to joint torques reflected the torque direction but, unexpectedly, often occurred as multiple bursts that encoded the rate of change of positive forces (dF/dt) even when force levels were high. All discharges also showed hysteresis (history dependence), as firing substantially decreased or ceased during transient force decrements. These findings have been tested in simulation in a mathematical model of the sensilla (Szczecinski NS, Dallmann CJ, Quinn RD, Zill SN. Bioinspir Biomim 16: 065001, 2021) that accurately reproduced the biological data. Our results suggest the hypothesis that sensory feedback from the femoro-tibial joint indicating force dynamics (dF/dt) can be used to counter the instability in traversing sloped surfaces in animals and, potentially, in walking machines.NEW & NOTEWORTHY Discharges of sensory receptors (campaniform sensilla) in the hind legs of stick insects can differentially signal forces that occur in walking uphill versus walking downhill. Unexpectedly, sensory firing most closely reflects the rate of change of force (dF/dt) even when the force levels are high. These signals have been replicated in a mathematical model of the receptors and could be used to stabilize leg movements both in the animal and in a walking robot.

Cryptic Sex: Female stick insects reproduce without males-but have a secret.

Cryptic Sex: Female stick insects reproduce without males-but have a secret.

Redescription of Hybusa occidentalis (Westwood) (Orthoptera: Proscopiidae: Hybusinae): new images, description of male genitalia, pictorial key of Hybusa species and morphological comparison of Hybusinae with Xeniinae and Proscopiinae.

Proscopiidae (Orthoptera: Caelifera) are grasshoppers found in Central and South America, often mistaken for true stick insects (Phasmatodea) due to their stick-like appearance. The family comprises three subfamilies: Hybusinae, Proscopiinae, and Xeniinae, with Hybusinae as the smallest subfamily, housing the genus Hybusa Erichson, which contains four species endemic to Chile. Considering that the most recent comprehensive taxonomic assessment of this genus was conducted approximately three decades ago, herein we redescribe the type species of Hybusa, H. occidentalis (Westwood), based on males and females. The study includes new images of H. occidentalis body and male genitalia, along with a comprehensive pictorial key for all Hybusa species using available type material. Additionally, a morphological comparison between Hybusa and other Proscopiidae taxa is presented.

Unraveling the Sexual Dimorphism of First Instar Nymphs of the Giant Stick Insect, Cladomorphus phyllinus Gray, 1835, from the Atlantic Forest, Brazil.

The first instar nymphs, both male and female, of the giant stick insect Cladomorphus phyllinus Gray, 1835 were carefully described and measured, revealing a remarkable sexual dimorphism that is considered rare among insects and is poorly explored in the order Phasmida. The studied F1 nymphs originated in captivity from eggs laid by a coupled female specimen collected in the Atlantic Forest in the vicinity of Petrópolis city, state of Rio de Janeiro, Brazil. The first instar nymphs of C. phyllinus were measured and illustrated in high-resolution photographs to show the general aspects and details of sexually dimorphic traits, making clear the phenotypic differences in the sexes. A total of 100 nymphs were kept alive until morphological sexual dimorphism was confirmed and quantified. All recently hatched first instar nymphs were separated based on the presumed male and female characteristics, i.e., the presence and absence of the suture in the metanotum in the males and females, respectively, had their sexes confirmed in 100% of the specimens as previously assigned. These results confirm this new morphological trait, which here is named "alar suture" as sex-specific in the first instar nymphs, a novelty in this stage of development of sexual differentiation. In addition, the distinct conformations of the last three abdominal sternites of both sexes were recorded.

DNA methylation differences between stick insect ecotypes.

Epigenetic mechanisms, such as DNA methylation, can influence gene regulation and affect phenotypic variation, raising the possibility that they contribute to ecological adaptation. Beginning to address this issue requires high-resolution sequencing studies of natural populations to pinpoint epigenetic regions of potential ecological and evolutionary significance. However, such studies are still relatively uncommon, especially in insects, and are mainly restricted to a few model organisms. Here, we characterize patterns of DNA methylation for natural populations of Timema cristinae adapted to two host plant species (i.e. ecotypes). By integrating results from sequencing of whole transcriptomes, genomes and methylomes, we investigate whether environmental, host and genetic differences of these stick insects are associated with methylation levels of cytosine nucleotides in the CpG context. We report an overall genome-wide methylation level for T. cristinae of ~14%, with methylation being enriched in gene bodies and impoverished in repetitive elements. Genome-wide DNA methylation variation was strongly positively correlated with genetic distance (relatedness), but also exhibited significant host-plant effects. Using methylome-environment association analysis, we pinpointed specific genomic regions that are differentially methylated between ecotypes, with these regions being enriched for genes with functions in membrane processes. The observed association between methylation variation and genetic relatedness, and with the ecologically important variable of host plant, suggests a potential role for epigenetic modification in T. cristinae adaptation. To substantiate such adaptive significance, future studies could test whether methylation can be transmitted across generations and the extent to which it responds to experimental manipulation in field and laboratory studies.

Phylogeographical evidence for historical long-distance dispersal in the flightless stick insect Ramulus mikado.

Exploring how organisms overcome geographical barriers to dispersal is a fundamental question in biology. Passive long-distance dispersal events, although infrequent and unpredictable, have a considerable impact on species range expansions. Despite limited active dispersal capabilities, many stick insect species have vast geographical ranges, indicating that passive long-distance dispersal is vital for their distribution. A potential mode of passive dispersal in stick insects is via the egg stage within avian digestive tracts, as suggested by experimental evidence. However, detecting such events under natural conditions is challenging due to their rarity. Therefore, to indirectly assess the potential of historical avian-mediated dispersal, we examined the population genetic structure of the flightless stick insect Ramulus mikado across Japan, based on a multifaceted molecular approach [cytochrome oxidase subunit I (COI) haplotypes, nuclear simple sequence repeat markers and genome-wide single nucleotide polymorphisms]. Subsequently, we identified unique phylogeographic patterns, including the discovery of identical COI genotypes spanning considerable distances, which substantiates the notion of passive long-distance genotypic dispersal. Overall, all the molecular data revealed the low and mostly non-significant genetic differentiation among populations, with identical or very similar genotypes across distant populations. We propose that long-distance dispersal facilitated by birds is the plausible explanation for the unique phylogeographic pattern observed in this flightless stick insect.

Independent wing reductions and losses among stick and leaf insects (Phasmatodea), supported by new Cretaceous fossils in amber

BackgroundPhasmatodea (stick and leaf insects) play a central role on the debate regarding wing reduction and loss, and its wings are putative reacquisition from secondarily wingless ancestors based solely on extant species. A pivotal taxon in this respect is the species-poor Timematodea, consisting of approximately 21 wingless extant species, which form the sister group of all remaining winged or wingless stick and leaf insects, the Euphasmatodea.ResultsHerein, the new fossils of Timematodea from mid-Cretaceous Kachin amber are reported, with winged and wingless species co-occurring. The palaeogeographic distributions of all fossils of Holophasmatodea are summarized, showing their wide paleo-distributions. The phylogenetic analysis based on morphological characters confirms the earliest-diverging lineage of winged Breviala cretacea gen. et sp. nov. in Timematodea, and the possible relationships among all families of Holophasmatodea. These are critical for the reconstruction of patterns of wing evolution in early Phasmatodea.ConclusionsThe new fossils suggest that Timematodea once had wings, at least during the mid-Cretaceous. The palaeogeographic occurrences imply that Timematodea probably have been widely distributed since at least the Jurassic. The phylogenetic analysis with the ancestral-state reconstruction of wings indicates that the common ancestors of Holophasmatodea were winged, the reductions and losses of wings among Timematodea and Euphasmatodea have occurred independently since at least the Cretaceous, and the reduction or loss of the forewing earlier than the hind wings.

A new record of a winged stick insect (Phasmatodea) from Mexico, with a checklist and key to the species of the family Pseudophasmatidae from continental North America

The winged stick insect Metriophasma iphicles (Redtenbacher, 1906) (Phasmatodea: Pseudophasmatidae) is recorded for the first time from Mexico (state of Veracruz), making this the northernmost record of both the species and genus. A checklist of species in the family Pseudophasmatidae from Mexico and the USA is presented, and a key to the species listed is proposed. With the current record, the number of continental North American species of Phasmatodea increases to 108, and the number of genera in the region increases to 23.

Evidence for cryptic sex in parthenogenetic stick insects of the genus Timema.

Obligately parthenogenetic species are expected to be short lived since the lack of sex and recombination should translate into a slower adaptation rate and increased accumulation of deleterious alleles. Some, however, are thought to have been reproducing without males for millions of years. It is not clear how these old parthenogens can escape the predicted long-term costs of parthenogenesis, but an obvious explanation is cryptic sex. In this study, we screen for signatures of cryptic sex in eight populations of four parthenogenetic species of Timema stick insects, some estimated to be older than 1 Myr. Low genotype diversity, homozygosity of individuals and high linkage disequilibrium (LD) unaffected by marker distances support exclusively parthenogenetic reproduction in six populations. However, in two populations (namely, of the species Timema douglasi and T. monikensis) we find strong evidence for cryptic sex, most likely mediated by rare males. These populations had comparatively high genotype diversities, lower LD, and a clear LD decay with genetic distance. Rare sex in species that are otherwise largely parthenogenetic could help explain the unusual success of parthenogenesis in the Timema genus and raises the question whether episodes of rare sex are in fact the simplest explanation for the persistence of many old parthenogens in nature.

Characterization of Morphologically Distinct Components in the Tarsal Secretion of Medauroidea extradentata (Phasmatodea) Using Cryo-Scanning Electron Microscopy.

Attachment to the substrate is an important phenomenon that determines the survival of many organisms. Most insects utilize wet adhesion to support attachment, which is characterized by fluids that are secreted into the interface between the tarsus and the substrates. Previous research has investigated the composition and function of tarsal secretions of different insect groups, showing that the secretions are likely viscous emulsions that contribute to attachment by generating capillary and viscous adhesion, leveling surface roughness and providing self-cleaning of the adhesive systems. Details of the structural organization of these secretions are, however, largely unknown. Here, we analyzed footprints originating from the arolium and euplantulae of the stick insect Medauroidea extradentata using cryo-scanning electron microscopy (cryo-SEM) and white light interferometry (WLI). The secretion was investigated with cryo-SEM, revealing four morphologically distinguishable components. The 3D WLI measurements of the droplet shapes and volumes over time revealed distinctly different evaporation rates for different types of droplets. Our results indicate that the subfunctionalization of the tarsal secretion is facilitated by morphologically distinct components, which are likely a result of different proportions of components within the emulsion. Understanding these components and their functions may aid in gaining insights for developing adaptive and multifunctional biomimetic adhesive systems.

A hierarchical model for external electrical control of an insect, accounting for inter-individual variation of muscle force properties.

Cyborg control of insect movement is promising for developing miniature, high-mobility, and efficient biohybrid robots. However, considering the inter-individual variation of the insect neuromuscular apparatus and its neural control is challenging. We propose a hierarchical model including inter-individual variation of muscle properties of three leg muscles involved in propulsion (retractor coxae), joint stiffness (pro- and retractor coxae), and stance-swing transition (protractor coxae and levator trochanteris) in the stick insect Carausius morosus. To estimate mechanical effects induced by external muscle stimulation, the model is based on the systematic evaluation of joint torques as functions of electrical stimulation parameters. A nearly linear relationship between the stimulus burst duration and generated torque was observed. This stimulus-torque characteristic holds for burst durations of up to 500ms, corresponding to the stance and swing phase durations of medium to fast walking stick insects. Hierarchical Bayesian modeling revealed that linearity of the stimulus-torque characteristic was invariant, with individually varying slopes. Individual prediction of joint torques provides significant benefits for precise cyborg control.

Descending interneurons of the stick insect connecting brain neuropiles with the prothoracic ganglion.

Stick insects respond to visual or tactile stimuli with whole-body turning or directed reach-to-grasp movements. Such sensory-induced turning and reaching behaviour requires interneurons to convey information from sensory neuropils of the head ganglia to motor neuropils of the thoracic ganglia. To date, descending interneurons are largely unknown in stick insects. In particular, it is unclear whether the special role of the front legs in sensory-induced turning and reaching has a neuroanatomical correlate in terms of descending interneuron numbers. Here, we describe the population of descending interneurons with somata in the brain or gnathal ganglion in the stick insect Carausius morosus, providing a first map of soma cluster counts and locations. By comparison of interneuron populations with projections to the pro- and mesothoracic ganglia, we then estimate the fraction of descending interneurons that terminate in the prothoracic ganglion. With regard to short-latency, touch-mediated reach-to-grasp movements, we also locate likely sites of synaptic interactions between antennal proprioceptive afferents to the deutocerebrum and gnathal ganglion with descending or ascending interneuron fibres. To this end, we combine fluorescent dye stainings of thoracic connectives with stainings of antennal hair field sensilla. Backfills of neck connectives revealed up to 410 descending interneuron somata (brain: 205 in 19 clusters; gnathal ganglion: 205). In comparison, backfills of the prothorax-mesothorax connectives stained only up to 173 somata (brain: 83 in 16 clusters; gnathal ganglion: 90), suggesting that up to 60% of all descending interneurons may terminate in the prothoracic ganglion (estimated upper bound). Double stainings of connectives and antennal hair field sensilla revealed that ascending or descending fibres arborise in close proximity of afferent terminals in the deutocerebrum and in the middle part of the gnathal ganglia. We conclude that two cephalothoracic pathways may convey cues about antennal movement and pointing direction to thoracic motor centres via two synapses only.

Evidence for direct oviposition into substrates by the New Zealand stick insect Spinotectarchus acornutus

ABSTRACT Stick insects (Phasmatodea) have many different oviposition strategies, reflecting a range of adaptive behaviours and morphologies to best place and secure eggs in their environments. Oviposition strategies in Aotearoa New Zealand phasmids are not well documented, but the literature so far suggests that they drop individual eggs to the ground from their position in the foliage. Here, we present evidence for an oviposition strategy unique among the Rō stick insects of Aotearoa New Zealand. Individual female Spinotectarchus acornutus were observed inserting their eggs in a range of substrates, in particular the bark of trees and in spaces within the textured surfaces of tree fern trunks. We also highlight how the specific morphology of their eggs may be an adaptation to assist in substrate attachment, while their elongated secondary ovipositor could aid in egg insertion into substrates.