Background Repetitive DNA sequences, including transposable elements (TEs) and tandemly repeated satellite DNA (satDNAs), collectively called the “repeatome”, are found in high proportion in organisms across the Tree of Life. Grasshoppers have large genomes, averaging 9 Gb, that contain a high proportion of repetitive DNA, which has hampered progress in assembling reference genomes. Here we combined linked-read genomics with transcriptomics to assemble, characterize, and compare the structure of repetitive DNA sequences in four chromosomal races of the morabine grasshopper Vandiemenella viatica species complex and determine their contribution to genome evolution. Results We obtained linked-read genome assemblies of 2.73–3.27 Gb from estimated genome sizes of 4.26–5.07 Gb DNA per haploid genome of the four chromosomal races of V. viatica. These constitute the third largest insect genomes assembled so far. Combining complementary annotation tools and manual curation, we found a large diversity of TEs and satDNAs, constituting 66 to 75% per genome assembly. A comparison of sequence divergence within the TE classes revealed massive accumulation of recent TEs in all four races (314–463 Mb per assembly), indicating that their large genome sizes are likely due to similar rates of TE accumulation. Transcriptome sequencing showed more biased TE expression in reproductive tissues than somatic tissues, implying permissive transcription in gametogenesis. Out of 129 satDNA families, 102 satDNA families were shared among the four chromosomal races, which likely represent a diversity of satDNA families in the ancestor of the V. viatica chromosomal races. Notably, 50 of these shared satDNA families underwent differential proliferation since the recent diversification of the V. viatica species complex. Conclusion This in-depth annotation of the repeatome in morabine grasshoppers provided new insights into the genome evolution of Orthoptera. Our TEs analysis revealed a massive recent accumulation of TEs equivalent to the size of entire Drosophila genomes, which likely explains the large genome sizes in grasshoppers. Despite an overall high similarity of the TE and satDNA diversity between races, the patterns of TE expression and satDNA proliferation suggest rapid evolution of grasshopper genomes on recent timescales.

No abstract

Insects have a gas-filled respiratory system, which provides a challenge for those that have become aquatic secondarily. Diving beetles (Dytiscidae) use bubbles on the surface of their bodies to supply O 2 for their dives and passively gain O 2 from the water. However, these bubbles usually require replenishment at the water's surface. A highly diverse assemblage of subterranean dytiscids has evolved in isolated calcrete aquifers of Western Australia with limited/ no access to an air-water interface, raising the question of how they are able to respire. We explored the hypothesis that they use cutaneous respiration by studying the mode of respiration in three subterranean dytiscid species from two isolated aquifers. The three beetle species consume O 2 directly from the water, but they lack structures on their bodies that could have respiratory function. They also have a lower metabolic rate than other insects. O 2 boundary layers surrounding the beetles are present, indicating that O 2 diffuses into the surface of their bodies via cutaneous respiration. Cuticle thickness measurements and other experimental results were incorporated into a mathematical model to understand whether cutaneous respiration limits beetle size. The model indicates that the cuticle contributes considerably to resistance in the O 2 cascade. As the beetles become larger, their metabolic scope narrows, potentially limiting their ability to allocate energy to mating, foraging and development at sizes above approximately 5 mg. However, the ability of these beetles to utilise cutaneous respiration has enabled the evolution of the largest assemblage of subterranean dytiscids in the world.

The river bug Aphelocheirus aestivalis is a 40 mg aquatic insect that, as an adult, relies totally on an incompressible physical gill to exchange respiratory gases with the water. The gill (called a 'plastron') consists of a stationary layer of air held in place on the body surface by millions of tiny hairs that support a permanent air-water interface, so that the insect never has to renew the gas at the water's surface. The volume of air in the plastron is extremely small (0.14 mm 3 ), under slightly negative pressure and connected to the gas-filled tracheal system through spiracles on the cuticle. Here, we measure P O2 of the water and within the plastron gas with O 2 -sensing fibre optics to understand the effectiveness and limitations of the gas exchanger. The difference in P O2 is highest in stagnant water and decreases with increasing convection over the surface. Respiration of bugs in water-filled vials varies between 33 and 296 pmol O 2 s −1 , depending on swimming activity. The effective thickness of the boundary layer around the plastron was calculated from respiration rate, P O2 difference and plastron surface area, according to the Fick diffusion equation and verified by direct measurements with the fibreoptic probes. In stagnant water, the boundary layer is approximately 500 μm thick, which nevertheless can satisfy the demands of resting bugs, even if the P O2 of the free water decreases to half that of air saturation. Active bugs require thinner boundary layers (∼100 μm), which are achieved by living in moving water or by swimming.

Flying insects have the highest mass-specific metabolic rate of all animals. Oxygen is supplied to the flight muscles by a combination of diffusion and convection along the internal air-filled tubes of the tracheal system. This study measured maximum flight metabolic rate (FMR) during tethered flight in the migratory locust under varying oxygen partial pressure ( ) in background gas mixtures of nitrogen (N), sulfur hexafluoride (SF) and helium (He), to vary O diffusivity and gas mixture density independently. With N as the sole background gas (normodiffusive-normodense), mass-independent FMR averaged 132±19 mW g at normoxia ( =21 kPa), and was not limited by tracheal system conductance, because FMR did not increase in hyperoxia. However, FMR declined immediately with hypoxia, oxy-conforming nearly completely. Thus, the locust respiratory system is matched to maximum functional requirements, with little reserve capacity. With SF as the sole background gas (hypodiffusive-hyperdense), the shape of the relationship between FMR and was similar to that in N, except that FMR was generally lower (e.g. 24% lower at normoxia). This appeared to be due to increased density of the gas mixture rather than decreased O diffusivity, because hyperoxia did not reverse it. Normoxic FMR was not significantly different in He-SF (hyperdiffusive-normodense) compared with the N background gas, and likewise there was no significant difference between FMR in SF-He (normodiffusive-hyperdense) compared with the SF background gas. The results indicate that convection, not diffusion, is the main mechanism of O delivery to the flight muscle of the locust when demand is high.

No abstract

Background: The repeatome, the collection of repetitive DNA sequences represented by transposable elements (TEs) and tandemly repeated satellite DNA (satDNAs), is found in high proportion in organisms across the tree of life. Grasshoppers have large genomes (average 9 Gb), containing large amounts of repetitive DNA which has hampered progress in assembling reference genomes. Here we combined linked-read genomics with transcriptomics to assemble, characterize, and compare the structure of the repeatome and its contribution to genome evolution, in four chromosomal races of the morabine grasshopper Vandiemenella viatica species complex. Results: We obtained linked-read genome assemblies of 2.73-3.27 Gb from estimated genome sizes of 4.26-5.07 Gb DNA per haploid genome of the four chromosomal races of V. viatica. These constitute the third largest insect genomes assembled so far (the largest being two locust grasshoppers). Combining complementary annotation tools and manual curation, we found a large diversity of TEs and satDNAs constituting 66 to 75 % per genome assembly. A comparison of sequence divergence within the TE classes revealed massive accumulation of recent TEs in all four races (314-463 Mb per assembly), indicating that their large genome size is likely due to similar rates of TE accumulation across the four races. Transcriptome sequencing showed more biased TE expression in reproductive tissues than somatic tissues, implying permissive transcription in gametogenesis. Out of 129 satDNA families, 102 satDNA families were shared among the four chromosomal races, which likely represent a repertoire of satDNA families in the ancestor of the V. viatica chromosomal races. Notably, 50 of these shared satDNA families underwent differential proliferation since the recent diversification of the V. viatica species complex. Conclusion: In-depth annotation of the repeatome in morabine grasshoppers provided new insights into the genome evolution of Orthoptera. Our TEs analysis revealed a massive recent accumulation of TEs equivalent to the size of entire Drosophila genomes, which likely explains the large genome sizes in grasshoppers. Although the TE and satDNA repertoires were rather similar between races, the patterns of TE expression and satDNA proliferation suggest rapid evolution of grasshopper genomes on recent timescales.

Many aquatic insects utilise air bubbles on the surface of their bodies to supply O 2 while they dive. The bubbles can simply store O 2 , as in the case of an 'air store', or they can act as a physical 'gas gill', extracting O 2 from the water. Backswimmers of the genus Anisops augment their air store with O 2 from haemoglobin cells located in the abdomen. The O 2 release from the haemoglobin helps stabilise bubble volume, enabling backswimmers to remain near neutrally buoyant for a period of the dive. It is generally assumed that the backswimmer air store does not act as a gas gill and that gas exchange with the water is negligible. This study combines measurements of dive characteristics under different exotic gases (N 2 , He, SF 6 , CO) with mathematical modelling, to show that the air store of the backswimmer Anisops deanei does exchange gases with the water. Our results indicate that approximately 20% of O 2 consumed during a dive is obtained directly from the water. Oxygen from the water complements that released from the haemoglobin, extending the period of near-neutral buoyancy and increasing dive duration.