A step-by-step method to quantify coloration with digital photography.

Coloration is often used in biological studies, for example when studying social signaling or antipredator defense. Yet, few detailed and standardized methods are available to measure coloration using digital photography. Here we provide a step-by-step guide to help researchers quantify coloration from digital images. We first identify the do's and don'ts of taking pictures for coloration analysis. We then describe how to i) extract reflectance values with the software ImageJ; ii) fit and apply linearization equations to reflectance values; iii) scale and select the areas of interest in ImageJ; iv) standardize pictures; and v) binarize and measure the proportion of different colors in an area of interest. We apply our methodological protocol to digital pictures of painted turtles (Chrysemys picta), but the approach could be easily adapted to any species. More specifically, we wished to calculate the proportion of red and yellow on the neck and head of turtles. With this protocol, our main aims are to make coloration analyses with digital photography:•More accessible to researchers without a background in photography.•More consistent between studies.

Effects of ophidiomycosis on movement, survival, and reproduction of eastern foxsnakes (Pantherophis vulpinus).

Ophidiomycosis (snake fungal disease) is caused by the fungal pathogen Ophidiomyces ophidiicola, which causes dermal lesions, occasional systemic infections, and in some cases, mortality. To better understand potential conservation implications of ophidiomycosis (i.e., population-level effects), we investigated its impacts on individual fitness in a population of endangered eastern foxsnakes (Pantherophis vulpinus). We tracked 38 foxsnakes over 6 years and quantified body condition, movement patterns, oviposition rates, and survival. Body condition, distance travelled, and oviposition rates were similar between snakes with and without ophidiomycosis. Interestingly, snakes that tested positive for the pathogen travelled farther, suggesting that movement through a greater diversity of habitats increases risk of exposure. Ophidiomycosis did not negatively affect survival, and most apparently infected snakes persisted in a manner comparable to snakes without ophidiomycosis. Only one mortality was directly attributed to ophidiomycosis, although infected snakes were overrepresented in a sample of snakes killed by predators. Overall, our results suggest that ophidiomycosis may have sublethal effects on eastern foxsnakes, but do not suggest direct effects on survival, ovipositioning, or viability of the study population.

SEASONAL AND INTERSPECIFIC VARIATION IN THE PREVALENCE OF OPHIDIOMYCES OPHIDIICOLA AND OPHIDIOMYCOSIS IN A COMMUNITY OF FREE-RANGING SNAKES.

Ophidiomycosis in snakes is caused by the fungus Ophidiomyces ophidiicola. Clinical signs associated with the disease range from minor skin lesions to severe swelling of the face. In some cases, the fungus invades the snake's underlying muscle and bone and internal organs; disease severity appears to peak during brumation. We quantified the prevalence of O. ophidiicola and ophidiomycosis in free-ranging snakes to explore seasonal variation in detection of the pathogen and disease. We collected skin swabs (n=464 samples) from seven species of free-ranging snakes (n=336) from Rondeau Provincial Park (Ontario, Canada) and tested the swabs for O. ophidiicola using quantitative PCR. We also assessed individuals for lesions consistent with ophidiomycosis and monitored changes in gross lesions over time in recaptured individuals. Eastern foxsnakes (Pantherophis vulpinus) had the highest prevalence of O. ophidiicola (24/84) and of lesions consistent with ophidiomycosis (34/84). On other species (Nerodia sipedon, Storeria dekayi, Thamnophis sirtalis, and Thamnophis sauritus), we detected the pathogen on only 4/229 snakes and observed gross lesions consistent with ophidiomycosis on 24/229 snakes. Body length of eastern foxsnakes was associated with detection of O. ophidiicola, suggesting that eastern foxsnakes' large size increases the risk of pathogen exposure relative to the other, smaller, species at our study site. Ophidiomyces ophidiicola and lesions consistent with ophidiomycosis were detected most frequently in eastern foxsnakes soon after emergence from brumation and less frequently later in the active season (O. ophidiicola: April=29.8%, October=3.9%; lesions: April=36.1%, October=5.5%). This decrease corresponded with resolution of lesions in 6/13 resampled eastern foxsnakes. Considering the seasonal cycle of O. ophidiicola and ophidiomycosis when planning disease surveillance research may improve detection probabilities for ophidiomycosis in Nearctic snakes.

Revisiting Ophidiomycosis (Snake Fungal Disease) After a Decade of Targeted Research.

Emerging infectious diseases (EIDs) are typically characterized by novelty (recent detection) and by increasing incidence, distribution, and/or pathogenicity. Ophidiomycosis, also called snake fungal disease, is caused by the fungus Ophidiomyces ophidiicola (formerly "ophiodiicola"). Ophidiomycosis has been characterized as an EID and as a potential threat to populations of Nearctic snakes, sparking over a decade of targeted research. However, the severity of this threat is unclear. We reviewed the available literature to quantify incidence and effects of ophidiomycosis in Nearctic snakes, and to evaluate whether the evidence supports the ongoing characterization of ophidiomycosis as an EID. Data from Canada remain scarce, so we supplemented the literature review with surveys for O. ophidiicola in the Canadian Great Lakes region. Peer-reviewed reports of clinical signs consistent with ophidiomycosis in free-ranging, Nearctic snakes date back to at least 1998, and retrospective molecular testing of samples extend the earliest confirmed record to 1986. Diagnostic criteria varied among publications (n = 33), confounding quantitative comparisons. Ophidiomycosis was diagnosed or suspected in 36/121 captive snakes and was fatal in over half of cases (66.7%). This result may implicate captivity-related stress as a risk factor for mortality from ophidiomycosis, but could also reflect reporting bias (i.e., infections are more likely to be detected in captive snakes, and severe cases are more likely to be reported). In contrast, ophidiomycosis was diagnosed or suspected in 441/2,384 free-ranging snakes, with mortality observed in 43 (9.8 %). Ophidiomycosis was only speculatively linked to population declines, and we found no evidence that the prevalence of the pathogen or disease increased over the past decade of targeted research. Supplemental surveys and molecular (qPCR) testing in Ontario, Canada detected O. ophidiicola on 76 of 657 free-ranging snakes sampled across ~136,000 km2. The pathogen was detected at most sites despite limited and haphazard sampling. No large-scale mortality was observed. Current evidence supports previous suggestions that the pathogen is a widespread, previously unrecognized endemic, rather than a novel pathogen. Ophidiomycosis may not pose an imminent threat to Nearctic snakes, but further research should investigate potential sublethal effects of ophidiomycosis such as altered reproductive success that could impact population growth, and explore whether shifting environmental conditions may alter host susceptibility.

Transcriptional host-pathogen responses of Pseudogymnoascus destructans and three species of bats with white-nose syndrome.

Understanding how context (e.g., host species, environmental conditions) drives disease susceptibility is an essential goal of disease ecology. We hypothesized that in bat white-nose syndrome (WNS), species-specific host-pathogen interactions may partly explain varying disease outcomes among host species. We characterized bat and pathogen transcriptomes in paired samples of lesion-positive and lesion-negative wing tissue from bats infected with Pseudogymnoascus destructans in three parallel experiments. The first two experiments analyzed samples collected from the susceptible Nearctic Myotis lucifugus and the less-susceptible Nearctic Eptesicus fuscus, following experimental infection and hibernation in captivity under controlled conditions. The third experiment applied the same analyses to paired samples from infected, free-ranging Myotis myotis, a less susceptible, Palearctic species, following natural infection and hibernation (n = 8 sample pairs/species). Gene expression by P. destructans was similar among the three host species despite varying environmental conditions among the three experiments and was similar within each host species between saprophytic contexts (superficial growth on wings) and pathogenic contexts (growth in lesions on the same wings). In contrast, we observed qualitative variation in host response: M. lucifugus and M. myotis exhibited systemic responses to infection, while E. fuscus up-regulated a remarkably localized response. Our results suggest potential phylogenetic determinants of response to WNS and can inform further studies of context-dependent host-pathogen interactions.

Road avoidance and its energetic consequences for reptiles.

Roads are one of the most widespread human-caused habitat modifications that can increase wildlife mortality rates and alter behavior. Roads can act as barriers with variable permeability to movement and can increase distances wildlife travel to access habitats. Movement is energetically costly, and avoidance of roads could therefore impact an animal's energy budget. We tested whether reptiles avoid roads or road crossings and explored whether the energetic consequences of road avoidance decreased individual fitness. Using telemetry data from Blanding's turtles (Emydoidea blandingii; 11,658 locations of 286 turtles from 15 sites) and eastern massasaugas (Sistrurus catenatus; 1,868 locations of 49 snakes from 3 sites), we compared frequency of observed road crossings and use of road-adjacent habitat by reptiles to expected frequencies based on simulated correlated random walks. Turtles and snakes did not avoid habitats near roads, but both species avoided road crossings. Compared with simulations, turtles made fewer crossings of paved roads with low speed limits and more crossings of paved roads with high speed limits. Snakes made fewer crossings of all road types than expected based on simulated paths. Turtles traveled longer daily distances when their home range contained roads, but the predicted energetic cost was negligible: substantially less than the cost of producing one egg. Snakes with roads in their home range did not travel further per day than snakes without roads in their home range. We found that turtles and snakes avoided crossing roads, but road avoidance is unlikely to impact fitness through energetic expenditures. Therefore, mortality from vehicle strikes remains the most significant impact of roads on reptile populations.

Tree lizard (Urosaurus ornatus) growth decreases with population density, but increases with habitat quality.

Habitat selection models can explain spatial patterns in the relative abundance of animals in different habitats based on the assumption that fitness declines as density in a habitat increases. Ectotherms, such as lizards, may not follow predictions of density-dependent habitat selection models because temperature, which is unaffected by density, strongly influences their habitat selection. If competition for limited resources decreases fitness, then crowding should cause a decrease in body size and growth rates. We used skeletochronology and body size data from tree lizards (Urosaurus ornatus) at six sites that each spanned two habitats varying in quality to test the hypothesis that habitat selection is density dependent because growth is limited by competition for resources and by habitat quality. First, we tested that the maximum body size of lizards decreased with higher densities in a habitat by comparing growth between sites. Second, we tested whether body size and growth were higher in the habitat with more resources by controlling for density in a habitat and comparing growth between habitats in different sites. We found evidence of density-dependent growth in females, but not in males. Females in more crowded sites reached a smaller maximum size. Females in the higher quality habitat also grew larger than females in the lower quality habitat after controlling for differences in density between the habitats. Therefore, we found partial support for our hypothesis that competition for resources limits growth and causes density-dependent habitat selection.

Do ectotherms partition thermal resources? We still do not know.

Partitioning of the niche space is a mechanism used to explain the coexistence of similar species. Ectotherms have variable body temperatures and their body temperatures influence performance and, ultimately, fitness. Therefore, many ectotherms use behavioral thermoregulation to avoid reduced capacities associated with body temperatures far from the optimal temperature for performance. Several authors have proposed that thermal niche partitioning in response to interspecific competition is a mechanism that allows the coexistence of similar species of ectotherms. We reviewed studies on thermal resource partitioning to evaluate the evidence for this hypothesis. In almost all studies, there was insufficient evidence to conclude unequivocally that thermal resource partitioning allowed species coexistence. Future studies should include sites where species are sympatric and sites where they are allopatric to rule out alternative mechanisms that cause differences in thermal traits between coexisting species. There is evidence of conservatism in the evolution of most thermal traits across a wide range of taxa, but thermal performance curves and preferred temperatures do respond to strong selection under laboratory conditions. Thus, there is potential for selection to act on thermal traits in response to interspecific competition. Nevertheless, more stringent tests of the thermal resource partitioning hypothesis are required before we can assess whether it is widespread in communities of ectotherms in nature.