Future-proofing the koala: Synergising genomic and environmental data for effective species management.

Climatic and evolutionary processes are inextricably linked to conservation. Avoiding extinction in rapidly changing environments often depends upon a species' capacity to adapt in the face of extreme selective pressures. Here, we employed exon capture and high-throughput next-generation sequencing to investigate the mechanisms underlying population structure and adaptive genetic variation in the koala (Phascolarctos cinereus), an iconic Australian marsupial that represents a unique conservation challenge because it is not uniformly threatened across its range. An examination of 250 specimens representing 91 wild source locations revealed that five major genetic clusters currently exist on a continental scale. The initial divergence of these clusters appears to have been concordant with the Mid-Brunhes Transition (~430 to 300 kya), a major climatic reorganisation that increased the amplitude of Pleistocene glacial-interglacial cycles. While signatures of polygenic selection and environmental adaptation were detected, strong evidence for repeated, climate-associated range contractions and demographic bottleneck events suggests that geographically isolated refugia may have played a more significant role in the survival of the koala through the Pleistocene glaciation than in situ adaptation. Consequently, the conservation of genome-wide genetic variation must be aligned with the protection of core koala habitat to increase the resilience of vulnerable populations to accelerating anthropogenic threats. Finally, we propose that the five major genetic clusters identified in this study should be accounted for in future koala conservation efforts (e.g., guiding translocations), as existing management divisions in the states of Queensland and New South Wales do not reflect historic or contemporary population structure.

Sperm Storage in a Family-Living Lizard, the Tree Skink (Egernia striolata).

The ability to produce viable offspring without recently mating, either through sperm storage or parthenogenesis, can provide fitness advantages under a suite of challenging ecological scenarios. Using genetic analysis, we demonstrate that 3 wild-caught female Tree Skinks (Egernia striolata) reproduced in captivity with no access to males for over a year, and that this is best explained by sperm storage. To the best of our knowledge, this is the first time female sperm storage has been documented in any monogamous family-living reptile, including social Australian egerniine skinks (from the subfamily Egerniinae). Furthermore, by using paternal reconstruction of genotypes we show that captive-born offspring produced by the same females in the preceding year, presumably without sperm storage, were sired by different males. We qualitatively compared aspects of these females' mates and offspring between years. The parents of each litter were unrelated, but paternal and offspring genotypes from litters resulting from stored sperm were more heterozygous than those inferred to be from recent matings. Family-living egerniine skinks generally have low rates of multiple paternity, yet our study suggests that female sperm storage, potentially from outside social partners, offers the real possibility of benefits. Possible benefits include increasing genetic compatibility of mates and avoiding inbreeding depression via cryptic female choice. Sperm storage in Tree Skinks, a family-living lizard with a monogamous mating system, suggests that females may bet-hedge through extra-pair copulation with more heterozygous males, reinforcing the idea that females could have more control on reproductive outcomes than previously thought.

Range size and growth temperature influence Eucalyptus species responses to an experimental heatwave.

Understanding forest tree responses to climate warming and heatwaves is important for predicting changes in tree species diversity, forest C uptake, and vegetation-climate interactions. Yet, tree species differences in heatwave tolerance and their plasticity to growth temperature remain poorly understood. In this study, populations of four Eucalyptus species, two with large range sizes and two with comparatively small range sizes, were grown under two temperature treatments (cool and warm) before being exposed to an equivalent experimental heatwave. We tested whether the species with large and small range sizes differed in heatwave tolerance, and whether trees grown under warmer temperatures were more tolerant of heatwave conditions than trees grown under cooler temperatures. Visible heatwave damage was more common and severe in the species with small rather than large range sizes. In general, species that showed less tissue damage maintained higher stomatal conductance, lower leaf temperatures, larger increases in isoprene emissions, and less photosynthetic inhibition than species that showed more damage. Species exhibiting more severe visible damage had larger increases in heat shock proteins (HSPs) and respiratory thermotolerance (Tmax ). Thus, across species, increases in HSPs and Tmax were positively correlated, but inversely related to increases in isoprene emissions. Integration of leaf gas-exchange, isoprene emissions, proteomics, and respiratory thermotolerance measurements provided new insight into mechanisms underlying variability in tree species heatwave tolerance. Importantly, warm-grown seedlings were, surprisingly, more susceptible to heatwave damage than cool-grown seedlings, which could be associated with reduced enzyme concentrations in leaves. We conclude that species with restricted range sizes, along with trees growing under climate warming, may be more vulnerable to heatwaves of the future.

Adaptation and conservation insights from the koala genome.

The koala, the only extant species of the marsupial family Phascolarctidae, is classified as 'vulnerable' due to habitat loss and widespread disease. We sequenced the koala genome, producing a complete and contiguous marsupial reference genome, including centromeres. We reveal that the koala's ability to detoxify eucalypt foliage may be due to expansions within a cytochrome P450 gene family, and its ability to smell, taste and moderate ingestion of plant secondary metabolites may be due to expansions in the vomeronasal and taste receptors. We characterized novel lactation proteins that protect young in the pouch and annotated immune genes important for response to chlamydial disease. Historical demography showed a substantial population crash coincident with the decline of Australian megafauna, while contemporary populations had biogeographic boundaries and increased inbreeding in populations affected by historic translocations. We identified genetically diverse populations that require habitat corridors and instituting of translocation programs to aid the koala's survival in the wild.

Unmasking the complexity of species identification in Australasian flying-foxes.

Pteropus (flying-foxes) are a speciose group of non-echolocating large bats, with five extant Australian species and 24 additional species distributed amongst the Pacific Islands. In 2015, an injured flying-fox with unusual facial markings was found in Sydney, Australia, following severe and widespread storms. Based on an initial assessment, the individual belonged to Pteropus but could not be readily identified to species. As a consequence, four hypotheses for its identification/origin were posited: the specimen represented (1) an undescribed Australian species; or (2) a morphological variant of a recognised Australian species; or (3) a hybrid individual; or (4) a vagrant from the nearby Southwest Pacific Islands. We used a combination of morphological and both mitochondrial- and nuclear DNA-based identification methods to assess these hypotheses. Based on the results, we propose that this morphologically unique Pteropus most likely represents an unusual P. alecto (black flying-fox) potentially resulting from introgression from another Pteropus species. Unexpectedly, this individual, and the addition of reference sequence data from newly vouchered specimens, revealed a previously unreported P. alecto mitochondrial DNA lineage. This lineage was distinct from currently available haplotypes. It also suggests long-term hybridisation commonly occurs between P. alecto and P. conspicillatus (spectacled flying-fox). This highlights the importance of extensive reference data, and the inclusion of multiple vouchered specimens for each species to encompass both intraspecific and interspecific variation to provide accurate and robust species identification. Moreover, our additional reference data further demonstrates the complexity of Pteropus species relationships, including hybridisation, and potential intraspecific biogeographical structure that may impact on their management and conservation.

Phylogeography of the Koala, (Phascolarctos cinereus), and Harmonising Data to Inform Conservation.

The Australian continent exhibits complex biogeographic patterns but studies of the impacts of Pleistocene climatic oscillation on the mesic environments of the Southern Hemisphere are limited. The koala (Phascolarctos cinereus), one of Australia's most iconic species, was historically widely distributed throughout much of eastern Australia but currently represents a complex conservation challenge. To better understand the challenges to koala genetic health, we assessed the phylogeographic history of the koala. Variation in the maternally inherited mitochondrial DNA (mtDNA) Control Region (CR) was examined in 662 koalas sampled throughout their distribution. In addition, koala CR haplotypes accessioned to Genbank were evaluated and consolidated. A total of 53 unique CR haplotypes have been isolated from koalas to date (including 15 haplotypes novel to this study). The relationships among koala CR haplotypes were indicative of a single Evolutionary Significant Unit and do not support the recognition of subspecies, but were separated into four weakly differentiated lineages which correspond to three geographic clusters: a central lineage, a southern lineage and two northern lineages co-occurring north of Brisbane. The three geographic clusters were separated by known Pleistocene biogeographic barriers: the Brisbane River Valley and Clarence River Valley, although there was evidence of mixing amongst clusters. While there is evidence for historical connectivity, current koala populations exhibit greater structure, suggesting habitat fragmentation may have restricted female-mediated gene flow. Since mtDNA data informs conservation planning, we provide a summary of existing CR haplotypes, standardise nomenclature and make recommendations for future studies to harmonise existing datasets. This holistic approach is critical to ensuring management is effective and small scale local population studies can be integrated into a wider species context.

Genetic Divergence among Regions Containing the Vulnerable Great Desert Skink (Liopholis kintorei) in the Australian Arid Zone.

Knowledge of genetic structure and patterns of connectivity is valuable for implementation of effective conservation management. The arid zone of Australia contains a rich biodiversity, however this has come under threat due to activities such as altered fire regimes, grazing and the introduction of feral herbivores and predators. Suitable habitats for many species can be separated by vast distances, and despite an apparent lack of current geographical barriers to dispersal, habitat specialisation, which is exhibited by many desert species, may limit connectivity throughout this expansive region. We characterised the genetic structure and differentiation of the great desert skink (Liopholis kintorei), which has a patchy, but widespread distribution in the western region of the Australian arid zone. As a species of cultural importance to local Aboriginal groups and nationally listed as Vulnerable, it is a conservation priority for numerous land managers in central Australia. Analysis of mitochondrial ND4 sequence data and ten nuclear microsatellite loci across six sampling localities through the distribution of L. kintorei revealed considerable differentiation among sites, with mitochondrial FST and microsatellite F'ST ranging from 0.047-0.938 and 0.257-0.440, respectively. The extent of differentiation suggests three main regions that should be managed separately, in particular the southeastern locality of Uluru. Current genetic delineation of these regions should be maintained if future intervention such as translocation or captive breeding is to be undertaken.

Microsatellite markers for the Chameleon grasshopper (Kosciuscola tristis) (Orthoptera: Acrididae), an Australian Alpine Specialist.

A set of polymorphic loci was characterised using an enrichment library for the Australian alpine specialist, the chameleon grasshopper (Kosciuscola tristis), an atypical grasshopper known for its remarkable temperature-controlled colour change. The number of alleles per locus ranged from three to 20 and observed heterozygosity from 0.16 to 0.76. These are the first microsatellite markers for a non-endangered Australian alpine animal and will inform questions of gene flow across the sky islands of this unique and threatened region.

Antifungal activity in thrips soldiers suggests a dual role for this caste.

The social insect soldier is perhaps the most widely known caste, because it often exhibits spectacular weapons, such as highly enlarged jaws or reinforced appendages, which are used to defend the colony against enemies ranging in size from wasps to anteaters. We examined the function of the enlarged forelimbs of soldiers (both male and female) of the eusocial, gall-inhabiting insect Kladothrips intermedius, and discovered that they have little impact on their ability to repel the specialized invading thrips Koptothrips species. While the efficacy of the enlarged forelimb appears equivocal, we show that soldiers secrete strong antifungal compounds capable of controlling the specialized insect fungal pathogen, Cordyceps bassiana. Our data suggest that these thrips soldiers have evolved in response to selection by both macro- and micro-organisms. While it is unknown whether specialized fungal pathogens have been major selective agents in the evolution of the soldier caste in general, they were probably present when sociality first evolved and may have been the primordial enemies of social insects.