Systematically identifying peer-reviewed studies to include

This systematic review followed the workflow proposed by the Preferred Reporting Items for Systematic Reviews and Meta-Analyses 2020 statement (Page et al. 2021). A literature search for peer-reviewed scientific articles on Pb exposure in wild mammals was conducted using the database Scopus (Elsevier B.V, Amsterdam, Netherlands) on 21 June 2022. Scopus was chosen over other well-known databases and online search engines, such as Web of Science, PubMed, or Google Scholar due to the ability of Scopus to allow the chemical lead (Pb) as a search term, an option not provided by other databases or online search engines. In addition, Scopus permitted the exclusion of articles containing the word “lead” when used as a verb in the absence of any occurrence of lead as a chemical in the source. Furthermore, Scopus only indexed peer-reviewed articles and did not have restrictions on the length of search queries.

In Scopus, different query combinations of the following searches were used: CASREGNUMBER (7439-92-1)/CHEMNAME (lead); wild/wildlife/ free-living/“free living”/free-ranging/“free ranging”/semi-wild/“semi wild”/free-roaming/“free roaming”/street/feral; captive/zoo/domestic/domesticated/ semi-captive/“semi captive”/semi-domestic/“semi domestic”/semi-domesticated/“semi domesticated”; exposure/expose/exposed/exposing/poisoning/poison/ poisoned; keywords for mammals, and scientific nomenclature of all mammalian orders, families and genera taken from the International Union for the Conservation of Nature (IUCN) red list of threatened species (IUCN 2021; see   Supplementary Material (JWD-D-23-00055R1_Hydeskov_SUPPL_Final.docx)).

The first author conducted the assessment of all records and screening of studies. After identifying records via Scopus, records were checked for duplicates. Unique records were then screened by reading the title and abstract. The selected studies were thoroughly screened by reading the entire article. Articles on Pb exposure in wild mammals that were cited in the reference sections of the Scopus articles passing the full screening were also screened to identify additional publications not found by the original literature searches (i.e., snowball search).

Inclusion criteria included the requirement that studies must be peer-reviewed original research studies reporting Pb concentrations in tissues or body fluids in a nondomestic mammalian (called “wild mammals” in this review) species. Studies written in any language were included, and language was not used as an exclusion criterion. Studies reporting Pb concentrations in captive wild mammals outside laboratory settings were included to illustrate the variety of wild mammalian species documented to be susceptible to Pb exposure from their environment. Exclusion criteria included articles not reporting Pb concentrations in wild mammalian tissues or blood, studies using purely experimental settings, and studies reporting Pb concentrations in free-ranging domestic mammalian species. Data from studies were excluded if the study reported Pb concentration in tissues or body fluids knowingly contaminated with Pb-based ammunition (i.e., from killing the animal). Some studies reported nonstandard units (i.e., liquid concentrations for tissues or units unknown to the authors of this study). In these cases or when there was a mismatch between the data reported in text and tables, the corresponding author was contacted via the email provided in the publication and given a maximum of 30 d to respond. Without a response, either the data in question or the entire study was excluded.

Data extracted from the studies

For every study included, the following information was extracted, where available: Taxonomy (order, family, common species name, and scientific species name); continent and country where the study was conducted; description of known pollution sources nearby the study site; type of living condition (free-ranging, captive, or semicaptive); year of sample collection; season; age; sex; tissue; sample size, Pb analysis method; reporting in wet weight or dry weight concentrations (when both were reported, the dry weight concentration was preferred); and Pb concentration arithmetic mean (geometric means and weighted arithmetic means were excluded), SD, SE, median, and minimum and maximum values. The Pb concentrations were converted to micrograms per kilogram or micrograms per liter, where applicable, for comparison. Whether the study had investigated and identified any health effects of Pb was also recorded. To investigate in which part of the mammalian food web Pb exposure is most frequently reported, the feeding type of the mammalian species (i.e., herbivore, omnivore, or carnivore) and whether or not the species scavenges on animal material was determined by following species descriptions available in a recently published mammalian textbook (Wilson and Mittermeier 2009–19). Articles from Russia were allocated to the European or Asia continent based on the geographic location of the sample collection. To investigate potential economic resource biases in the publications, the Organisation for Economic Co-operation and Development's designation of developed and developing countries was used (Organisation for Economic Co-operation and Development 2022).

Different parts of a tissue or body fluid (e.g., whole kidney versus kidney cortex or whole blood versus plasma) were considered separate tissues for the purpose of the review. In addition, cortical bone and antler were differentiated, as cortical bone is permanent bone, whereas antler is deciduous bone (representing only short-term or annual exposure).

Characteristics of the studies

A total of 254 records were retrieved from Scopus (Fig. 1), with no duplicates identified. After the initial screening of the title and abstract, 125 studies remained for full screening, resulting in 69 studies from the Scopus search queries meeting the inclusion criteria. In addition, 114 studies were identified for inclusion through a snowball search, giving a total of 183 studies for systematic review (see   Supplementary Material (JWD-D-23-00055R1_Hydeskov_SUPPL_Final.docx)).

Figure1

Modified Preferred Reporting Items for Systematic Reviews and Meta-Analyses 2020 flow diagram (Page et al. 2021) for systematic reviews, indicating the number of studies that were included and excluded in the screening process for studies on lead (Pb) exposure in wild mammals.

The studies were published over 62 yr (1961 to June 2022), with most of the studies (158/183, 86%) published in the second half of this period (1992 to 2022; Fig. 2). The majority of studies (162/183, 89%) were conducted in Europe (108 studies, 59%) and North America (54 studies, 30%); some studies were conducted in multiple countries (but always within the same continent). The US was the country in which most studies were conducted (38 studies), followed by Canada (17 studies), Poland (16 studies), Germany (12 studies), and the UK (11 studies). Only 6% (11/183) of the studies were conducted in developing countries (Fig. 3).

Figure2

The number of studies published per year on lead (Pb) exposure in wild mammals included in this systematic review (n=183). Studies were published in the period 1961 to June 2022.

Figure3

The number of studies (n=183) conducted on lead (Pb) exposure in wild mammals between 1961 and June 2022 per country. Studies were conducted in 35 countries across six continents. Darker color indicates a higher number of studies conducted. Created using Datawrapper (Datawrapper GmbH 2022).

Pb exposure

Pb was present in all mammalian species studied, with concentrations reported in a total of 153 wild mammalian species belonging to 45 families and 11 orders. The most studied order, family, and species were Cetartiodactyla, Cervidae, and red deer (Cervus elaphus), with 67, 46, and 20 studies, respectively (Table 1). In terms of feeding type, herbivores, omnivores, and carnivores were researched in 92, 79, and 49 studies, respectively; 49 studies researched scavenging species.

Table 1

Top 5 most studied mammalian orders, families and species by the number of studies published on lead (Pb) exposure in wild mammals between 1961 and June 2022.

A total of 33 different tissues and body fluids were tested. The five most studied tissues were the liver (123 studies), kidney (92 studies), skeletal muscle (54 studies), bone (32 studies), and blood (25 studies). At 1,506,000 lg/kg dry weight, the highest Pb concentration reported across all 11 mammalian orders studied was recorded in the kidney of a Northern short-tailed shrew (Blarina brevicauda; Table 2).

Table 2

Highest individual (where n=1) or mean lead (Pb) concentration reported in each wild mammalian order studied in studies published between 1961 and June 2022. Solid tissue concentrations are reported in either wet weight (ww) or dry weight (dw).

Health effects of Pb

A total of 45 of 183 (25%) studies investigated the health effects of Pb. Of these, 28 (62%) studies identified a negative health effect in 57 species. No studies identified a positive health effect. Negative health effects were reported in 32 herbivores, 21 omnivores, and four carnivores and ranged in severity from “interference with developing permanent teeth” to “fatality” (Table 3).

Table 3

Negative health effects associated with lead (Pb) exposure in wild mammals by organ system affected in studies published between 1961 and June 2022.

Variation in methodology

The 183 studies used a large variety of sampling approaches and Pb analysis techniques and reported the results in different ways, making it hard to compare findings. For example, some studies only reported mean values, while others reported mean, SD or SE (or both), median, and range of Pb concentration. Full details on sex and age of the animals tested and on whether the sampling was undertaken in an area with known Pb pollution, were not provided in 158 (86%), 134 (73%), and 101 (55%) studies, respectively.

Coverage of Pb exposure in wild mammals in the literature

The number of publications of Pb studies on wild mammalian species has increased over the last three decades. Nevertheless, the frequency of Pb publications on wild avian species still appears to be much higher, indicated by the large number of avian reviews published in recent years (e.g., Pain et al. 2019; Monclús et al. 2020; Ives et al. 2022).

The Pb concentrations were reported in 153 wild mammalian species, highlighting that this class of animals is exposed to Pb on a large scale. Despite this clear evidence of Pb exposure across many taxa, based on IUCN taxonomic classification, only 41% (11/27) of orders, 28% (45/162) of families, and 3% (153/ 5,968) of species were studied, which evidences a large group of mammals with no information on Pb exposure (IUCN 2021). Understandably, there has been a tendency to study species that are easiest to acquire (i.e., hunted species and small species that are easy to trap without chemical immobilization). Consequently, the studies identified in this review are skewed toward herbivores and omnivores. It is not possible to determine whether tissues were selected due to easy access or because the researcher hypothesized that the selected tissue would provide the most information in the study species. However, liver, kidney, skeletal muscle, bone, and blood are commonly collected and stored for further investigations during wildlife necropsies (McAloose et al. 2018).

Health effects of Pb in wild mammals

Only 28 studies, 62% of those in which health effects were investigated, identified negative health effects of Pb in wild mammals. As expected, no positive effects were found. From the breadth of health effects identified and organ systems involved, it is clear that Pb affects many parts of the body in wild mammals, as in other species (Pain et al. 2019; Bergdahl and Skerfving 2022). Many of the identified health effects would most likely be subclinical, so it is not possible to determine how exactly these affect wild mammalian populations.

It is expected that due to feeding behaviors, some mammal species are more likely than some bird species to be exposed to smaller amounts of Pb, relative to body mass, but over extended periods of time. For example, waterfowl are known to pick up large numbers of Pb-based shotgun pellets or fishing weights from the bottom of wetland areas when foraging (Pain et al. 2019), yet most wild mammals would not be exposed to such large amounts of Pb at once, as they forage using different methods (e.g., Carnivora species). This difference in feeding behavior would reduce the risk of acute poisonings, seen in many waterfowl and scavenging and predatory bird species, but may cause more chronic and often subclinical health effects (e.g., on the nervous and immune systems). Chronic effects tend to be much less apparent than acute effects in free-ranging wildlife so may be overlooked unless investigated explicitly. Ecke et al. (2017) concluded that sublethal Pb exposure in golden eagles (Aquila chrysaetos) might impair flight performance and thus increase the risk of mortality from other causes. Yet, other eagle studies could not find evidence that subclinical Pb concentrations are linked to “increased risk of trauma” (Isomursu et al. 2018) or “other causes of mortality” (Franson and Russell 2014). These examples demonstrate the difficulty in investigating subclinical and chronic health effects in wildlife, especially when studies do not classify effects similarly. Furthermore, ethical permits to conduct Pb toxicological effect studies on wild mammals and other wildlife are, rightly, unlikely to be granted. Therefore, subclinical effects are difficult to confirm in wildlife.

Although the impacts of Pb exposure on human, laboratory mammalian, and wild avian health are well-documented, there is still much to learn about the impacts on wild mammal health. In contrast to important infectious diseases in animals, the World Organisation for Animal Health does not have any reference laboratories for any noninfectious diseases such as Pb poisoning, which could guide or help standardize how the diagnostic steps should be conducted (World Organisation for Animal Health 2022). Potential strategies to research the health effects of Pb on wild mammals include behavioral studies, reproductive success analyses, and immune system function assessments; these are all challenging to conduct in free-ranging animals. Research that would be easier to achieve in free-ranging settings includes correlations between blood Pb concentrations and hematologic and biochemical parameters, investigations of blood and tissue Pb concentrations to understand the distribution of Pb in the body, and investigations of tissue pathologies in Pb-exposed populations. Ideally, studies should use the most sensitive techniques available for measuring Pb, such as inductively coupled plasma mass spectrometry (Apostoli 2002) and report the results in dry weight for easier comparisons. Regardless of the approach taken by future studies to investigate Pb exposure and health effects in wild mammals, studies should report details of the study area, sex and age of animals, and analytic methods, as well as providing an appropriate range of descriptive statistics (e.g., sample sizes, SD or SE), which would facilitate future evidence-based policies.

Methodologies, kinetics, and thresholds

Our systematic review found a lack of consistency in estimating Pb concentrations, health effects, and how this is reported. The different Pb analytic methods used in the literature reviewed (e.g., atomic absorption spectroscopy and inductively coupled plasma mass spectrometry) vary in sensitivity and differ in susceptibility to matrix effects (Apostoli 2002). Therefore, some studies might have failed to detect Pb, if present in low concentrations. In addition, some studies reported tissue concentrations by dry weight and others by wet weight, impeding direct comparison of such concentrations. Furthermore, Pb uptake and storage distribution potentially may differ between species: As there are no studies available on Pb kinetics in wild mammals, this remains unknown. Variations in methodologies and target species make it difficult to compare Pb concentrations between studies and between species. Consequently, it is not possible to establish toxicity thresholds for wild mammals based on the current body of literature. However, although thresholds may be a valuable tool for some professions (e.g., veterinarians in clinical practice), a pragmatic reason for not attempting to establish threshold values for different wild mammalian tissues or species is the risk of creating the false impression of acceptable or safe levels of Pb exposure in wild mammals. Therefore, we recommend ceasing the use of threshold values for Pb concentrations in wildlife research, as already suggested by Pain et al. (2019).

Designing future studies on Pb exposure and health effects

From a One Health perspective (FAO et al. 2022), human, animal and ecosystem health are interlinked. Although it would be beneficial to study Pb concentrations in species at differing trophic levels in the wild, the cost of periodic screening programs means that these are likely to be restricted to only a few selected species. Because Pb bioaccumulates in both plants and animals (Clemens 2006; Radomyski et al. 2018), testing for Pb in carnivorous and scavenging mammalian species might not only aid monitoring of environmental Pb exposure but also indicate the potential exposure of humans to Pb, as species at these higher trophic levels often have long life spans similar to humans (O'Brien et al. 1993). Using higher trophic-level species as sentinels is already being used in avian Pb research (e.g., Monclús et al. 2020) and in mammalian research on infectious diseases and antimicrobial resistance (e.g., Millán et al. 2014; Sacristán et al. 2020). Another benefit of studying top predatory or scavenging mammals, which are often iconic and charismatic (Albert et al. 2018), is that these mammals might attract more attention from stakeholders and policymakers involved in the regulation of Pb, a phenomenon already seen in conservation, with charismatic species receiving the most attention (Sitas et al. 2009).

To assess or monitor short-term and recent Pb exposure using reduced sampling effort and noninvasive sampling, antlers or hairs might be suitable tissues to consider in the future. Antlers represent Pb exposure of about 6 mo, from spring to autumn, and have previously been used to assess Pb contamination (Ludolphy et al. 2022). Antlers do, however, introduce potential biases into a Pb monitoring program. For most cervid species, antlers are only available from adult and subadult males; thus, detecting differences between age classes and sexes would not be possible using this tissue. Hair is often easily available even from museum specimens (i.e., on pelts), and it can be collected from free-ranging mammals noninvasively using hair traps. The period of Pb exposure can be estimated using laser ablation techniques, provided that potential Pb surface contamination is removed, the particular species' hair growth cycle is known, and the sampling is carried out in a consistent manner (Sela et al. 2007).

To gain information on where Pb is stored in a particular wild mammalian species, researchers would have to analyze multiple tissues and blood, all obtained from the same individual at the same time (Krone 2018; Bergdahl and Skerfving 2022). However, obtaining this quantity of samples would likely only be possible in freshly dead or mildly autolyzed individuals (McAloose et al. 2018). Also, extra expenses from analyzing multiple samples from each individual could be cost prohibitive. Therefore, more controlled studies on the storage and clinical pathology of Pb in captive individuals could help understand the health effects of Pb in wild mammals.

With studies written in any language being included, this review highlights that few studies on Pb exposure in wild mammals have been conducted in developing countries. However, note that detection of studies, in general, always depends on how journals have indexed articles in various databases; therefore, it cannot be ruled out that studies written in other languages than English will be missed. Moreover, it is important to consider that fewer studies published in developing countries does not mean the problem is less prevalent in these countries. Indeed, the lack of studies could be explained by lower access to research funding (Kpokiri et al. 2022). On the contrary, environmental exposure to Pb might be greater in developing countries due to fewer legislative controls on Pb use in products, as seen with Pb-based paints (WHO 2022a). Indeed, children in developing countries are more exposed to Pb than children in developed countries (Hwang et al. 2019; Bergdahl and Skerfving 2022). Therefore, more research on Pb exposure in wild mammals must be conducted and published in these countries to understand the levels of environmental exposure to Pb, the risk to human and animal health, and to study whether legislation on Pb in products affects the Pb present in the environment.understand the levels of environmental exposure to Pb, the risk to human and animal health, and to study whether legislation on Pb in products affects the Pb present in the environment.

As all organisms are exposed to Pb, studies helping to determine the impacts of Pb on different species will add to current knowledge of this toxic metal. Collectively, this increase in knowledge will create stronger evidence to influence policymakers involved in the regulation of Pb in products that continue to act as sources of environmental contamination and be a major One Health issue. Overall, Pb exposure in wild mammalian species must be recognized as an issue, as this systematic review demonstrates. Control and regulation of Pb need to take a One Health approach and not focus on single taxa groups when developing policies.