This protocol describes the quantification of heat transmission through a flat skinned avian specimen using a thermal camera and hot water bath. The method allows obtaining of quantitative, comparative data about the thermal performance of feather coats across species using dried flat skin specimens.
Feathers are essential to insulation, and therefore to the cost of thermoregulation, in birds. There is robust literature on the energetic cost of thermoregulation in birds across a variety of ecological circumstances. However, few studies characterize the contribution of the feathers alone to thermoregulation. Several previous studies have established methods for measuring the insulation value of animal pelts, but they require destructive sampling methods that are problematic for birds, whose feathers are not distributed evenly across the skin. More information is needed about 1) how the contribution of feathers to thermoregulation varies both across and within species and 2) how feather coats may change over space and time. Reported here is a method for rapidly and directly measuring the thermal performance of feather coats and the skin using dried whole skin specimens, without the need to destroy the skin specimen. This method isolates and measures the thermal gradient across a feather coat in a way that measurements of heat loss and metabolic cost in live birds, which use behavioral and physiological strategies to thermoregulate, cannot. The method employs a thermal camera, which allows the rapid collection of quantitative thermal data to measure heat loss from a stable source through the skin. This protocol can easily be applied to various research questions, is applicable to any avian taxa, and does not require destruction of the skin specimen. Finally, it will further the understanding of the importance of passive thermoregulation in birds by simplifying and accelerating the collection of quantitative data.
Feathers are the defining characteristic of birds and serve many functions, among the most crucial being insulation1. Birds have the highest average core temperatures of any vertebrate group, and feathers insulating them from environmental temperature changes are a vital part of energy balance, especially in cold environments2. Despite the importance of feathers, the majority of literature on changes in thermal condition in birds has focused on metabolic responses to temperature variation rather than the function of feathers as insulation3,4,5,6,7,8,9,10 (for further details, refer to Ward et al.)11,12,13. However, feathers themselves may vary across time, individuals, and species.
The method presented here is useful for quantifying the overall thermal value of the feather coat alone. It removes confounding factors in live birds, such as behavioral thermoregulation1 and varying amounts of insulating fat. More widespread measurement of the thermal performance of feather coats is necessary to improve the understanding of how feathers contribute to insulation and how this varies among and within species throughout a bird’s life history and annual cycle.
Feathers insulate birds by trapping air both between the skin and feathers as well as within the feathers, and they create a physical barrier to heat loss14. Feathers consist of a central feather shaft, called a rachis, with projections called barbs14. Barbules are small, secondary projections on barbs that interlock with adjacent barbs together to “zip” up the feather and give it structure. Furthermore, down feathers lack a central rachis and have few barbules, therefore forming a loose, insulative mass of barbs over the skin14. Feather coats vary across species15,16, within species17,18, and within comparable individuals2,19,20,21,22,23,24. However, there is little quantitative information about how variations in number of feathers, the relative abundance of different types of feathers on a bird, or changes in the numbers of barbs/barbules affect the overall thermal performance of a feather coat. Previous studies have focused on determining a single mean value of insulation and thermal conductivity for a given species11,12,13.
The feather coat is known to vary among species. For example, most birds have distinct areas of skin from which feathers do, or do not, grow called the pterylae and apteria, respectively14. The placement of the pterylae (sometimes called “feather tracts”) varies across species and has some value as a taxonomic character14. However, some birds (i.e., ratites and penguins) have lost this pterylosis and have a uniform distribution of feathers across the body14. Additionally, different species, especially those inhabiting different environments, have different proportions of feather types15. For example, birds inhabiting colder climates have more down feathers15 and contour feathers with a larger plumulaceous portion16 than species inhabiting warmer environments.
The microstructure of certain types of feathers may also have an effect on insulation across species25,26. Lei et al. compared the microstructure of contour feathers of many Chinese Passerine sparrows and found that species inhabiting colder environments have a higher proportion of plumulaceous barbs in each contour feather, longer barbules, higher node density, and larger nodes than species inhabiting warmer environments25. D’alba et al. compared the microstructure of down feathers of common eiders (Somateria mollissima) and graylag geese (Anser anser) and described how these differences affect both the cohesive ability of the feathers and ability of the feathers to trap air26. Quantitative comparative data about how these variations in feathering affect the overall thermal performance of the feather coat across species is limited (for more details, refer to Taylor and Ward et al.)11,13.
Within a species, the feather coat’s thermal performance may vary. Some species, such as the monk parakeet (Myiopsitta monachus)17, inhabit very large and diverse geographical ranges. The different thermal stresses posed by these different environments may affect the feather coats of birds within a species regionally, but there are currently no data available on this topic. Additionally, Broggi et al. compared two populations of great tits (Parus major L.) in the northern hemisphere. They demonstrated that contour feathers of the more northern population were denser but shorter and less proportionally plumulaceous than those of the more southern population. However, these differences disappeared when birds from both populations were raised in the same place18.
Furthermore, Broggi et al. explained these findings as a plastic response to differing thermal conditions, but they did not measure the insulation values of these different feather coats18. The results also suggest that contour feather density is more important to insulation than the proportion of plumulaceous barbs in contour feathers, but Broggi et al. suggested that northern populations may be unable to produce optimal feathers due to a lack of adequate nutrients18. Quantitative measurements of the overall thermal performance of these feather coats would further the understanding of the significance of plumage differences.
Over time, the feather coats of individual birds vary. At least once a year, all birds molt (replace all of their feathers)19. As the year goes on, feathers become worn2,20 and less numerous18,21,22,23. Some birds molt more than once a year, giving them multiple distinct feather coats each year19. Middleton showed that American goldfinches (Spinus tristis), which molt twice a year, have a higher number of feathers and higher proportion of downy feathers in their basic plumage in winter months than they do in their alternate plumage during summer months24. These annual differences in the feather coat may allow birds to conserve more heat during colder periods passively or shed more heat passively during warm seasons, but no studies have tested this conclusively.
Although birds thermoregulate behaviorally1,27 and can acclimate metabolically to different thermal conditions3,4,5,6,7,8,9,10,26, feathers play an important role in thermoregulation by providing a constant layer of insulation. The method described here is designed to answer questions about the feather coat alone and its role in passive thermoregulation (i.e., how much heat does a living bird retain without modifying its behavior or metabolism?) by isolating the feathers. While active and physiological thermoregulation are ecologically important, it is also important to understand how the feathers alone aid in insulation and how they influence the need for active behavioral and physiological thermoregulation.
Previous studies have established methods for quantifying thermal conductivity and thermal insulation of animal pelts11,12,13,28. The method presented here is an extension of the “guarded hot plate” method11,12,13,28. However, the method described here measures the temperature at the outer boundary of the feather coat using a thermal camera, rather than thermocouples. The guarded hot plate method gives very precise estimates of energy flow through a pelt, but it requires construction of a multi-material hot plate, some familiarity with the use of thermocouples and thermopiles, and destructive use of a pelt that must be cut into small pieces. These pieces are then greased to eliminate air between the sample and hot plate apparatus. With the exception of the few birds that lack apteria (e.g., penguins), cutting small squares from bird skins is problematic for comparative purposes, since the location of the cut has large effects on the number of feathers actually attached to (and overlying) the skin. This problem is exacerbated by the variation among taxa in the presence, size, and placement of ptyerlae14.
Furthermore, while museum specimens can be a potentially rich resource for assessing the variation in insulation among birds, in general, permission to cut and grease skin specimens in scientific collections is unattainable. Additionally, specimens taken from the wild for guarded hot plate measurements cannot be subsequently used as museum specimens. The method presented here differs from the guarded hot plate method in that it can be used with whole dried bird skins, without 1) requiring the destruction of the specimen and 2) greasing the underside of the skin. It uses thermal cameras, which are increasingly affordable (though still relatively expensive), precise, and used for live bird measurements of thermal relations.
This method does not measure energy flow (and therefore thermal conductivity or insulation value) through the skin and feathers directly as the guarded hot plate method does. Instead, it measures the temperature at the outer boundary of a feather coat using a thermal camera. The resulting values represent an integrated measure of the heat lost passively through the skin, feathers, and air trapped between them (compared to a heat source underneath). Specimens prepared as flat skins and measured using the described technique can be stored in collections, and indefinitely provide value for future research. This method provides a standardized, comparable, and relatively simple way to measure feather coat thermal performance in any flat skinned specimen, which is especially useful in inter- and intra-specific comparisons.
This work did not involve any work with live animals and was therefore exempt from animal care review.
1. Set-up and materials (Figure 1)
2. Performance of measurements
3. Data collection from thermal images
4. Calculation of thermal performance
Representative results from a series of one individual of each of five species, measured at six temperatures, are presented in Figure 4 and Figure 5. These show that small variations in the placement of the skin can result in variations in the readings of up to 1.7 °C. Figure 4 shows how training of an investigator increases repeatability of the measurements. For example, the same individual house sparrow (Passer domesticus) was measured five times at a single target temperature by an inexperienced investigator (Figure 4A). After training on a variety of specimens of different species, one investigator (J.G.) measured the same specimen five times at the same target temperature (Figure 4B). The estimate of the relationship between the temperature of the acrylic glass and temperature at the surface of the feathers changed by only a small (but perhaps important) amount. As a result, repeatability of the measurements themselves changed by almost four-fold. Repeated practice is therefore highly recommended for operators on a non-sample skin (before taking measurements that will be analyzed) until measurements converge and variation in the measurements stabilize (i.e., no further improvement in reproducibility is seen with additional practice). This is important to achieve before conducting analyses on repeated measures (or the mean of repeated measures) in each specimen.
The data shown in Figure 5 represents a small pilot sample but suggests that this method for measuring thermal performance of the feather coat is likely to yield important insight into the thermal ecology of birds. To reduce measurement error, only one investigator (J.G.) trained and took the measurements. Although these data represent only a single individual of each of the species listed (house sparrow, eastern phoebe [Sayornis phoebe], gray catbird [Dumetella carolinensis], eastern bluebird [Sialia sialis], and tufted titmouse [Baeolophus bicolor]), variation in the slopes of the resulting data demonstrates that the thermal performance of feather coats varies among these individuals. Furthermore, the magnitude of these differences suggests that the variation may be due to species differences.
Moreover, given that a single trained investigator performed all the measurements in Figure 5, investigator skill alone does not control for variation in R2 values. For example, it was particularly difficult to obtain measurements repeatedly in the house sparrow, even after training, compared to the eastern phoebe and eastern bluebird (Figure 4, Figure 5). The latter two birds were both hatch-year individuals. Thus, their age class might influence the evenness of their insulation (although, that is speculation without further study), but there is no reason to expect that placement of their skins for the measurement should be any easier to repeat than that of the house sparrow. Thus, some incompletely understood quality of feather coats in the house sparrow may demand further investigation. Similarly, variation in slopes of the lines in Figure 5 suggests that measuring thermal performance across temperature ranges (e.g., the thermoneutral zone of a species) may be more biologically informative than using a single reference heat level.
Figure 1: Diagram of the complete thermal camera and hot water bath set-up. Please click here to view a larger version of this figure.
Figure 2: Thermal images representing a method for replicating the same placement over multiple trials. The ellipse has already been placed on the heated area on the acrylic glass. These images show movement of the skin and not the ellipse. Gently and briefly touch a fingertip to the target measurement area on the feather coat. The fingertip will leave a heat mark on the skin for a few seconds. (A) Shown is the heat mark outside the ellipse, meaning that the target measurement area is not exposed to the heat. Being careful not to move the foam or acrylic glass (this would cause the ellipse to inaccurately represent the area of heat exposure), adjust the placement of the flat skin and touch the target measurement area again. Continue this process until (B) the heat mark is contained within the ellipse. Please click here to view a larger version of this figure.
Figure 3: Relationship between temperature of the hot water bath (display reading) and temperature at the acrylic glass surface (e.g., the actual heat source the flat skin is exposed to). It should be noted that the temperature at the surface of the acrylic glass is consistently slightly higher than the temperature displayed by the hot water bath. Use this figure only to gain an understanding of this relationship, and always measure the temperature at the surface of the acrylic glass for each trial (section 2.6). Please click here to view a larger version of this figure.
Figure 4: Improvement in the repeatability of temperature measurements at the surface of a feather coat in a single bird. These values were obtained from an individual house sparrow (A) before and (B) after repeatability training by the investigator for measurement performance. Please click here to view a larger version of this figure.
Figure 5: Relationship between temperature at the acrylic glass surface and temperature at the feather surface in single specimens of five bird species. Points on any single graph represent repeated measurements in the same individual at six different target temperatures. It should be noted that although measurements at the reference heating point of 40 °C are similar, the slope of these lines varies. This suggests that the thermal performance of feather coats in these birds differs (with a slope of 0 being a perfect insulator and slope of 1 being completely non-insulative). It should also be noted that measurement repeatability varies. Even after measurement training of the investigator, variance in repeated measures is highest for the house sparrow and lowest for eastern phoebe and eastern bluebird. Please click here to view a larger version of this figure.
This paper provides a protocol for repeatable, standardized thermal imaging measurements of avian flat skin specimens. This method makes it possible to compare thermal performance of the feather coat among species, within species, between comparable individuals, and at different locations on the bodies of individuals, all without destruction of the specimen.
The availability of necessary materials and equipment may be a limitation of this method. Although thermal cameras are rapidly becoming more accessible and affordable, research-grade thermal cameras still cost tens of thousands of dollars37. However, thermal cameras can be used for many practical applications in biology. McCaffery advocates for the use of thermal cameras to investigate ecological questions28. Thermal cameras are especially useful for collecting data on free-living organisms in the field because they are long-distance and non-invasive tools. The method presented here makes possible the integration of field and laboratory studies with measurements in the same units, as performed by the same equipment.
The use of thermal camera software other than that used here may necessitate modifications to this protocol, but such changes will only affect the set-up stage (section 1). Studies of smaller birds or certain questions about larger species of birds may require differently sized holes in the foam layer.
Similarly, the temperature of the hot water bath (step 2.1) may need modification for some species with higher or lower core temperatures if the goal is to measure temperatures with direct biological relevance to the experimental question. In general, water bath temperature standardization at 40 °C across studies will facilitate comparative analysis of the relative thermal performance of different kinds of intact plumages and feather structures. If precise measurement of the energy flux across skin and feathers is required, the guarded hot plate method11,12,13,28 is likely a better approach, since it 1) eliminates air between the heat source and skin and 2) measures temperature at the inner surface of the skin directly. However, while this method does not measure or calculate energy transfer directly, it is designed to facilitate rapid and repeatable measurements of whole specimens. Finally, the results demonstrate ample precision in detecting patterns of variation in plumage thermal performance.
This method uses flat skins, which are not currently widely available in most museum collections. Round skins, which are abundantly available in most natural history collections, could be used with this method if demounted, softened, flattened, and redried. However, curators are unlikely to approve such remounting in most cases. To increase the resources for comparative studies of the thermal values of bird feathers, we advocate for widespread adoption of flat skinning in as many species as possible. Added benefits of flat skinning are that flat skins do not require the partial destruction of the skeleton and musculature of a specimen that round skinning does, and higher numbers of flat skins can be stored in the same space that round skins require.
In skins of any particular species, it is essential to develop a technique for accurately placing the skin in the same spot over the heat hole every time. The results obtained here suggest that the technique (as described in step 2.7) minimizes measurement error more rapidly and effectively than practice in skin placement alone. However, it is plausible that especially dense plumages (e.g., penguins11) may not lose sufficient heat through the feathers to make it possible to visualize heat holes through the skin and feathers on the thermal image.
Because of the presence of pterylae in most bird species, the arrangement of the feathers over the skin on a specimen will affect the pattern of heat transfer across the feather coat. Therefore, it is important that the feathers are positioned in as close to their natural position in a live bird as possible. Preening the feathers into a neat, natural position is the last step in the protocol for flat skinning a specimen29. Thus, if specimens are prepared properly, feather placement should be appropriate to species across specimens. The amount of ptiloerection of the feathers will also affect thermal performance of the feather coat by trapping insulating air in the feather coat. In contrast, in flat skinned specimens, the feathers lay flat on the skin29, so ptiloerection should effectively be comparable across all specimens.
Although this study focuses on birds, this method may be equally useful for mammal skins. Boonstra et al. asserted that bird feathers are more insulative than mammal fur, but this study was a qualitative assessment based on visual analysis of thermal videos39 rather than a quantitative measure of the heat escaping from comparable body areas. It is believed that the method described here will contribute to an expansion of comparative thermal research and yield great insight into the evolution and ecology of thermoregulatory structures such as feathers40.
The authors have nothing to disclose.
This research was funded in part by a University of Connecticut Research Advisory Council Faculty Large Grant to M. Rubega. K. Burgio was supported on National Science Foundation NRT- IGE grant #1545458 to M. Rubega. The manuscript was significantly improved by the thoughtful feedback of two anonymous reviewers.
Aluminum Foil | Reynolds Wrap | 109000831 | 30 square ft.; this exact model need not be used. |
Foam Core Board | Foamular | 20WE | 1 in. x 4 ft. x 8 ft; this exact model need no be used. |
General Purpose Water Bath | PolyScience | WB02 | Ambiet +5 °C to 100 °C; ±.01 °C |
PDF Data logger | Elitech | RC-51H | Built in temperature and humidity sensor |
Plexiglass | AdirOffice | 1212-3-C | Acrylic glass; 12 in. x 12 in. x 1/8 in.; this exact model need not be used. |
Thermal Image Analysis Software | FLIR | ResearchIR Max v4.40.7.26 (64-bit) | Allows collection of precise, quantitative thermal data |
Thermal Imaging Camera | FLIR | SC655 | 680×480-pixel resolution, ±2 °C or ±2% accuracy, 40 cm minimum focusing distance |
Tripod | The Audubon Shop | The Birder Tripod with Manfrotto 700RC2 Rapid Release Head | 65" maximum height; this exact model need not be used. |