Allorecognition in the Tasmanian devil (Sarcophilus harrisii), an endangered marsupial species with limited genetic diversity

Tasmanian devils (Sarcophilus harrisii) are on the verge of extinction due to a transmissible cancer, devil facial tumour disease (DFTD). This tumour is an allograft that is transmitted between individuals without immune recognition of the tumour cells. The mechanism to explain this lack of immune recognition and acceptance is not well understood. It has been hypothesized that lack of genetic diversity at the Major Histocompatibility Complex (MHC) allowed the tumour cells to grow in genetically similar hosts without evoking an immune response to alloantigens. We conducted mixed lymphocyte reactions and skin grafts to measure functional MHC diversity in the Tasmanian devil population. The limited MHC diversity was sufficient to produce measurable mixed lymphocyte reactions. There was a wide range of responses, from low or no reaction to relatively strong responses. The highest responses occurred when lymphocytes from devils from the east of Tasmania were mixed with lymphocytes from devils from the west of Tasmania. All of the five successful skin allografts were rejected within 14 days after surgery, even though little or no MHC I and II mismatches were found. Extensive T-cell infiltration characterised the immune rejection. We conclude that Tasmanian devils are capable of allogeneic rejection. Consequently, a lack of functional allorecognition mechanisms in the devil population does not explain the transmission of a contagious cancer.     

MHC gene copy number variation in Tasmanian devils: implications for the spread of a contagious cancer

Tasmanian devils face extinction owing to the emergence of a contagious cancer. Devil facial tumour disease (DFTD) is a clonal cancer spread owing to a lack of major histocompatibility complex (MHC) barriers in Tasmanian devil populations. We present a comprehensive screen of MHC diversity in devils and identify 25 MHC types and 53 novel sequences, but conclude that overall levels of MHC diversity at the sequence level are low. The majority of MHC Class I variation can be explained by allelic copy number variation with two to seven sequence variants identified per individual. MHC sequences are divided into two distinct groups based on sequence similarity. DFTD cells and most devils have sequences from both groups. Twenty per cent of individuals have a restricted MHC repertoire and contain only group I or only group II sequences. Counterintuitively, we postulate that the immune system of individuals with a restricted MHC repertoire may recognize foreign MHC antigens on the surface of the DFTD cell. The implication of these results for management of DFTD and this endangered species are discussed.     

Low major histocompatibility complex diversity in the Tasmanian devil predates European settlement and may explain susceptibility to disease epidemics

The Tasmanian devil (Sarcophilus harrisii) is at risk of extinction owing to the emergence of a contagious cancer known as devil facial tumour disease (DFTD). The emergence and spread of DFTD has been linked to low genetic diversity in the major histocompatibility complex (MHC). We examined MHC diversity in historical and ancient devils to determine whether loss of diversity is recent or predates European settlement in Australia. Our results reveal no additional diversity in historical Tasmanian samples. Mainland devils had common modern variants plus six new variants that are highly similar to existing alleles. We conclude that low MHC diversity has been a feature of devil populations since at least the Mid-Holocene and could explain their tumultuous history of population crashes.     

Infectious disease and sickness behaviour: tumour progression affects interaction patterns and social network structure in wild Tasmanian devils

Infectious diseases, including transmissible cancers, can have a broad range of impacts on host behaviour, particularly in the latter stages of disease progression. However, the difficulty of early diagnoses makes the study of behavioural influences of disease in wild animals a challenging task. Tasmanian devils (Sarcophilus harrisii) are affected by a transmissible cancer, devil facial tumour disease (DFTD), in which tumours are externally visible as they progress. Using telemetry and mark–recapture datasets, we quantify the impacts of cancer progression on the behaviour of wild devils by assessing how interaction patterns within the social network of a population change with increasing tumour load. The progression of DFTD negatively influences devils'' likelihood of interaction within their network. Infected devils were more active within their network late in the mating season, a pattern with repercussions for DFTD transmission. Our study provides a rare opportunity to quantify and understand the behavioural feedbacks of disease in wildlife and how they may affect transmission and population dynamics in general.     

Seasonal,demographic and density‐related patterns of contact between Tasmanian devils (Sarcophilus harrisii): Implications for transmission of devil facial tumour disease

Abstract Devil facial tumour disease (DFTD), is an emerging infectious cancer thought to be spread by biting. It is causing ongoing, severe population decline of the Tasmanian devil (Sarcophilus harrisii), the largest surviving marsupial carnivore and there are concerns that DFTD may lead to extinction of the devil. Whether extinction is likely depends on contact rates and their relationship to host density. We investigated contact rates using two different datasets. The first consisted of field observations of contact and biting behaviour around prey carcasses and, the second was a 3‐year longitudinal series of injuries in a marked devil population. During feeding interactions at carcasses, contact rates were significantly positively associated with population density and subadults delivered more bites than adult males and females. Injuries from the marked devil population did not differ between adult males and females. In two of the three years, penetrating biting (resulting in injury) increased markedly during the mating season and was more frequent in adults than in subadults. Among injured devils with wounds penetrating the dermal layer, adults were more frequently bitten in the head (the location of primary tumours) in the mating season than in other seasons, and had more head bites than subadults. Our results suggest that the mating season may be the key period for disease transmission. If most penetrating bites occur during mating interactions, DFTD transmission is likely to be frequency dependent, which means that there would be no threshold host density for disease persistence, and disease‐induced extinction is possible.     

Vincristine Chemotherapy Trials and Pharmacokinetics in Tasmanian Devils with Tasmanian Devil Facial Tumor Disease

Tasmanian Devil Facial Tumor Disease (DFTD) is a transmissible cancer threatening to cause the extinction of Tasmanian Devils in the wild. The aim of this study was to determine the susceptibility of the DFTD to vincristine. Escalating dosage rates of vincristine (0.05 to 0.136 mg/kg) were given to Tasmanian devils in the early stages of DFTD (n = 8). None of these dosage rates impacted the outcome of the disease. A dosage rate of 0.105 mg/kg, a rate significantly higher than that given in humans or domestic animals, was found to the highest dosage rate that could be administered safely. Signs of toxicity included anorexia, vomiting, diarrhea and neutropenia. Pharmacokinetic studies showed that, as with other species, there was a rapid drop in blood concentration following a rapid intravenous infusion with a high volume of distribution (1.96 L/kg) and a relatively long elimination half life (11 h). Plasma clearance (1.8 ml/min/kg) was slower in the Tasmanian devil than in humans, suggesting that pharmacodynamics and not pharmacokinetics explain the Tasmanian devil’s ability to tolerate high dosage rates of vincristine. While providing base-line data for the use of vincristine in Tasmanian devils and possibly other marsupials with vincristine susceptible cancers, these findings strongly suggest that vincristine will not be effective in the treatment of DFTD.     

Extreme Telomere Length Dimorphism in the Tasmanian Devil and Related Marsupials Suggests Parental Control of Telomere Length

Telomeres, specialised structures that protect chromosome ends, play a critical role in preserving chromosome integrity. Telomere dynamics in the Tasmanian devil (Sarcophilus harrisii) are of particular interest in light of the emergence of devil facial tumour disease (DFTD), a transmissible malignancy that causes rapid mortality and threatens the species with extinction. We used fluorescent in situ hybridisation to investigate telomere length in DFTD cells, in healthy Tasmanian devils and in four closely related marsupial species. Here we report that animals in the Order Dasyuromorphia have chromosomes characterised by striking telomere length dimorphism between homologues. Findings in sex chromosomes suggest that telomere length dimorphism may be regulated by events in the parental germlines. Long telomeres on the Y chromosome imply that telomere lengthening occurs during spermatogenesis, whereas telomere diminution occurs during oogenesis. Although found in several somatic cell tissue types, telomere length dimorphism was not found in DFTD cancer cells, which are characterised by uniformly short telomeres. This is, to our knowledge, the first report of naturally occurring telomere length dimorphism in any species and suggests a novel strategy of telomere length control. Comparative studies in five distantly related marsupials and a monotreme indicate that telomere dimorphism evolved at least 50 million years ago.     

Comparative landscape genetics reveals differential effects of environment on host and pathogen genetic structure in Tasmanian devils (Sarcophilus harrisii) and their transmissible tumour

Genetic structure in host species is often used to predict disease spread. However, host and pathogen genetic variation may be incongruent. Understanding landscape factors that have either concordant or divergent influence on host and pathogen genetic structure is crucial for wildlife disease management. Devil facial tumour disease (DFTD) was first observed in 1996 and has spread throughout almost the entire Tasmanian devil geographic range, causing dramatic population declines. Whereas DFTD is predominantly spread via biting among adults, devils typically disperse as juveniles, which experience low DFTD prevalence. Thus, we predicted little association between devil and tumour population structure and that environmental factors influencing gene flow differ between devils and tumours. We employed a comparative landscape genetics framework to test the influence of environmental factors on patterns of isolation by resistance (IBR) and isolation by environment (IBE) in devils and DFTD. Although we found evidence for broad‐scale costructuring between devils and tumours, we found no relationship between host and tumour individual genetic distances. Further, the factors driving the spatial distribution of genetic variation differed for each. Devils exhibited a strong IBR pattern driven by major roads, with no evidence of IBE. By contrast, tumours showed little evidence for IBR and a weak IBE pattern with respect to elevation in one of two tumour clusters we identify herein. Our results warrant caution when inferring pathogen spread using host population genetic structure and suggest that reliance on environmental barriers to host connectivity may be ineffective for managing the spread of wildlife diseases. Our findings demonstrate the utility of comparative landscape genetics for identifying differential factors driving host dispersal and pathogen transmission.     

Transmissible cancer in Tasmanian devils: localized lineage replacement and host population response

Tasmanian devil facial tumour disease (DFTD) is a clonally transmissible cancer threatening the Tasmanian devil (Sarcophilus harrisii) with extinction. Live cancer cells are the infectious agent, transmitted to new hosts when individuals bite each other. Over the 18 years since DFTD was first observed, distinct genetic and karyotypic sublineages have evolved. In this longitudinal study, we investigate the associations between tumour karyotype, epidemic patterns and host demographic response to the disease. Reduced host population effects and low DFTD infection rates were associated with high prevalence of tetraploid tumours. Subsequent replacement by a diploid variant of DFTD coincided with a rapid increase in disease prevalence, population decline and reduced mean age of the population. Our results suggest a role for tumour genetics in DFTD transmission dynamics and epidemic outcome. Future research, for this and other highly pathogenic emerging infectious diseases, should focus on understanding the evolution of host and pathogen genotypes, their effects on susceptibility and tolerance to infection, and their implications for designing novel genetic management strategies. This study provides evidence for a rapid localized lineage replacement occurring within a transmissible cancer epidemic and highlights the possibility that distinct DFTD genetic lineages may harbour traits that influence pathogen fitness.     

Towards a Case Definition for Devil Facial Tumour Disease: What Is It?

In the mid 1990s an emerging disease characterised by the development of proliferative lesions around the face of Tasmanian devils (Sarcophilus harrisii) was observed. A multi-disciplinary approach was adopted to define the condition. Histopathological and transmission electron microscopic examination combined with immunohistochemistry help define Devil Facial Tumour Disease (DFTD) as a neoplastic condition of cells of neuroendocrine origin. Cytogenetic analysis of neoplastic tissue revealed it to be markedly different from normal devil tissue and having a consistent karyotype across all tumours examined. Combined with evidence for Major histocompatability (MHC) gene analysis there is significant evidence to confirm the tumour is a transmissible neoplasm.     

Evidence that disease-induced population decline changes genetic structure and alters dispersal patterns in the Tasmanian devil

Infectious disease has been shown to be a major cause of population declines in wild animals. However, there remains little empirical evidence on the genetic consequences of disease-mediated population declines, or how such perturbations might affect demographic processes such as dispersal. Devil facial tumour disease (DFTD) has resulted in the rapid decline of the Tasmanian devil, Sarcophilus harrisii, and threatens to cause extinction. Using 10 microsatellite DNA markers, we compared genetic diversity and structure before and after DFTD outbreaks in three Tasmanian devil populations to assess the genetic consequences of disease-induced population decline. We also used both genetic and demographic data to investigate dispersal patterns in Tasmanian devils along the east coast of Tasmania. We observed a significant increase in inbreeding (F_IS pre/post-disease −0.030/0.012, P<0.05; relatedness pre/post-disease 0.011/0.038, P=0.06) in devil populations after just 2–3 generations of disease arrival, but no detectable change in genetic diversity. Furthermore, although there was no subdivision apparent among pre-disease populations (θ=0.005, 95% confidence interval (CI) −0.003 to 0.017), we found significant genetic differentiation among populations post-disease (θ=0.020, 0.010–0.027), apparently driven by a combination of selection and altered dispersal patterns of females in disease-affected populations. We also show that dispersal is male-biased in devils and that dispersal distances follow a typical leptokurtic distribution. Our results show that disease can result in genetic and demographic changes in host populations over few generations and short time scales. Ongoing management of Tasmanian devils must now attempt to maintain genetic variability in this species through actions designed to reverse the detrimental effects of inbreeding and subdivision in disease-affected populations.     

Genomic restructuring in the Tasmanian devil facial tumour: chromosome painting and gene mapping provide clues to evolution of a transmissible tumour

Devil facial tumour disease (DFTD) is a fatal, transmissible malignancy that threatens the world's largest marsupial carnivore, the Tasmanian devil, with extinction. First recognised in 1996, DFTD has had a catastrophic effect on wild devil numbers, and intense research efforts to understand and contain the disease have since demonstrated that the tumour is a clonal cell line transmitted by allograft. We used chromosome painting and gene mapping to deconstruct the DFTD karyotype and determine the chromosome and gene rearrangements involved in carcinogenesis. Chromosome painting on three different DFTD tumour strains determined the origins of marker chromosomes and provided a general overview of the rearrangement in DFTD karyotypes. Mapping of 105 BAC clones by fluorescence in situ hybridisation provided a finer level of resolution of genome rearrangements in DFTD strains. Our findings demonstrate that only limited regions of the genome, mainly chromosomes 1 and X, are rearranged in DFTD. Regions rearranged in DFTD are also highly rearranged between different marsupials. Differences between strains are limited, reflecting the unusually stable nature of DFTD. Finally, our detailed maps of both the devil and tumour karyotypes provide a physical framework for future genomic investigations into DFTD.     

Isotopic niche variation in Tasmanian devils Sarcophilus harrisii with progression of devil facial tumor disease

Devil facial tumor disease (DFTD) is a transmissible cancer affecting Tasmanian devils Sarcophilus harrisii. The disease has caused severe population declines and is associated with demographic and behavioral changes, including earlier breeding, younger age structures, and reduced dispersal and social interactions. Devils are generally solitary, but social encounters are commonplace when feeding upon large carcasses. DFTD tumors can disfigure the jaw and mouth and so diseased individuals might alter their diets to enable ingestion of alternative foods, to avoid conspecific interactions, or to reduce competition. Using stable isotope analysis (δ¹³C and δ¹⁵N) of whiskers, we tested whether DFTD progression, measured as tumor volume, affected the isotope ratios and isotopic niches of 94 infected Tasmanian devils from six sites in Tasmania, comprising four eucalypt plantations, an area of smallholdings and a national park. Then, using tissue from 10 devils sampled before and after detection of tumors and 8 devils where no tumors were detected, we examined whether mean and standard deviation of δ¹³C and δ¹⁵N of the same individuals changed between healthy and diseased states. δ¹³C and δ¹⁵N values were generally not related to tumor volume in infected devils, though at one site, Freycinet National Park, δ¹⁵N values increased significantly as tumor volume increased. Infection with DFTD was not associated with significant changes in the mean or standard deviation of δ¹³C and δ¹⁵N values in individual devils sampled before and after detection of tumors. Our analysis suggests that devils tend to maintain their isotopic niche in the face of DFTD infection and progression, except where ecological conditions facilitate a shift in diets and feeding behaviors, demonstrating that ecological context, alongside disease severity, can modulate the behavioral responses of Tasmanian devils to DFTD.     

A preliminary study assessing risk to Tasmanian devils from poisoning for red foxes

The recent introduction of red foxes (Vulpes vulpes) to Australia's island state of Tasmania represents a major threat to native fauna. In response, the Tasmanian government has begun a fox eradication program using Foxoff®, a bait containing the poison sodium monofluoroacetate (commonly known as 1080). The bait is potentially attractive to native Tasmanian carnivores as well as to foxes. Of particular concern is the endangered Tasmanian devil (Sarcophilus harrisii), which is already at risk from an emergent infectious disease, devil facial tumor disease (DFTD). In both a captive and a field study using non-toxic Foxoff bait, we assessed bait palatability and possible effects of demographics, hunger level, bait age, and bait burial method on the likelihood of bait uptake by Tasmanian devils. Captive devils showed varying interest in the bait, but wild devils appeared to find it uniformly palatable. In the captive study, males and younger, captive-born animals were more likely to excavate and remove bait. Subterranean burial at 15 cm was the most effective deterrent to bait excavation; effectiveness decreased at shallower depths and with surface-level bait buried beneath soil mounds. Our findings suggest that the current fox-baiting campaign may negatively impact individual devils. More extensive study is necessary to assess potential risk at the population level. © 2011 The Wildlife Society.     

Conservation implications of limited genetic diversity and population structure in Tasmanian devils (<Emphasis Type="Italic">Sarcophilus harrisii</Emphasis>)

Tasmanian devils face a combination of threats to persistence, including devil facial tumor disease (DFTD), an epidemic transmissible cancer. We used RAD sequencing to investigate genome-wide patterns of genetic diversity and geographic population structure. Consistent with previous results, we found very low genetic diversity in the species as a whole, and we detected two broad genetic clusters occupying the northwestern portion of the range, and the central and eastern portions. However, these two groups overlap across a broad geographic area, and differentiation between them is modest (\({{F}_{\text{ST}}}\)?=?0.1081). Our results refine the geographic extent of the zone of mixed ancestry and substructure within it, potentially informing management of genetic variation that existed in pre-diseased populations of the species. DFTD has spread across both genetic clusters, but recent evidence points to a genomic response to selection imposed by DFTD. Any allelic variation for resistance to DFTD may be able to spread across the devil population under selection by DFTD, and/or be present as standing variation in both genetic regions.     

Landscape genetics of the Tasmanian devil: implications for spread of an infectious cancer

Emerging infectious diseases are increasingly recognized in species’ declines and extinctions. Landscape genetics can be used as a tool to predict disease emergence and spread. The Tasmanian devil is threatened with extinction by a nearly 100% fatal transmissible cancer, which has spread across 95% of the species’ geographic range in 20 years. Here, we present a landscape genetic analysis in the last remaining uninfected parts of the Tasmanian devil’s geographic range to: describe population genetic structure, characterize genetic diversity, and test the influence of landscape variables on Tasmanian devil gene flow to assess the potential for disease spread. In contrast to previous genetic studies on Tasmanian devils showing evidence for two genetic populations island-wide, our genetic based assignment tests and spatial principal components analyses suggest at least two, and possibly three, populations in a study area that is approximately 15% of the size of the overall species’ geographic range. Positive spatial autocorrelation declined at about 40 km, in contrast to 80 km in eastern populations, highlighting the need for range-wide genetic studies. Strong genetic structure was found between devils in the northern part of the study area and those found south of Macquarie Harbor, with weaker structure found between the northeastern and northwestern portion of our study area. Consistent with previous work, we found low overall genetic diversity, likely owing to a combination of founder effects and extreme weather events thousands of years ago that likely caused large-scale population declines. We also found possible signs of recent bottlenecks, perhaps resulting from forest clearing for dairy farming in the central part of the study area. This human disturbance also may have contributed to weak genetic structuring detected between the northeastern and northwestern part of the study area. Individual-based least cost path modeling showed limited influence of landscape variables on gene flow, with weak effects of variation in elevation in the northeast. In the northwest, however, landscape genetic models did not perform better than the null isolation-by-distance model. At the larger spatial scale of the northern part of the study area, elevation and temperatures were negatively correlated with gene flow, consistent with low dispersal suitability of higher elevation habitats that have lower temperatures and dense, wet vegetation. Overall, Tasmanian devils are a highly vagile species for which dispersal and gene flow appear to be influenced little by landscape features, and spread of devil facial tumor disease to the remaining portion of the devil’s geographic range seems imminent. Nonetheless, strong genetic structure found between the northern and southern portions of our study area, combined with low densities and limited possible colonization of DFTD from the east suggest there is some time for implementation of management strategies.     

Contagious cancer: lessons from the devil and the dog

Cancer is generally defined as uncontrollable growth of cells caused by genetic aberrations and/or environmental factors. Yet contagious cancers also occur. The recent emergence of a contagious cancer in Tasmanian devils has reignited interest in transmissible cancers. Two naturally occurring transmissible cancers are known: devil facial tumour disease and canine transmissible venereal tumour. Both cancers evolved once and have then been transmitted from one individual to another as clonal cell lines. The dog cancer is ancient; having evolved more than 6,000 years ago, while the devil disease was first seen in 1996. In this review I will compare and contrast the two diseases focusing on the life histories of the clonal cell lines, their evolutionary trajectories and the mechanisms by which they have achieved immune tolerance. A greater understanding of these contagious cancers will provide unique insights into the role of the immune system in shaping tumour evolution and may uncover novel approaches for treating human cancer.     

Infection of the fittest: devil facial tumour disease has greatest effect on individuals with highest reproductive output

Emerging infectious diseases rarely affect all members of a population equally and determining how individuals’ susceptibility to infection is related to other components of their fitness is critical to understanding disease impacts at a population level and for predicting evolutionary trajectories. We introduce a novel state‐space model framework to investigate survival and fecundity of Tasmanian devils (Sarcophilus harrisii) affected by a transmissible cancer, devil facial tumour disease. We show that those devils that become host to tumours have otherwise greater fitness, with higher survival and fecundity rates prior to disease‐induced death than non‐host individuals that do not become infected, although high tumour loads lead to high mortality. Our finding that individuals with the greatest reproductive value are those most affected by the cancer demonstrates the need to quantify both survival and fecundity in context of disease progression for understanding the impact of disease on wildlife populations.     

A review of parasites in the Tasmanian devil (<Emphasis Type="Italic">Sarcophilus harrisii</Emphasis>)

Threatened by devil facial tumour disease, the Tasmanian devil (Sarcophilus harrisii), a carnivorous marsupial confined to Tasmania, Australia, is the subject of conservation management under the Save the Tasmanian Devil Program. Conservation actions such as captive breeding and translocation may impact upon parasite ecology, presenting risk of increased disease through stress and impaired immunity, and by exposing hosts to parasites to which they are immunologically naïve. Given the importance of parasites to ecosystem function, it has been argued from a biodiversity perspective that parasites should be conserved in their own right. In this review we describe current knowledge, and limitations in our knowledge, of Tasmanian devil parasites. We then discuss the potential for changes in host–parasite interactions as a result of host-population decline and conservation management, both generally and with examples from the Tasmanian devil. The review closes with a recommendation for a systematic evaluation of parasites in captive and wild devils to aid conservation of this host–parasite system in its entirety.     

Space use and temporal partitioning of sympatric Tasmanian devils and spotted‐tailed quolls

Sympatric species can minimise interspecific competition by spatial avoidance or by altering their temporal activity to reduce encounter rates. The Tasmanian devil (Sarcophilus harrisii), the largest carnivorous marsupial, coexists with the smaller spotted‐tailed quoll (Dasyurus maculatus) in Tasmania, Australia. Quolls may be susceptible to interspecific competition from devils, because they utilise similar habitats, consume similar prey species and are displaced by devils at food sources. Such competition might cause quolls to spatially or temporally avoid devils. To investigate whether spatial or temporal avoidance occurred, we deployed GPS collars on sympatric devils and quolls and conducted a camera survey at a site in northwest Tasmania where the devil population was not affected by devil facial tumour disease. GPS tracking coincided with the lactation period when devils and quolls had young in dens and continued until weaning occurred. We found little spatial segregation of home range and core area placement between devils and quolls and among devils. Quolls showed more spatial segregation within the sexes than between them. Devils had larger home ranges than quolls. Male devils had larger home ranges than females, but there was no difference in home range size between the sexes of quolls. Females of both species travelled significantly further per night than did males. There was moderate temporal partitioning between the two species: devil activity peaked after dusk and devils remained active until the early morning, while quoll activity showed distinct peaks around dusk and dawn. In conclusion, quolls did not spatially avoid devils but moderate temporal partitioning occurred. It is plausible that quolls are active at different times of the diel cycle to reduce encountering devils, but further studies are needed to resolve the cause of this temporal partitioning.