No Ranaviral DNA Found in Australian Freshwater Turtles, 2014-19, Despite Previous Serologic Evidence.

Ranaviruses are pathogens of ectothermic vertebrates (fish, amphibians, and reptiles). Turtles are the most common group of reptiles reported with ranaviral infections. However, there have been no surveys for wild ranaviral infection in any turtles from the suborder Pleurodira, despite ranaviral distributions and experimentally susceptible pleurodiran turtle populations overlapping in several areas, including Australia. We assayed 397 pooled blood samples from six Australian freshwater turtle species collected from five different sites in northern Australia between 2014 and 2019. Historical serologic surveys in the area had found antiranaviral antibodies; however, we did not detect any ranaviral DNA in our samples. Discrepancies between historical serologic and our molecular results may be explained by low viral prevalence during the years that these samples were collected, survivorship bias, or possibly an age class bias in sampling.

How turtles keep their cool: Seasonal and diel basking patterns in a tropical turtle.

Behavioural thermoregulation by ectotherms is an important mechanism for maintaining body temperatures to optimise physiological performance. Experimental studies suggest that nocturnal basking by Krefft's river turtles (Emydura macquarii krefftii) in the tropics may allow them to avoid high water temperatures, however, this hypothesis has yet to be tested in the field. In this study, we examined the influence of environmental temperature on seasonal and diel patterns of basking in E. m. krefftii in tropical north Queensland, Australia. Wildlife cameras were used to document turtle basking events for seven consecutive days and nights for each month over a year (April 2020-March 2021). Air and water temperatures were recorded simultaneously using temperature loggers. We used a negative binomial mixed effects model to compare mean basking durations (min) occurring among four environmental temperature categories based on population thermal preference (26 °C): 1) air temperature above and water temperature below preferred temperature; 2) air temperature below and water temperature above preferred temperature; 3) air and water temperatures both above preferred temperature; and 4) air and water temperatures both below preferred temperature. Basking behaviour was influenced significantly by the relationship between air and water temperature. During the day, turtles spent significantly less time basking when both air and water temperatures were above their preferred temperatures. Conversely, at night, turtles spent significantly more time basking when water temperatures were warm and air temperatures were cool relative to their preferred temperature. This study adds to the growing body of work indicating pronounced heat avoidance as a thermoregulatory strategy among tropical reptile populations.

Comparison of home range size, habitat use and the influence of resource variations between two species of greater gliders (Petauroides minor and Petauroides volans).

Knowledge of the spatial requirements of a species is fundamental to understanding its environmental requirements. However, this can be challenging as the size of a species' home range can be influenced by ecological factors such as diet and size-dependent metabolic demands, as well as factors related to the quality of their habitat such as the density and distribution of resources needed for food and shelter. Until recently, the genus Petauroides was thought to include only a single species with a widespread distribution across eastern Australia. However, a recent study has provided genetic and morphological evidence supporting Petauroides minor as a distinct northern species. Previous studies have focused on the ecology of P. volans, but there has been inadequate research on P. minor. Data on home range and habitat use were obtained for both species using a combination of techniques including GPS collar locations, radiotelemetry, and spotlighting and comparisons were made using consistent methodology. Home range sizes of P. minor (4.79 ha ± 0.97 s.d., KUD .95) were significantly larger than those of P. volans (2.0 ha ± 0.42 s.d., KUD .95). There were no significant differences between male and female home range sizes in either species. Both species showed site-specific preferences for tree species and for larger diameter trees for both forage and shelter. Tree size and biomass/ha were significantly greater in the P. volans study sites than the P. minor study sites and there was a negative correlation between home range size and eucalypt biomass. Larger home range size is likely driven by the substantial differences in biomass between northern (tropical) and southern (temperate) eucalypt-dominated habitats affecting the quality and quantity of resources for food and shelter. Understanding landscape use and habitat requirements within each species of Petauroides can provide important information regarding limiting factors and in directing conservation and management planning.

Invasive wild deer exhibit environmental niche shifts in Australia: Where to from here?

Invasive species have established populations around the world and, in the process, characteristics of their realized environmental niches have changed. Because of their popularity as a source of game, deer have been introduced to, and become invasive in, many different environments around the world. As such, deer should provide a good model system in which to test environmental niche shifts. Using the current distributions of the six deer species present in Australia, we quantified shifts in their environmental niches that occurred since introduction; we determined the differences in suitable habitat between their international (native and invaded) and their Australian ranges. Given knowledge of their Australian habitat use, we then modeled the present distribution of deer in Australia to assess habitat suitability, in an attempt to predict future deer distributions. We show that the Australian niches of hog (Axis porcinus), fallow (Dama dama), red (Cervus elaphus), rusa (C. timorensis), and sambar deer (C. unicolor), but not chital deer (A. axis), were different to their international ranges. When we quantified the potential range of these six species in Australia, chital, hog, and rusa deer had the largest areas of suitable habitat outside their presently occupied habitat. The other three species had already expanded outside the ranges that we predicted as suitable. Here, we demonstrate that deer have undergone significant environmental niche shifts following introduction into Australia, and these shifts are important for predicting the future spread of these invasive species. It is important to note that current Australian and international environmental niches did not necessarily predict range expansions, thus wildlife managers should treat these analyses as conservative estimates.

The role of ecdysis in repair of an attachment system: a case study using geckos.

Skin provides functions such as protection and prevention of water loss. In some taxa, the outer surface of skin has been modified to form structures that enable attachment to various surfaces. Constant interaction with surfaces is likely to cause damage to these attachment systems and reduce function. It seems logical that when skin is shed via ecdysis, its effectiveness will increase, through repair of damage or other rejuvenating mechanisms. We address two questions using three diplodactylid geckos as model species. (1) Does repeated mechanical damage affect clinging ability in geckos to the point that they cannot support their own body weight? (2) Does use without induced damage reduce effectiveness of the attachment system, and if so, does ecdysis restore clinging ability? We found that repeated damage reduced clinging ability in all three species, although at different rates. Additionally, use reduced clinging ability over time when no apparent damage was incurred. Clinging ability increased after ecdysis in all three species, both when damage was specially induced, and when it was not. After normal use without induced damage, the increase in clinging ability after ecdysis was statistically significant in two of three species. Our findings show that use decreases clinging ability, and mechanical damage also effects geckos' capacity to exert shear forces consistently. Thus, ecdysis improves clinging ability both in scenarios where damage is induced and more generally. In addition to the physiological functions provided by skin, our study highlights an important function of ecdysis in a speciose vertebrate group.

Constant-temperature predictions underestimate growth of a fungal amphibian pathogen under individual host thermal profiles.

Ectotherm body temperatures fluctuate with environmental variability and host behavior, which may influence host-pathogen interactions. Fungal pathogens are a major threat to ectotherms and may be highly responsive to the fluctuating thermal profiles of individual hosts, especially cool-loving fungi exposed to high host temperatures. However, most studies estimate pathogen thermal performance based on averages of host or surrogate environmental temperatures, potentially missing effects of short-term host temperature shifts such as daily or hourly heat spikes. We recorded individual thermal profiles of Australian rainforest frogs using temperature-sensitive radio-transmitters. We then reproduced a subset of individual thermal profiles in growth chambers containing cultures of the near-global amphibian pathogen Batrachochytrium dendrobatidis (Bd) to investigate how realistic host temperature profiles affect Bd growth. We focused on thermal profiles that exceed the thermal optimum of Bd because the effects of realistic heat spikes on Bd growth are unresolved. Our laboratory incubation experiment revealed that Bd growth varied in response to relatively small differences in heat spike characteristics of individual frog thermal profiles, such as a single degree or a few hours, highlighting the importance of individual host behaviors in predicting population-level disease dynamics. The fungus also grew better than predicted under the most extreme and unpredictable frog temperature profile, recovering from two days of extreme (nearly 32 °C) heat spikes without negative effects on overall growth, suggesting we are underestimating the growth potential of the pathogen in nature. Combined with the previous finding that Bd reduces host heat tolerance, our study suggests that this pathogen may carry a competitive edge over hosts in the face of anthropogenic climate change.

Pathogenesis of Bohle iridovirus infection in Krefft's freshwater turtle hatchlings (Emydura macquarii krefftii).

Ranaviruses have been detected in over 12 families of reptiles including many genera of turtles, tortoises, and terrapins, but the pathogenesis of these infections is still poorly understood. Krefft's river turtle hatchlings (N = 36; Emydura macquarii krefftii) were inoculated intramuscularly with Bohle iridovirus (BIV, Ranavirus, isolate) or saline, and euthanized at 9 timepoints (3 infected and 1 control per timepoint) over a 24-day period. Samples of lung, liver, kidney, and spleen were collected for quantitative polymerase chain reaction (PCR); internal organs, skin, and oral cavity samples were fixed for histopathological examination. The earliest lesions, at 8 days postinoculation (dpi), were lymphocytic inflammation of the skin and fibrinoid necrosis of regional vessels at the site of inoculation, and mild ulcerative necrosis with lymphocytic and heterophilic inflammation in the oral, nasal, and tongue mucosae. Fibrinonecrotic foci with heterophilic inflammation were detected in spleen and gonads at 16 dpi. Multifocal hepatic necrosis, heterophilic inflammation, and occasional basophilic intracytoplasmic inclusion bodies were observed at 20 dpi, along with ulcerative lymphocytic and heterophilic tracheitis and bronchitis. Tracheitis, bronchitis, and rare bone marrow necrosis were present at 24 dpi. Of the viscera tested for ranaviral DNA by PCR, the liver and spleen had the highest viral loads throughout infection, and thus appeared to be major targets of viral replication. Testing of whole blood by qPCR was the most-effective ante-mortem method for detecting ranaviral infection compared with oral swabs. This study represents the first time-dependent pathogenesis study of a ranaviral infection in turtles.

Dancing to a different tune: changing reproductive seasonality in an introduced chital deer population.

Male and female reproductive behaviour is typically synchronised. In species such as those in the family Cervidae, reproductive timing is often cued by photoperiod, although in females, it can be dependent on body condition. When a species is introduced to a novel environment, the environment changes, or responses of the sexes to such cues differ, asynchronous reproductive behaviour between males and females may occur. We investigated the seasonality of reproductive behaviour in introduced chital deer in northern Queensland by examining male antler phase in relation to female conception rates. We then analysed the influence of different variables likely to affect the timing of male and female reproductive physiology. The lowest percentage of chital in hard antler in any 1 month in this study was 35% (Fig. 1), but the average value was closer to 50%, thus there was a seasonal peak in antler phase linked with photoperiod. Females conceived at any time of year, but were strongly influenced by the amount of rainfall 3 months prior to conception. This resulted in varying conception peaks year-to-year that often did not correspond to the male's peak in hard antler. In this system, a proportion of males and females were physiologically and behaviourally ready to mate at any time of the year. We predict that differences in the timing of the peaks between the males and females will lead to increased reproductive skew (variation in reproductive success among individual males). This pattern may select for different mating strategies or physiological mechanisms to increase reproductive success. Fig. 1 The average percentage of male chital deer in hard antler by month from 2014 to 2019 in north Queensland. Values above the bars indicate the total number of males that were sampled in each month and the error bars indicate the standard error. In the month with the lowest % males in hard antler in the entire study (November, 2017), 35% of males were in hard antler.

Variation in density, but not morphology, of cutaneous sensilla among body regions in nine species of Australian geckos.

Skin sense organs, cutaneous sensilla, are a well-known feature of the integument of squamate reptiles and particularly geckos. They vary widely in morphology among species and are thought to be mechanosensitive, associated with prey capture and handling, tail autotomy and placement of the adhesive toepads in pad-bearing species. Some authors suggest that they may also sense abiotic environmental features, such as temperature or humidity. Here, we describe the morphology and distribution of cutaneous sensilla among body regions of nine Australian gecko species, in four genera. We hypothesised that if sensilla morphology was distinct, or sensilla density high, around the mouth, on the tail and on extremities, sensilla were likely used for these direct tactile functions. We found that sensilla morphology was uniform among body regions within species, but varied among species, while sensilla densities varied among species and body regions. In all species studied, sensilla density was highest on the labials and the dorsal tail scales and low on the feet, head and body, providing strong support for the hypothesis that sensilla serve tactile mechanoreceptive functions for prey capture and handling and for predator avoidance, but not for toepad placement. We suggest sensilla density may be explained by mechanoreception, whereas structure may be influenced by other factors.

Using citizen science to test for acoustic niche partitioning in frogs.

The acoustic niche hypothesis proposes that to avoid interference with breeding signals, vocal species should evolve to partition acoustic space, minimising similarity with co-occurring signals. Tests of the acoustic niche hypothesis are typically conducted using a single assemblage, with mixed outcomes, but if the process is evolutionarily important, a pattern of reduced acoustic competition should emerge, on average, over many communities. Using a continental-scale dataset derived from audio recordings collected by citizen scientists, we show that frogs do partition acoustic space. Differences in calls were predominately caused by differences in spectral, rather than temporal, features. Specifically, the 90% frequency bandwidths of observed frog assemblages overlapped less than expected, and there was greater distance between dominant frequencies than expected. To our knowledge, this study is the first to use null models to test for acoustic niche partitioning over a large geographic scale.

Increased rates of dispersal of free-ranging cane toads (Rhinella marina) during their global invasion.

Invasions often accelerate through time, as dispersal-enhancing traits accumulate at the expanding range edge. How does the dispersal behaviour of individual organisms shift to increase rates of population spread? We collate data from 44 radio-tracking studies (in total, of 650 animals) of cane toads (Rhinella marina) to quantify distances moved per day, and the frequency of displacement in their native range (French Guiana) and two invaded areas (Hawai'i and Australia). We show that toads in their native-range, Hawai'i and eastern Australia are relatively sedentary, while toads dispersing across tropical Australia increased their daily distances travelled from 20 to 200 m per day. That increase reflects an increasing propensity to change diurnal retreat sites every day, as well as to move further during each nocturnal displacement. Daily changes in retreat site evolved earlier than did changes in distances moved per night, indicating a breakdown in philopatry before other movement behaviours were optimised to maximise dispersal.

Leech removal is not the primary driver of basking behavior in a freshwater turtle.

Leaving the water to bask (usually in the sun) is a common behavior for many freshwater turtles, with some species also engaging in "nocturnal basking." Ectoparasite removal is an obvious hypothesis to explain nocturnal basking and has also been proposed as a key driver of diurnal basking. However, the efficacy of basking, day or night, to remove leeches has not been experimentally tested. Therefore, we examined the number of leeches that were removed from Krefft's river turtles (Emydura macquarii krefftii) after experimentally making turtles bask at a range of times of day, durations, and temperatures. Turtles had high initial leech loads, with a mean of 32.1 leeches per turtle. Diurnal basking under a heat lamp for 3 hr at ~28°C significantly reduced numbers of leeches relative to controls. In diurnal trials, 90.9% of turtles lost leeches (mean loss of 7.1 leeches per turtle), whereas basking for 30 min under the same conditions was not effective (no turtles lost leeches, and all turtles were still visibly wet). Similarly, "nocturnal basking" at ~23°C for 3 hr was not effective at removing leeches. Only 18% of turtles lost leeches (one turtle lost one leech and another lost four leeches). Diurnal basking outdoors under direct sunlight for 20 min (mean temp = 34.5°C) resulted in a small reduction in leeches, with 50% of turtles losing leeches and an average loss of 0.7 leeches per turtle. These results indicate basking can remove leeches if temperatures are high or basking durations are long. However, it was only effective at unusually long basking durations in this system. Our data showed even the 20-min period was longer than 70.1% of natural diurnal basking events, many of which took place at cooler temperatures. Therefore, leech removal does not appear to be the purpose of the majority of basking events.

Population growth lags in introduced species.

When introduced to new ecosystems, species' populations often grow immediately postrelease. Some introduced species, however, maintain a low population size for years or decades before sudden, rapid population growth is observed. Because exponential population growth always starts slowly, it can be difficult to distinguish species experiencing the early phases of slow exponential population growth (inherent lags) from those with actively delayed growth rates (prolonged lags). Introduced ungulates provide an excellent system in which to examine lags, because some introduced ungulate populations have demonstrated rapid population growth immediately postintroduction, while others have not. Using studies from the literature, we investigated which exotic ungulate species and populations (n = 36) showed prolonged population growth lags by comparing the doubling time of real ungulate populations to those predicted from exponential growth models for theoretical populations. Having identified the specific populations that displayed prolonged lags, we examined the impacts of several environmental and biological variables likely to influence the length of lag period. We found that seventeen populations (47%) showed significant prolonged population growth lags. We could not, however, determine the specific factors that contributed to the length of these lag phases, suggesting that these ungulate populations' growth is idiosyncratic and difficult to predict. Introduced species that exhibit delayed growth should be closely monitored by managers, who must be proactive in controlling their growth to minimize the impact such populations may have on their environment.

Geckos cling best to, and prefer to use, rough surfaces.

BACKGROUND: Fitness is strongly related to locomotor performance, which can determine success in foraging, mating, and other critical activities. Locomotor performance on different substrates is likely to require different abilities, so we expect alignment between species' locomotor performance and the habitats they use in nature. In addition, we expect behaviour to enhance performance, such that animals will use substrates on which they perform well. METHODS: We examined the associations between habitat selection and performance in three species of Oedura geckos, including two specialists, (one arboreal, and one saxicolous), and one generalist species, which used both rocks and trees. First, we described their microhabitat use in nature (tree and rock type) for these species, examined the surface roughnesses they encountered, and selected materials with comparable surface microtopographies (roughness measured as peak-to-valley heights) to use as substrates in lab experiments quantifying behavioural substrate preferences and clinging performance. RESULTS: The three Oedura species occupied different ecological niches and used different microhabitats in nature, and the two specialist species used a narrower range of surface roughnesses compared to the generalist. In the lab, Oedura geckos preferred substrates (coarse sandpaper) with roughness characteristics similar to substrates they use in nature. Further, all three species exhibited greater clinging performance on preferred (coarse sandpaper) substrates, although the generalist used fine substrates in nature and had good performance capabilities on fine substrates as well. CONCLUSION: We found a relationship between habitat use and performance, such that geckos selected microhabitats on which their performance was high. In addition, our findings highlight the extensive variation in surface roughnesses that occur in nature, both among and within microhabitats.

Skin hydrophobicity as an adaptation for self-cleaning in geckos.

Hydrophobicity is common in plants and animals, typically caused by high relief microtexture functioning to keep the surface clean. Although the occurrence and physical causes of hydrophobicity are well understood, ecological factors promoting its evolution are unclear. Geckos have highly hydrophobic integuments. We predicted that, because the ground is dirty and filled with pathogens, high hydrophobicity should coevolve with terrestrial microhabitat use. Advancing contact-angle (ACA) measurements of water droplets were used to quantify hydrophobicity in 24 species of Australian gecko. We reconstructed the evolution of ACA values, in relation to microhabitat use of geckos. To determine the best set of structural characteristics associated with the evolution of hydrophobicity, we used linear models fitted using phylogenetic generalized least squares (PGLS), and then model averaging based on AICc values. All species were highly hydrophobic (ACA > 132.72°), but terrestrial species had significantly higher ACA values than arboreal ones. The evolution of longer spinules and smaller scales was correlated with high hydrophobicity. These results suggest that hydrophobicity has coevolved with terrestrial microhabitat use in Australian geckos via selection for long spinules and small scales, likely to keep their skin clean and prevent fouling and disease.

Infection dynamics, dispersal, and adaptation: understanding the lack of recovery in a remnant frog population following a disease outbreak.

Emerging infectious diseases can cause dramatic declines in wildlife populations. Sometimes, these declines are followed by recovery, but many populations do not recover. Studying differential recovery patterns may yield important information for managing disease-afflicted populations and facilitating population recoveries. In the late 1980s, a chytridiomycosis outbreak caused multiple frog species in Australia's Wet Tropics to decline. Populations of some species (e.g., Litoria nannotis) subsequently recovered, while others (e.g., Litoria dayi) did not. We examined the population genetics and current infection status of L. dayi, to test several hypotheses regarding the failure of its populations to recover: (1) a lack of individual dispersal abilities has prevented recolonization of previously occupied locations, (2) a loss of genetic variation has resulted in limited adaptive potential, and (3) L. dayi is currently adapting to chytridiomycosis. We found moderate-to-high levels of gene flow and diversity (Fst range: <0.01-0.15; minor allele frequency (MAF): 0.192-0.245), which were similar to previously published levels for recovered L. nannotis populations. This suggests that dispersal ability and genetic diversity do not limit the ability of L. dayi to recolonize upland sites. Further, infection intensity and prevalence increased with elevation, suggesting that chytridiomycosis is still limiting the elevational range of L. dayi. Outlier tests comparing infected and uninfected individuals consistently identified 18 markers as putatively under selection, and several of those markers matched genes that were previously implicated in infection. This suggests that L. dayi has genetic variation for genes that affect infection dynamics and may be undergoing adaptation.

Impacts of artificial light on food intake in invasive toads.

Artificial light at night (ALAN) is a major form of anthropogenic disturbance. ALAN attracts nocturnal invertebrates, which are a food source for nocturnal predators, including invasive species. Few studies quantify the effects of increased food availablity by ALAN on invasive vertebrate predators, and enhancement of food intake caused by ALAN may also be influenced by various environmental factors, such as proximitity to cities, moon phase, temperature, rainfall and wind speed. Revealing the potential impacts on invasive predators of ALAN-attracted invertebrates, and the influence of other factors on these effects, could provide important insights for the management of these predators. We constructed and supplied with artificial light field enclosures for invasive toads, and placed them at locations with different levels of ambient light pollution, in northeastern Australia. In addition, we determined the effect of rainfall, temperature, wind speed, and lunar phase on food intake in toads. We found that ALAN greatly increased the mass of gut contents of invasive toads compared to controls, but that the effect was increased in dark lunar phases, and when there were low ambient light pollution levels. Effects of rainfall, temperature and wind speed on food intake were comparatively weak. To avoid providing food resources to toads, management of ALAN in rural areas, and during dark lunar phases may be advisable. On the contrary, to effectively capture toads, trapping using lights as lures at such times and places should be more successful.

Nonlinear variation in clinging performance with surface roughness in geckos.

Understanding the challenges faced by organisms moving within their environment is essential to comprehending the evolution of locomotor morphology and habitat use. Geckos have developed adhesive toe pads that enable exploitation of a wide range of microhabitats. These toe pads, and their adhesive mechanisms, have typically been studied using a range of artificial substrates, usually significantly smoother than those available in nature. Although these studies have been fundamental in understanding the mechanisms of attachment in geckos, it is unclear whether gecko attachment simply gradually declines with increased roughness as some researchers have suggested, or whether the interaction between the gekkotan adhesive system and surface roughness produces nonlinear relationships. To understand ecological challenges faced in their natural habitats, it is essential to use test surfaces that are more like surfaces used by geckos in nature. We tested gecko shear force (i.e., frictional force) generation as a measure of clinging performance on three artificial substrates. We selected substrates that exhibit microtopographies with peak-to-valley heights similar to those of substrates used in nature, to investigate performance on a range of smooth surfaces (glass), and fine-grained (fine sandpaper) to rough (coarse sandpaper). We found that shear force did not decline monotonically with roughness, but varied nonlinearly among substrates. Clinging performance was greater on glass and coarse sandpaper than on fine sandpaper, and clinging performance was not significantly different between glass and coarse sandpaper. Our results demonstrate that performance on different substrates varies, probably depending on the underlying mechanisms of the adhesive apparatus in geckos.

Spinal arthritis in invasive cane toads is linked to rate of dispersal as well as to latitude.

Initial research on the spread of cane toads (Rhinella marina) through tropical Australia reported a high incidence of spinal arthritis (spondylosis) in toads at the invasion front (where toads disperse rapidly), but not in areas colonized earlier (where toads are more sedentary). The idea that spondylosis was a cost of rapid dispersal was challenged by wider spatial sampling which linked rates of spondylosis to hot (tropical) climates rather than to dispersal rates. Here, the authors of these competing interpretations collaborate to reinterpret the data. Our reanalysis supports both previous hypotheses; rates of spondylosis are higher in populations established by fast-dispersing toads, and are higher in tropical than in temperate environments; they are also higher in larger toads. The functional reason for climatic effects is unclear, but might involve effects on the soil-living bacteria involved in the induction of spondylosis; and/or may reflect higher movement (as opposed to dispersal) or more pronounced dry-season aggregation rates of toads in tropical conditions.

Dose-dependent morbidity of freshwater turtle hatchlings, Emydura macquarii krefftii, inoculated with Ranavirus isolate (Bohle iridovirus, Iridoviridae).

Ranaviral infections cause mass die-offs in wild and captive turtle populations. Two experimental studies were performed to first determine the susceptibility of an Australian turtle species (Emydura macquarii krefftii) to different routes of infection and second examine the effect of viral titre on the morbidity in hatchlings. All inoculation routes (intracoelomic, intramuscular and oral) produced disease, but the clinical signs, histopathology and time to onset of disease varied with the route. The median infectious and lethal doses for intramuscularly inoculated hatchlings were 102.52 (1.98-2.93) and 104.43 (3.81-5.19) TCID50 ml-1, respectively. Clinical signs began 14 to 29 days post-inoculation and the median survival time was 22 days (16-25) across all dose groups. For every 10-fold increase in dose, the odds of developing any clinical signs or severe clinical signs increased by 3.39 [P<0.01, 95 % confidence interval (CI): 1.81-6.36] and 3.71 (P<0.01, 95 % CI: 1.76-7.80), respectively. Skin lesions, previously only reported in ranaviral infection in lizards, were observed in the majority of intramuscularly inoculated hatchlings that developed ranaviral disease. The histological changes were consistent with those in previous reports for reptiles and consisted of necrosis at or near the site of injection, in the spleen, liver and oral cavity. Systemic inflammation was also observed, predominantly affecting necrotic organs. The estimates reported here can be used to model ranaviral disease and quantify and manage at-risk populations.