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Catastrophic wildfires impacted large areas of western Kangaroo Island (KI), South Australia in 2019–2020, burning habitat for many species, including large proportions of the distributional range of the KI micro-trapdoor spider Moggridgea rainbowi, which led to it being listed as Endangered under Australia’s Environment Protection and Biodiversity Conservation Act (EPBC Act). In order to assess population genetic structure in M. rainbowi and detect diagnosable lineages and their distributional patterns across KI, 28 individuals were genotyped for 2495 loci, sampling from all known populations of the species. Population genetic and phylogenetic analyses of nuclear and mitochondrial sequence data provided strong support for three Evolutionarily Significant Units (ESUs) within M. rainbowi; two populations on eastern KI and a heavily fire-impacted western population. High levels of divergence and fixed allelic differences between 5 and 16% indicate a lack of gene flow between ESUs and long periods of isolation. Distributional patterns of these lineages match likely locations of isolation events caused by successive changes to sea level during the Quaternary (2.58 million years ago to present), which led to KI being intermittently connected to the mainland or separated into one or more islands. Our findings have strong conservation implications for M. rainbowi and highlight the importance of inclusion of population genetic structure to inform conservation strategies and to conserve lineage biodiversity at the species level and below.

The Scirtidae Fleming, 1821 has been identified as one of the earliest diverging groups of Polyphagan beetles and is particularly speciose in Australia. However, very little is known about the origin of the Australian scirtids and there is a need for a robust, well-supported phylogeny to guide the genus and species descriptions and understand the relationships among taxa. In this study we carried out a phylogenetic analysis of the Australian Scirtinae Fleming, 1821, using DNA sequence data from ultraconserved elements (UCEs) and included representative taxa from New Zealand, New Caledonia, South America, South Africa and Eurasia in the analysis. Bayesian analyses of a concatenated dataset from 79 taxa recovered four major Southern Hemisphere groupings and two Australian–Eurasian groupings. The Veronatus group mainly consisted of genera from New Zealand, with the three Australian representatives only distantly related to each other. Relaxed molecular clock analyses, using the estimated age of the crown node of the Polyphaga for calibration, support a Gondwanan history for four of the groups of Australian Scirtinae and a northern origin for two groups. Our results highlight the value of commercially available UCEs for resolving the phylogenetic history of ancient groups of Coleoptera.

Amphipod crustaceans comprise a significant and enigmatic component of Australian groundwater ecosystems, particularly in the Pilbara region of Western Australia. Many amphipod species in the Pilbara, including species in the genus Nedsia Barnard & Williams, 1995, are considered short range endemics, poorly or contentiously defined by taxonomic treatments based on morphology alone and have uncertain distributions as a consequence of this taxonomy. A modern systematic revision of Nedsia is presented here, utilising both molecular and morphological analyses alongside distributional data to delineate species. We describe 13 new species of Nedsia, confirm three existing species and synonymise eight previously described species. Nedsia species are confirmed to be functionally morphologically cryptic, with COI divergences at the 5–20% level. We present comparatively reduced taxonomic descriptions for these cryptic amphipod species in an effort to provide an accelerated pathway for future taxonomic work. The research provides the basis for future environmental impact assessments involving Nedsia species and ongoing monitoring of the groundwater communities these form part of in the resource-rich Pilbara region.

Chemical cues in subterranean habitats differ highly from those on the surface due to the contrasting environmental conditions, such as absolute darkness, high humidity or food scarcity. Subterranean animals underwent changes to their sensory systems to facilitate the perception of essential stimuli for underground lifestyles. Despite representing unique systems to understand biological adaptation, the genomic basis of chemosensation across cave-dwelling species remains unexplored from a macroevolutionary perspective. Here, we explore the evolution of chemoreception in three beetle tribes that underwent at least six independent transitions to the underground through a phylogenomics spyglass. Our findings suggest that the chemosensory gene repertoire varies dramatically between species. Overall, no parallel changes in the net rate of evolution of chemosensory gene families were detected prior, during, or after the habitat shift among subterranean lineages. Contrarily, we found evidence of lineage-specific changes within surface and subterranean lineages. Our results reveal key duplications and losses shared between some of the lineages transitioning to the underground, including the loss of sugar receptors and gene duplications of the highly conserved ionotropic receptors IR25a and IR8a, involved in thermal and humidity sensing among other olfactory roles in insects. These duplications were detected both in independent subterranean lineages and their surface relatives, suggesting parallel evolution of these genes across lineages giving rise to cave-dwelling species. Overall, our results shed light on the genomic basis of chemoreception in subterranean beetles and contribute to deepen our understanding of the genomic underpinnings of adaptation to the subterranean lifestyle at a macroevolutionary scale.

Information on the extent of genetic differentiation among seabird populations is essential for conservation planning. Here we used, for the first time in little penguins (Eudyptula minor), a next generation sequencing approach to investigate population genetic structure, gene flow and inbreeding among eight colonies in South Australia. We found that Troubridge Island individuals were genetically distinct from the other sampled colonies, and that colonies on all islands were connected by moderate levels of gene flow possibly due to migration from the Kangaroo Island colonies. While further genetic sampling is needed, these data support the idea that South Australia may be a zone of subtle and rapid genetic change for little penguins.

This study investigates two parasitic reptile ticks — Bothriocroton hydrosauri and Amblyomma limbatum — of the sleepy lizard (Tiliqua rugosa) that abut at a 1–2 km wide parapatric boundary in South Australia. Long‐term research has investigated potential mechanisms to explain the maintenance of this boundary but has not uncovered why the distribution of A. limbatum does not extend further south. It has been previously hypothesised that pathogens may be responsible for maintaining parapatric boundaries. Rickettsia spp. has previously been reported in B. hydrosauri ticks. This study explored whether Rickettsia spp. occurs in co‐occurring A. limbatum. We observed that Rickettsia spp. was absent from all A. limbatum ticks and that 83% of examined B. hydrosauri were found to be positive with a spotted fever group Rickettsia strain. This study puts forward the hypothesis that Rickettsia spp. could contribute to the maintenance of the Mt Mary parapatric boundary between these two tick species. Further work is required to determine whether Rickettsia spp. can be transmitted from B. hydrosauri to A. limbatum and — if transmission can occur — to explore whether Rickettsia is lethal to A. limbatum ticks.

In the framework of neutral theory of molecular evolution, genes specific to the development and function of eyes in subterranean animals living in permanent darkness are expected to evolve by relaxed selection, ultimately becoming pseudogenes. However, definitive empirical evidence for the role of neutral processes in the loss of vision over evolutionary time remains controversial. In previous studies, we characterized an assemblage of independently-evolved water beetle (Dytiscidae) species from a subterranean archipelago in Western Australia, where parallel vision and eye loss have occurred. Using a combination of transcriptomics and exon capture, we present evidence of parallel coding sequence decay, resulting from the accumulation of frameshift mutations and premature stop codons, in eight phototransduction genes (arrestins, opsins, ninaC and transient receptor potential channel genes) in 32 subterranean species in contrast to surface species, where these genes have open reading frames. Our results provide strong evidence to support neutral evolutionary processes as a major contributing factor to the loss of phototransduction genes in subterranean animals, with the ultimate fate being the irreversible loss of a light detection system.

Amphipod crustaceans comprise a significant and enigmatic component of Australian groundwater ecosystems, particularly in the Pilbara region of Western Australia. Many amphipod species in the Pilbara, including species in the genus Nedsia Barnard & Williams, 1995, are considered short range endemics, poorly or contentiously defined by taxonomic treatments based on morphology alone and have uncertain distributions as a consequence of this taxonomy. A modern systematic revision of Nedsia is presented here, utilising both molecular and morphological analyses alongside distributional data to delineate species. We describe 13 new species of Nedsia, confirm three existing species and synonymise eight previously described species. Nedsia species are confirmed to be functionally morphologically cryptic, with COI divergences at the 5–20% level. We present comparatively reduced taxonomic descriptions for these cryptic amphipod species in an effort to provide an accelerated pathway for future taxonomic work. The research provides the basis for future environmental impact assessments involving Nedsia species and ongoing monitoring of the groundwater communities these form part of in the resource-rich Pilbara region.

Mitochondrial heteroplasmy is the occurrence of more than one type of mitochondrial DNA within a single individual. Although generally reported to occur in a small subset of individuals within a species, there are some instances of widespread heteroplasmy across entire populations.

ContextIn the era of the Anthropocene, habitat loss and environmental change threaten the persistence of many species. Genotyping-By-Sequencing (GBS) is a useful molecular tool for understanding how patterns of gene flow are associated with contemporary habitat distributions that may be affected by environmental change. Two parapatric subspecies of the threatened thick-billed grasswren (TBGW; Amytornis modestus) more frequently occur in different plant communities. As such, a preference for plant community type could reduce subspecific introgression and increase genetic diversity at the parapatric boundary.AimsWe aimed to measure gene flow within and among two TBGW subspecies and tested whether divergent genomic markers were associated with plant community type.MethodsWe sequenced 118 individuals from either of the two TBGW subspecies or in the region of parapatry and identified 7583 SNPs through ddRADseq.Key resultsWe found evidence of asymmetric gene flow and a genomic pattern of isolation-by-distance. There were sixteen genomic outliers correlated with plant community type (regardless of location).ConclusionsThese findings show that plant community type does not prevent introgression in one subspecies (A. m. raglessi), but low dispersal and habitat heterogeneity could contribute to the maintenance of distinct subspecific morphotypes. Local adaptation in different plant community types could also provide a mechanism for future divergence.ImplicationsWe suggest subspecific introgression could increase genetic variation and the adaptive potential of the species, facilitating species persistence under conditions of climate change.Introgression between grasswren subspeciesCharacterising gene flow facilitates conservation management. This study used genomic markers to measure gene flow between thick-billed grasswren subspecies and found results that support taxonomic identification of the two subspecies and suggests grasswrens have low dispersal and may benefit from increased genetic diversity. Recognition of models of divergence with gene flow will be necessary for future conservation management.

BackgroundThe 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 grasshopperVandiemenella viaticaspecies complex.ResultsWe 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 ofV. 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 theV. viaticachromosomal races. Notably, 50 of these shared satDNA families underwent differential proliferation since the recent diversification of theV. viaticaspecies complex.ConclusionIn-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 entireDrosophilagenomes, 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.

Most subterranean animals are assumed to have evolved from surface ancestors following colonisation of a cave system, however very few studies have raised the possibility of 'subterranean speciation' in underground habitats (i.e. obligate cave-dwelling organisms (troglobionts) descended from troglobiotic ancestors). Numerous endemic subterranean diving beetle species from spatially-discrete calcrete aquifers in Western Australia (stygobionts) have evolved independently from surface ancestors; however, several cases of sympatric sister species raises the possibility of subterranean speciation. We tested this hypothesis using vision (phototransduction) genes that are evolving under neutral processes in subterranean species and purifying selection in surface species. Using sequence data from 32 subterranean and five surface species in the genus Paroster (Dytiscidae), we identified deleterious mutations in: long wavelength opsin (lwop), arrestin 1 (arr1), and arrestin 2 (arr2) shared by a sympatric sister-species triplet, arr1 shared by a sympatric sister-species pair, and lwop and arr2 shared among closely related species in adjacent calcrete aquifers. In all cases, a common ancestor possessed the function-altering mutations, implying they were already adapted to aphotic environments. Our study represents one of the first confirmed cases of subterranean speciation in cave insects. The assessment of genes undergoing pseudogenisation provides a novel way of testing modes of speciation and the history of diversification in blind cave animals. This article is protected by copyright. All rights reserved.

BackgroundRepetitive 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 grasshopperVandiemenella viaticaspecies complex and determine their contribution to genome evolution.ResultsWe 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 ofV. 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 theV. viaticachromosomal races. Notably, 50 of these shared satDNA families underwent differential proliferation since the recent diversification of theV. viaticaspecies complex.ConclusionThis 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 entireDrosophilagenomes, 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.

The major histocompatibility complex (MHC) is a hypervariable gene family that plays an essential role in the recognition of pathogens and immune response. Research on the reptilian MHC has lagged behind other vertebrate groups. Here, we genotyped individuals of an Australian agamid lizard species at MHC class I loci using a recently developed clustering method and family group data. Our method allowed identification of low amplification efficiency alleles and estimation of both type I and II genotyping error rates. The number of MHC class I alleles per individual varied within populations and together with allele segregation patterns, suggests either natural copy number variation or allele dropout. Genotypes from individuals across five populations revealed shared alleles among populations and low allelic diversity in an island population. Finally, we identified sites under selection and designated them putative peptide binding regions. Our results provide a foundation for future work on the MHC class I region of agamid lizards.

The endangered subspecies of Southern Emu-Wren (Stipiturus malachurus intermedius) in the Mount Lofty Ranges (MLR), South Australia has a fragmented distribution and poor dispersal capabili...

A recent DNA barcoding study of Australian microgastrines (Hymenoptera: Braconidae) sought to use next-generation sequencing of the cytochrome c oxidase subunit 1 (COI) barcoding gene region, the wingless (WG) gene and the internal transcribed spacer 2 (ITS2) to delimit molecular species in a highly diverse group of parasitic wasps. Large intragenomic distances between ITS2 variants, often larger than the average interspecific variation, caused difficulties in using ITS2 for species delimitation in both threshold and tree-based approaches, and the gene was not included in the reported results of the previous DNA barcoding study. We here report on the intragenomic, and the intra- and interspecies, variation in ITS2in the microgastrine genus Diolcogasterto further investigate the value of ITS2as a marker for species delimitation and phylogenetics of the Microgastrinae. Distinctive intragenomic variant patterns were found in different species of Diolcogaster, with some species possessing a single major variant, and others possessing many divergent variants. Characterizing intragenomic variation of ITS2is critical as it is a widely used marker in hymenopteran phylogenetics and species delimitation, and large intragenomic distances such as those found in this study may obscure phylogenetic signal.

Next-generation sequencing has revolutionised molecular ecology. Its key advantages are a more accurate representation of genetic variation made possible by the generation of large volumes of data, more quickly and at a lower price per sequence than traditional sequencing methods. Yet these benefits come with a cost. For example, next-generation sequencing is error prone and requires increased quality control compared with traditional methods. Problems associated with next-generation sequencing may be exacerbated when sequencing gene complexes such as the major histocompatibility complex (MHC). Although not eliminated, significant progress has been made in addressing some of those problems and there is an increasing literature utilising this technology for studies of the MHC. However, what is generally lacking is detailed documentation of the methods used, and clear reasoning, for each step. Here we document detailed methodology, using an Australian lizard, Egernia stokesii, as a case study, with ...

The Microgastrinae are a hugely diverse subfamily of endoparasitoid wasps of lepidopteran caterpillars. They are important in agriculture as biological control agents and play a significant ecological role in the regulation of caterpillar populations. Whilst the group has been the focus of intensive rearing and DNA barcoding studies in the Northern Hemisphere, the Australian fauna has received little attention. In total, 99 species have been described from or have been introduced into Australia, but the real species diversity for the region is clearly much larger than this. In this study, museum ethanol samples and recent field collections were mined for hundreds of specimens of microgastrine wasps, which were then barcoded for the COI region, ITS2 ribosomal spacer and the wingless nuclear genes, using a pooled sequencing approach on an Illumina Miseq system. Full COI sequences were obtained for 525 individuals which, when combined with 162 publicly available sequences, represented 417 haplotypes, and a total of 236 species were delimited using a consensus approach. By more than doubling the number of known microgastrine wasp species in Australia, our study highlights the value of DNA barcoding in the context of employing high-throughput sequencing methods of bulk ethanol museum collections for biodiversity assessment.

The origin of the mammalian order Eulipotyphla has been debated intensively with arguments around whether they began diversifying before or after the Cretaceous-Palaeogene (K-Pg) boundary at 66 Ma. Here, we used an in-solution nucleotide capture method and next generation DNA sequencing to determine the sequence of hundreds of ultra-conserved elements (UCEs), and conducted phylogenomic and molecular dating analyses for the four extant eulipotyphlan lineages—Erinaceidae, Solenodontidae, Soricidae, and Talpidae. Concatenated maximum-likelihood analyses with single or partitioned models and a coalescent species-tree analysis showed that divergences among the four major eulipotyphlan lineages occurred within a short period of evolutionary time, but did not resolve the interrelationships among them. Alternative suboptimal phylogenetic hypotheses received consistently the same amount of support from different UCE loci, and were not significantly different from the maximum likelihood tree topology, suggesting the prevalence of stochastic lineage sorting. Molecular dating analyses that incorporated among-lineage evolutionary rate differences supported a scenario where the four eulipotyphlan families diversified between 57.8 and 63.2 Ma. Given short branch lengths with low support values, traces of rampant genome-wide stochastic lineage sorting, and post K-Pg diversification, we concluded that the crown eulipotyphlan lineages arose through a rapid diversification after the K-Pg boundary when novel niches were created by the mass extinction of species. We are grateful to Rafael Borroto-Páez, Jorge L. Delgado-Labañino,José Alvarez-Lemus for their help in sampling of solenodons in Cuba.We thank Kieren Mitchell for his advice on our next generation se-quencing experiments. We also appreciate Junji Moribe and AkioShinohara for providing the animal photos ofCrocidura dsinezumiandMogera wogura, respectively. This study was supported by theLeadership Fund of Hokkaido University (no grant number to SDO) andJSPS, Grants-in-Aid for Scientific Research (16H05655 to MK). Scopus

Three new species of Koonunga were discovered in surface and subterranean waters in southern Australia, and were defined using mtDNA analyses and morphology. The new species are: Koonunga hornei Leijs & King; K. tatiaraensis Leijs & King and K. allambiensis Leijs & King. Molecular clock analyses indicate that the divergence times of the species are older than the landscape that they currently inhabit. Different scenarios explaining this apparent discrepancy are discussed in the context of the palaeography of the area. A freshwater epigean origin for Koonunga is considered the most likely hypothesis, whereby some lineages made the transition to the subterranean environment within the last few million years influenced by significant climatic cooling/drying. We discuss the possibility that one stygobitic lineage secondarily regained some of its body pigmentation as adaptation to increased photic conditions after cave collapse and forming of cenotes during the last glacial maximum.