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Aim We explored lineage diversification within desert-dwelling fauna. Our goals were (1) to determine whether phylogenetic lineages and population expansions were consistent with younger Pleistocene climate fluctuation hypotheses or much older events predicted by pre-Pleistocene vicariance hypotheses, (2) to assess concordance in spatial patterns of genetic divergence and diversity among species and (3) to identify regional evolutionary hotspots of divergence and diversity and assess their conservation status. Location Mojave, Colorado, and Sonoran Deserts, USA. Methods We analysed previously published gene sequence data for twelve species. We used Bayesian gene tree methods to estimate lineages and divergence times. Within each lineage, we tested for population expansion and age of expansion using coalescent approaches. We mapped interpopulation genetic divergence and intra-population genetic diversity in a GIS to identify hotspots of highest genetic divergence and diversity and to assess whether protected lands overlapped with evolutionary hotspots. Results In seven of the 12 species, lineage divergence substantially predated the Pleistocene. Historical population expansion was found in eight species, but expansion events postdated the Last Glacial Maximum (LGM) in only four. For all species assessed, six hotspots of high genetic divergence and diversity were concentrated in the Colorado Desert, along the Colorado River and in the Mojave/Sonoran ecotone. At least some proportion of the land within each recovered hotspot was categorized as protected, yet four of the six also overlapped with major areas of human development. Main conclusions Most of the species studied here diversified into distinct Mojave and Sonoran lineages prior to the LGM – supporting older diversification hypotheses. Several evolutionary hotspots were recovered but are not strategically paired with areas of protected land. Long-term preservation of species-level biodiversity would entail selecting areas for protection in Mojave and Sonoran Deserts to retain divergent genetic diversity and ensure connectedness across environmental gradients.

Wildlife translocation is increasingly used to mitigate disturbances to animals or habitat due to human activities, yet little is known about the extent to which translocating animals causes stress. To understand the relationship between physiological stress and translocation, we conducted a multiyear study (2007–2009) using a population of desert tortoises (Gopherus agassizii) near Fort Irwin, California. Blood samples were collected from adult tortoises in three treatment groups (resident, translocated and control) for 1 year prior to and 2 years after translocation. Samples were analyzed by radioimmunoassay for plasma total corticosterone (CORT), a glucocorticoid hormone commonly associated with stress responses in reptiles. CORT values were analyzed in relation to potential covariates (animal sex, date, behavior, treatment, handling time, air temperature, home-range size, precipitation and annual plant production) among seasons and years. CORT values in males were higher than in females, and values for both varied monthly throughout the activity season and among years. Year and sex were strong predictors of CORT, and translocation explained little in terms of CORT. Based on these results, we conclude that translocation does not elicit a physiological stress response in desert tortoises.

We translocated 120 Agassiz's desert tortoises to 5 sites in Nevada and Utah to evaluate the effects of translocation on tortoise survivorship, reproduction, and habitat use. Translocation sites included several elevations, and extended to sites with vegetation assemblages not typically associated with desert tortoises in order to explore the possibility of moving animals to upper elevation areas. We measured survivorship, reproduction, and movements of translocated and resident animals at each site. Survivorship was not significantly different between translocated and resident animals within and among sites, and survivorship was greater overall during non-drought years. The number of eggs produced by tortoises was similar for translocated and resident females, but differed among sites. Animals translocated to atypical habitat generally moved until they reached vegetation communities more typical of desert tortoise habitat. Even within typical tortoise habitat, tortoises tended to move greater distances in the first year after translocation than did residents, but their movements in the second or third year after translocation were indistinguishable from those of resident tortoises. Our data show that tortoises translocated into typical Mojave desert scrub habitats perform well; however, the large first-year movements of translocated tortoises have important management implications. Projects that employ translocations must consider how much area will be needed to contain translocated tortoises and whether roads need fencing to prevent the loss of animals. 2012 The Wildlife Society.

Heterogeneity in habitat often influences how organisms traverse the landscape matrix that connects populations. Understanding landscape connectivity is important to determine the ecological processes that influence those movements, which lead to evolutionary change due to gene flow. Here, we used landscape genetics and statistical models to evaluate hypotheses that could explain isolation among locations of the threatened Mojave desert tortoise (Gopherus agassizii). Within a causal modeling framework, we investigated three factors that can influence landscape connectivity: geographic distance, barriers to dispersal, and landscape friction. A statistical model of habitat suitability for the Mojave desert tortoise, based on topography, vegetation, and climate variables, was used as a proxy for landscape friction and barriers to dispersal. We quantified landscape friction with least-cost distances and with resistance distances among sampling locations. A set of diagnostic partial Mantel tests statistically separated the hypotheses of potential causes of genetic isolation. The best-supported model varied depending upon how landscape friction was quantified. Patterns of genetic structure were related to a combination of geographic distance and barriers as defined by least-cost distances, suggesting that mountain ranges and extremely low-elevation valleys influence connectivity at the regional scale beyond the tortoises’ ability to disperse. However, geographic distance was the only influence detected using resistance distances, which we attributed to fundamental differences between the two ways of quantifying friction. Landscape friction, as we measured it, did not influence the observed patterns of genetic distances using either quantification. Barriers and distance may be more valuable predictors of observed population structure for species like the desert tortoise, which has high dispersal capability and a long generation time.

Understanding predator-prey relationships can be pivotal in the conservation of spe- cies. For 2 decades, desert tortoise Gopherus agassizii populations have declined, yet quantitative evidence regarding the causes of declines is scarce. In 2005, Ft. Irwin National Training Center, California, USA, implemented a translocation project including 2 yr of baseline monitoring of desert tortoises. Unusually high predation on tortoises was observed after translocation occurred. We conducted a retrospective analysis of predation and found that translocation did not affect the proba- bility of predation: translocated, resident, and control tortoises all had similar levels of predation. However, predation rates were higher near human population concentrations, at lower elevation sites, and for smaller tortoises and females. Furthermore, high mortality rates were not limited to the National Training Center. In 2008, elevated mortality (as high as 43%) occurred throughout the listed range of the desert tortoise. Although no temporal prey base data are available for analysis from any of the study sites, we hypothesize that low population levels of typical coyote Canis latrans prey (i.e. jackrabbits Lepus californicus and other small animals) due to drought conditions influenced high predation rates in previous years. Predation may have been exacerbated in areas with high levels of subsidized predators. Many historical reports of increased predation, and our observation of a range- wide pattern, may indicate that high predation rates are more common than generally considered and may impact recovery of the desert tortoise throughout its range.

Hypolithic microbes, primarily cyanobacteria, inhabit the highly specialized microhabitats under translucent rocks in extreme environments. Here we report findings from hypolithic cyanobacteria found under three types of translucent rocks (quartz, prehnite, agate) in a semiarid region of tropical Australia. We investigated the photosynthetic responses of the cyanobacterial communities to light, temperature and moisture in the laboratory, and we measured the microclimatic variables of temperature and soil moisture under rocks in the field over an annual cycle. We also used molecular techniques to explore the diversity of hypolithic cyanobacteria in this community and their phylogenetic relationships within the context of hypolithic cyanobacteria from other continents. Based on the laboratory experiments, photosynthetic activity required a minimum soil moisture of 15% (by mass). Peak photosynthetic activity occurred between approximately 8 degrees C and 42 degrees C, though some photosynthesis occurred between -1 degrees C and 51 degrees C. Maximum photosynthesis rates also occurred at light levels of approximately 150-550 micromol m(-2) s(-1). We used the field microclimatic data in conjunction with these measurements of photosynthetic efficiency to estimate the amount of time the hypolithic cyanobacteria could be photosynthetically active in the field. Based on these data, we estimated that conditions were appropriate for photosynthetic activity for approximately 942 h (approximately 75 days) during the year. The hypolithic cyanobacteria community under quartz, prehnite and agate rocks was quite diverse both within and between rock types. We identified 115 operational taxonomic units (OTUs), with each rock hosting 8-24 OTUs. A third of the cyanobacteria OTUs from northern Australia grouped with Chroococcidiopsis, a genus that has been identified from hypolithic and endolithic communities from the Gobi, Mojave, Atacama and Antarctic deserts. Several OTUs identified from northern Australia have not been reported to be associated with hypolithic communities previously.

Assessing the recovery of the federally listed Mojave population of desert tortoises Gopherus agassizii requires detecting subtle changes in population size over a period of many years. The methods that have been employed by the US Fish and Wildlife Service to estimate population density of desert tortoises are inadequate for detecting modest trends in population density, partly due to a hidden variance in a parameter (g0) that corrects for the proportion of tortoises that are inactive and thus unavailable for sampling when population density is being assessed. We used small dataloggers to record the activity of tortoises throughout their active season, and derived daily esti- mates of g0, which we compared with the 2004 estimate published by the US Fish and Wildlife Ser- vice. Due to the substantial variation in animal activity that we found within and among days during the active season, we used a daily method to estimate density. We found that this method, while pro- viding a more accurate assessment of g0, translates into estimates of density with coefficients of vari- ation that are 4" larger than previously reported due to the variation in g0 and n (number of animals encountered). This discrepancy could adversely influence managers' perceptions of population recovery for desert tortoises, and could undermine any ability to monitor the efficacy of recovery actions for populations.

The use of dogs (Canis lupus familiaris) trained to locate wildlife under natural conditions may increase the risk of attracting potential predators or alter behavior of target species. These potentially negative effects become even more problematic when dealing with threatened or endangered species, such as the Mojave Desert tortoise (Gopherus agassizii). We addressed three concerns regarding use of dogs trained to locate desert tortoises in the wild. First, we looked at the potential for dogs to attract native and non-native predators to sites at a greater rate than with human visitation alone by comparing presence of predator sign before and after visitation by dogs and by humans. We found no significant difference in predator sign based upon type of surveyor. Second, we looked at the difference in risk of predation to desert tortoises that were located in the wild by humans versus humans with wildlife-detector dogs. Over a 5-week period, during which tortoises were extensively monitored and a subsequent period of 1 year during which tortoises were monitored monthly, there was no predation on, nor sign of predator-inflicted trauma to tortoises initially encountered either by humans or wildlife-detector dogs. Third, we looked at movement patterns of tortoises after encounter by either humans or wildlife-detector dogs. Movement of desert tortoises was not significantly different after being found by a human versus being found by a wildlife-detector dog. Based upon these initial results we conclude that use of trained wildlife-detector dogs to survey for desert tortoises in the wild does not appear to increase attraction of predators, increase risk of predation, or alter movement patterns of desert tortoises more than surveys conducted by humans alone.

This research examined the onset, duration, and termination of hibernation in Desert Tortoises (Gopherus agassizii) over several years at multiple sites in the northeastern part of their geographic range, and recorded the temperatures experienced by tortoises during winter hibernation. The timing of hibernation by Desert Tortoises differed among sites and years. Environmental cues acting over the short-term did not appear to influence the timing of the hibernation period. Different individual tortoises entered hibernation over as many as 44 days in the fall and emerged from hibernation over as many as 49 days in the spring. This range of variation in the timing of hibernation indicates a weak influence at best of exogenous cues hypothesized to trigger and terminate hibernation. There do appear to be regional trends in hibernation behavior as hibernation tended to begin earlier and continue longer at sites that were higher in elevation and generally cooler. The emergence date was generally more similar among study sites than the date of onset. While the climate and the subsequent timing of hibernation differed among sites, the average temperatures experienced by tortoises while hibernating differed by only about five degrees from the coldest site to the warmest site.

The federally listed desert tortoise (Gopherus agassizii) is currently monitored using distance sampling to estimate population densities. Distance sampling, as with many other techniques for estimating population density, assumes that it is possible to quantify the proportion of animals available to be counted in any census. Because desert tortoises spend much of their life in burrows, and the proportion of tortoises in burrows at any time can be extremely variable, this assumption is difficult to meet. This proportion of animals available to be counted is used as a correction factor (g0) in distance sampling and has been estimated from daily censuses of small populations of tortoises (6-12 individuals). These censuses are costly and produce imprecise estimates of g0 due to small sample sizes. We used data on tortoise activity from a large (N = 1 50) experimental population to model activity as a function of the biophysical attributes of the environment, but these models did not improve the precision of estimates from the focal populations. Thus, to evaluate how much of the variance in tortoise activity is apparently not predictable, we assessed whether activity on any particular day can predict activity on subsequent days with essentially identical environmental conditions. Tortoise activity was only weakly correlated on consecutive days, indicating that behavior was not repeatable or consistent among days with similar physical environments.

For several species, refuges (such as burrows, dens, roosts, nests) are an essential resource for protection from predators and extreme environmental conditions. Refuges also serve as focal sites for social interactions including mating, courtship and aggression. Knowledge of refuge use patterns can therefore provide information about social structure, mating and foraging success, as well as the robustness and health of wildlife populations, especially for species considered to be relatively solitary. In this study, we construct networks of burrow use to infer social associations in a threatened wildlife species typically considered solitary - the desert tortoise. We show that tortoise social networks are significantly different than null networks of random associations, and have moderate spatial constraints. We next use statistical models to identify major mechanisms behind individual-level variation in tortoise burrow use, popularity of burrows in desert tortoise habitat and test for stressor-driven changes in refuge use patterns. We show that seasonal variation has a strong impact on tortoise burrow switching behavior. On the other hand, burrow age and topographical condition influence the number of tortoises visiting a burrow in desert tortoise habitat. Of three major population stressors affecting this species (translocation, drought, disease), translocation alters tortoise burrow switching behavior, with translocated animals visiting fewer unique burrows than residents. In a species that is not social, our study highlights the importance of leveraging refuge use behavior to study the presence of and mechanisms behind non-random social structure and individual-level variation. Our analysis of the impact of stressors on refuge-based social structure further emphasizes the potential of this method to detect environmental or anthropogenic disturbances.Significance statement: Adaptive and social behavior that affects fitness is now being increasingly incorporated in the conservation and management of wildlife species. However, direct observations of social interactions in species considered to be solitary are difficult, and therefore integration of behavior in conservation and management decisions in such species has been infrequent. For such species, we propose quantifying refuge use behavior as it can provide insights towards their (hidden) social structure, establish relevant contact patterns of infectious disease spread, and provide early warning signals of population stressors. Our study highlights this approach in a long-lived and threatened species, the desert tortoise. We provide evidence towards the presence of and identify mechanisms behind the social structure in desert tortoises formed by their burrow use preferences. We also show how individuals burrow use behavior responds to the presence of population stressors.

Most directly transmitted infections require some form of close contact between infectious and susceptible hosts to spread. Often disease models assume contacts are equal and use mean field estimates of transmission probability for all interactions with infectious hosts. Such methods may inaccurately describe transmission when interactions differ substantially in their ability to cause infection. Understanding this variation in transmission risk may be critical to properly model and manage some infectious diseases. In this study, we investigate how varying exposure and transmission may be key to understanding disease dynamics in the threatened desert tortoise Gopherus agassizii. We created heterogeneity in Mycoplasma agassizii exposure (the putative bacterial agent of a respiratory disease) by varying the duration of interactions between naturally infected and uninfected captive desert tortoises. Using qPCR, we identified new infections and compared models of transmission probability as a function of contact duration and pathogen load. We then examined the contact patterns of a wild tortoise population using proximity loggers to identify heterogeneity in contact duration. The top-ranked model predicting M. agassizii transmission included a dose term defined as the product of the number of days in proximity to an infected host and the infection level of that host. Models predicted low transmission probability for short interactions, unless the infectious host had a high load of M. agassizii: such hosts were predicted to transmit infection at higher rates with any amount of contact. We observed predominantly short-lived interactions in a free-ranging tortoise population and thus, expect transmission patterns in this population to vary considerably with the frequency and duration of high infection levels. Mean field models may misrepresent natural transmission patterns in this and other populations depending on the distribution of high-risk contact and shedding events. Rapid outbreaks in generally solitary species may result from changes to their naturally low-risk contact patterns or due to increases in the frequency of severe infections or super-shedding events - population characteristics that should be further investigated to develop effective management strategies.

Many of the threats to the persistence of populations of sensitive species have physiological or pathological mechanisms, and those mechanisms are best understood through the inherently integrative discipline of physiological ecology. The desert tortoise was listed under the Endangered Species Act largely due to a newly recognized upper respiratory disease thought to cause mortality in individuals and severe declines in populations. Numerous hypotheses about the threats to the persistence of desert tortoise populations involve acquisition of nutrients, and its connection to stress and disease. The nutritional wisdom hypothesis posits that animals should forage not for particular food items, but instead, for particular nutrients such as calcium and phosphorus used in building bones. The optimal foraging hypothesis suggests that, in circumstances of resource abundance, tortoises should forage as dietary specialists as a means of maximizing intake of resources. The optimal digestion hypothesis suggests that tortoises should process ingesta in ways that regulate assimilation rate. Finally, the cost-of-switching hypothesis suggests that herbivores, like the desert tortoise, should avoid switching food types to avoid negatively affecting the microbe community responsible for fermenting plants into energy and nutrients. Combining hypotheses into a resource acquisition theory leads to novel predictions that are generally supported by data presented here. Testing hypotheses, and synthesizing test results into a theory, provides a robust scientific alternative to the popular use of untested hypotheses and unanalyzed data to assert the needs of species. The scientific approach should focus on hypotheses concerning anthropogenic modifications of the environment that impact physiological processes ultimately important to population phenomena. We show how measurements of such impacts as nutrient starvation, can cause physiological stress, and that the endocrine mechanisms involved with stress can result in disease. Finally, our new syntheses evince a new hypothesis. Free molecules of the stress hormone corticosterone can inhibit immunity, and the abundance of "free corticosterone" in the blood (thought to be the active form of the hormone) is regulated when the corticosterone molecules combine with binding globulins. The sex hormone, testosterone, combines with the same binding globulin. High levels of testosterone, naturally occurring in the breeding season, may be further enhanced in populations at high densities, and the resulting excess testosterone may compete with binding globulins, thereby releasing corticosterone and reducing immunity to disease. This sequence could result in physiological and pathological phenomena leading to population cycles with a period that would be essentially impossible to observe in desert tortoise. Such cycles could obscure population fluctuations of anthropogenic origin.

Upper respiratory tract disease (URTD) caused by Mycoplasma agassizii is considered a threat to desert tortoise popula- tions that should be addressed as part of the recovery of the species. Clinical signs can be intermittent and include serous or mucoid nasal discharge and respiratory difficulty when nares are occluded. This nasal congestion may result in a loss of the olfactory sense. Turtles are known to use olfaction to identify food items, predators, and conspecifics; therefore, it is likely that URTD affects not only their physical well-being but also their behavior and ability to perform necessary functions in the wild. To determine more specifically the impact nasal discharge might have on free-ranging tortoises (Gopherus agassizii), we compared the re- sponses of tortoises with and without nasal discharge and both positive and negative for M. agassizii antibodies to a visually hidden olfac- tory food stimulus and an empty control. We found that nasal discharge did reduce sense of smell and hence the ability to locate food. Our study also showed that moderate chronic nasal discharge in the absence of other clinical signs did not affect appetite in desert tortoises.

Diet quality can influence many aspects of digestion, but the links between diet quality and resting metabolism are poorly understood. In nature, it might be beneficial to reduce energy expenditure when only poor quality diets are available. Alternatively, animals might increase the processing capacity of the gut to more thoroughly extract energy. If maintaining the processing capacity of the gut is energetically expensive, then increasing gut size or function might result in higher resting metabolism. In ectotherms, most digestive functions are temperature dependent, thus another strategy to maintain energy balance might be to alter selected body temperatures. We tested whether differing concentrations of dietary fiber affected the resting metabolic rate or body temperatures selected by chuckwallas (Sauromalus obesus) – lizards that naturally experience marked variation in dietary fiber. Resting metabolic rates measured at two temperatures and over three time intervals did not differ between groups of lizards force-fed low- (30% neutral-detergent fiber; NDF) and high-fiber (45% NDF) diets, nor did these diet differences influence body temperatures selected in a thermal gradient. We conclude that ecologically relevant differences in diet quality may have negligible effects on resting metabolic rates and body temperatures selected by chuckwallas.

Tortoises are susceptible to a wide variety of environmental stressors, and the influence of human disturbances on health and survival of tortoises is difficult to detect. As an addition to current diagnostic methods for desert tortoises, we have developed the first leukocyte gene transcription biomarker panel for the desert tortoise (Gopherus agassizii), enhancing the ability to identify specific environmental conditions potentially linked to declining animal health. Blood leukocyte transcript profiles have the potential to identify physiologically stressed animals in lieu of clinical signs. For desert tortoises, the gene transcript profile included a combination of immune or detoxification response genes with the potential to be modified by biological or physical injury and consequently provide information on the type and magnitude of stressors present in the animal's habitat. Blood from 64 wild adult tortoises at three sites in Clark County, NV, and San Bernardino, CA, and from 19 captive tortoises in Clark County, NV, was collected and evaluated for genes indicative of physiological status. Statistical analysis using a priori groupings indicated significant differences among groups for several genes, while multidimensional scaling and cluster analyses of transcription C T values indicated strong differentiation of a large cluster and multiple outlying individual tortoises or small clusters in multidimensional space. These analyses highlight the effectiveness of the gene panel at detecting environmental perturbations as well as providing guidance in determining the health of the desert tortoise.

Our experiments were designed to test the hypotheses that dietary lipids can affect whole-animal physiological processes in a manner concordant with changes in the fluidity of cell membranes. We measured (1) the lipid composition of five tissues, (2) body temperatures selected in a thermal gradient (T(sel)), (3) the body temperature at which the righting reflex was lost (critical thermal minimal [CTMin]), and (4) resting metabolic rate (RMR) at three body temperatures in desert iguanas (Dipsosaurus dorsalis) fed diets enriched with either saturated or unsaturated fatty acids. The composition of lipids in tissues of the lizards generally reflected the lipids in their diets, but the particular classes and ratios of fatty acids varied among sampled organs, indicating the conservative nature of some tissues (e.g., brain) relative to others (e.g., depot fat). Lizards fed the diet enriched with saturated fatty acids selected warmer nighttime body temperatures than did lizards fed a diet enriched with unsaturated fatty acids. This difference is concordant with the hypothesis that the composition of dietary fats influences membrane fluidity and that ectotherms may compensate for such changes in fluidity by selecting different body temperatures. The CTMin of the two treatment groups was indistinguishable. This may reflect the conservatism of some tissues (e.g., brain) irrespective of diet treatment. The RMR of the saturated treatment group nearly doubled between 30 degrees and 40 degrees C. Here, some discrete membrane domains in the lizards fed the saturated diet may have been in a more-ordered phase at 30 degrees C and then transformed to a less-ordered phase at 40 degrees C. In contrast, the RMR of the unsaturated treatment group exhibited temperature independence in metabolic rate from 30 degrees to 40 degrees C. Perhaps the unsaturated diet resulted in membranes that developed a higher degree of disorder (i.e., a certain phase) at a lower temperature than were membranes of lizards fed the saturated diet. Our study demonstrates links between dietary fats and whole-animal physiology; however, the mechanistic basis of these links, and the general knowledge of lipid metabolism in squamate reptiles, remain poorly understood and warrant further study.

Predicting wildfires that affect broad landscapes is important for allocating suppression resources and guiding land management. Wildfire prediction in the south-western United States is of specific concern because of the increasing prevalence and severe effects of fire on desert shrublands and the current lack of accurate fire prediction tools. We developed a fire risk model to predict fire occurrence in a north-eastern Mojave Desert landscape. First we developed a spatial model using remote sensing data to predict fuel loads based on field estimates of fuels. We then modelled fire risk (interactions of fuel characteristics and environmental conditions conducive to wildfire) using satellite imagery, our model of fuel loads, and spatial data on ignition potential (lightning strikes and distance to roads), topography (elevation and aspect) and climate (maximum and minimum temperatures). The risk model was developed during a fire year at our study landscape and validated at a nearby landscape; model performance was accurate and similar at both sites. This study demonstrates that remote sensing techniques used in combination with field surveys can accurately predict wildfire risk in the Mojave Desert and may be applicable to other arid and semiarid lands where wildfires are prevalent.

Spatially explicit decision support systems are assuming an increasing role in natural resource and conservation management. In order for these systems to be successful, however, they must address real-world management problems with input from both the scientific and management communities. The National Training Center at Fort Irwin, California, has expanded its training area, encroaching U.S. Fish and Wildlife Service critical habitat set aside for the Mojave desert tortoise (Gopherus agassizii), a federally threatened species. Of all the mitigation measures proposed to offset expansion, the most challenging to implement was the selection of areas most feasible for tortoise translocation. We developed an objective, open, scientifically defensible spatially explicit decision support system to evaluate translocation potential within the Western Mojave Recovery Unit for tortoise populations under imminent threat from military expansion. Using up to a total of 10 biological, anthropogenic, and/or logistical criteria, seven alternative translocation scenarios were developed. The final translocation model was a consensus model between the seven scenarios. Within the final model, six potential translocation areas were identified.