Measuring health and frailty in wild insects

Grantee: Jelle Boonekamp

Institutions: University of Glasgow, United Kingdom

Grant amount: $63,536

Grant type: Ad hoc

Focal species: Field cricket (Gryllinae)

Conservation status: Least concern

Disciplines: Physiology, entomology

Research location: United Kingdom

---

Project summary

From an evolutionary perspective, health and well-being is best expressed in terms of fitness variation, the logic being that animals who are optimally adapted to their local environment should be healthy. Hence, health and fitness are intimately entwined on a conceptual level. Morphological, behavioral, and physiological traits that are predictive of fitness could provide potential biomarkers of health, and such approaches have been developed for many different wild vertebrate species. However, due to their small size and mobility, it has proven exceedingly challenging to follow individual insects longitudinally in the wild, let alone their descendants, to measure individual performance and fitness. Consequently, there is a paucity of literature on wild insect health and well-being.

The aim of this project is to address this knowledge gap by developing a non-invasive frailty index that measures the health of individual insects in their natural habitat and that can be applied across species. Analogizing to frailty indices used in human patients, this project will develop an insect frailty index using relevant traits applicable to many different insect species. In humans, the frailty index is highly predictive of morbidity and remaining life expectancy. Similarly, this project seeks to validate the insect frailty index against mortality and fitness data in an insect population by testing whether “frail” individuals have increased mortality risk and reduced fitness. 

This project is made possible by the work that we have done to establish a long-term field study of a natural population of crickets (WildCrickets.org). Using a network of 140 video cameras, all the adults in the population are longitudinally monitored from their emergence in early spring to their natural death in late summer. All their movements, reproductive behaviors, fights, and predation events are recorded, and fitness is measured by genotyping and counting the number of surviving genetic descendants in the next year.

Why we funded this project

Willingness to investigate the welfare of insects is somewhat rare, as are creative ways to assess their welfare. This project both proposed a potentially usable metric, and has access to an unusually useful resource through the WildCrickets.org project. While any particular approach to assessing welfare in insects is unlikely to work, we are hoping to seed enough approaches and strategies that others take up the call, eventually drawing in enough different approaches to produce usable strategies for assessing insect welfare in the wild. Finally, because many of the project resources were already acquired through other sources, we were to fund exclusively the welfare-focused aspects of the project.

---

Photos

---

Integrating nonlethal field and lab assessments of wild fish welfare in the Colorado River in Grand Canyon

Grantee: Isaac Schuman

Institutions: Oregon State University, United States

Grant amount: $30,000

Grant type: Seed grants

Focal species: Flannelmouth sucker (Catostomus latipinnis), humpback chub (Gila cypha), rainbow trout (Oncorhynchus mykiss)

Conservation status: Least concern

Disciplines: Physiology, ichthyology, infectious disease

Research location: United States

---

Project summary

This project will monitor the welfare of three species of fish in the Grand Canyon using a battery of metrics including body condition, gut microbiome, parasite load, and cortisol in skin mucus. The researchers will attempt to validate these as welfare indicators by testing whether they correlate with one another within individual fish, and whether they follow consistent patterns across the three focal species. Finally, they will use long-term population monitoring data to investigate potential correlations between these individual-based welfare indicators and environmental and demographic characteristics of individual study sites, such as food availability and age structure. 

Why we funded this project

We appreciate the variety of potential welfare indicators that this study will measure, and that there will be an explicit attempt to test the validity of the indicators for these particular species. It is exciting to see such a comprehensive analysis of wild fish welfare. We are also interested in the project’s comparison of individual welfare indicators to population-level demographic parameters, as better understanding those relationships could help with both identifying welfare threats from more readily available population data, and with predicting indirect impacts of welfare interventions.

---

Are populations that are well-adapted to their environment less stressed than those that are not?

Grantee: Ryan S. Mohammed

Institutions: Auburn University, United States

Grant amount: $30,374

Grant type: Seed grants

Focal species: Guppy (Poecilia reticulata), jumping guabine (Anablepsoides hartii)

Conservation status: Least concern

Disciplines: Physiology, ichthyology

Research location: Trinidad and Tobago, United States

---

Project summary

In this project, wild-caught guppies (Poecilia reticulata) will be exposed to an apparent threat of predation by transferring water from a tank containing a common predator (pike cichlid) into their tank. The amount of water transferred from the predators’ tank will be varied to simulate a range of predator densities. Then, cortisol levels will be measured in both the tissue of guppies and the water they are kept in. These tissue and water cortisol measurements will be compared to develop a protocol for inferring the cortisol levels of fish based on non-invasive water measurements. The researchers hypothesize that the strength of the guppies’ physiological stress response (cortisol) will vary with apparent predation risk, and they intend to eventually build on this pilot study by using the developed non-invasive protocol to compare guppies from populations the are adapted to varying intensities of predation. 

Why we funded this project

This project will develop a protocol for inferring the cortisol levels of fish based on non-invasive water measurements, which should allow researchers seeking to use cortisol as a metric of physiological stress to avoid needing to kill fish in order to measure the chemical in their tissues. In terms of broader wild animal welfare theory, we also appreciate the project’s focus on the indirect effects of predator-induced fear, which are likely ubiquitous. This project is also intended as a pilot for a larger project that would investigate the impact of evolutionary adaptation to avoid predation on the stress response to the presence of predators.

---

How do human activities impair the welfare of highly social fish?

Grantee: Joachim Frommen

Institutions: Manchester Metropolitan University, United Kingdom

Grant amount: $28,960

Grant type: Seed grants

Focal species: The princess of Zambia (Neolamprologus pulcher)

Conservation status: Least concern

Disciplines: Ichthyology, physiology, genetics/genomics

Research location: United Kingdom, Zambia

---

Project summary

This project will investigate the welfare of a common fish species in Lake Tanganyika, the princess cichlid (Neolamprologus pulcher), as they are exposed to varying levels of human activity. Across eight populations representing a range of distances to human settlements and shipping routes, the researchers will monitor behaviors indicative of stress or aggression, and measure body condition and the brain tissue expression of five genes involved in stress physiology (glucocorticoid response pathway; crf, cyp11b, gr1, gr2, mr). These welfare indicators will be compared with specific environmental characteristics, including boat noise, water visibility (sedimentary and algal load), human fishing intensity, temperature stress, and structural complexity of the local environment.

Why we funded this project

By focusing on an established model system (cichlids), this project is able to benefit from background knowledge of the species’ ecology and behavior and proceed to more neglected welfare questions, as well as potentially engaging a ready audience of cichlid researchers. An especially interesting component of this project is its investigation of brain gene expression to potentially better understand how stress physiology relates to an animal’s subjective experience.

---

Determination of Fecal Tri-iodothyronine and Cortisol as Physiological Proxies of Animal Welfare

Grantees: Michael Cherry, Joe Hediger

Institutions: Caesar Kleberg Wildlife Research Institute, Texas A&M University, University of Massachusetts Dartmouth, United States

Grant amount: $25,860

Grant type: Small grants

Focal species: White-tailed deer (Odocoileus virginianus)

Conservation status: Least concern

Disciplines: Physiology, animal welfare science

Research location: United States

---

Project summary

Wildlife are susceptible to both direct and indirect effects of thermal stress imposed by a warming climate. Direct effects include increased energetic costs to maintain a healthy body temperature and immune system impairment, while indirect effects include changes in food availability and increases in disease transmission. White-tailed deer (Odocoileus virginianus) in southern Texas are on the front lines of this environmental challenge. This project aims to assess the reliability of fecal tri-iodothyronine (T3) and fecal glucocorticoids (FGC) as non-invasive physiological metrics for monitoring the health of white-tailed deer. The use of T3 as an indicator of wildlife health and welfare is relatively novel, and the researchers will attempt to refine the use of these metrics through controlled experiments, correlating the T3 measurements with a more widely used indicator in FGC. Both physiological indicators will also be validated against behavioral observations of the same deer that are thought to reflect their emotional state.

Why we funded this project

The study findings will help in understanding how wild animals cope with increasing temperatures and the impact of thermal stress on their welfare and health. Notably, previous work has suggested that T3 measurements in ungulates are especially sensitive to thermal stress, and so comparing T3 with other indicators based on different physiological pathways, such as glucocorticoids, could help researchers to diagnose the relative significance of different environmental stressors an animal is facing. The project’s behavioral metrics are also crucial for realizing that potential. A secondary reason for our interest in this project is that it has near-term policy implications, potentially highlighting the value of preserving or promoting specific landscape features for the ecosystem service they offer, in the form of shade, to wild ungulates.

---

Photos

---

Validation and efficacy of faecal glucocorticoid metabolites as indicators of animal welfare

Grantee: Miriam Zemanova

Institution: University of Fribourg, Animalfree Research, Switzerland, Oxford Centre for Animal Ethics, United Kingdom

Grant amount: $28,500

Grant type: Small grants

Disciplines: Physiology, animal welfare science

Research location: Switzerland, United Kingdom

---

Project summary

One of the central components of the stress response is the production of glucocorticoids (GC). The measurement of glucocorticoid levels in blood serum is therefore often used in animal welfare studies. However, it is not always apparent how valuable these measurements are for understanding stress reactions and their relationship to animal welfare. Moreover, blood sampling causes discomfort and is impossible without restraint or immobilization, which can be harmful to the animal and distort the experimental results. Fecal glucocorticoid metabolites (FGM) may be a suitable non-invasive alternative to blood analysis. While there have been several reviews written on fecal glucocorticoids, no systematic review of their validity as an animal welfare indicator is currently available. Therefore, through a systematic review process, this project seeks to provide an evidence base for the efficacy of non-invasive measurement of stress levels in wild animals using fecal glucocorticoids. Evidence will be compiled from studies that have assessed the correlation between fecal glucocorticoids and at least one other credible animal welfare indicator or factor that may be assumed to lead to impaired welfare. The project also aims to compare the performances of blood and fecal samples to assess stress levels. 

Why we funded this project

This study will review the evidence base for the validity and efficacy of non-invasively obtained glucocorticoid measurements and identify both challenges and best practices for working with fecal samples across multiple animal taxa. This is important because WAI is supporting multiple projects utilizing FGM as a physiological welfare indicator. Invasive measurements of stress, such as blood-based GCs, are disadvantageous because, without proper strategy and training, the stress induced by the sampling procedure can influence the measurement. Therefore, non-invasive measurement techniques are not only better for the animals, but may make the science less expensive and more reliable. Another reason we are supporting this study is that it will help to address a general need for better validation of the link between glucocorticoids and animal welfare, which is related to but conceptually distinct from physiological stress.

---

Photos

---

Characterization of fecal oxytocin and immunoglobulin A in lions (Panthera leo): Exploring a multi-biomarker approach in animal welfare research

Grantee: Paula Serres Corral

Institution: Universitat Autònoma de Barcelona, Spain

Grant amount: $25,385

Grant type: Small grants

Focal species: Lion (Panthera leo)

Conservation status: Vulnerable

Disciplines: Physiology, animal welfare science

Research location: Spain

---

Project summary

Glucocorticoids (GCs) remain the most common physiological indicators of stress and, by extension, of animal welfare. However, welfare means more than stress, and therefore there is a need to develop complementary biomarkers to expand our knowledge of animals’ overall welfare states, both positive and negative. This project will consider oxytocin (OT) and secretory immunoglobulin A (IgA), markers of neuroendocrine and immune function respectively, which have been proposed as potential indicators of positive affective states. So far, both indicators have been reliably measured in the saliva and urine of a limited set of wild mammalian species. However, previous work on their analysis in fecal samples has been even more limited. This project aims to validate the measurement of OT and IgA in feces of lions (Panthera leo) as a model for social carnivores and evaluate their applicability as additional physiological indicators in wildlife welfare studies. The researchers will evaluate baseline levels for the species based on a small number of captive individuals, and assess how these biomarkers are correlated with GCs and behavior.

Why we funded this project

With a multi-biomarker approach, these potential biomarkers, in combination with GCs, will enable a more robust interpretation of findings in welfare assessments. We are especially interested in the potential for these physiological indicators to support identification of positive welfare states in wild animals, considering the crucial role of oxytocin, for example, in social bonding. Although the project itself is limited to a small number of captive individuals, we expect that this project will represent a significant step towards validating these putative indicators of positive welfare thanks to the detailed behavioral monitoring that the captive environment allows, including a Quantitative Behavioral Assessment (QBA) approach.

---

Photos

---

Thermal imaging as a non-invasive welfare assessment tool for tracking the impact of environmental stressors across wild animal populations

Grantee: Ross MacLeod

Institution: Liverpool John Moores University, United Kingdom

Grant amount: $29,810

Grant type: Small grants

Focal species: Wild birds

Conservation status: Least concern

Disciplines: Physiology, animal welfare science, population ecology, ornithology

Research location: United Kingdom

---

Project summary

This project aims to test and validate a standardized multi-species approach to monitoring physiological stress in wild birds by using thermal imaging cameras to measure the animals’ body surface temperature, which could enable tracking of chronic stress in wild populations facing different levels of environmental stressors. Building on pilot work, the project will focus on wild bird populations to develop a standardized thermal imaging methodology capable of monitoring surface temperature of a wide range of free-living wild animals. The methodology will be validated using bird communities visiting 54 standardized feeding and drinking stations spread across three urban-rural gradients in the UK, to quantify how changes in surface temperature are linked to starvation risk, predation risk, and human disturbance.

Why we funded this project

This approach to measuring wild animals’ physiological stress levels through thermal imaging analysis has great potential for scalable monitoring of large numbers of individuals and is usable for cross-species comparisons. We appreciated that this team was interested in addressing non-anthropogenic causes of suffering in starvation and predation, and in extending their method to other species that tend to be neglected (e.g., wild rodents). This work also relates to the project by Paul Jerem that we previously funded, creating a longer-term relationship between Wild Animal Initiative and project participants.

Find Ross’ other project, studying house sparrows, here.

---

Photos

---

Developing loop-mediated isothermal amplification (LAMP) assays for detecting pathogens in wild animal populations

Grantee: Cameron Semper

Institution: University of Calgary, University of Lethbridge, Canada

Grant amount: $30,000

Grant type: Small grants

Focal species: Deer mice (Peromyscus sp.)

Conservation status: Least concern

Disciplines: Infectious disease, physiology, animal welfare science

Research location: Canada

---

Project summary

Loop-mediated isothermal amplification (LAMP) is a low-cost technique that amplifies specific DNA to levels that can enable visual detection. LAMP has been extensively applied as a point-of-care diagnostic tool for human health, but its application in wild animal populations remains underexplored. This project aims to develop LAMP assays for rapid, in situ detection of representative viral and bacterial pathogens as well as parasitic worm infections in wild animals. This project will develop two methods for detecting viral and bacterial infections, respectively. The viral test will focus on viruses that have caused epidemics in wild frigatebird and sooty tern populations, while the bacterial test will focus on tick-borne illnesses and parasitic worms which infect deer mice. After developing the LAMP assays, the researchers will validate their potential contribution to monitoring wild animal welfare in a non-invasive manner by using them to test for pathogenic load in fecal samples from deer mice. 

Why we funded this project

This research will serve as a proof-of-concept for the applicability of LAMP for monitoring infectious disease, a key determinant of wild animal welfare. Additionally, the project is specifically targeting diseases that affect a large number of animals. LAMP is simple to perform, and results can be interpreted visually without the need for sophisticated equipment, reducing the cost in time and materials for assessing disease in wild animals. Because it can be carried out in the field, follow-up treatment or further study can be immediately given to the same animal.

---

Photos

---

Raman spectroscopy as a novel non-invasive technique to assess wildlife welfare

Grantee: Teresa Romero

Institution: University of Lincoln, University of Portsmouth, United Kingdom

Grant amount: $19,100

Grant type: Small grants

Focal species: Tufted capuchin monkey (Cebus apella)

Conservation status: Critically endangered

Disciplines: Physiology, animal welfare science

Research location: United Kingdom

---

Project summary

Steroid hormones are routinely used as biomarkers of stress and can be measured in different biological matrices, such as serum, saliva, feces, and urine. However, commonly used collection methods in wildlife are challenging; the stress they induce in the animals may affect hormone levels and also presents ethical issues. More recently, analysis of hormones accumulated in the hair has emerged as a non-invasive tool for measuring chronic stress in wildlife, but current limitations of hair analyses include variation in cortisol concentrations depending on sample preparation and the amount of hair required for cortisol extraction. This project will test whether Raman spectroscopy, which is suitable for the field and does not require sample pre-treatment, is a better way to use hair samples to assess steroid hormones. The planned methodology will make use of samples of mammalian hair with a known cortisol content using a control technique — liquid chromatography mass spectrometry — for comparison. The samples will be subjected to analysis by Raman to validate the technique and establish a robust analytical methodology for the non-invasive analysis of welfare biomarkers in wildlife.

Why we funded this project

Although this project is focused on development of chemical methods, we expect that the method, if validated, could make glucocorticoid assessment in the hair of wild animals much easier, increasing the future quantity and quality of that type of data. Hair is an especially interesting medium for glucocorticoid analysis because it integrates glucocorticoid levels in the body over time, causing the measurements to be more stable over time and potentially more indicative of baseline welfare, since the values are less sensitive to an animal’s most recent activities.

---

Photos

---