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Aim: After environmental disasters, species with large population losses may need urgent protection to prevent extinction and support recovery. Following the 2019–2020 Australian megafires, we estimated population losses and recovery in fire-affected fauna, to inform conservation status assessments and management. Location: Temperate and subtropical Australia. Time period: 2019–2030 and beyond. Major taxa: Australian terrestrial and freshwater vertebrates; one invertebrate group. Methods: From > 1,050 fire-affected taxa, we selected 173 whose distributions substantially overlapped the fire extent. We estimated the proportion of each taxon’s distribution affected by fires, using fire severity and aquatic impact mapping, and new distribution mapping. Using expert elicitation informed by evidence of responses to previous wildfires, we estimated local population responses to fires of varying severity. We combined the spatial and elicitation data to estimate overall population loss and recovery trajectories, and thus indicate potential eligibility for listing as threatened, or uplisting, under Australian legislation. Results: We estimate that the 2019–2020 Australian megafires caused, or contributed to, population declines that make 70–82 taxa eligible for listing as threatened; and another 21–27 taxa eligible for uplisting. If so-listed, this represents a 22–26% increase in Australian statutory lists of threatened terrestrial and freshwater vertebrates and spiny crayfish, and uplisting for 8–10% of threatened taxa. Such changes would cause an abrupt worsening of underlying trajectories in vertebrates, as measured by Red List Indices. We predict that 54–88% of 173 assessed taxa will not recover to pre-fire population size within 10 years/three generations. Main conclusions: We suggest the 2019–2020 Australian megafires have worsened the conservation prospects for many species. Of the 91 taxa recommended for listing/uplisting consideration, 84 are now under formal review through national processes. Improving predictions about taxon vulnerability with empirical data on population responses, reducing the likelihood of future catastrophic events and mitigating their impacts on biodiversity, are critical.

More than a third of the world’s amphibian species are listed as Threatened or Extinct, with a recent assessment identifying 45 Australian frogs (18.4% of the currently recognised species) as ‘Threatened’ based on IUCN criteria. We applied structured expert elicitation to 26 frogs assessed as Critically Endangered and Endangered to estimate their probability of extinction by 2040. We also investigated whether participant experience (measured as a self-assigned categorical score, i.e. ‘expert’ or ‘non-expert’) influenced the estimates. Collation and analysis of participant opinion indicated that eight species are at high risk (>50% chance) of becoming extinct by 2040, with the disease chytridiomycosis identified as the primary threat. A further five species are at moderate–high risk (30–50% chance), primarily due to climate change. Fourteen of the 26 frog species are endemic to Queensland, with many species restricted to small geographic ranges that are susceptible to stochastic events (e.g. a severe heatwave or a large bushfire). Experts were more likely to rate extinction probability higher for poorly known species (those with

The budding of intralumenal vesicles (ILVs) at endosomes requires membrane scission by the ESCRT-III complex. This step is negatively regulated in yeast by Doa4, the ubiquitin hydrolase that deubiquitinates transmembrane proteins sorted as cargoes into ILVs. Doa4 acts non-enzymatically to inhibit ESCRT-III membrane scission activity by directly binding the Snf7 subunit of ESCRT-III. This interaction inhibits the remodeling/disassembly of Snf7 polymers required for the ILV membrane scission reaction. Thus, Doa4 is thought to have a structural role that delays ILV budding while it also functions enzymatically to deubiquitinate ILV cargoes. In this study, we show that Doa4 binding to Snf7 in vivo is antagonized by another ESCRT-III subunit, Vps20. Doa4 is restricted from interacting with Snf7 in yeast expressing a mutant Vps20 allele that constitutively binds Doa4. This inhibitory effect of Vps20 is suppressed by overexpression of another ESCRT-III-associated protein, Bro1. We show that Bro1 binds directly to Vps20, suggesting that Bro1 has a central role in relieving the antagonistic relationship that Vps20 has toward Doa4.

It may be possible to avert threatened species declines by protecting refuges that promote species persistence during times of stress. To do this, we need to know where refuges are located, and when and which management actions are required to preserve, enhance or replicate them. Here we use a niche-based perspective to characterise refuges that are either fixed or shifting in location over ecological time scales (hours to centuries). We synthesise current knowledge of the role of fixed and shifting refuges, using threatened species examples where possible, and examine their relationships with stressors including drought, fire, introduced species, disease, and their interactions. Refuges often provide greater cover, water, food availability or protection from predators than other areas within the same landscapes. In many cases, landscape features provide refuge, but refuges can also arise through dynamic and shifting species interactions (e.g., mesopredator suppression). Elucidating the mechanisms by which species benefit from refuges can help guide the creation of new or artificial refuges. Importantly, we also need to recognise when refuges alone are insufficient to halt the decline of species, and where more intensive conservation intervention may be required. We argue that understanding the role of ecological refuges is an important part of strategies to stem further global biodiversity loss.

Accurate detection of the amphibian fungal pathogen Batrachochytrium dendrobatidis (Bd) is critical for wildlife disease research; however, false negatives in detection do occur. Here we compared different DNA extraction methods to determine the threshold for Bd detection and identify an optimal extraction method to improve detection and quantification of the pathogen. We extracted both lab-created cell suspension standards using PrepMan Ultra, Chelex resin, and 3 spin column DNA extraction kits (Qiagen DNeasy Blood and Tissue, Zymo Quick DNA miniprep, and IBI gMAX mini kit), and further compared extraction methods using field-collected samples. We found that when extracting Bd DNA from cells in lab-created culture, the spin column extraction methods and PrepMan Ultra were equivalent, while the resin method detected higher Bd DNA quantities, especially at higher loads. However, when swabs from live animals were analyzed, low Bd quantities were more than twice as likely to be detected using a spin column extraction than with the PrepMan Ultra extraction method. All tested spin column extraction methods performed similarly across both field and lab samples. Samples containing low Bd quantities yielded inconsistent detection and quantification of Bd DNA copies regardless of extraction method. To manage imperfect detection of Bd, we suggest that presence/absence analyses are more informative than attempting to quantify Bd DNA when quantities are low. Overall, we recommend that a cost-benefit analysis of target species susceptibility and epidemiology be taken into consideration when designing an experiment to determine the most appropriate DNA extraction method to be used, because sometimes detecting low Bd quantities is imperative to the study, whereas in other situations, detecting low DNA quantities is less important.

The ESCRT-III protein complex executes reverse-topology membrane scission. The scission mechanism is unclear but is linked to remodeling of ESCRT-III complexes at the membrane surface. At endosomes, ESCRT-III mediates the budding of intralumenal vesicles (ILVs). In Saccharomyces cerevisiae, ESCRT-III activity at endosomes is regulated through an unknown mechanism by Doa4, an ubiquitin hydrolase that deubiquitylates transmembrane proteins sorted into ILVs. We report that the non-catalytic N-terminus of Doa4 binds Snf7, the predominant ESCRT-III subunit. Through this interaction, Doa4 overexpression alters Snf7 assembly status and inhibits ILV membrane scission. In vitro, the Doa4 N-terminus inhibits association of Snf7 with Vps2, which functions with Vps24 to arrest Snf7 polymerization and remodel Snf7 polymer structure. In vivo, Doa4 overexpression inhibits Snf7 interaction with Vps2 and also with the ATPase Vps4, which is recruited by Vps2 and Vps24 to remodel ESCRT-III complexes by catalyzing subunit turnover. Our data suggest a mechanism by which the deubiquitylation machinery regulates ILV biogenesis by interfering with ESCRT-III remodeling.

Practical conservation normally requires making decisions in the face of uncertainty. Our attitude toward that uncertainty, and the risks it entails, shape the way conservation decisions are made. Stochastic dominance (SD), a method more commonly used in economics, can be used to rank alternative conservation actions by comparing the probability distributions of their outcomes, making progressive simplified assumptions about the preferences of decision makers. Here, we illustrate the application of SD to conservation decisions using the recovery plan for an endangered frog species in Australia as a case study. SD is simple and intuitively appealing for conservation decisions; its broader application may encourage conservation decision makers to consider probabilistic uncertainty in light of their preferences, which may otherwise be difficult to recognize and assess transparently. A better treatment of attitudes towards uncertainty and risk may help ensure rational decision making in conservation and remove potential causes of stakeholder conflict.

Ex situ conservation strategies for threatened species often require long-term commitment and financial investment to achieve management objectives. We present a framework that considers the decision to adopt ex situ management for a target species as the end point of several linked decisions. We used a decision tree to intuitively represent the logical sequence of decision making. The first decision is to identify the specific management actions most likely to achieve the fundamental objectives of the recovery plan, with or without the use of ex-situ populations. Once this decision has been made, one decides whether to establish an ex situ population, accounting for the probability of success in the initial phase of the recovery plan, for example, the probability of successful breeding in captivity. Approaching these decisions in the reverse order (attempting to establish an ex situ population before its purpose is clearly defined) can lead to a poor allocation of resources, because it may restrict the range of available decisions in the second stage. We applied our decision framework to the recovery program for the threatened spotted tree frog (Litoria spenceri) of southeastern Australia. Across a range of possible management actions, only those including ex situ management were expected to provide >50% probability of the species' persistence, but these actions cost more than use of in situ alternatives only. The expected benefits of ex situ actions were predicted to be offset by additional uncertainty and stochasticity associated with establishing and maintaining ex situ populations. Naively implementing ex situ conservation strategies can lead to inefficient management. Our framework may help managers explicitly evaluate objectives, management options, and the probability of success prior to establishing a captive colony of any given species.

Understanding why related species have differing population-level responses to threats can be key to identifying conservation options for declining populations. However, this is difficult when multiple threats are implicated. Chytrid fungus (Batrachochytrium dendrobatidis: Bd) is implicated in at least 500 amphibian population declines globally, although few tangible options exist to mitigate pathogen impacts. Other threatening processes also invariably operate on most amphibians. Non-native fish, for example, can contribute to amphibian declines and may exacerbate Bd impacts. We disentangled the impacts of Bd and non-native fish upon two stream breeding frog species with differing conservation statuses to reveal vital rates that are crucial for species persistence. Litoria spenceri are threatened and historically occurred at elevations between 300–1100 m asl in southeastern Australia. Litoria lesueurii are non-threatened and sympatric with L. spenceri at some sites. Using mark-recapture derived demographic rates known to correlate with climate and elevation, discrete-time deterministic population models were constructed for each species at high, moderate and low elevation sites, and multiple management scenarios. Our study reveals that age to maturation, clutch size and egg-year 1 survival influenced interspecific and intraspecific population-level responses of the two frogs to Bd impacts. Importantly, our results highlight that an amphibian population’s capacity to counteract Bd-mediated adult mortality is clearly constrained by other threats and environment interactions that moderate species recruitment. Furthermore, understanding limits to recruitment may help identify Bd mitigation strategies. In our study, disease mitigation may be best achieved at some sites by enhancing recruitment through non-native fish management.

Anthropogenic climate change is a key threat to global biodiversity. To inform strategic actions aimed at conserving biodiversity as climate changes, conservation planners need early warning of the risks faced by different species. The IUCN Red List criteria for threatened species are widely acknowledged as useful risk assessment tools for informing conservation under constraints imposed by limited data. However, doubts have been expressed about the ability of the criteria to detect risks imposed by potentially slow-acting threats such as climate change, particularly because criteria addressing rates of population decline are assessed over time scales as short as 10 years. We used spatially explicit stochastic population models and dynamic species distribution models projected to future climates to determine how long before extinction a species would become eligible for listing as threatened based on the IUCN Red List criteria. We focused on a short-lived frog species (Assa darlingtoni) chosen specifically to represent potential weaknesses in the criteria to allow detailed consideration of the analytical issues and to develop an approach for wider application. The criteria were more sensitive to climate change than previously anticipated; lead times between initial listing in a threatened category and predicted extinction varied from 40 to 80 years, depending on data availability. We attributed this sensitivity primarily to the ensemble properties of the criteria that assess contrasting symptoms of extinction risk. Nevertheless, we recommend the robustness of the criteria warrants further investigation across species with contrasting life histories and patterns of decline. The adequacy of these lead times for early warning depends on practicalities of environmental policy and management, bureaucratic or political inertia, and the anticipated species response times to management actions.

Residual limb pain (RLP) and phantom limb pain (PLP) can be debilitating and can prevent functional gains following amputation. High correlations have been reported between RLP and the stump neuromas following amputation. Many treatment methods including physical therapy, medications, and interventions, have been used with limited success. Pulsed radiofrequency ablation (PRFA) has shown promise in treating neuropathic pain because of the inhibition of evoked synaptic activity. We present 4 amputees who were treated with PRFA after failing conservative management for their RLP and PLP. All 4 patients underwent PRFA and demonstrated at least 80% relief of RLP for over 6 months. One patient reported a complete resolution of phantom sensation while another patient had significantly decreased frequency of spontaneous PLP and resolution of evoked PLP. In addition, all patients reported improved overall function including increased prosthetic tolerance and decreased oral pain medications. This case series suggests that PRFA is a viable treatment option which might be used for long-term relief of intractable RLP and/or PLP.

Although graphite is the most commonly used anode material for Li-ion batteries, its low capacity (372 mAh/g) limits the energy storage performance. Recently, graphene has demonstrated great promise as a candidate to replace graphite due to its high power and high energy densities.1-3 Here, we prepared Li-ion battery anodes based on reduced graphene oxide (rGO) powder obtained from an established company and investigated effects of oxygen contents in the rGO on the energy storage properties. The oxygen content of rGO could be controlled by heat-treating the rGO at 500 oC and 800 oC under Ar condition. The O/C atomic ratios from XPS were 0.208, 0.126, and 0.031 for the pristine rGO, the rGO heat-treated at 500 oC (rGO-HT500) and the 800 oC (rGO-HT800), respectively. We employed Dr. Blade method to prepare the electrodes using the pristine and heat-treated rGO. While the loading density of the electrodes were kept in the range of 1.68−1.75 mgrGO/cm2 electrode, the density gradually decreased with the heat-treatment from 0.62 to 0.21 g/cm3. We then fabricated a Li coin-type cell (CR2032) with the rGO electrode in an Ar-filled glovebox. Li metal foil, Celgard 2500 membrane, and 1 M LiPF6 (in EC/DMC) were used as a counter electrode, a separator, and an electrolyte, respectively. For evaluating energy storage performance of the rGO-based electrode, we conducted galvanostatic charge/discharge test at different current densities in the range of 37.2–3720 mA/g. At the current density of 37.2 mA/g, both electrodes based on the pristine rGO and rGO-HT500 showed ~1000 mAh/grGO, which is greatly enhanced capacity by 2−3 folds relative to the theoretical capacity of graphite (Fig. 1). However, the rGO-HT800-based electrode exhibited only showed a capacity of 579 mAh/grGO, indicating the minor oxygen content could contribute to the charge capacity of rGO in the Li-ion battery. J. Hassoun et al., An advanced lithium-ion battery based on a graphene anode and a lithium iron phosphate cathode. Nano letters 14, 4901-4906 (2014). G. Wang, X. Shen, J. Yao, J. Park, Graphene nanosheets for enhanced lithium storage in lithium ion batteries. Carbon 47, 2049-2053 (2009). F. J. Sonia et al., Understanding the Li-storage in few layers graphene with respect to bulk graphite: experimental, analytical and computational study. Journal of Materials Chemistry A 5, 8662-8679 (2017). Figure 1. (a) Charge/discharge profiles of pristine rGO at difference current densities from 37.5 to 3750 mA/g. (b) Comparison of charge/discharge profiles of pristine and heat-treated rGO electrodes obtained at 37.5 mA/g. Figure 1

Ex situ conservation strategies for threatened species often require long-term commitment and financial investment to achieve management objectives. We present a framework that considers the decision to adopt ex situ management for a target species as the end point of several linked decisions. We used a decision tree to intuitively represent the logical sequence of decision making. The first decision is to identify the specific management actions most likely to achieve the fundamental objectives of the recovery plan, with or without the use of ex-situ populations. Once this decision has been made, one decides whether to establish an ex situ population, accounting for the probability of success in the initial phase of the recovery plan, for example, the probability of successful breeding in captivity. Approaching these decisions in the reverse order (attempting to establish an ex situ population before its purpose is clearly defined) can lead to a poor allocation of resources, because it may restrict the range of available decisions in the second stage. We applied our decision framework to the recovery program for the threatened spotted tree frog (Litoria spenceri) of southeastern Australia. Across a range of possible management actions, only those including ex situ management were expected to provide50% probability of the species' persistence, but these actions cost more than use of in situ alternatives only. The expected benefits of ex situ actions were predicted to be offset by additional uncertainty and stochasticity associated with establishing and maintaining ex situ populations. Naïvely implementing ex situ conservation strategies can lead to inefficient management. Our framework may help managers explicitly evaluate objectives, management options, and the probability of success prior to establishing a captive colony of any given species.

Anthropogenic climate change is a key threat to global biodiversity. To inform strategic actions aimed at conserving biodiversity as climate changes, conservation planners need early warning of the risks faced by different species. The IUCN Red List criteria for threatened species are widely acknowledged as useful risk assessment tools for informing conservation under constraints imposed by limited data. However, doubts have been expressed about the ability of the criteria to detect risks imposed by potentially slow-acting threats such as climate change, particularly because criteria addressing rates of population decline are assessed over time scales as short as 10 years. We used spatially explicit stochastic population models and dynamic species distribution models projected to future climates to determine how long before extinction a species would become eligible for listing as threatened based on the IUCN Red List criteria. We focused on a short-lived frog species (Assa darlingtoni) chosen specifically to represent potential weaknesses in the criteria to allow detailed consideration of the analytical issues and to develop an approach for wider application. The criteria were more sensitive to climate change than previously anticipated; lead times between initial listing in a threatened category and predicted extinction varied from 40 to 80 years, depending on data availability. We attributed this sensitivity primarily to the ensemble properties of the criteria that assess contrasting symptoms of extinction risk. Nevertheless, we recommend the robustness of the criteria warrants further investigation across species with contrasting life histories and patterns of decline. The adequacy of these lead times for early warning depends on practicalities of environmental policy and management, bureaucratic or political inertia, and the anticipated species response times to management actions.

To protect Australian amphibian biodiversity, we have identified and prioritised frog species at an imminent risk of extinction from chytridiomycosis, and devised national management and research priorities for disease mitigation. Six Australian frogs have not been observed in the wild since the initial emergence of chytridiomycosis and may be extinct. Seven extant frog species were assessed as needing urgent conservation interventions because of (1) their small populations and/or ongoing declines throughout their ranges (southern corroboree frog (Pseudophryne corroboree, New South Wales), northern corroboree frog (Pseudophryne pengilleyi, Australian Capital Territory, New South Wales), Baw Baw frog (Philoria frosti, Victoria), Litoria spenceri (spotted tree frog, Victoria, New South Wales), Kroombit tinkerfrog (Taudactylus pleione, Queensland), armoured mist frog (Litoria lorica, Queensland)) or (2) predicted severe decline associated with the spread of chytridiomycosis in the case of Tasmanian tree frog (Litoria burrowsae, Tasmania). For these species, the risk of extinction is high, but can be mitigated. They require increased survey effort to define their distributional limits and to monitor and detect further population changes, as well as well-resourced management strategies that include captive assurance populations. A further 22 frog species were considered at a moderate to lower risk of extinction from chytridiomycosis. Management actions that identify and create or maintain habitat refugia from chytridiomycosis and target other threatening processes such as habitat loss and degradation may be effective in promoting their recovery. Our assessments for some of these species remain uncertain and further taxonomical clarification is needed to determine their conservation importance. Management actions are currently being developed and trialled to mitigate the threat posed by chytridiomycosis. However, proven solutions to facilitate population recovery in the wild are lacking; hence, we prioritise research topics to achieve this aim. Importantly, the effectiveness of novel management solutions will likely differ among species due to variation in disease ecology, highlighting the need for species-specific research. We call for an independent management and research fund of AU$15 million over 5 years to be allocated to recovery actions as determined by a National Chytridiomycosis Working Group of amphibian managers and scientists. Procrastination on this issue will likely result in additional extinction of Australia’s amphibians in the near future.

About 10 per cent of birds’ eggs fail to hatch, but\ud the incidence of failure can be much higher in\ud endangered species.Most studies fail to distinguish\ud between infertility (due to a lack of sperm) and\ud embryo mortality as the cause of hatching failure,\ud yet doing so is crucial in order to understand the\ud underlying problem. Using newly validated techniques\ud to visualize sperm and embryonic tissue,\ud we assessed the fertility status of unhatched eggs\ud of five endangered species, including both wild\ud and captive birds. All eggs were classified as ‘infertile’\ud when collected, but most were actually fertile\ud with numerous sperm on the ovum. Eggs of captive\ud birds had fewer sperm and were more likely\ud to be infertile than those of wild birds. Our findings\ud raise important questions regarding the\ud management of captive breeding programmes.

Residual limb pain (RLP) and phantom limb pain (PLP) can be debilitating and can prevent functional gains following amputation. High correlations have been reported between RLP and the stump neuromas following amputation. Many treatment methods including physical therapy, medications, and interventions, have been used with limited success. Pulsed radiofrequency ablation (PRFA) has shown promise in treating neuropathic pain because of the inhibition of evoked synaptic activity. We present 4 amputees who were treated with PRFA after failing conservative management for their RLP and PLP. All 4 patients underwent PRFA and demonstrated at least 80% relief of RLP for over 6 months. One patient reported a complete resolution of phantom sensation while another patient had significantly decreased frequency of spontaneous PLP and resolution of evoked PLP. In addition, all patients reported improved overall function including increased prosthetic tolerance and decreased oral pain medications. This case series suggests that PRFA is a viable treatment option which might be used for long-term relief of intractable RLP and/or PLP.

Porcine FSH/LH stimulation successfully induced development of multiple large (≥4 mm) antral follicles in 10 of 11 common wombats. A mean of 5.5 metaphase II (MII) oocytes were aspirated from wombats that were stimulated during the follicular phase of the oestrous cycle (n = 3) or after pouch young removal (n = 3). Three subadults (n = 3) and two anoestrus adults did not produce MII oocytes despite pFSH/pLH administration. In vitro maturation of immature oocytes at the time of aspiration doubled the number of MII oocytes that could be collected from pFSH/pLH stimulated wombats. Immature oocytes with cumulus attached, matured more readily to the MII stage than immature oocytes without cumulus. Following intracytoplasmic sperm injection (ICSI), approximately 5% of the oocytes that were MII at the time of collection cleaved. Approximately 5% of those that were matured by in vitro maturation (IVM) formed two polar bodies following ICSI, although they not cleave. Parthenogenesis cannot be excluded. This demonstrates that assisted reproductive technologies may be applicable to the common wombat.

Assisted reproductive techniques, such as in vitro oocyte maturation in conjunction with in vitro fertilisation, may be used as a tool to manipulate reproduction. Using the common wombat as a model for the critically endangered northern hairy-nosed wombat, the present study examined whether oocyte maturation could be achieved under field conditions. At the time of collection, no oocytes were at the metaphase II (MII) stage (0/42). After 60 h culture using the submarine incubation system, 34% of oocytes (24/70) matured to telophase/MII, as indicated by the presence of a polar body. The proportion of oocytes that reached MII was higher for oocytes collected from follicles >2 mm in diameter compared with follicles 2 mm in diameter had the highest maturation rate (58%). Maturation was not affected by the reproductive status of the common wombat or a delay of up to 5 h before oocyte collection. In conclusion, the present study demonstrated that oocytes collected from non-stimulated common wombats can mature to MII in culture.

Wild-caught female common wombats from Victoria, Australia, were studied in captivity to investigate the oestrous cycle by assessing vaginal cytology and peripheral plasma progesterone concentrations. Eight wombats, five adults (21–29 kg) and three subadults (19–23 kg), which were held for between 2 weeks and 11 months did not cycle in captivity. Their progesterone concentrations were consistently low (≤6.9 nmol L–1) and vaginal smears contained predominantly superficial epithelial cells. Three wombats (21–27 kg), held in captivity for >1 year, regularly cycled (when bodyweights exceeded 23.5 kg). Information gathered from four consecutive cycles in each of these three wombats revealed a follicular phase with low progesterone concentrations (≤6.9 nmol L–1) and vaginal smears with a high percentage of superficial epithelial cells alternating with periods of high progesterone concentrations (range 41.6–123.8 nmol L–1) and smears in which parabasal–intermediate epithelial cells predominated. The average length of the monitored oestrous cycles was 47.2 days (35–60 days). The follicular phase lasted ~19 days and the luteal phase lasted ~28 days. In conclusion, wombats can cycle regularly in captivity even under conditions of intensive monitoring.